UNITED STATES OF AMERICA

P-R-O-C-E-E-D-I-N-G-S

(8:37 a.m.)

DR. FREAS: Good morning. Before we officially start the meeting, I would like to go around and introduce to the members of the public those committee members that are seated at the head table.

Starting on the right-hand side of the room, we have Dr. David Markovitz, Professor, Division of Infectious Diseases, University of Michigan Medical Center. Sitting next to him is Dr. Walter Dowdle, Senior Public Health Consultant, The Task Force for Child Survival and Development. Next we have Dr. Judith Goldberg, Director, Division of Biostatistics, New York University School of Medicine.

Next we have Dr. Ruth Karron, Associate Professor, Johns Hopkins University School of Hygiene and Public Health. Next we have Dr. Walter Royal, Associate Professor of Medicine, Morehouse School of Medicine. Next we have Dr. Monica Farley, Professor of Medicine, Emory University School of Medicine.

Next we have Pamela McInnes, Deputy Director, Division of Microbiology and Infectious Diseases, NIH. Next we have Ms. Cindy Lyn Province, Associate Director, Bioethics Center of St. Louis. Next we have Dr. Bruce Gellin, Director, National Vaccine Program.

Next we have ‑‑ joining with us this morning Dr. Stephen Phillips, Director, Deployment Medicine and Surveillance, Office of the Assistant Secretary of Defense. Next, to the Chairman of this committee, Dr. Gary Overturf, Professor, Pediatrics and Pathology, University of New Mexico School of Medicine.

Next we have Dr. Philip LaRussa, Professor of Clinical Pediatrics, Columbia Presbyterian Hospital. Next we have Dr. Martin Myers, Co-Director, Public Health Policy and Education, Sealy Center for Vaccine Development, University of Texas Medical Branch. Next we have Dr. Bonnie Word, Assistant Professor of Pediatrics, Baylor College of Medicine, Texas Children's Hospital.

Next we have Dr. Peter Palese, Chairman and Professor, Department of Microbiology, Mount Sinai School of Medicine. Next we have Dr. Arnold Monto, Professor at the University of Michigan. Next we have Dr. Ted Eickhoff, Professor of Medicine, University of Colorado Health Sciences Center.

Next is our non-voting industry representative, Dr. Michael Decker, Vice President, Scientific and Medical Affairs, Aventis Pasteur. Next we have Dr. Nancy Cox, Chief, Influenza Branch, Center for Disease Control and Prevention. And at the end of the table we have Dr. Reverend Levandowski ‑‑ Dr. Roland Levandowski ‑‑ excuse me ‑‑

(Laughter.)

‑‑ okay, from FDA.

I would like to welcome all of you here.

CHAIRMAN OVERTURF: I'd like to call the ‑‑ I'd like to welcome you back for the second day of this VRBPAC meeting.

At this time, we have an opportunity for an open public hearing, and I request that anybody who would like to make a statement to please make their way to a microphone. Please announce who you are and any groups that you may represent.

Okay. Since we have nobody who is approaching the microphone, I assume that there is nobody wanting to address the committee during the open public hearing. So we will proceed to the questions for this committee meeting, and that is the options for the strain selection for the 2004-2005 influenza vaccine, and I'll turn the meeting over to Dr. Levandowski.

DR. LEVANDOWSKI: Okay. Thank you.

Good morning, everybody. I'll just plunge right into this, and actually this should say options for Influenza A H1N1 and H1N2, or just Influenza A H1.

Just to summarize, I'll try to summarize information from yesterday as a reminder of what we were looking at for each of the strains, and then go through options that are possible for strain selection.

The H1N1 and H1N2 viruses ‑‑ of course there have been relatively few of these viruses isolated during this season. The isolates have come mainly from sporadic cases that have been in the Americas, Asia, and Europe, but there have been some outbreaks. I don't know if we emphasized that. There have been outbreaks of Influenza A H1N1 and H1N2 in Iceland and the Ukraine during this past season.

The HA on most of the H1 strains antigenically are similar to the current vaccine strain, which is A/New Caledonia/20/99, and the H1 virus is generally seen to be well inhibited by antisera from people who have been immunized with the current vaccines that contain A/New Caledonia/20/99.

In addition to that, we have high growth reassortants for A/New Caledonia/20/99 that are available or used in current manufacturing, and all of that is very well worked out at this time. So the first option for this ‑‑ for the H1 strain is to retain the New Caledonia/20/99 virus.

And in favor of that, as I mentioned, most of the recent H1 viruses are antigenically very close in nature to the hemagglutinins. They're very close in nature antigenically to the A/New Caledonia vaccine strain.

The current vaccines, of course, appear to be well matched to the HA of the current strains, and the manufacturing has worked out and the yield is very predictable. I'm not in favor of that. The con here is that there have been relatively few strains to look at for analysis.

The second option for this, of course, is to change to use a more recent H1N1 virus, and in favor of that there is the possibility that a more recent strain might provide a closer match with hemagglutinins and the neuraminidases of the contemporary strains.

But against that the new strain is not likely to be superior in terms of immunogenicity or efficacy compared to the current vaccine strain, and there aren't ‑‑ the manufacturing issues have not really been worked out for any strains at this point.

And the third option, of course, always is to defer the recommendation. In favor of that might be that there could be analysis of more contemporary strains if those were identified, and maybe that would make a closer match for the hemagglutinin and neuraminidase of next year's vaccine. But against that is the fact that it appears that there will be little new information that's forthcoming, since there are so few H1N1 viruses that are causing disease.

So moving on for the H3N2 viruses, by way of review, of course everyone knows that the Influenza A H3N2 viruses have predominated globally since September of 2003. The HA on most of these strains are antigenically distinguishable from the current vaccine strain, which is A/Panama/2007/99, and they are more closely related to the A/Fujian/411/2002 strains.

At this point, the influenza season appears to be declining in the northern hemisphere, and, of course, we also know that the majority of the strains that are ‑‑ that have been isolated are not that well inhibited by antisera produced against the current vaccines in people.

There are ‑‑ at this point, there are high growth reassortants of the ‑‑ of A/Fujian/411/2002-like strains, and that includes, as we heard yesterday, reassortants for A/Wyoming/3/2003 and A/Kumamoto/102/2002. And we know that those grow reasonably well.

So for H3N2, again, the first option would be to retain the A/Panama strain as the vaccine strain. And in favor of that, of course, the manufacturing is very well worked out, and it's very highly predictable. But against that we know that the HA of most of the H3N2 viruses are antigenically distinguishable from the current vaccine strain. And serological tests with current vaccines indicate that the majority of strains are not very well inhibited by antiserum from people who are immunized with the Panama strain.

And also, as we know, the H3N2 influenza viruses are often responsible for significant morbidity and mortality, and I think that was highlighted in Maria Zambon's talk.

So the second option for H3N2 is to choose a more recent H3N2 virus, and in favor of that a more recent strain could provide a closer match with the hemagglutinin and neuraminidase of contemporary strains. There are high growth reassortants for Fujian/411-like viruses available for manufacturing, and they appear to be reasonable in terms of their yield. And, again, reason to do this is because H3N2 viruses are often responsible for more significant morbidity and mortality.

Against this we don't really know that a new strain would actually provide superior immunogenicity or efficacy compared to the current vaccine strain, and we wouldn't know that until the vaccine was actually made and used.

The third option, of course, would be to defer the recommendation. In favor of that it might give us time to look at some other strains, and a different, more recent strain might provide a closer match with other contemporary H3N2 viruses in terms of the hemagglutinin and neuraminidase.

And, again, because the H3N2 viruses are associated with significant morbidity and mortality, that might be something to consider. Against that there is already a lot of information about how the H3N2 viruses have evolved during this current season, and it seems very unlikely that we're going to get any additional information in the next few weeks that would really add to what we already know about the current ‑‑ the majority of the current strains. And we don't really know whether another strain would be superior in terms of efficacy for vaccine production.

So for Influenza B ‑‑ again, to summarize ‑‑ the Influenza B viruses that have been circulating are in both of the known HA ‑‑ two known HA lineages. Strains that are in the vaccine HA lineage ‑‑ that is, the Victoria/287-like viruses ‑‑ seem to be antigenically very similar to the current vaccine strain, which is B/Hong Kong/330/2001-like virus.

Strains in the other HA lineage ‑‑ that is, the B/Yamagata/16/88 lineage, however, are the predominant viruses, and those have been represented by the reference strain B/Shanghai/361/2002.

Even though the Influenza B viruses have not been the predominant viruses in the recent months, strains similar to the B/Shanghai/361 virus have been isolated in many parts of the world. So they're out there circulating widely.

And although this B/Shanghai/361/2002 virus is related to previous vaccine strains, that and similar viruses are antigenically distinguishable from the previous B/Yamagata HA lineage viruses that were used in vaccines.

There is a little bit of evidence that the Influenza B viruses that are similar to B/Shanghai/361 are less well inhibited by antisera from people who have been immunized with vaccines that contained the virus in the same lineage, but even more so it's very apparent that the current vaccines do not inhibit these newer viruses as well.

And it's particularly obvious in looking at antisera from children where there's a true dichotomy, where there's a pretty reasonable vaccine response to the vaccine strain, but really no cross-reactive antibodies against the other HA lineage.

And we also have heard that there are some ‑‑ there are a number of candidate viruses in the B/Shanghai/361/2002 category that are available for potential vaccine use. But we don't really have a lot of information about those at this point, because they've only just recently been distributed to ‑‑ some of them have been recently distributed to manufacturers for evaluation.

So we don't really know exactly what their growth characteristics are going to be, although we heard from Greg Slusaw that the B/Jilin strain at least seemed to be a moderate strain in terms of its growth on early ‑‑ very early examination.

So the options for Influenza B ‑‑ one, we could retain the B/Hong Kong/330/2001-like virus, which is the current vaccine strain. For that manufacturing is very well defined, and it's very predictable. Against that at the moment the predominant strains are not in the same HA lineage, and those strains have been found in many parts of the world at this point.

Also against that, Influenza B viruses that are not in the vaccine HA lineage are not that well inhibited by post-infection and post-immunization antisera. And in particular, as I mentioned, the immunologically naive young children ‑‑ sera from immunologically naive young children who get the current vaccine don't seem to inhibit the B/Yamagata lineage viruses at all.

So the second option is to use a more recent Influenza B virus, and in favor of that the vaccines might provide better coverage for the currently circulating Influenza B viruses. There are several candidate strains that have been identified, and those are being examined right now.

Against that a new strain, you know, might not provide superior immunogenicity, and use of a new Influenza B strain could cause some difficulties in manufacturing, since we don't really have a lot of information at this point.

And then, finally, the third option, of course, is to defer the recommendation. And in favor of that that would provide more time to evaluate these candidate vaccine strains, and that might permit us to find the best match for a strain that matches the contemporary hemagglutinin and neuraminidases. Against that we don't really know whether a new strain really would be ‑‑ how much better that would be in terms of its actual use.

And so I'll stop there, and just remind the committee that the question is, of course, what strains should be recommended for the antigenic composition of the 2004-2005 influenza virus vaccine that we'll be using in the United States?

And the answer to this question should, of course, be based on the epidemiology and antigenic characteristics of the viruses, serologic responses, and availability of candidate strains for use. And I can answer a few questions, if there are ‑‑ if you want me to.

CHAIRMAN OVERTURF: Are there any questions for Dr. Levandowski? Yes.

DR. FARLEY: This is on the same track as yesterday's questions about B virus. This is for H1 virus, Influenza A H1. The numbers that we had in the handout were very, very small, as you've referred to. I think we only had 10 that showed the responsiveness to the previous vaccine sera.

If we look at the other WHO sites, does that give us reassurance that everything ‑‑ the additional numbers that that might bring in were also responsive?

DR. LEVANDOWSKI: Maybe Nancy can ‑‑

DR. FARLEY: Sort of a question for Nancy.

DR. LEVANDOWSKI: Maybe Nancy Cox would like to answer that.

DR. COX: Sure. This is Nancy Cox. The only significant outbreaks that have occurred since October have been in the Ukraine and Iceland. Those ‑‑ isolates from those outbreaks were analyzed by the WHO Collaborating Center in London, and the antigenic characteristics of those viruses were very similar or identical to the antigenic characteristics of the viruses that we've displayed in our table.

So I think that although the numbers are really small, if you look back at the previous time interval, which was during the southern hemisphere influenza season, that data also is reassuring that, you know, what we have ‑‑ we've looked at what is available, and it is very consistent data that we're seeing from all centers with small numbers of viruses, and the viruses are very well inhibited by antiserum to the New Caledonia vaccine strain.

CHAIRMAN OVERTURF: Are there other questions for Dr. Levandowski?

At this time, I think I should open up the discussion. And what I would prefer to do is to have a discussion regarding each selected strain, and we'll open it up with the H1N1 strain. And then after that we can vote on that strain and the recommendations, and then that will be followed by a discussion by each strain in sequence.

Is there any discussion regarding H1N ‑‑ further discussion about H1N1 strains? Any comments?

Well, the options ‑‑ the options for the H1N1 strain is to either retain the current New Caledonian strain or to go to a more recent strain or to defer the recommendation. And so those are your three options when we polled the committee on a vote.

And so I will start with David and have you go around the table and all state your vote.

DR. MARKOVITZ: Yes. David Markovitz. I would like to indeed vote to retain the current New Caledonia strain. Strains like the current circulating viruses are all very similar to this, and it's up and running and in production, and so it seems fairly clear that that's the way we should go.

DR. DOWDLE: Walter Dowdle. And I would vote to retain the current Caledonian strain.

DR. GOLDBERG: Judith Goldberg. I vote to retain it.

DR. KARRON: Ruth Karron. I'd vote to retain the A/New Caledonia strain.

DR. ROYAL: Walter Royal. I vote to retain the A/Caledonia strain.

DR. FARLEY: Monica Farley. I vote to retain it.

DR. McINNES: Pamela McInnes. I vote to retain the A/New Caledonia/20/99-like virus in the current vaccine.

MS. PROVINCE: Cindy Province. I vote to retain the current strain.

DR. GELLIN: Bruce Gellin. I also vote to retain the current New Caledonia strain.

DR. PHILLIPS: Steve Phillips. On behalf of Department of Defense, I vote to retain.

CHAIRMAN OVERTURF: This is Gary Overturf. I also would retain the current New Caledonia strain.

DR. LaRUSSA: Phil LaRussa. I vote to retain.

DR. MYERS: Martin Myers. Retain.

DR. WORD: Bonnie Word. Vote to retain.

DR. PALESE: Peter Palese. Retain, please.

DR. MONTO: Arnold Monto. Retain.

DR. EICKHOFF: Ted Eickhoff. I think there's an echo in this room.

(Laughter.)

Vote to retain. This is the easy one.

CHAIRMAN OVERTURF: Nobody wanted to buck a trend.

DR. FREAS: Just for the record, I'd like to announce that we're at the end of the voting members of the table, but we do ask industry for their opinion.

DR. DECKER: The industry applauds the wisdom of the committee.

(Laughter.)

CHAIRMAN OVERTURF: Is there any further comment? I think there's a unanimous vote to retain the New Caledonian strain for the H1N1 Influenza A.

The second issue is regarding the H3N2 strain, and I'll open this issue up to discussion again. And, again, there are three alternatives for that. One is, of course, to ‑‑ to use a more recent influenza A/Fujian strain or to retain the A/Panama strain or to defer this decision as well.

So I think those are the three options that we'll be voting on, and I'll open this up for discussion at this point. Is there any discussion?

Yes, Martin.

DR. MYERS: I guess it's a ‑‑ like on the last one, I have a question for Nancy. Some of the recent isolates were ‑‑ that you showed yesterday in your table were less reactive against the Fujian. Is there further drift that you anticipate occurring?

DR. COX: We have looked very carefully at recent viruses, and we really concentrated on looking at viruses that are in slight ‑‑ those slightly different genetic groups that have the changes at 126 in one group, or the change ‑‑ and the changes at 197 ‑‑ sorry, 193 and 227 in the other group.

And we haven't found that those viruses are antigenically distinguishable from the Fujian-like viruses. So we're seeing low avid viruses have a tendency to group together in the 193/227 group, but some of those viruses that are not as well inhibited by the Fujian antiserum are also not well inhibited by any of the other antisera. So we just think those are low avid viruses.

So I don't ‑‑ in answer to your question, the short answer is we don't, at the moment, see anything that looks like a definable antigenic variant from Fujian.

CHAIRMAN OVERTURF: The one thing that was troubling to me yesterday a little bit in the data ‑‑ and maybe I was misinterpreting it ‑‑ it seemed to me that there was a fair amount of heterogeneity in the serological neutralization of A/Fujian. Is that ‑‑ was that a correct interpretation? Or do you feel it was fairly uniform among the A/Fujian-like isolates that in the sera was fairly representative, fairly neutralizing? Do you understand the question?

DR. LEVANDOWSKI: You mean from the human serologies.

CHAIRMAN OVERTURF: Yes.

DR. LEVANDOWSKI: Well, I think we saw some variability. I tried to point that out. Maybe I didn't do such a good job. I think generally what we saw for most of the sera examined in most of the laboratories was that the majority of the A/Fujian-like viruses are not well inhibited by current vaccine antisera.

CHAIRMAN OVERTURF: Okay.

DR. LEVANDOWSKI: But they ‑‑ I think we can see that there is ‑‑ there is some heterogeneity there, too, that is a little bit reflective of ‑‑ of what is seen with the ferret serum. And we didn't emphasize that, but there are some ‑‑ there were some of the low reactor type strains that were included in the serologies. And just as they were low reacting in ‑‑ with the ferret sera, they were very low reacting with the human sera as well.

So it's ‑‑ there is that heterogeneity that is out there, and I don't know that that can answer your concern except to say that, yes, I think you are correct in interpreting it as ‑‑ as there is some variability in the results, not only from lab to lab but between the viruses themselves.

CHAIRMAN OVERTURF: Is there any further discussion regarding the issue of the H3N2 strain selection? Again, I will repeat, I think the three options is either to defer this, to retain the A/Panama strain, or to elect to proceed to an A/Fujian-like H3N2 strain for the 2004-2005 season.

So with that, I think we can probably, again, poll the committee for a vote. And this time, to give David a break, I will start with Ted.

DR. EICKHOFF: Thank you. I truly think this is the other easy one. The next one won't be this easy. But I think certainly based on the data that Roland and Nancy have presented, I think it's time for an update. So I would recommend that we switch strains and move to an A/Fujian-like strain. There are two candidates available that are said to grow moderately well or grow well in eggs.

There's another issue that I'll just put out on the table. I think there is an issue of public perception here that we should be aware of. If we voted to retain the current A/Panama strain, I think there would be a certain loss of credibility ‑‑

(Laughter.)

‑‑ on the part of this panel. Hence, I would vote to update to an A/Fujian-like strain.

DR. MONTO: Arnold Monto. I agree. We need to move to the A/Fujian-like strains. I think that everything is in place for their manufacture, which I think is probably going to come on second. And, again, I totally agree that if we didn't do it, we'd have a lot of explaining to do.

DR. PALESE: Peter Palese. I vote for switching to the Fujian strain, which we should have probably done last year.

(Laughter.)

DR. WORD: This is Bonnie Word. I would agree for the switch to the Fujian-like strain based on the information presented here.

DR. MYERS: I agree.

DR. LaRUSSA: Phil LaRussa. I vote to switch.

CHAIRMAN OVERTURF: I also vote to switch. I also think there is not just the issue of credibility, but I think there is a broader issue in terms of vaccine acceptability. I think when one vaccine seems to perform poorly, or that's the public perception, I think it hurts all vaccines. And being a vaccine advocate, I would really like to make sure that we choose the optimal strain. So I also would vote for a change to A/Fujian.

DR. PHILLIPS: Steve Phillips. We vote to change.

DR. GELLIN: Bruce Gellin. I also vote to change to the Fujian.

MS. PROVINCE: Cindy Province. I vote to change to A/Fujian.

DR. McINNES: Pamela McInnes. Given that we are supposed to take into account data, along with a degree of pragmatism in our decisionmaking, I vote to change to an A/Fujian-like strain.

DR. FARLEY: Monica Farley. I vote to change to the Fujian-like strain.

DR. ROYAL: Walter Royal. I vote to switch to the Fujian strain.

DR. KARRON: Ruth Karron. I vote to change to an A/Fujian-like strain.

DR. GOLDBERG: Judith Goldberg. I vote to switch to the A/Fujian strain.

DR. DOWDLE: Well, I'm not going to start a new trend at this late date. But ‑‑ but ‑‑ this is Walter Dowdle. I would like to point out, though, that I think the data that we've seen so far it's just a little too early to decide how well the Panama strain performed last year.

You know, I think we shouldn't leave here without recognizing that we've got a lot more data coming in, and the studies that we've seen in this country at least were not designed to really evaluate the current strain. So we need to keep this in mind. I don't think that it ‑‑ based on past evidence, there's no reason to believe that this was a disaster at all, because past evidence just simply didn't point that direction.

Having said that, I think we also always agree to try to have the closest match we can get, and it's clearly Fujian.

DR. MARKOVITZ: Yes. David Markovitz. I'd like to agree completely with what Walter said and vote to switch to the Fujian-like strain.

CHAIRMAN OVERTURF: Dr. Decker, would you like to make a comment for industry?

DR. DECKER: We're ready to get to work making it.

CHAIRMAN OVERTURF: Again, that's another unanimous vote to change to the A/Fujian strain. And we'll proceed onward, then, to the selection for an Influenza B strain.

It seems to me, again, there are really three options here. Again, one is to retain the current strain, the second one is to proceed to substitute with a B/Yamagata-like virus, or, again, to defer the recommendation. And so I will now open this issue up to discussion.

Dr. McInnes.

DR. McINNES: Thank you. I'd like to ask a question, please, of Nancy.

Nancy, the data that you shared about the B strains ‑‑ I think it was pretty clear that of what you have, the B/Yamagata-like strains are dominating, and you showed that the two ‑‑ that the strains you have are most either like the B/Shanghai/361 or the B/Jilin/20. And I still don't know if it's 2003 or 2002. Is it 2003 at the end? That's not my question, but is it B/Jilin/20/2003?

DR. COX: 2003.

DR. McINNES: Okay. And that the data from the October '03 through what you currently have from the three collaborating centers, you had 49 B isolates total, of which 46 were B/Yamagata and three would be Victoria-like.

DR. COX: That's correct. And I can add a bit more data that is from Canada. And so the numbers now are 57 total B's, 52 of which are Yamagata lineage and five of which are Victoria lineage.

DR. McINNES: Thank you. So my question was, given, I think, you also said there wasn't much B activity in, we presume, the collaborating centers, is it ‑‑ is it a fair assumption that the collaborating centers have all of the B data we're going to be able to get for the foreseeable future? Or is there a likelihood ‑‑ are there viruses that have ‑‑ are in the collaborating centers or could be obtained that have not yet been looked at?

DR. COX: We have a few at CDC. But, again, it's relatively few that are in the pipeline. And certainly whatever data we can develop within the next few weeks would be available for the committee, should that be necessary.

It doesn't look to me ‑‑ we have ‑‑ I was checking the log-in sheets before I left. And although we've received a lot of packages, the majority of the viruses are actually A's, H3N2's. So it would be a limited amount of information.

CHAIRMAN OVERTURF: Nancy, of the additional strains over the 49 that you mentioned yesterday, and now the ones from Canada, what were the dates of recovery of those viruses?

DR. COX: They were all isolated between October and the current time. So they're all in that most recent time interval that you see on ‑‑ on our tables that we present.

DR. KARRON: One more ‑‑ one more question for Nancy. I know that sometimes that when we have an A year, we often have a ‑‑ we have B late in the season. Would we ‑‑ if we were going to have that happen in the northern hemisphere, would we already have seen that?

DR. COX: Influenza is full of surprises, as we all know. But we've been watching very closely for that second wave of activity, and we have ‑‑ and we sort of expected that we might see it for Influenza B viruses this year, since we had Victoria-like viruses circulating last year, quite a few school outbreaks, and so on and so forth.

But we haven't seen that increase in the proportion of the total isolates that are Influenza B. In fact, in past weeks we've seen very few B's isolated in the U.S., so we're not getting ‑‑ we're not getting that signal that we usually do when we have that second wave.

CHAIRMAN OVERTURF: Dr. Palese.

DR. PALESE: Could I ask whether we could fine-tune one of the options, so one is retaining it, one is changing, and the third one was to defer ‑‑ if I remember correctly, looking for the most recent isolate, the most ‑‑ I think that was the option in terms of deferring.

I would like to fine-tune this and say we defer, but really have only the choice between the Victoria and the Yamagata. That would give the manufacturer a chance to develop and gear up. They have the old strain ‑‑ to gear up the new one, and then a deferment would ‑‑ deferring would mean that we chose to wait, what we get in in the next couple of weeks or more ‑‑ three weeks or four weeks.

So, in other words, not sort of leaving the option completely open in terms of deferring, but rather say at the end of that period a decision will be made for either switching or retaining.

CHAIRMAN OVERTURF: Yes. This is very similar to what we did with the A strain last year, actually. Yes.

DR. DECKER: Just a suggestion of the refinement of that, or maybe this is what Peter was saying and I didn't understand it. But it would seem to me that the data are sufficiently well in hand to make a provisional choice, which would stand unless CDC, FDA, and the Chair elected to call another committee meeting.

In other words, instead of putting off your decision, I wonder if you can make a decision now, recognizing that you've got X period of time to revisit that, because we are waiting for reassortants to be made, and so on.

DR. PALESE: No, that was not what I meant. I really meant to leave it open, not to make a provisional decision.

DR. DECKER: Well, then, my impression was right, and Peter didn't say that. But I wanted to.

(Laughter.)

CHAIRMAN OVERTURF: Dr. Monto.

DR. MONTO: Yes. I would agree with what I thought was a refinement by Dr. Decker of the ‑‑ of Peter's remarks, because I think this is a critical decision, given the fact that, number one, there is so much divergence between the two lineages, and also the fact that we've experienced in the past situations where one lineage predominated in one part of the world and the other lineage predominated in another part of the world.

Correct me if I'm wrong, Nancy, but I think the B ‑‑ the B/Vics persisted in ‑‑ in Asia when we were having B/Yamagata a few years back. So the question I would have is: how many of these isolates are actually from the western hemisphere?

DR. COX: That would take me some time to dig through.

DR. MONTO: Just an impression.

DR. COX: But the majority of these isolates are from China and Japan and Thailand, and so on, rather than from the western hemisphere. So what we're seeing there is ‑‑ well, we ‑‑ you're absolutely correct in recalling that the B/Victoria viruses persisted in Asia, and particularly in China, for a number of years when they were detected nowhere else.

And then we had the continuing evolution of the Vic-like lineage, and then had its reemergence in the rest of the world. It's really ‑‑ because we've had co-circulation of the two lineages, it's really difficult to know exactly what's going to happen, particularly based on relatively small numbers. But the signal is there that the Yamagata lineages ‑‑ lineage is predominating actually globally.

CHAIRMAN OVERTURF: Dr. Goldberg.

DR. GOLDBERG: Yes. I mean, actually, what you're observing from the data that you have is that 91 percent of the observed isolates are Yamagata lineage. And if you just put a 95 percent confidence interval on that crudely, it's compatible with something between 83 percent and 100 percent, as the true underlying rate would lie within that interval.

So I think that with the data we have it would take a lot to overturn it. And if you don't expect very many more of these isolates, I think we're dealing with an issue that we have to make some informed judgment, recognizing all of the biases in the way we get the data.

CHAIRMAN OVERTURF: Yes. If you look at the current data, it sounds like we've only had three or four B isolates per week over the last season. And if we were to wait another two or three weeks, we might have another half dozen at best, unless something starts up, obviously, but I really think the chances that we will see new isolates is probably small. And I'm a little bit perplexed about how you would make a decision, unless you had five out of six isolates suddenly turn to A/Victoria ‑‑ I mean, B/Victoria.

Yes, Peter.

DR. MONTO: No, I think that's the dilemma, because we have seen, and almost consistently seen, late B seasons. But the problem is they may be, as Nancy is saying, that since nothing much is in the pipeline, it may be too late, even though we do ‑‑ we do see B activity in March and going into April. It's going to be too late, really, to ‑‑ given the need for time for analysis and the rest to ‑‑ to be able to say anything that we haven't been able to say today.

CHAIRMAN OVERTURF: Yes, Dr. Palese.

DR. PALESE: I just wonder whether these numbers ‑‑ I feel uncomfortable with 50 isolates when we are told that there are 20,000 isolates per year. So is that the number I heard, that overall 20,000 isolates are made per year? So if you only have 10 percent ‑‑ or, let's say, five percent B viruses per year, we should have 1,000 isolates.

So I don't understand why we have so few isolates. And is there a possibility of asking WHO and other countries to help us? I mean, I can see that we have basically no B in the States. But out of those 20,000 isolates, which are ‑‑ have been ‑‑ are being made worldwide, I think we should be able to get some more B virus isolates.

CHAIRMAN OVERTURF: Dr. Cox, do you want to respond to that?

DR. COX: I didn't think that was a question.

(Laughter.)

DR. PALESE: No, it is a question.

CHAIRMAN OVERTURF: I think that was a ‑‑

DR. PALESE: If we have 20,000 isolates per year, and we get a conservative number that we have five percent of Influenza B virus, there should be 1,000 isolates of B around, and then we could make a more informed decision.

DR. COX: Unfortunately, this year less than one percent of the isolates have been B, so ‑‑ and we don't necessarily have 20,000 isolates every year. So, and not all of the isolates are sent to the WHO collaborating centers for analysis, so we receive a subset of the viruses. You know, we have analyzed what we've had. We have beaten the bushes, and we have actually developed ferret sera, as soon as we saw the switch in lineages.

So we ‑‑ we have really worked hard to develop as much data as we could within the timeframe between the time we realized that we were seeing the shift in lineages with relatively few B viruses coming in.

I personally don't know of the existence of B viruses anywhere outside of the U.S. that we could ask for. There just has been quite little activity. I can see if there is a possibility to get some information, but I certainly couldn't promise that in three weeks or four weeks, that we would have a significant amount of new information.

CHAIRMAN OVERTURF: Dr. LaRussa.

DR. LaRUSSA: I realize you get a select and limited group of isolates. But do you have any information on the lineage of the most severe clinical presentations ‑‑ the deaths and very severe disease?

DR. COX: We do not, and the information that comes with the isolates that are sent globally does not include information about severe cases. The information that we get from the U.S. sometimes includes that information, and, of course, when we get information like that we look at those isolates very carefully.

And, in fact, this year for the H3N2 isolates that have come from fatal cases, or serious cases, we have tried to sequence the entire genomes to see if there's anything unusual about those. So if we do get that information, we look very carefully. Most of the time we do not get that information accompanying the isolates.

DR. LaRUSSA: What about the B viruses? Do you have any information for the B's?

DR. COX: We have no information this year that any of these isolates came from fatal cases or particularly severe cases. Obviously, all of these cases ‑‑ all of the isolates come from patients who sought medical attention, and that's all we can say.

CHAIRMAN OVERTURF: Dr. Markovitz.

DR. MARKOVITZ: Yes. Just, Peter, you were suggesting that we get data from WHO. But the WHO has already selected Yamagata-like strain, right? So anything they have would it seems would support that, I would infer.

DR. GELLIN: Yes. I have two questions, one is for Nancy, and one is for Michael representing industry. The Nancy question is to ‑‑ is there something about the B viruses or these B viruses that would lead them to be unrepresented in the surveillance system? Do they not grow well, or is there some reason why the ‑‑ they may be underrepresented? Or do you think that the numbers out there reflects the circulation?

DR. COX: The Influenza B viruses that have been circulating over the past few years have grown relatively well in tissue culture. And MDCK cells are actually used most widely for isolation of influenza viruses generally. And Influenza B viruses of both lineages have grown reasonably well, so we ‑‑ we wouldn't have any reason to believe that the B/Vic-like ‑‑ or B/Vic lineage viruses are underrepresented in our ‑‑ in our sample.

DR. GELLIN: And my Michael question is a deja vu from last year. Given that manufacturing timelines ‑‑ if we were going to entertain a delay decision ‑‑ what's the drop-dead date by which you would need to know to proceed with manufacturing strain three?

DR. DECKER: Well, I ‑‑ let me turn it around, because I don't think I can answer it the way you posed it. I have no reason to think that, say, a three-week delay in making a decision would have any ‑‑ any plausible risk of an adverse effect on supply or ‑‑ time and/or availability of supply.

You start going out beyond that and my anxiety grows, because of the cascade of factors that have to occur. I'm reasonably confident that we could spend the next three weeks cheerfully making H3N2 and worrying about recombinants, and so on. But eventually you're going to have questions about developing the high growth reassortants, generating the reagents, and on top of that at some point you're going to start manufacturing the monovalent, and so on.

And since I have the open mike, let me go on with a comment I wanted to make. It seems to me not only is it true that the remaining absentee ballots out, if you will, can't ‑‑ are not likely to be able to change the election, because there are already so many ballots counted ‑‑ that is to say, B's already looked at ‑‑ that you're not going to be able to shift the vote tally.

But also, if that were to happen, let's suppose we got five more and they were all Vic, I think we would still sit here saying, you know, not only is the majority vote still for the Yamagata, but in addition we've got a population that hasn't seen the Yamagata vaccine for a long time.

And if we're unsure, we've got to go for the new strain, because there's ‑‑ if we're wrong, there's at least the hope of some benefit from prior years' campaigns with the other strain, whereas if we stick with the current strain you don't have that going with you. So it's hard for me to imagine that delay will actually buy anything useful.

CHAIRMAN OVERTURF: Yes. Dr. Goldberg was first.

DR. GOLDBERG: If we assumed ‑‑ to continue what I brought up before, if we assume that we got 10 more B isolates, and that they were all not Yamagata, you still would have observed 63 percent of them, and you'd still be more than 50 ‑‑ pretty certain if the true rate was over 50 percent would fall within that interval.

So that I think it would be very hard to overturn these data in a short amount of time, unless the rate completely changes of accrual of the isolates. I mean, that's really the hooker, and so I ‑‑ I mean, I guess it was really the drop-dead date, with an expectation of what you might assume.

You think five. I was more generous and said 10, to see how far we could push it. But it doesn't seem like you would really overturn the view in this short amount of time.

CHAIRMAN OVERTURF: Yes, Dr. Karron.

DR. KARRON: Just a comment I wanted to make to address something that Michael Decker said. I am particularly concerned, given the point that Marty Myers made yesterday, about responsive of young, naive children to Influenza B.

So whether there has been prior experience in the population with the other strain or not I don't think is as relevant for those young children who are naive. So I think it's ‑‑ it is ‑‑ if we can, particularly incumbent on us to choose the right strain to protect that population.

CHAIRMAN OVERTURF: Is there any ‑‑ yes.

DR. McINNES: And just following up on Ruth, because I also was very concerned about the pediatric population that, Marty, you and Dr. Turley put on the table, because the mean age was pretty low, and they really were the younger infants.

And I just want to confirm, because my ‑‑ dendrogam reading is sometimes impaired.

(Laughter.)

On the table that you showed, the HI antibody response to the B component in that pediatric population, you had an n of 25, and then you show ‑‑ one, two, three, four, five different antigens. Am I reading this correctly, that only the B/Hong Kong, which is close to the vaccine strain, comes from the B/Victoria, and that those other four are B ‑‑ or B/Sichuan/259/2003 is also from the B/Victoria, right?

So that the B/Jilin, the B/Washington, and the B/Jiatsu are all from the B/Yamagata, which matches, then, the very poor cross-reactivity that we're seeing.

Thank you.

CHAIRMAN OVERTURF: Yes. Dr. Levandowski.

DR. LEVANDOWSKI: And I should emphasize that the other data that we had from the other pediatric study showed the same thing. It was identical in the respect that there was a good response against the vaccine type strain, but not against the B/Yamagata lineage strain that was ‑‑ it was the same kind of night and day, even though the children were somewhat older. And a few of them might have been immunologically primed, but the data show the same thing, really.

CHAIRMAN OVERTURF: Dr. Eickhoff.

DR. EICKHOFF: Ted Eickhoff. Basically, I subscribe to Judith's way of looking at this. I think the numbers now clearly weigh in favor of a switch to B/Yamagata. However, if indeed we were to see a late B outbreak this spring, and it were 100 percent A/Vic, that would make me worry a great deal.

I think the odds of that happening are, frankly, remote. But who knows?

CHAIRMAN OVERTURF: Dr. Palese.

DR. PALESE: Can I just address also ‑‑ I mean, flu has its bottlenecks genetically. So if suddenly, really, 10 ‑‑ I mean, hopefully it won't be more ‑‑ of the B/Victoria would come up, I think that would weigh much heavier despite statistics, because it would genetically say that one which makes it, and, therefore, I think it would be very worrisome, and suggest that the next wave would be a B/Victoria rather than B/Yamagata.

CHAIRMAN OVERTURF: Dr. Monto I think was first.

DR. MONTO: And just to continue the statistical discussion, I think we are dealing with clusters ‑‑ with cluster sampling here.

DR. GOLDBERG: Yes. It's very biased accrual and sampling as well, so that what you don't know is whether you're getting them all in the right sequence from the same location. So that I think that we're in a small numbers business.

And, unfortunately, it all rests on what Dr. Eickhoff brought up, which is really, what is the likelihood that we will really see an outbreak of B somewhere, that we will have some new isolates clustered in time and in place, so that we would really be able to interpret the data. I think that's the crux of the issue, and anything beyond that is ‑‑ it's personal probabilities of what you feel the likelihood of that is.

CHAIRMAN OVERTURF: Dr. Karron.

DR. KARRON: A question back for Michael Decker, which is, how helpful would it be to industry to make a provisional recommendation but with a mandated, you know, rereview by the committee at X time of what the choice of strain is. Is that helpful in any way? Is that not helpful?

DR. DECKER: The only thing that I was thinking was that if we think it's probably a settled issue, we'll want to reserve a chance to ‑‑ for a change, leaving it at ‑‑ at the administrative level, rather than having to convene the committee in a conference call, which depends upon schedules and will be on a fixed time, might have some advantages. That's all.

Because if ‑‑ if, for example, over the next two and a half weeks the answer becomes obvious, either we need to have a meeting or we don't need to have a meeting, obviously, the CDC and FDA, they can just transmit the ‑‑ if it's not have a meeting, they can transmit the information to industry, go ahead and start making this one, whatever was chosen.

If you ‑‑ if you set a meeting date, if you think it's necessary to set a meeting date, I think the consequence of that will be that people will not take action until that meeting date. I mean, otherwise there is ‑‑ the fact that you chose to do it that way sends the signal that anything produced in that period of time is at risk.

So it's a small difference. But if ‑‑ if ‑‑ in a case like this where I think we're reasonably sure, I just want to raise that possibility as one option for the committee.

CHAIRMAN OVERTURF: Dr. Levandowski.

DR. LEVANDOWSKI: Just so you know, there was the anticipation that there might be a possibility of deferral, as there was last year, and so there is a meeting date that's already been set. And I think it has already been announced. Maybe Bill Freas could fill us in on the details on that.

DR. FREAS: Administratively, I believe the date that has been selected is March 17th as the latest date that this committee could convene and make a decision. And that would be by teleconference.

CHAIRMAN OVERTURF: Dr. Gellin.

DR. GELLIN: I mean, we're here ‑‑ we're here at a fixed point in time, and our ‑‑ you know, weighing a decision ‑‑ the viruses continue to circulate. Surveillance continues to be ongoing. So I guess I wonder if we should have a brief discussion about plan B. Should there be a plan B?

And if ‑‑ I mean, this is, I mean, Michael's worst nightmare, but I think we should have a discussion about if something were to change later in the year, for which we wanted to either reconsider or have another discussion, what would we do? I mean, if ‑‑ sort of it ‑‑ and pick your number of isolates or outbreaks that occur, and then there is some shift for which we think that there might be a need to have a different virus to provide better protection next fall, recognizing that the large ‑‑ large manufacturing processes may have begun.

What would we do? Are we nimble enough to do that? Or are these all or nothing decisions, and we're stuck with our decision, whether it's today or March 17th, and then potentially watch another season go by where we would have wished we might have done something different.

CHAIRMAN OVERTURF: Yes, I'd like to hear a discussion regarding that from industry. Each ‑‑ this is now the second year this has come up, and the issue always is: is it an all or none decision? It sounds like we make the decision at a point in time A, and we make another decision at a point in time B, but it didn't sound like last year ‑‑ and I'm not sure this year ‑‑ that anything is really reversed by the second ‑‑ by the second call, because it's just too short a period of time to make a lot of changes and decisionmaking.

So it sounds to me like somewhat an irreversible decision. The only thing that happens is is that there's a delay in manufacturing ‑‑ in starting the B process, if we don't make a final decision today.

DR. DECKER: Yes. Dr. Gellin raises an interesting question with a number of angles to it. First of all, let me comment again. I don't see any problem with our putting off the B decision for three to four weeks, because I think we'll be fully occupied with H3N2 in the meanwhile.

So this year, this time, I don't see a particular problem with that. That's not always true. But the question really is, what if a freight train comes running down the tracks in July? What happens then? Well, either we've got ‑‑ either you're talking about a pandemic freight train, in which case, Bruce, you know as well as anybody in the country what we do then, and how ‑‑ what we figured out what to do.

But the simple answer to that is that there are some high-level meetings that made some fundamental decisions. Do we ‑‑ do we stop all current production and devote all resources to generating monocomponent pandemic strain? If it's a serious enough threat, that's what we'll do.

And then, everything that's in the hopper at that point, a decision will be made, between the government and academia and industry of what to do with what's in the hopper. Do we distribute it? Do we not? But you'll stop making it. That's if it's a pandemic. I think that's pretty clear.

But let's suppose you have something else. It's not the pandemic, but ‑‑ but you have a situation, as we had a decade or a decade and a half ago, where you've got this new strain that comes on very strong, and there's a desire to do something about it.

I think the simple fact of the matter is you've got no answers that work. There are lots of choices. They've all been tried, and none of them were satisfactory. For example, you can make a monovalent supplemental strain, and by the time you get it made and distributed nobody wants it.

You can alter your current formula, in which case you're throwing away months of production, and that will impair both the timing and availability of vaccine, and you'll end up with a better vaccine that arrives to people later in lower quantity.

So if you do some type of net calculus of the benefit to the country, you've not clearly benefitted. And we faced a variant of that last year, where this committee agonized over the choice of strain, and everybody here wanted to use Fujian, but there was ‑‑ but the majority of the committee felt that it wasn't practical to try to go that way, because of the impact on timing and supply of vaccine.

So we reluctantly went with the next best choice, and we ended up with what was predictable, which is a vaccine that didn't prevent ILI very well, but which ‑‑ which, if past history is any guide, will turn out a year from now when we have all of the data to have been pretty good at preventing death and serious morbidity, which is basically what we hire the vaccine for in the first place.

So was our decision last year wrong? Probably not, but we don't know yet. So what if the same thing happened again this year? What would we do? I just don't think we've got a good answer, because we're ‑‑ the virus moves on its timeline, which is not necessarily consistent with the manufacturing timeline.

If it rises to a pandemic and you want to stop making the regular vaccine, that can be done. But when it falls in that gray zone, and you don't want to throw away the current vaccine, but you'd like something more, you probably can't get something more done using any technology known to man in the timeframe available before the virus comes and goes.

CHAIRMAN OVERTURF: I'm concerned that ‑‑ still, that the argument that we will not see a difference in three or four weeks is probably very good. I would prefer, if we plan a meeting three to four weeks from now, that it only be to rereview what we've looked at.

But I can't ‑‑ I can't conceive of ‑‑ of unless ‑‑ unless we have a major outbreak of B/Vic someplace in the next three weeks, I can't imagine how this is going to change the decision. And we're just ‑‑ I think we're stuck with a low numbers game for this year, and that's ‑‑ that's the game.

Yes.

DR. DOWDLE: Walter Dowdle. I tend to agree with that, that you already have now a Victoria ‑‑ I mean, sorry, a Hong Kong strain that grows well, I mean, in the current vaccine. You've already got that, so that's nothing new. I mean, you could have that available.

But to me it's inconceivable that this committee would not want you to start right now and to work as hard as ‑‑ I mean, to send a strong signal that we need to start adapting the Yamagata-like strains for growth of a vaccine. I don't see how we can send any other signal except we need to start right now.

So the question is, three or four weeks from now, is ‑‑ is you either with Yamagata or you go with Victoria. And we're all saying the likelihood of going with ‑‑ with the Hong Kong-like strains, Victoria-like strains, are pretty slim.

So I ‑‑ I would agree. I'm not sure what ‑‑ waiting is going to do that much good, except to say ‑‑ just to say, well, in the last minute there's possibility to reverse, but that's pretty remote.

CHAIRMAN OVERTURF: Yes, Dr. Markovitz.

DR. MARKOVITZ: Yes. Just following up with what Walter just said. I wanted to just specifically ask Roland and Nancy ‑‑ sorry ‑‑ I think you guys have made it fairly clear, but where do you come down on the issue of the likelihood of any significant real data emerging in the next four weeks?

I think, Nancy, you said very unlikely, but I'd just like to hear from both of you how likely it is that we're going to find anything new in the next four weeks.

DR. COX: I think that unless, as others have discussed, unless there is a large outbreak somewhere, large outbreak of Victoria lineage viruses somewhere, it is unlikely that we would be changing our view of what's going on. And it doesn't appear at this time that there is a lot of B activity out there.

I suspect that we could do something such as develop an understanding that if such an outbreak did occur that information would be transmitted to the committee immediately. I don't know if we've had such an arrangement in the past, but that I think might help reassure the committee that if anything did happen they would have that information as soon as possible and then the committee could reconvene or ‑‑ or do what would be necessary to look at that new data and reconsider the options.

CHAIRMAN OVERTURF: Actually, that's a plan that I would actually favor, would be to leave it up to the individuals who continue to do the surveillance to make the committee aware of any significant changes. If we want to define a "drop-dead date," or industry wants to define a drop-dead date when we can't change, although we do already know we have the current Vic strains.

So I don't think that's a huge issue, if we make a decision to change. But the only decision would be predicated on extremely good surveillance that changes that picture, and I ‑‑ I have difficulty knowing that that would happen. But it would seem better to leave it up to the surveillance folks to give us that information.

Yes, Dr. McInnes.

DR. McINNES: I wonder if I might ‑‑ I'm a little bit uncomfortable with a decision and then, if necessary, to be sent the data and review it. And I wonder if there is the consideration of a provisional recommendation, which sends the strong signal, and a meeting on March 17th with whatever data we have in hand, to either confirm that recommendation or to change that provisional recommendation.

I'm a little bit uncomfortable with a recommendation, unless I hear something otherwise. And so I wondered if there was ‑‑ would be an opportunity to convene and get whatever data there would be by March 17th, and either then, you know, confirm that provisional recommendation or change it.

CHAIRMAN OVERTURF: I'd have to hear from Roland or Bill on this. I would think there would be several ways to transmit the information to the committee, either via mail or e-mail or a variety of other ways. And unless ‑‑ and we would have to convene an emergency meeting if there was ‑‑ people could leave the date open.

DR. LEVANDOWSKI: If you want me to answer, I ‑‑ I think we have had similar situations where we've had, at the ‑‑ when we used to routinely have two meetings, we were trying to avoid having multiple meetings for this purpose, but anticipated that it might be necessary.

In the past when we had such experiences, there were provisional recommendations that were made, and then they were either confirmed at the next meeting, and usually they were confirmed at the next meeting, in fact. The original decisions of the committee ‑‑ recommendations of the committee I think actually were the ones that were the ones that stuck.

But I think that this ‑‑ it's already set up that ‑‑ it's the possibility of having that meeting, and unless the committee didn't want to do it, I think it would probably be better for us to have that ‑‑ have that meeting in such a situation so that we do just transmit the information and make sure that it was understood.

CHAIRMAN OVERTURF: Dr. Goldberg.

DR. GOLDBERG: I would support the provisional recommendation and having some kind of confirmatory meeting, because we all know from events of the recent years, the unthinkable can happen. And it's a protection against that in the decisionmaking, as long as that date ‑‑ the March 17th date or whatever the committee date is will still meet the manufacturing deadline. It's to protect against the unthinkable.

CHAIRMAN OVERTURF: Dr. Karron, yes.

DR. KARRON: I'm trying to just figure out whether what industry ‑‑ their interpretation. Clearly, there is much interest in the issue of Yamagata. It's hard for me to imagine that whether we did provisional or if we even just formally deferred, that industry isn't seeing the positive signal towards Yamagata and wouldn't be looking at it as the strong candidate. And would they not be doing the preparation anyway?

And I'm just wondering how important it is, the actual category we put this into, because I think I agree having some protection against an overwhelming outbreak should be there. But does it matter where we say "deferred" with a strong feeling towards Yamagata or provisional Yamagata in terms of what ‑‑ what you will do in the next three or four weeks.

DR. DECKER: No. This is actually ‑‑ this aspect of it is relatively easy this year. I mean, I'm feeling pretty comfortable with the idea ‑‑ more comfortable than in some prior ‑‑ the idea of deferring the B decision until the March 17th date, or somewhere around that.

My comment before was more just towards the ‑‑ being reasonable to everybody's lives and schedule. For example, if you knew right now today that there would be no further isolates, would you still schedule a March 17th meeting? Probably not.

So if it turns out on March 17th there were no further isolates, do you still want to have a meeting? Or if CDC communicated that by e-mail, might we elect not to? That was the only nature of my question. It wasn't an industry capacity issue.

But let me turn it around for a second, just so that we've ‑‑ although I don't think it's ‑‑ although it's not applicable this year, it could be applicable next year. I'm feeling very comfortable from the industrial point of view about our deferring, if we wish, the B decision until the 17th, because we've got a clear-cut H ‑‑ we've got two clear-cut decisions, and the one we decided on that involved a choice ‑‑ a change ‑‑ the H3, we've got everything we need in hand to get to work on it.

But let's suppose that the facts were transposed, and you knew exactly what you wanted to do with B, but we didn't have the materials to work with yet. And you didn't know what you wanted to do with A H3N2, where we have the materials. That would be a real problem for us, because then you would be telling us we couldn't actually make stuff for three or four weeks. This year that's not a problem; we can make stuff.

So we ‑‑ I ‑‑ I believe there is only a threat to production if the ‑‑ if something happens that delays a decision beyond the point where we would actually start manufacturing rather than getting ready.

CHAIRMAN OVERTURF: Dr. Monto.

DR. MONTO: Let me create a strawman for a minute, and say that in the next week we have, in some place which has not had flu for the last few weeks, which is rather unheard of in the middle of the winter, a major outbreak in the United States of a virus in the B/Vic lineage.

We still have the situation where we know that in children, who are the main ones who get Type B infection and get in trouble with Type B infection, the situation that you either produce antibody to one or you produce antibody to the other. And you're not going to be producing antibody to both.

How would we move at that time? And will it ‑‑ will we move any differently if that happens?

CHAIRMAN OVERTURF: We'll discuss a quadravalent vaccine again, which we do every year also.

(Laughter.)

DR. MONTO: That's why this is more biology and ‑‑ than policy.

CHAIRMAN OVERTURF: Yes.

DR. DECKER: We had a wonderful discussion on quadravalent last year. But I just want to remind you ‑‑ I want to join the mathematical group with Dr. Goldberg and point out that 80 million doses at three serotypes equals 60 million doses at four serotypes. So had we made that decision last year, we would have had a more interesting vaccine for which 20 million Americans would have been told, "You can't have it," who actually got vaccine last year.

CHAIRMAN OVERTURF: Yes. It's a very important point to reiterate each year, that you reduce your doses considerably when you go to the quadravalent vaccine.

Yes.

DR. MARKOVITZ: Although if we really ‑‑ if we really had quadravalent vaccine every year, you'd ultimately increase production probably, right? So ‑‑

CHAIRMAN OVERTURF: Hearing no answer to Dr. Monto's question, I think we should probably proceed with a vote. And I ‑‑ to me, I think we have three options still, and the only issue is whether you wish to preface the change to a B/Yamagata with a provisional change, or whether you wish to simply vote for a change.

Obviously, the other two options are to retain the current strain, and the third option is to defer completely any decision today and plan for a review in the future.

So, yes, one ‑‑

DR. KARRON: I just want to make sure that I'm clear. So that if we vote for a provisional change, there will be a mandatory meeting on March 17th at which we will review the data?

CHAIRMAN OVERTURF: There will either be a mandatory meeting or there will be information conveyed that would ‑‑ would negate the need for that meeting, one or the other.

Is there any further discussion? Yes, Dr. Levandowski.

DR. LEVANDOWSKI: I think I'd like to reiterate that if there's a provisional recommendation, I think we'd want to have the next meeting. I don't think we want to just do things as an informal sending of information. I think we'd rather have it, as we're doing this, as an open public kind of presentation.

CHAIRMAN OVERTURF: I guess it would be a very short meeting if there was no new data, or if the data confirmed the selection.

Yes.

DR. PALESE: If one of the choices is deferring a decision, could that be also tied into with a request for more data? In other words, I don't know whether we have a way of doing this, but to really ask that a major effort is made ‑‑ and I don't know by whom ‑‑ that we get worldwide data, particularly from the summer season in the southern hemisphere.

CHAIRMAN OVERTURF: Well, I ‑‑ to me, the deferral is an automatic request for more data. But I ‑‑ in terms of getting more data, part of it is ‑‑ is whether isolates are available, and they simply may not be.

Yes.

DR. COX: Just ‑‑ we have the isolates from the southern hemisphere for the previous season. Their season ‑‑ their upcoming season doesn't actually peak until July, and so there is ‑‑ so we do have all of the information from the previous southern hemisphere season, and it's in the package. But we don't and won't have the new information in time to really help us out.

CHAIRMAN OVERTURF: Dr. Eickhoff.

DR. EICKHOFF: May I make a motion to ‑‑

CHAIRMAN OVERTURF: You can make a motion.

DR. EICKHOFF: ‑‑ get us started? I move that we recommend ‑‑ we provisionally recommend switch to a B/Shanghai/361/02-like virus, and that the target date for making that final is a teleconference on March 17th.

CHAIRMAN OVERTURF: Do I hear a second for that motion? Okay. Dr. Markovitz has made a second.

With that, I think we'll just go around the room from Dr. Eickhoff and request discussion. Dr. Dowdle.

DR. DOWDLE: I wonder if he would accept a slight amendment.

(Laughter.)

And that is that we recommend the Yamagata-like strain, and reserve the right to reconsider that at the March meeting, which would give a very strong signal, and would say that's the way we're going. And unless something comes up that strongly disagrees with this decision, then we can take that step.

It is ‑‑ I admit, it's the same thing, except that I think it does have some ‑‑ some implications here for the manufacturers and for FDA.

DR. EICKHOFF: I have no problem with that.

CHAIRMAN OVERTURF: So the motion has been made to vote on a recommendation to change to the B/Shanghai-like virus in the Yamagata lineage, and with an option to review that and change it, if necessary, at the March 17th meeting. Is that correct?

This time we'll start with you again, David, and ask for your poll.

DR. MARKOVITZ: I think it's pretty easy. I agree with Drs. Eickhoff and Dowdle that we should do exactly that. So I think it's probably being slightly overcautious, because it seems very unlikely that we're going to end up changing our mind.

But as caution is a very good thing to do with influenza, I think it's a very good motion. And hopefully we'll ‑‑ or probably we'll have a very short teleconference that day. Everyone has it blocked off on their calendar anyway, so it's a good idea.

DR. DOWDLE: Thank you. Walter Dowdle. What can I say but agree?

(Laughter.)

DR. GOLDBERG: Judith Goldberg. I agree. I think that there is an extremely low likelihood that the decision will be changed, though, but it is a good protection.

DR. KARRON: Ruth Karron. I agree.

DR. ROYAL: Walter Royal. I agree with the change to the B/Shanghai strain with the teleconference on March 17th.

DR. FARLEY: Monica Farley. I slightly prefer it to be called a provisional decision, but setting that aside I vote in favor of the recommendation to change with the option of reconsidering.

DR. McINNES: Pamela McInnes. I prefer the philosophy behind provisional recommendation, with an urging for whatever data can be brought to the table on March 17th to be done, and a reconsideration of those data with either an affirmation of that provisional recommendation or a change.

MS. PROVINCE: Cindy Province. I agree with Dr. Eickhoff's motion.

DR. GELLIN: This is Bruce Gellin. I agree with the motion but would like to put in a placeholder that if there is a lot of new data that is unveiled between now and then, we have a better understanding of where it has been.

DR. PHILLIPS: This is Steve Phillips, and we agree with the recommendation.

CHAIRMAN OVERTURF: I also agree with the recommendation. I ‑‑ I can't foresee that we're going to change this, but I ‑‑ I think it's a reasonable precaution.

DR. LaRUSSA: Phil LaRussa. I think we know what we're going to know. But given that the only harm is to the ‑‑ inconveniencing the committee members with another vote, I agree.

DR. MYERS: Martin Myers. I agree with Dr. Eickhoff's recommendation.

DR. WORD: Bonnie Word. I agree with Dr. Eickhoff's and Dr. Dowdle's recommendation. I'm just a little confused as we went around the room. I thought there was only one question on the table, and then I start hearing these other responses. So I don't understand how we're ‑‑ how we're ‑‑

CHAIRMAN OVERTURF: I'm interpreting it as one question, so ‑‑

DR. WORD: Okay. Because I was, like, there was one question on the floor.

CHAIRMAN OVERTURF: I can restate that if you want, yes.

Dr. Palese.

DR. PALESE: I would like to sort of see the word "provisional" decision, and to me that is like deferring. But I think it would ‑‑ I think we should really make the final decision on March 17th.

Now, I don't know what the Chair ‑‑ how he counts my vote then.

(Laughter.)

CHAIRMAN OVERTURF: I can discuss it with you later.

DR. MONTO: Arnold Monto. I agree with Dr. Dowdle's revision of Dr. Eickhoff's original motion.

CHAIRMAN OVERTURF: Dr. Eickhoff.

DR. EICKHOFF: I agree.

CHAIRMAN OVERTURF: If you understand this better than we do, Dr. Decker, you are willing to go ahead and try and interpret it. We'd like your opinion.

DR. DECKER: No better, but hopefully no worse. The ‑‑ once again, industry is grateful to the committee for its efforts.

CHAIRMAN OVERTURF: Well, I think we are actually scheduled for a break, and we are about 45 minutes early. The break was scheduled for 15 minutes, so I will plan to go ahead and take a break now and reconvene at 10:15.

Thank you very much.

(Whereupon, the proceedings in the foregoing matter went off the record at 9:59 a.m. and went back on the record at 10:25 a.m.)

DR. FREAS: As we take our seats, I just want to let the committee members know that, again, we have to schedule meetings months in advance. And so on your desk will be the Federal Register announcement of the teleconference on March 17th, and, of course, it's scheduled from 1:00 to 5:00 ‑‑ 1:00 a.m.?

(Laughter.)

It's scheduled from 1:00 p.m. to 5:00 p.m., and it's on this little calendar as well. So the date and the time are set. It will not last as long as we initially anticipated last February, but please mark that on your individual calendars.

CHAIRMAN OVERTURF: Dr. Gellin would like to clarify an issue that he brought up.

MR. GELLIN: As far as data that may be hiding, I think that the ‑‑ I think that what I said before may have been misinterpreted, because we had a pretty robust discussion about how the world gets its data, where it's been, the processing thereof. And, therefore, the idea that there is data out there hiding seemed to me, you know, remote at best. So what we're really talking about is, you know, the review of incremental data that comes forward.

So my intention was, really, to ‑‑ to clarify that I thought that we would have little to talk about at the next meeting, not that there was somebody with a big closet full of stuff out there. And I won't elaborate on people who do find closets full of things.

Thank you.

CHAIRMAN OVERTURF: The other discussion that came up during the informal discussion during the break was the issue about the quadravalent vaccine again, and I guess sometime we need to ‑‑ to reintroduce it.

It sounds to me like if we were to consider a quadravalent vaccine we'd have to first of all make that for a recommendation for a future year, and make it a generalized recommendation, and then would have to consider strains each time, if we were going to maintain production as we have in the past.

Yes, Dr. Decker.

DR. DECKER: Well, and if you're serious about it, the other thing you would have to have is ‑‑ I think is a commitment that you would then stick with a quadravalent for a decade, or something like that, because as Dr. Markovitz pointed out, if industry knows that the demand will be there and has time to adapt, the capacity can be built. But nobody builds a factory that's not going to be used year after next. So there's got to be the commitment to sustain the capacity.

CHAIRMAN OVERTURF: I assume there is no further discussion, then.

The next issue on the topic is, again, an issue that's been brought up a number of times and was discussed recently, again, at the recent NVAC meetings, and that's the use of mammalian tissue culture for reference strain isolation.

So the FDA has invited a discussion about that this year, and we'll start out with Dr. Ye.

DR. YE: Good morning. I think probably we remember that almost exactly one year ago A/Fujian/411 virus was not chosen as the vaccine strain. Part of the reason is that this virus was isolated from cell culture rather than from eggs.

So today's talk we tried to ‑‑ and I try to explain why this virus was not chosen, and is there any possible solution for not using this cell culture driven virus for the vaccine.

And this presentation actually was based on a preparation ‑‑ a position paper initiated by our Division Director, Jerry Weir, and had input by others. And this ‑‑ my ‑‑ the title of my presentation is, "The Preparation of Reference Influenza Viruses in Mammalian Cells," which is what ‑‑ FDA's perspective.

There are two objectives for this presentation. The first one is to review the issues associated with the source of the reference influenza viruses. Another objective is to discuss the possible strategies for use in ‑‑ for using mammalian cell lines to prepare influenza reference viruses.

I think since the 1940s inactivated influenza viruses vaccine has been prepared in eggs. The reason is that by using eggs we have a good production of influenza vaccine, which eggs provide the reasonable virus yield.

The candidate vaccine viruses originally isolate in surveillance labs use eggs as opposed to use mammalian cells, which is a global consensus within WHO national regulatory authorities, as well as manufacturers.

However, over the time, many surveillance labs has come to rely on ‑‑ use cells for isolation of influenza viruses from clinical samples, for the reasons of robustness and convenience, even though such isolation are not used as reference viruses.

And this slide shows the isolation of circulation viruses from clinical samples in national influenza centers all over the world. There are two methods for isolation of influenza viruses from clinical samples. One is use cell culture.

Such cells are not qualified or certified for ‑‑ there are three cells that have been used for isolation of influenza viruses from clinical samples. One is MDCK cells, which is canine kidney cells. And these cells have been used for quite some time for isolation of propagation of the influenza viruses.

The second one is VERO cells. This cell is monkey kidney cells, recently introduced for isolation or propagation of the influenza viruses. Only a few labs ‑‑ surveillance labs ‑‑ use such cells for isolation of influenza viruses from clinical specimens.

The last ‑‑ for the MDCK cells and the VERO cells are continuous cell lines. And the last one is the chicken embryo kidney cells, which is the primary cell lines. Only a few labs use these cells for isolation of a virus.

And the last one ‑‑ next one is eggs. In the beginning of the influenza vaccine production, the majority of the labs used eggs for isolation of influenza viruses, because of the cell ‑‑ the eggs that you have to ‑‑ the limited supply of the eggs and also the convenience of isolation of the influenza viruses.

So fewer and fewer labs use this method for isolation of the viruses. Actually, in the United States, there are no labs or surveillance labs that use this method for isolation of influenza viruses from clinical samples.

This slide shows the isolation rate of influenza viruses from clinical samples. This clinical sample has been prescreened by using more sensitive methods, such as immunofluorescence and rtPCR. And the table shows the isolation rate of the influenza viruses from prescreened clinical samples.

And the Y bar shows the percentage of isolation, and X bar shows the different method. The first two shows the ‑‑ from MDCK or VERO cells, and the last one is for eggs. And the red bar shows the isolation rate for Type A influenza viruses, and the yellow bar shows Type B.

As you can see, the MDCK cells and the VERO cells ‑‑ there is not too much difference in terms of the sensitivity of isolation of the virus, even though MDCK cells is relatively sensitive than VERO cells. But overall the two cell lines looks similar for isolation of both viruses.

The yellow bar shows the B viruses isolation. As you can see, there is not too much difference between cell lines and eggs for isolation of clinical samples. Where there is dramatic difference in isolation of Type A influenza viruses, you can see that only less than 20 percent of the isolation rate for A in eggs, where more than 60 percent of the isolation rate for Influenza A viruses. So this is why majority of surveillance labs use cell line for isolation rather than use eggs.

Because we use ‑‑ we use cells for isolation of influenza viruses from clinical samples, there are some consequences. Yearly strain selection can be negatively affected by requirement for egg isolates as seed viruses, which resulted in delaying recommendation for vaccine composition, such as last year, and also resulted in altering composition for vaccine strain because of availabilities.

In addition, a delay in availability of strain selection during the pandemic situation, it could be disastrous consequences.

This slide shows the licensed influenza vaccine in United States. The two licensed influenza vaccines in the United States ‑‑ first one is inactivated influenza virus vaccine, which commercially available for more than 50 years. And this vaccine is produced in eggs and delivered by parenteral injection.

And the next one is a live attenuated influenza virus vaccine, which is recently licensed. And this vaccine produced in specific pathogen-free eggs rather than regular eggs, and delivered by intranasal inoculation.

This slide shows the influenza vaccine under development, divided into two major categories. One is a cell-based, inactivated virus vaccine in MDCK cells or in VERO cells, which I just mentioned before. And another one is DNA vaccine or recombinant protein such as to ‑‑ to express the protein in virus protein in insect cells.

Okay. Now I'll just go through the reference and seed virus development. The first WHO global surveillance system, such as national influenza virus ‑‑ influenza centers in ‑‑ all over the world to provide the reference of viruses which isolated from clinical samples use eggs as opposed to using cells.

Then, international influenza center would provide reference viruses to WHO collaborating centers, such as CDC in Atlanta, London, Melbourne in Australia, or Tokyo in Japan. In WHO collaborating center, they provide reference viruses, include high growth reassortant viruses to manufacturers.

Then, manufacturers will develop preparatory seed viruses from egg isolate, which are recommended by WHO or local authorities. And, finally, regulatory agencies will accept the reference of viruses and approve seed viruses for use in licensed vaccine preparation.

And this slide shows the summary of the preparation of reference and seed viruses. As you can see from here, to provide a seed ‑‑ reference of viruses or seed viruses, it required a teamwork of globally. The first one ‑‑ WHO surveillance centers in national influenza center will provide the virus which isolated from clinical samples from using eggs.

Then, they sent this virus ‑‑ reference viruses to WHO collaborating centers. If this reference virus recommended is ‑‑ the vaccines ‑‑ vaccine candidates, then they will send this to manufacturers as well as to regulatory agents.

Manufacturers then will develop their own preparatory seed viruses, and then sent back to WHO collaborating centers or back to ‑‑ or to regulatory agents for evaluation of the seed viruses.

So this slide shows that, really, to develop seed viruses or reference viruses it requires a lot of, you know, organizations work.

And this slide shows why we use eggs for isolation of influenza seed strain. As I mentioned that this is a safety ‑‑ egg isolates has a safety record for more than 50 years, and possible exclusion of some adventitious agents from clinical samples, because using eggs rather than some ‑‑ adventitious agents may grow well in tissue cultures but not grow well in eggs, and also leading to high yield in vaccine production.

In addition, this method has been validated for inactivation of common chicken viruses in vaccine process.

Now I show to you the ‑‑ the pros and cons for consideration of mammalian cell lines for isolation of seed strain. The problem for using mammalian cells for isolation of influenza viruses from clinical specimens is ‑‑ are following. The first shows the pros, the advantage. It's more robust than eggs for primary isolation of Influenza A viruses from clinical samples ‑‑ I mentioned to you earlier.

Secondly, it's less selective pressure than in eggs for isolation of influenza viruses from ‑‑ with alternative receptor specificity. In general, influenza virus, like other viruses, it does not like switch from one species to another one. They have a sacrifice to adapt in the different species.

So by ‑‑ by ‑‑ because of human sample, the virus originally grow in the human samples, will more likely grow well in the same mammalian cells, so they have less pressure for the antigenicity variation.

Another reason is a practical reason, because of majority of WHO global surveillance centers have such cells for isolation. So they are familiar with and they are comfortable for use in such cells for isolation of the virus.

Now the problem ‑‑ the problem is, of course, that we have limited experience in influenza vaccine development and production. One such as the isolation ‑‑ the isolation of the virus or the isolates may not necessarily grow well in eggs, because they are in mammalian cells first.

Then, more importantly, there are some issues related to the use of mammalian cells for isolation of seed viruses for the possible contamination of adventitious reagents, as I mentioned that a lot of a virus will grow well in mammalians cells rather than in eggs.

And this slide shows the regulatory requirement and guidance for ‑‑ for the related ‑‑ related to use of mammalian cell lines. This ‑‑ the first one is 21 CFR 610.18 regarding cell ‑‑ use of cell lines for manufacturing biological products.

Even ‑‑ although this guidance is ‑‑ is for using cell lines for vaccine production, but what I'm ‑‑ I'm talking about is the use of cells for isolation of influenza viruses, but they share same concern, because of use ‑‑ even though you use a cell for isolation viruses rather than use the cells for production of flu vaccine.

The general requirement for the cell line is to have to ‑‑ the cell line has to be identified by the history, such as where the cells come from from a dog or from a monkey, and also have to show the pathogenic history, what condition of the cell is being passed, is there any possibility introduce advantageous into it, what kind of lab condition you are using to maintain such cell line.

Another thing is to have ‑‑ to describe with respect to the cytogenic characteristics, such as the chromosomes ‑‑ how many chromosomes and the morphology of such cells as well as tumorigenicity, and also includes oncogenicity of the DNA from these cells, and also have to be characterized with respect to in vitro growth characterization and the type ‑‑ life potential such as, you know, it takes how many hours to divide it and what's the time expended, how many passages you pass before the cell dies or it can be passed forever, so on and so forth, and have a test for the percent of detectable adventitious agents.

And more details of the regulation ‑‑ regulatory requirement and guidance is the point to consider in characterization and cell line used for production for the product ‑‑ for producing biologicals. And this is published in 1993; however, currently under the revision by Office of Vaccine ‑‑ Vaccines and Research and Review, and which provide relatively detailed and additional guidance on preparations and standards for characterization and the qualification of cell substrates and virus seeds.

An additional regulatory requirement and guidance is as follows. One is WHO technical reports series regarding requirement for biological substance, and also the ICH guidance, QC5, has some details regarding the ‑‑ using cells for production of influenza vaccine or the evaluation of the cell lines.

Another one is European Medicine Evaluation Agency guidance, and also provides some information regarding the ‑‑ using the cell line for influenza vaccine production.

Okay. Now we move on to the strategies for consideration of use of mammalian cell lines for isolation of reference viruses. The first one actually kind of ‑‑ we are ‑‑ is we are using a similar method for H5 ‑‑ H5N1 virus isolation. First is retain the clinical samples of the initial isolation in national influenza centers, and then forward the samples to select ‑‑ select WHO collaborating center for reisolation in a qualified cell line.

As I mentioned, this is similar to the current procedure, except for using a cell line for reisolation rather than use eggs.

Of course, in order to achieve this, we have to have a suitable cell line for reisolation, so preferably to cell bank would be produced and qualified for distribution to selected centers for reisolation.

And since the original samples directly cultured or in the ‑‑ in the cells, so any contamination in the original samples may amplify in the cell lines. So the further purification and adaptation of a cell line for vaccine production in eggs could be considered, if necessary.

The second one is the use of reference viruses direct isolated on the cell line which are not characterized or qualified. So as you can imagine, the cell line is not qualified, neither qualified or characterized, so any contamination will be there will be ‑‑ will be amplified. So it would require the purification of reference viruses to be acceptable.

For instance, you know, possible plaque purification on the specific pathogen-free chicken kidney cells, and have to be evaluated if the contaminate or potential viruses can be eliminated by such purification. And, of course, the resources and the infrastructure for purification would be necessary, because who is going to do it and going to evaluate such method.

The third one is to use the reference viruses isolate in characterized and qualified cell line. Of course, in ‑‑ to achieve this, it would require the production and qualification of preferably the cell bank for distribution to national influenza center.

Remember that national influenza center, their purpose is not just for isolation of influenza viruses, those other viruses, and maybe have some other viruses was, you know, handled in the same lab. So the contamination would be quite high if they do not handle cell well.

So the first rule is we have to choose the cell line such as MDCK cells or VERO cells, other cells such as MRC5 cells for qualification, and then, more importantly, the maintenance of the quality control of the distribution to avoid additional ‑‑ introduce adventitious agents into the qualified cell line.

Then, the further purification or adaptation of cell isolates for vaccine production in eggs wouldn't be necessary. It would be considered if it's necessary.

And the last one is to use the reverse genetic method and qualified cell line to generate new reference viruses. You may remember yesterday from Nancy and Phil, as well as Linda Lambert's talk, to use reverse genetics for preparation of 85 N 1 for pandemic preparation. They used this ‑‑ such method.

So this ‑‑ of course this would require the production and qualification of cell bank for reverse genetics, because we use these cells for licensed vaccine production rather than for pandemic preparation.

Another one is by using this method, I think it's ‑‑ because when you extract viral RNA for ‑‑ during reverse genetics, you really use a very harsh way to purify the RNA from the viruses. So a lot of contamination usually is limited by this procedure. So once you purify the RNA for construct ‑‑ the construct for reverse genetics, so usually it's quite a good way to eliminate concern regarding the contamination of the original sources.

And, of course, the problem will be accepted on the technology and intellectual property issues, which may not permit near ‑‑ in the near term.

All right. The conclusion is that the use of influenza seeds viruses obtained from non-egg sources may be acceptable if adventitious agents in the original isolation or in the cell lines used for isolation can be addressed.

A second one is investment in the resources and infrastructure is necessary.

And the last ‑‑ lastly, because ‑‑ I mentioned these are global issues, you know, to ‑‑ we have to have consensus ‑‑ global consensus, which we seen WHO, national regulatory authorities, as well as the manufacturers, regard ‑‑ regarding use ‑‑ regarding the issues to be addressed for implementation of this method.

Thank you.

CHAIRMAN OVERTURF: Are there questions or discussion for ‑‑ yes, Dr. Markovitz.

DR. MARKOVITZ: Yes. I was curious if you could just describe in a little more scientific detail exactly how you certify a cell line. I mean, I understood the general principles, but what do you actually do? I mean, what are the techniques involved?

DR. YE: To certify a cell line, as I mentioned in the guidance, first you have to ‑‑ to show the cells is really what you're talking about. If it's from a canine, then you have to show this is from a canine.

In some cases in the lab or in the ‑‑ in organization, when they pass the cells sometimes they release some cells which are contaminated and ‑‑ but not from canines, some cells such as the contaminant, and they have to show that, really, the cells you're talking about.

And, secondly, if ‑‑

DR. MARKOVITZ: I'm sorry. Do you do that genetically? I mean, you ‑‑

DR. YE: You can ‑‑

DR. MARKOVITZ: Do you sequence part of the DNA of the cell?

DR. YE: You can sequence it, and there is some enzyme you can detect. They have a difference in ‑‑ between different species, so you can tell the ‑‑ you know, some enzyme approach, and you can identify to tell difference. And also, have to be addressed, any contaminations because some viruses may not be detectable by current method, but is hidden in the cell lines, but may be passed on through the vaccine or the virus ‑‑ the seeds viruses.

So you have to show the ‑‑ you really handle the cell lines well. And also, and the ‑‑ if the cells cause a tumor, or the DNA causes a tumor as well, and also there are other viruses, you know, in ‑‑ integrated into the DNA chromosome of the cells, you have to ‑‑

DR. MARKOVITZ: How do you do that, though? I mean, how ‑‑ because which viruses do people look for, and how do you do it?

DR. YE: Here I cannot ‑‑ it's this thick ‑‑ page by page to go through that. I ‑‑

DR. MARKOVITZ: No. But, I mean, just in general, do you do a PCR looking for known viral sequences? Or what do you actually do to eliminate it? Because that's obviously a huge concern with this.

DR. YE: There is a huge protocol to evaluate it ‑‑ say, use animal ‑‑ newborn animal to show if this causes tumor, just by inject the cells or you isolate it, DNA to the animals, see if anything happened, or culture in the cells, sensitive cells, depending upon which viruses you are looking for. So you have to use the sensible ‑‑ suitable cell lines for particular virus.

DR. MARKOVITZ: Maybe ‑‑ but which viruses, though, are people actually looking for when they ‑‑ it can't be that long of a list. What ‑‑

DR. YE: I probably have to refer this to my colleagues.

(Laughter.)

DR. MARKOVITZ: Oh, I guess it can be a long list.

DR. YE: There's a lot ‑‑ a lot of ‑‑

DR. MARKOVITZ: So how long is the list?

DR. YE: I have a short memory, but I cannot remember at all.

DR. MARKOVITZ: How long is the list exactly? And how do you do it?

DR. YE: As long as you can list.

DR. MARKOVITZ: So what do you do practically. I mean ‑‑

DR. YE: Yes.

CHAIRMAN OVERTURF: I think Dr. Levandowski would like to try to clarify it.

DR. LEVANDOWSKI: Right. I think Zhiping on his slide where he indicated the CFR position on this, it sort of shows you generally what needs to be done. It doesn't tell you specifically, because there may be more ‑‑ more than one way to skin a cat. But generally identified by history ‑‑ as you mentioned, you start out by saying where you got the cell line and how it was handled before you got it, if there's anything known about it.

Most places ‑‑ most people get their cell lines either from another laboratory or they get it from ATCC or from the European culture collection, or one of the standard culture collections. So that's how you start out.

But there's a lot of information that goes with that in the passage history as to what media were used to propagate the cells throughout their life, what other excipients might have been present, and all of those things. And all of those things in the documentation are part of the information that is being looked for to understand what might have happened to these cells or where they actually came from.

And it can't ‑‑ simply that information can be enough to tell you whether the cell line is appropriate or not. But he also mentioned or went over the fact that these need to be described by cytogenic characteristics, chromosome analysis, isoenzyme analysis to differentiate species. There are a number of ways to do that chemically and biochemically, and these days you probably could use genetic techniques to try to get to some of that answer.

And then, you know, finally, the information about detectable microbial agents ‑‑ how long is the list? The list is as long as you'd like to make it. There are both biological and other types of methods like PCR to try to get to information about what type of contaminating microbial agents might be present.

So some non-specific tests are done that are meant to catch any kind of virus that could replicate in a tissue culture. Usually there are several tissue cultures ‑‑ certified tissue culture cell lines that are used to try to replicate viruses that might be present.

And there are ways to try to show that viruses that are inferring might be there, you know, that don't necessarily show cytopathic effect but may be able to show, by another method, that they interfere with a marker virus.

And then you can use the ‑‑

DR. MARKOVITZ: How do you do that? Excuse me. How do you do that?

DR. LEVANDOWSKI: You inoculate the tissue culture with your interfering virus, and then see whether you get ‑‑ actually, you inoculate the tissue culture with your non-interfering virus and see if you get heme absorption ‑‑ or hemagglutination, for example, is one way to do it for that particular marker virus that you're using, to see whether there's interference.

There are also inoculation of animals. I mean, it's a fairly extensive protocol, as Zhiping said. It's not a simple matter to do this qualification or certification.

DR. MARKOVITZ: Yes. I mean, the reason I'm asking ‑‑ I'm not an alarmist about this or anything, but you know when you hear things are certified the immediate question is, well, what makes these really safer than anything else? And obviously, there are going to be some viruses nobody ever knows about, but ‑‑ but the question I guess really is, which ‑‑ you know, I mean, apparently this is something other people know, but, I mean, how many viruses really ‑‑ I mean, so there's two questions.

One is you're actually, you know, trying to look for any virus, any cytopathic activity, right? So, but the other question is, what viruses in particular might one actually find in these cells that would eliminate them? I mean, is there like ‑‑ are there five ‑‑

DR. YE: I think ‑‑

DR. MARKOVITZ: ‑‑ or how does that work?

DR. YE: In general, it's by using current method you can ‑‑ as much as you can to detect as much as possible of the virus you could do it. So I think to make it safe down the road.

You may not know now, and later after a few years we have a new method and say, "Hey, there is X virus in there, and it causes this in humans." So I think we'd rather do more complex work in the beginning to ensure down the road we ‑‑ you know, that we do not introduce some viruses to the human by ‑‑ through the vaccine.

DR. MARKOVITZ: What do you want to see in the chromosomes, by the way, Roland? Were you hoping for 46 or what ‑‑ what's the ‑‑ what are you looking for in chromosomes?

DR. LEVANDOWSKI: You're hoping for the number that it's supposed to have for the particular species it should be. So if it's not the right number for the species ‑‑

DR. YE: Right.

DR. LEVANDOWSKI: ‑‑ it's not the right cell line.

DR. YE: Right. Different cells have different ‑‑ usually, the cell line is a continued cell line. That ‑‑ you know, it's totally off. It's not regular ‑‑ they don't have regular chromosomes there.

DR. MARKOVITZ: A lot of cell lines, though, have ‑‑ you know, that are in common use have a lot of chromosomes. So that's I guess ‑‑ sure, you're trying to check to make sure it's the right species, but do you want ‑‑ I mean, how much ‑‑ when you're certifying something, do you tolerate extra chromosomes?

DR. LEVANDOWSKI: It need to be explained. I mean, there may be other things that happened in handling the cells as well, so there ‑‑ if there are extra chromosomes out of what should be expected for the description of the cell, then you, again, question whether it's actually what it's supposed to be.

DR. MARKOVITZ: No, I don't mean that. I mean, when you're talking about safety issues, one of the questions is, you know, is there some sort of oncogenic virus or whatever, or transformed cell that in some way might be dangerous? And the question is: are there ‑‑ you know, is that okay to have extra chromosomes, as long as it matches what the cell line is supposed to be.

DR. YE: Well, I ‑‑

CHAIRMAN OVERTURF: Sorry. There's ‑‑ Doctor? Yes. There's probably a switch at the top there.

DR. EGAN: Okay. Sorry. Bill Egan from Office of Vaccines. I think one of the problems is one can't give you a simple exact answer about what you need to do that's going to cover every single situation. Obviously, it's impossible to test for every single virus that ever existed. That you can't do.

But if you're, for example, going to use a mammalian cell line, you're certainly going to test for Hepatitis A, Hepatitis B, Hepatitis C, HIV. You're probably going to do ‑‑ and you're going to be doing those by PCR.

We generally are going to test for retroviruses. You're going to do reverse transcriptase PCR analysis to look for that. There are ‑‑ you'll be doing tissue culture to look for any cytopathic effects for unknowns. So there's a variety of things. Some cells are continuous, and they're going to have a distribution of chromosomes, and they'll be in a certain range because tumorigenicity, for example, of VERO cells are going to change by the number of passages.

So you ‑‑ so what you're going to look at, and the level of concern, is going to be specific to a particular situation. Now, in addition to all ‑‑ you know, if you have a vaccine produced in the cell and you can't guarantee some certain clearance of cells from the final product, then you certainly want to know something about the tumorigenicity of those cells and the oncogenicity of those cells, looking at, you know, the cells, and looking at DNA.

They're probably looking at DNA as a function of size. There's a whole variety of things, you know, that you're going to do depending on those cells, what they're used for, what you can do during the manufacturing process. Whether something is inactivated or not inactivated, what you can validate would be clear during that inactivation.

That's one of the reasons why we're very concerned also about this history. You know, what have they been through will give you some idea of what they could be contaminated with that you might want to look at. What else has been in the lab?

We're also ‑‑ and this wasn't mentioned ‑‑ we're very ‑‑ one of the things we're very concerned about are the agents of the transmissible spongiform encephalopathies. So if you have a cell line that was grown that was using calf serum or fetal calf serum, where did that come from? Can you document it or not?

What else do you know about the cell line? If it were contaminated, is that cell line able to propagate the agent of that TSE? And on and on and on.

But the only way to really answer your question is to give ‑‑ come in very, very specific. For example, I want to use VERO cells four, and then we can discuss it. Otherwise, you know ‑‑ and the bottom line, it's going to be all driven by the science.

DR. MARKOVITZ: Thanks. That's what I was wanting to hear. Thanks.

CHAIRMAN OVERTURF: Dr. Dowdle, and then Dr. Monto.

DR. DOWDLE: In regard to the logistics of providing characterized or certified cells to the laboratory, so that the laboratories have those available, I would just like to ‑‑ to remind you that the polio virus network of 150 laboratories throughout the world have such a system in place now, so that every laboratory has a liquid nitrogen bank ‑‑ cell bank. Many of these laboratories, by the way, are in developing countries. In fact, most are in developing countries.

The cells are provided by NIBSC, and Phil, in fact, may want to comment on this later in his discussion. These are highly characterized cells. They are set up ‑‑ the labs are set up in such a way that there's a very high QC component of the work, so that the cells are actually used only for 15 passages, discarded, they start again to cut down mycoplasma. All of these issues have been taken into consideration.

Now, you'd have to notch it up a little bit in terms of the strains that are provided, but all I'm saying is the logistics are there. In fact, even now you could start using that through the polio system. But it's very highly controlled, and it can work. And the logistics ‑‑ the cost of this is relatively small. It's not as big as we might think that it is.

DR. YE: Thank you. I think you addressed a good question for practical reasons. It's achievable if the lab do what it's supposed to do to maintain the cells, the cell bank for the isolation. So it is achievable, so ‑‑ but in order to do it, we have to evaluate it to make sure the cells are properly handled and no additional virus is going to be introduced into it.

CHAIRMAN OVERTURF: Did you respond specifically to that, Dr. Egan?

DR. EGAN: Stick this thing back on. Bill Egan. Yes. The issue, of course, here that may be a little bit different is we're talking about a virus that's going to be used for further manufacture. I mean, what you said is perfectly reasonable, and it is doable. But, you know, it's ‑‑ you have to set it up, and you have to continue to monitor.

So the other approach is to have, you know, a small group of centers that would just simply reisolate. And if you're talking about plaque purification or something like that, it's not unduly burdensome to set that up, and, you know, which becomes the easier system, you know, would ‑‑ that might be possible. You know, we could just, you know, see.

CHAIRMAN OVERTURF: Dr. Monto.

DR. MONTO: I think in examining the various options we need to bear in mind the logistics of reisolation, retaining of specimens, sending specimens for reisolation to central laboratories, presence of certified cell lines at national influenza centers, with the assurance that they are being handled appropriately for isolation, etcetera.

I think it's clear that we do have one option which would be ideal if intellectual property issues are taken care of, which is, I don't think, our consideration right now. But there is another one that is on the table ‑‑ number two ‑‑ that I think we need some further information about how easy it is taking strains directly isolated in cell cultures, unapproved cell cultures, to purify them using existing approved techniques and readying them for vaccine production. How easy would that be? What would the timeline be in your estimation?

DR. YE: I think we're really ‑‑ you're talking about option number two, right?

DR. MONTO: Yes.

DR. YE: Because I have ‑‑ over here it's hard to hear the question. As you can see, the cell is not certified, and then there's a possibility the contamination will be not only from the original sample, but also the cell itself. You don't know what ‑‑ what's going on in that cell, so extensive purification needs to be validated to make sure, say, if you spike a model virus you can show this virus can be eliminated by plaque purification.

So it's ‑‑ a lot of work needs to be done to ensure that no ‑‑ the virus has not, you know, contaminated in your seed virus. Probably is a short answer.

CHAIRMAN OVERTURF: Dr. Levandowski.

DR. LEVANDOWSKI: I just wanted to remind everybody that we are anxious to hear from the committee all their ideas and options. We're not sure that we have all of the answers here. We've laid out some considerations, but maybe there are others that the committee would have also. And, you know, just ‑‑ we really do want to hear what you all are thinking about in terms of what we've laid out here.

DR. YE: I think, as Roland Levandowski is ‑‑ as he pointed out, we are not just providing answers. Actually, we are, you know, asking for more questions or input. So it's maybe beyond that four options. Maybe if you have more reasonable, you know, strategies can be, you know, under discussion.

CHAIRMAN OVERTURF: Do you have a question, Dr. Karron?

DR. KARRON: Actually, a couple of things. First, I just wanted to clarify, Arnold, what you said when you said that there's an ideal option, but there are IP issues. I assume you're talking about number four, but I want to, one, make sure that that's the case.

DR. MONTO: Yes, I was.

DR. KARRON: Okay. And then I guess my second question has to do with, as you mentioned, we're currently using strategy number 1 for H5N1. You did say that in your talk. Is that ‑‑

DR. YE: Yes. But I don't know if it was the right thing to say or not. But it's kind of different, because you use the qualified cell line for pandemic ‑‑ okay. Go ahead, sorry.

DR. KARRON: I understand that. I guess my question would be I'd like to know, one, how well that system is working for isolation of pandemic strains in practice. And to have some discussion about what the barriers would be. I assume key barriers for implementation of that kind of a strategy for use for non-pandemic influenza.

DR. YE: I cannot answer the question on the IP issues. But in terms of using reverse genetics, I think it's ‑‑ as Phil Minor talked about yesterday, that is a wonderful method, and you can get what ‑‑ any kind of ‑‑ the virus in any condition can ‑‑ you can isolate ‑‑ go ahead.

DR. EGAN: I was going to say maybe have Dr. Minor give his presentation and the NIBSC perspective, and then come back to this issue more globally on discussion of what people think they might be able to do, what things might be acceptable, what are the pitfalls of various technologies, whether it's even possible to consider using an isolate that came through an unqualified cell line or something like that.

DR. MONTO: Just a point of information. Aren't we using for the H5 number four as well, the reverse genetics?

DR. YE: As I understand it, yes. But that's ‑‑ it's for pandemic rather than use the licensed vaccine.

Nancy?

DR. COX: Just to clarify, for the H5 vaccine reference strain production, we started out with an egg isolate that was sent to us from one of the national influenza centers in Hong Kong. And so we had an egg isolate, and it's also possible in many cases to get the original clinical material and then obtain an egg isolate or a cell culture isolate from that original clinical material.

So it takes extra time and there are extra logistics involved, but then you can start out with something that you're not concerned about at all and go on to the next steps.

CHAIRMAN OVERTURF: Dr. Royal.

DR. ROYAL: Thank you. In discussing the various options for how to incorporate mammalian cell lines into isolating viruses, are we also looking at how the flow chart that you've shown in slide 11 would be impacted?

It appears as though ‑‑ well, I guess it's the fact that industry receives seed virus from both the coordinating centers and from the FDA I guess, from the regulatory agencies, and given the fact that in your cell lines you'll ‑‑ you have a more permissive system, you'll have potentially lots more isolates to deal with, how might the flow of material be impacted as it's collected and distributed?

DR. YE: This is ‑‑ the flow chart actually is for ‑‑ refers to the egg isolate. For cell isolates, maybe the flow chart will be different.

DR. ROYAL: Should that be discussed?

CHAIRMAN OVERTURF: I'm going to ask for one more question, and I think we should then probably proceed to Dr. Minor's presentation. But Dr. Myers has one more comment.

DR. MYERS: Yes, it's a comment more than a question. I think it's clear we need to expand our repertoire for being able to develop candidate seed viruses. But I think strategy number two, which was the one that we discussed at such length last year, is really not a practical, pragmatic strategy for the development of the seed virus in a timely ‑‑ timely fashion or without compromising the potential safety of a vaccine.

And I think that's ‑‑ so I don't think that's really a practical consideration as a strategy for seed virus development. I didn't think that last year either.

CHAIRMAN OVERTURF: With that thought, why don't we hear Dr. Minor's presentation, and then we can readdress the issue.

DR. MINOR: Okay. Thanks. Thanks very much, indeed. Vaccines are extraordinarily sensitive kind of biological agents, as I'm sure you're all aware. I mean, they are given to people who are healthy, and the intention at least is to keep them healthy.

So there's an enormously conservative approach to how vaccines are actually produced. And even a fairly straightforward kind of vaccine, like, say, extracellular pertussis or something like that, if you call that straightforward, is really approached with extreme caution and extreme care along the line.

And it's quite simple, if you like, compared to what you see with influenza, because an influenza vaccine is ‑‑ firstly, it's a new product every year, because sooner or later somebody is going to update something at some stage, so you really have to consider the whole thing one a yearly basis. So many of the vaccines that are around at the moment have been going for 20, 30 years, and so on. Okay? So you have a lot of experience in how to do them.

Flu vaccines are far more complicated and far more ‑‑ far more delicate, if you'd like. So you really have to be aware of the fact that there is a deep conservatism in the vaccine industry.

As a truism, influenza vaccines depend on the isolates. If you haven't got the isolates, you haven't got the vaccines, and that's the way it works at the moment. And they depend on viable isolates at the moment as well. It may be that with reverse genetics you could actually get away from viable isolates, but you're not in that stage yet by any means. Okay?

The isolates that you get may have other viruses present in them, and they may have come out of it because of viruses that are coexisting in the nose of the patient from which you got the original swab. They may also have viruses in them resulting from their passage as they've been carried in the field and as they've appeared in the various laboratories that you just saw in the ‑‑ on some of the previous slides. They may have picked something up along the way. Okay.

Now, the reason why I think people are quite happy with flu vaccines is that there's a great deal of experience with them. Over about 50 years they've been produced in eggs, and there really doesn't seem to be much of a problem with them. I remember having a serious discussion in a bar with a manufacturer once where I said, you know, the good thing about flu vaccines is that, you know, they seem to be reasonably okay.

And he thought this was quite funny. It was a good, appetizing jingle. You know, flu vaccines, we haven't killed many people, you know.

(Laughter.)

I'm not sure it would be quite so good as that. I mean, it's better than that. It's better than that.

I mean, they have been used for over 50 years, and they have a good track record. I'm surprised, you know, they don't do much damage. So whatever you're doing it's okay. Right?

The fact is that, quite honestly, you don't know what you're doing. You're taking materials from patients who are out in the field who ‑‑ who have Lord knows what living up their noses, and you're injecting that into eggs, and most of these eggs that the isolates would be done in are not specified pathogen-free, so there may be other things growing in there. Okay?

There may be contamination of the product by other viruses, but so far as you can tell this has not been a problem. Okay? And by the time you get to the end product, it's not a problem. Fifty years worth of clinical experience says that this is a very safe product. Potentially it's a very effective product that you don't actually see the kind of difficulties that you have.

But it's a very, very complicated and complex chain of events that leads from somebody having flu to somebody actually having a vaccine which then goes into somebody else along the line. Okay?

And the regulatory view ‑‑ at least the NIBSC perspective ‑‑ is that if that's what you've got, a very complicated system where you're not quite sure what's going on, if you change anything in the procedure you've really got to be very, very careful indeed. You must be very cautious about any changes that you put in there. Okay?

Now, in the United Kingdom we had a certain amount of discussion in the clinical virology area about what kind of cells or systems people were actually using for detecting viral ‑‑ virus and disease, if you'd like. And you have to bear in mind that many of these clinical labs at least are not just concerned with isolating flu.

I mean, they are also interested in isolating other kind of respiratory viruses as well. And it may be that there are other kind of cells which are more suitable for isolating other respiratory viruses. There are certainly other kinds of cells which are more suitable for isolating enteric viruses.

There's a cell line called L20B, which is one of the things used in the polio network that Walt just mentioned, which is a mouse cell transected with a polio receptor site. Really, it's extreme good for isolating polio, but it's pretty useless for isolating anything else. Okay?

So depending on what you're trying to do, you will use a different kind of cell type. And in the United Kingdom what used to happen was that there were primary monkey kidney cells that were actually prepared and distributed from a central source, more or less at least, as a very, very good, highly sensitive catch-all kind of cell culture system that clinical virology labs could use for anything. Okay?

I think that supply has more or less dried up now, but, I mean, I may be wrong on that. Maybe Marie can comment on that if necessary.

There are other things ‑‑ MDCKs and VEROs, which were just mentioned, and possibly other cells as well. This is just from the United Kingdom where we have a rough idea of what we can actually look at to see what's being isolated. Out in the wide world I don't know what they use, although MDCK and VEROs are clearly in there, and kidneys as well. There may be other things as well. Okay?

And I'm not sure that I would want to handle a virus which had been isolated on a baboon kidney somewhere in the middle of Africa, for example. I'm not quite sure what else would actually be in there. Okay? So I think that's an issue that you might want to bear in mind.

The other thing is, as well as saying you have different cells to actually look at, the way the cells are carried is ‑‑ is really quite variable, if you'd like. It was pointed out that you have to ‑‑ if you're looking at a certified cell line, you have to have some characterization on it to show that it's what you think it is.

Cells lose sensitivity on passage. This is well documented and well known, and it may well be a real phenomenon. It may also be that they lose sensitivity because they get contaminated with things like mycoplasma, for example. There are viruses which may be present in serum, which they could actually pick up and become infected with in a totally subclinical way, if you'd like. So in a way that you wouldn't actually see that they were infected, but nonetheless the virus would be in there. Things like BVDV, for example, can possibly do this.

And there's certainly a porcine kidney line, which people have used for titrating yellow fever vaccine in ‑‑ which is chronically infected with the hog cholera virus. Okay. Nobody knows ‑‑ nobody cares about this, because they know about it. But there may be cell lines out there which ‑‑ which have similar kinds of viruses in them which you don't know about. Okay.

What's more, my cell is not necessarily the same as your cell. My MDCK cell is not necessarily the same as yours. My VERO is not the same as yours, and so on. Okay?

They also require a great deal of skill in maintenance, and this was something which emerged I think in the UK network. I mean, that ‑‑ it really was a ‑‑ quite an effort for the individual labs to maintain their own cell lines. And what has been tried at least is to send out cell cultures from a central facility and really that will improve things enormously. So it really does say that there are difficulties in maintaining these cell lines in real life.

Now, the polio network that Walt referred to really seems to have quite a good handle on this and how you actually deal with it, and you go back to your reference preparations repeatedly. When the issue of flu isolates first came to my attention, which was about 10 years ago, I mean, I thought, why don't we just tap into the polio network. And I suggested this to WHO who said, "Well, there are actually different labs." And so it's like, you know, I mean, you ‑‑ so there's a certain amount of turf wars to go on here. Okay?

I mean, it does seem to me that you could use the polio network in precisely this way, and it seems a shocking waste to me that this beautiful system has been set up, okay, and you're not going to use it for other things as well. And maybe it will be in the long run, but I don't think it is at the moment, which is a bit of a shame. Okay.

Right. And they vary in quality, if not in name, as I say here.

There are a number of different vaccine types which are potentially in the offering. There are three here which I will talk about. The egg-grown inactivated vaccines are the most common, at least in Europe, and I guess over here as well. So eggs are the basis of the production, and you have to bear in mind that these eggs are almost certainly not specified pathogen-free.

So they will come from commercial flocks, if you'd like, they are not examined carefully to make sure that they are totally free of any virus that may be there. Okay? So this is actual routine production of influenza vaccine. And this is the way it's been going on for 50 years, and, as I said in my initial opening remarks, there will be no issues with that. There is no difficulty with that. It's perfectly okay to do this, because the clinical results are satisfactory. Okay?

I have to say, if this kind of product came out as a brand-new kind of product nowadays, I think it would have a bit of a rough time actually.

There are other issues. Cell culture-grown vaccines, which have been inactivated, which again was referred to in the previous presentation, the issue is here slightly different, because your cell cultures may actually be growing viruses that your eggs are not. Okay? I mean, the assumption that people have worked on is that the eggs act as a sort of microbiological filter to some extent, which may or may not be true, but nonetheless it may be true.

So if you're growing your materials in cell cultures, such as VEROs, you may be growing viruses which you wouldn't normally grow in an egg system, and, therefore, you would worry about it. At least you'd want to consider it very carefully.

And, finally, there's the egg-grown live vaccines, which I won't talk about.

One of the arguments that frequently comes back is, well, the product is inactivated anyway, so what's the problem? Now, the inactivation process that's used most commonly involves formaldehyde, and regulators who say inactivation of viruses by formaldehyde have a little shudder and they think SV40, which I'm going to mention in a minute. Okay?

But it's very good at inactivating influenza. In general, in my experience at least, the processes have not really been validated, at least in data that has been presented to me, to show that they will inactivate other viruses as well. Okay? And where it has been done it's rather less impressive than you'd like it to be. Okay?

So, I mean, there are precedents for this kind of partial activation of potential virus contaminants. Okay? So you can't necessarily rely on the fact that you've inactivated your virus with formaldehyde to say you got rid of every virus that might be there. Okay? You can't trust it. It helps, without a doubt, but you can't trust it. At least I don't trust it. You may, if you wish.

Finally, there are a couple of examples where ‑‑ where this I think becomes a real issue. I mean, you can say, well, it's a pretend issue because there may be viruses in there. But who cares? I mean, chickens aren't going to affect me too much, are they?

And here are two issues which repeatedly come back to bite the regulators on a regular basis more or less. In the 1950s, inactivated vaccine was produced according to the Jonas Salk method, where you treat the live virus preparation with limiting concentrations of formalin over a long period of time. Okay? And you have to be very careful how you balance the formalin and the treatment with inactivating the virus and not destroying the antigenic properties. So it's a very carefully controlled process. Okay?

The virus was at that time grown on rhesus macaques kidneys, and what was not known was that a virus called SV40, which is a polyomavirus, which causes tumors in rodents, and so on, was actually a very frequent silent contamination of these kind of monkeys. Okay? When you start moving toward the monkeys you get a cytopathic effect, and it becomes obvious that you shouldn't use that cell culture. Okay? But at this time, with the rhesus macaques, it's a silent infection and you couldn't actually see anything in particular.

SV40 is less easily inactivated by formalin, and that is polio. And what the consequence of that was that ‑‑ from about 1955 to 1962, probably everybody who had three shots of polio vaccine in this country, or in my country, or anywhere else, probably had a bit of live SV40 stuffed into them as well. Okay?

Now, you could say that's a plus or it's a minus. Okay? Because if the SV40 is inactivated, what you've done is you've basically immunized them against SV40, right? So they're not going to get a tumor, right? You could say it's a bad thing because they've been given live SV40 and they weren't banking on that. Okay.

For a long time there were some quite serious epidemiological studies done on this, which suggested that there was no issue involved, and that the studies weren't that terrific, frankly. They were too short term, and they involved too few people, and they didn't look at the people who have actually had the contaminated material. They looked at other stuff. Okay?

But nonetheless, there's a certain element of contentment about it that made it look like everybody got away with it. And then, in the 1990s, there were publications from Michele Carbone and others, many others in fact, to say that you can identify SV40 sequence in certain kinds of human tumors. And they include things like mesotheliomas and ependymomas and osteosarcomas, and stuff like that. Okay?

It's still unclear what this actually means, and it's still highly unclear where it came from. I personally doubt very much whether it came from the polio vaccines that we used in the 1950s, and they certainly don't come, in my view, from polio vaccines or anything else that's actually made now, because we have evidence that they are clean. Okay?

But nonetheless, it's an issue which has now come back to bite everybody all over again. Okay? And, frankly, if back in the 1950s you could have said, "We're going to use vaccines which have no SV40 in them," you would have saved yourself an awful lot of trouble over the last, what, 40, 50 years or so. Okay?

So regulators look at this and they shudder. Whenever you say formalin and SV40, they'll shudder at you. Okay?

The second thing is simian cytomegalovirus ‑‑ again, this is a ‑‑ this is an issue which has cropped up from time to time where it seemed unlikely somehow that simian cytomegalovirus was a common contaminant of polio. It seemed unlikely that if it was present it could possibly infect human beings. And on the other hand, when you start looking at polio vaccines, some of them at least, you can find simian cytomegalovirus sequences present in them, simply because of the cell cultures they are growing on primary monkey kidney cells that they are growing on. Okay?

And this was something which had been going on for some time that nobody really picked up on. Okay? But none of this is there, and it also appears that some of these viruses, for example, will infect human diploid cells, so they are able to infect human cells of some description or other. And that's really quite a recent event.

I don't, again, think that this is a public health issue, insofar as there was no infectivity associated with them. But nonetheless, this is a major issue, and I guess it's a public issue as well, a public opinion issue apart from anything else.

So regulatory approaches are really very, very cautious when it comes to the possible virology of what's going on here. And when you have a complex system such as the influenza isolate production and leading on to vaccine production, it will be examined extremely carefully I think to make sure that nothing too bad is going to happen.

And the conclusion, really, I have is here. That if you have a great deal of experience at something, based on the clinical experience, such as you had with an influenza vaccine over the last 50 years where you make your isolates on eggs and you pulse everything on eggs, and then you make your vaccine on eggs and you inactivate it, and so on, okay, and you know that you don't have much of a problem with that, that's a very, very good system to actually stick with, because you know you don't have much problem with it.

If you start changing things a little bit, I think you've got to make sure that you know what you're doing. So you either do it by clinical experience or by knowing what you're doing. Okay?

That's it. Thank you very much.

(Applause.)

CHAIRMAN OVERTURF: We can continue the discuss at this point, and Dr. Minor is available also for questions. Bruce.

DR. GELLIN: Can you buy qualified VERO cells? I mean, is there somebody who makes this stuff and you just go out and purchase it? Or does everybody have to do their own? How does this work?

DR. MINOR: Manufacturers have qualified VERO cells. Okay. You can probably go to a manufacturer and try and buy them from him. A lot of effort goes into making them qualified. You have to bear in mind that it's qualified when it leaves the manufacturer's premises, and you can wreck it just like that. Right?

(Laughter.)

Right? So just because you bought it as a qualified doesn't mean that by the time you've got it laid down as a bank in your system that it's qualified again. Right? Okay? Right?

CHAIRMAN OVERTURF: Yes, Dr. Markovitz.

DR. MARKOVITZ: I'm just curious ‑‑ this is probably naive, but is it not possible to either just buy or make qualified cell lines, get them to all of the centers that collect samples of influenza, and then at least hope that they don't, you know, goof them up, and then you would have the whole chain.

I know there's proprietary issues, but can't somebody develop ‑‑ I mean, can't the government develop its own qualified cell line? This may be naive, but it seems too straightforward to not at least ask.

DR. MINOR: I think ‑‑ I think you have to bear in mind there is a large number of labs involved in this, right, that you really want to be sure that you know what they're all doing. Okay? I think the example that Walter Dowdle referred to earlier of the polio network is actually a good one, because I think there you've got a real seriously disciplined bunch of people, actually, who are ‑‑ so in principle it can be done I guess. Okay?

If you think that a qualified cell line will actually last the 15 passages, which it probably will do, maybe you could do that. But it would require a lot of investment and effort to actually go there, and you'd have to make sure that you actually trusted the results I think.

DR. ZAMBON: I'm not sure if I can switch this on. It's on, is it? Okay. Can everyone hear? I have a few remarks to make, which may be relevant in the context of the discussion, partly as running a national influenza center and partly also as being, if you like, one of the people ‑‑ advisors on diagnostic virology strategies in the United Kingdom.

I think, firstly, it's important to plan for change. Diagnostic virology techniques, of which isolating influenza viruses can't be considered in isolation, we are changing. We're in a changing world, and things we can see already from the discussions that have been had that the amount of egg isolation has diminished over the last decade or so. And very simply, that's a question of economics.

How do you use your resources? You do things which are much more efficient, so it's more efficient to work in cell culture than it is to work in eggs.

Now, that doesn't mean to say that you have to eliminate eggs from the vaccine production system, but you do have to plan how you might incorporate it using national influenza centers, which is where you get your isolates from.

And the business of isolating a flu virus very rarely comes in as a separate category. You're usually trying to do that alongside other things.

The second thing to take into account when you consider your options is where do most of the flu viruses which end up in vaccine come from? They don't come from American labs. They don't come from European labs. They don't come from the developed world. The majority of them come from the developing world, and the issues of resources there are much more critical than they are in the developed world.

And the reason I make that point is because although the example of the polio laboratories using accredited cell lines for the isolation of polio viruses is indeed a very good example, the polio viruses are not isolated for the purposes of making a vaccine. They are isolated for epidemiological work. So, therefore, if there is a model, or if there is some problem with the way that the cell lines are treated, it actually doesn't matter at the end of the day.

So my point is I think you need to be very careful about a strategy which says we're going to put our accredited cell lines in 110 national influenza centers and expect the quality control to be maintained in the most resource-poor countries. I know how we would struggle in the United Kingdom to do that, alongside a raft of other responsibilities. And personally, I don't think it's actually an option.

But I do think that you have to plan for change, and you have to look at new diagnostic techniques and how they may impact on the characterization of influenza isolates.

Those are remarks based on experience, really. And I hope they contribute to the discussion.

CHAIRMAN OVERTURF: Dr. Royal.

DR. ROYAL: I just have a quick comment regarding what was said about SV40 virus. I was under the impression that actually there is evidence that malignancies are starting to appear in sort of emerging data 40 years later to suggest that vaccinees in fact may have been at greater risk for developing cancers that he mentioned, as well as a few others. At least at the point we are it might warrant some additional discussion concerning the issue.

DR. MINOR: I don't think there's an issue about discussion. Okay? I mean, I'm not convinced that the evidence stacks up to say that ‑‑ that there is an increase in tumor incidence in these ‑‑ in these particular people.

I think the ‑‑ the issue arose because there were SV40 sequences present in them, and the question was, well, where did they come from, are they causal, all that kind of stuff. And the debate continues. I mean, there is no question that the debate continues, okay, and there is no question that it gives a lot of people a lot of headaches. Okay?

But I'm ‑‑ I'm not sure that there's a causal link actually being convincingly demonstrated there, frankly.

CHAIRMAN OVERTURF: Dr. Monto, you had a comment?

DR. MONTO: I just want to thank Maria Zambon for her comments, her very practical comments, because I think we have to view the various options in a practical way. One of the problems that we're going to have with the increasing use of rapid diagnostics is a decrease in the number of flu isolates from various parts of the world.

What we want to do is to increase the number of flu isolates, so we don't have to play the small numbers game in terms of making vaccine decisions. It is totally impractical to think that many, if not most, national influenza centers will be able to handle certified cell lines in a way that we can be comfortable that isolates were made following good laboratory practices, or whatever, techniques.

And, therefore, I think we need to focus on the other options, and I can't think of any additional options in terms ‑‑ one of the things that ‑‑ and, frankly, I think we need to back off for a minute and remember that we are in many ways consumers of what is going on out there in influenza surveillance.

This is a relatively poorly funded, or has been poorly funded activity in the past. Many of the developing countries, where most of our strains come from, have never identified influenza as a high priority pathogen, and, therefore, have not really put their own funds into developing influenza surveillance, influenza identification techniques. Until this changes, we're going to have certain generic problems, since we are consumers rather than ‑‑ rather than suppliers in this ‑‑ in this area.

CHAIRMAN OVERTURF: Dr. Cox.

DR. COX: I think now that I've kind of got a flavor of some of the thoughts around the table, I'd like to just make a few clarifying comments as well. And I'll be echoing some of the things that have already been said.

We were actually about a year ago tasked with reisolating SARS coronavirus in certified cells, so that they ‑‑ this material could be made available to people who were interested in developing SARS vaccine.

And so we actually received some certified viral cells from one of the manufacturers, and then invited an FDA consultant, actually an ex-FDA employee, to go over all of the paperwork and everything that would be required in order to make sure that what we produced at the end of the day would be acceptable in terms of the regulatory requirements.

It's actually extremely onerous, and I would doubt very, very much if the national influenza centers ‑‑ very many national influenza centers at least, not more than one or two, would be able to put systems in place so that they could actually provide the documentation that would be necessary.

So I think that we're really faced with a situation where some collaborating centers or other centers that are set up to do this type of work would be responsible for soliciting the original clinical specimens from the originators.

And I must ‑‑ and I want to mention here that the military has for us been an extremely excellent source of original clinical material, and we've actually ‑‑ last year in response to what was going on with the Fujian-like viruses, we started a very concerted effort to solicit original clinical specimens from originators around the world and then to try to isolate those viruses in eggs.

We had very poor yield in eggs, so we then had to go to SPAFAS chick kidney cells to get the original isolate, and then move on to growth in eggs. So there are ways that we can do this, but it will require a lot of sensitivity, as Maria said, to the need to change methods. There will be a need for additional resources within the system to make this happen, but I think there are some solutions available.

CHAIRMAN OVERTURF: To me, it sounds like the system is ‑‑ that actually the surveillance issues are very linked to the eventual vaccine production. Since so many of the ‑‑ obviously, I would think the committee would want to spend a lot of time actually, first of all, enhancing surveillance. But in order to do that, it sounds like it's going to be linked to the issue of cell culture, since those are going to be the cell cultures.

So I think eventually they are going to have to come up with a system to clean up these ‑‑ I guess I can use that term ‑‑ these cell cultures, because that's probably what's going to be used for the primary isolation.

I think the other issue that's going to come up is that with the ‑‑ this is a problem for laboratories worldwide in a whole lot of issues. But the continued use of rapid testing to solidify people's diagnostic clinical impressions is also another thing that is going to decrease the number of isolations which are necessary ultimately ‑‑ as Dr. Monto pointed out, we as consumers need those ‑‑ need those isolates.

So, unfortunately, I'm not sure I heard anything that was too encouraging about cell cultures, but it does seem to me that one of the things that we really need to do is to ‑‑ is to really, first of all, enhance surveillance by any methods. But if that means that we're going to have more cell culture isolates to deal with, we also are going to have to deal with the second issue of cleaning those up.

Yes.

DR. COX: I'm not sure that I was actually very clear, and so I'll try once again. We definitely do need to enhance surveillance, and we'll continue to work with WHO to improve surveillance in countries ‑‑ in developing countries as well as in developed countries. And I'm hopeful that over the next few years CDC might have some additional funds that could go into enhancing surveillance, particularly in developing countries.

Having said that, I'm sure that the laboratories in these developing countries will want to use MDCK cells. But we can still go back to them and get the original clinical material, have that shipped to CDC, or other central laboratories, where those original clinical samples can then be reisolated in a clean environment and clean cells or in eggs, as appropriate for the technology that's being used.

And I ‑‑ I really think that that's probably about as good as it gets, because having had quite a lot of experience training people in these countries, and seeing the resources that they have available, I think it's just unlikely that they could ‑‑ could ‑‑ that we would be very comfortable working with the cell culture-derived material.

CHAIRMAN OVERTURF: Thank you, Dr. Cox. That's very useful clarification.

Yes, Dr. Karron.

DR. KARRON: So just to follow up, Nancy, and make sure I understand. What you would see as a barrier right now is actually not the quality of our surveillance perhaps, but the availability of these qualified cell lines in WHO ‑‑ select WHO reference laboratories.

DR. COX: I think there are gaps in surveillance. There is no doubt about it. There are ‑‑ there are gaps ‑‑ for example, right now there are outbreaks of Avian H5N1 in poultry in Laos and Cambodia, and we have no WHO collaborating national influenza centers in these two countries. And as far as I know, there aren't trained individuals who can isolate influenza viruses.

So there are gaps, and we'll continue to work with WHO to kind of plug those gaps. But in terms of the material that we do get, and that we are are getting, I think we can ‑‑ we've been working to get more original clinical isolates ‑‑ specimens from around the world. We have several very good sources, including the military surveillance network.

And I think we can really expand on what we've been doing in the past few years and try to make this ‑‑ the system work. And if there are any other good ideas that come up, certainly we can work to make it happen by incorporating others into it.

But, yes, we need certified cell lines, and we need more isolates.

CHAIRMAN OVERTURF: Dr. Markovitz.

DR. MARKOVITZ: Yes. Thank you, Nancy. That's extremely helpful information. I was curious, though ‑‑ one other thing I might address to you or others who actively work with flu in the lab. In following up on a discussion I had the other ‑‑ yesterday with some of our industrial colleagues, the issue arose of if you are actually going to end up growing these things in mammalian cells.

Can you get high enough titres? You know, in other words, if it ‑‑ if the final process were making ‑‑ you know, you were going to use this all the way to a vaccine, can you grow it efficiently enough in mammalian cells versus eggs? And I'm asking if you could answer that.

DR. COX: I can only answer a very small part of that question. We can grow the viruses well enough to characterize them and hand them over to the manufacturers. Whether they grow well enough for the manufacturers to develop a viable product with ‑‑ or not, I ‑‑ you know, that would have to be answered by the manufacturers.

DR. EGAN: Maybe if I could just try and rephrase the question a little bit and the discussion. Let's assume ‑‑ you know, given that an isolate ‑‑ a flu isolate has gone through a cell line, mammalian cell line, MDCK or VERO, of various degrees of qualification, it may have started out as very, very well qualified, and then may be of questionable or unknown qualification later on.

But given that you've got this cell isolate, this flu isolate that has gone through these cells, and there may be some questions, there may or may not be problems, but certainly questions and concerns, can that isolate be cleaned up, rescued if you will, by subsequent techniques ‑‑ for example, through a plaque purification on the cells, the qualified cells that you are going to use for production, or through a reverse genetics?

I think this was the issue that faced us last year. So we want to come back to that question.

And then perhaps for Dr. Decker I'd like to ask, you know, if this were done, these were rescued/cleaned up, would the manufacturers have any concerns, particular concerns, about the acceptability of a seed that were produced this way?

CHAIRMAN OVERTURF: Dr. Decker.

DR. DECKER: I'll end up with your question, Bill, not start with it. Just before Dr. Egan spoke, I was going to comment on a matter that he kind of subsumed. I was going to say, given what I've heard here, I want to go back and readdress the comment that was made earlier that clearly option two is not satisfactory. I don't recall who said that, and maybe I misphrased it, but that's the sense I carried away from it.

And I don't ‑‑ and I questioned that. I was questioning that before Bill spoke, because it seems to me that what we want here are as many viable options as possible. What I think I see over and over again is that we don't know what nature will hand us. We don't know where it will hand it, and we want all of the tools available that will work in our toolbox.

For that reason we ‑‑ we want reverse genetics techniques, but that doesn't mean we throw away classic reassortant techniques. They've probably remained the most useful most of the time.

For production, it's wonderful to have cell culture technologies. I don't think anybody thinks we're going to throw away eggs, because each will have its appropriate purpose.

And, similarly, here it may be that the single best technique may not be growing it in unqualified cell lines and then rescuing them through plaque purification. But knowing that we can do that, if that's what actually happens some day, it might be enormously valuable.

So I would ‑‑ I would not at all turn my back on option two. Even if it's not the way we hope things play out, it may be the way things actually do play out. And so I think that would be very valuable.

Coming to Dr. Egan's question, he ought to know when he asks a question like that I'm going to turn it right back at him. I cannot contemplate, as a manufacturer, having FDA hand us a strain and say, "We hope it's safe."

(Laughter.)

DR. EGAN: We certainly appreciate that vote of confidence.

(Laughter.)

CHAIRMAN OVERTURF: Further questions? Discussion? Yes, Dr. McInnes.

DR. McINNES: Thank you. The reason I was staggering over all the bags back here to get to the table was to try to make a comment, exactly what Dr. Decker did far more eloquently.

I think what I was hearing before was sort of this all or none. It had to either go all the way or another way. And I think the balance was the system that we have is working extremely well, but it doesn't always work. And so I think what FDA was trying to lay out for us were some options for strategies when that optimum isn't working.

We saw the value of it in ‑‑ with the SARS experience. We're going to see the value of it in the pandemic experience. And last year in the interpandemic, an approach was in fact ‑‑ could be possible to have solved some of the dilemma that we were faced with last year.

So I would plead not to throw out what technology will offer us in these very difficult circumstances when there may be no other option.

CHAIRMAN OVERTURF: Yes. Dr. Minor.

DR. MINOR: My difficulty with option two is, how do you know that it has worked, right? I mean, how do you know that you got ‑‑ you got rid of whatever stuff you had in there in the first place, if you had stuff in there in the first place.

I think if you go through reverse genetics, I think you can be reasonably certain that you've cleaned it up. Right? If you do the egg thing, I think you don't care whether you've cleaned it up. but in the case of option two, how do you know that you've actually done what you meant to do, right?

CHAIRMAN OVERTURF: Yes, Dr. Decker.

DR. DECKER: True. Noted. But at the same time, let's look at another option, which would be let us suppose that there were ‑‑ we had a wonderful supply of a qualified cell line with no IP barriers, there was widespread distribution to every lab that wanted it. As has been pointed out, the day it leaves the door it's not qualified anymore.

And to the extent that you think that you're following that approach, aren't you really in option two? And hadn't you better have developed, to the extent you can, your techniques for subsequent purification?

If you go the reverse genetics road, or you work in eggs, you may sidestep that. But we saw last year, we could have wished all we wanted that they had worked in eggs last year, but the Fujian isolates were in unqualified cell lines, not in eggs.

And had we had in our hands at that time a ‑‑ a procedure that we had some faith in for ‑‑ I'll call it plaque neutralization, but it could be some entirely different technology, but some way of taking that next step in a known timeframe, had this committee known a year ago in two weeks we'll have an isolate that can be used for manufacture, I think we would have behaved very differently than we did.

And so I'm all for our developing broader techniques here, even though no one technique may be perfect.

CHAIRMAN OVERTURF: Yes. I think I would agree. I think at this point it seems too early in this whole issue to throw out all ‑‑ any option, really, until they have been more fully explored. And I also think there may be a certain urgency in certain situations.

A true pandemic of a very lethal influenza might incite a different level of caution than ‑‑ and also might ‑‑ might be backed by certain guarantees from manufacturers, and so forth, against liability, etcetera, that would not occur, for instance, in the annual routine immunization procedures.

So I think it's probably too early to throw out all ‑‑ all options at this point.

CHAIRMAN OVERTURF: Dr. Gellin, you had a ‑‑

DR. McINNES: Just to address the comment. I think there's a ‑‑ there may be a range of qualification on the cell lines that are under consideration, all the way from fully qualified cell line that is in the hands of the manufacturer to perhaps a ‑‑ such a cell line that has been transitioned to a very competent and capable lab, and, in fact, maybe the full requalification process hasn't gone, all the way to mashed up dog kidney cells sitting somewhere out there.

And, I mean, this is a full range. And so perhaps what that has to be tied to is, in fact, in such circumstances there would be a balance of data to support, in fact, almost a relief on ‑‑ and characterization of the reference virus, which I know right now is not done in interpandemic times. So I think it's, again, a balance of data that might need to be put forward for consideration as a reference virus.

CHAIRMAN OVERTURF: Dr. Karron.

DR. KARRON: I guess the one other comment I'd like to make, and my only concern about what Michael said, is that I do think it would be useful to have a hierarchy of preferences, because I do think that it may influence how you move forward.

If you ‑‑ if you say that any of these options are fine, and they may be, or there may be situations where we want to be able to exercise all of them, it doesn't perhaps push us to see if we can get qualified cell lines into the hands of, you know, WHO reference laboratories, or certain other activities.

So I think it might be useful at least to say we will consider all of these options, but these ‑‑ this is the order in which we would like to be able to have isolates.

CHAIRMAN OVERTURF: Dr. Myers.

DR. MYERS: I think there ‑‑ we're talking about a couple of apples and oranges issues. I think when we talk about pandemics and we talk about SARS and we talk about emergency circumstances, we may be talking about a different set of circumstances.

But what, Mike, you're talking about is a routine year-in and year-out isolation of seed viruses for ‑‑ and the issue we were talking about last year. I think the issue of perception of vaccine safety ‑‑ we are obligated in the routine set of circumstances to have the safest potential seed viruses that are available to us.

And although I understand what Nancy is saying about current good laboratory practices not being currently a common ‑‑ commonly used in diagnostic laboratories or in many of the laboratories around the world where these isolates are obtained, I think strategies of trying to bring the clinical specimen to a CGLP setting with documented cells is a strategy that is worth developing to increase repertoire on a year-in and year-out basis.

And I think that should be a priority, a lot higher priority than trying to clean up undocumented, unqualified, potentially contaminated cells.

CHAIRMAN OVERTURF: And I think, actually, the strategy that Dr. Cox had suggested, which was to link this to your surveillance program, so that you go back to the original isolates, and ‑‑ and use appropriate techniques to isolate the initial seed viruses, is probably going to be in the short term a very good answer.

But I ‑‑ I also would say that probably there is going to have to be a priority for some of these other techniques for pandemic situations and other kinds of isolates. I would agree that for the routine yearly isolation of viruses, and production of vaccine, I think we have an obligation to make the safest seed lots.

I'm sure ‑‑ and I'm sure the manufacturers are not going to be happy if we give them anything that's ‑‑ anything less than absolutely pure and good.

DR. HJORTH: Yes. I was interested in Nancy's comment about the SPAFAS chick kidney cells. And I think that might be another option. These ‑‑ these might not ‑‑ these might be a little easier to deal with than certified cell lines, and I was wondering ‑‑ Nancy mentioned the isolation rate was higher.

I just wonder, is it substantially higher, so that it would really be worthwhile considering these instead of eggs? But perhaps isolates from these cells might be considered as though they were egg isolates.

DR. COX: They are considered as egg isolates now. And they do give a substantially higher isolation rate for the current H3N2 viruses than eggs do. So ‑‑ so if we take the original clinical specimens and put them into eggs and SPAFAS chick kidney cells, we're more likely to get an isolate out of the SPAFAS chick kidney cells, which can subsequently be passaged in eggs.

So, but they are not ‑‑ I mean, they are a little bit finicky and fiddly, so you have to kind of know what you're doing. And it works best if you've got them going on a routine basis, and you've got somebody who is accustomed to looking at them week after week. And they are actually more difficult to deal with than the MDCK cells that are used in most of the reference labs.

CHAIRMAN OVERTURF: Yes, sir.

DR. EGAN: Yes. Dr. Overturf, I certainly appreciate your comment, but would like to ask you to maybe clarify it a little bit more, maybe other people that are here. Do you consider an isolate that had gone through, for example, an MDCK cell or a VERO cell, and then subsequently go through a reverse genetics procedure or go through several rounds of plaque purification to be any less safe?

You know, and, if so, you know, what ‑‑ you know, what ‑‑ what do you see as the safety concerns? Maybe to have a little more discussion of this.

CHAIRMAN OVERTURF: Well, and I ‑‑ some of us here are not experts in this area. But I ‑‑ I think it has been pretty clearly laid out that they cannot be at the present time considered as safe as those that have been primarily isolated in eggs, although I think we've taken a lot for granted about eggs for years, so I'm not sure that we have all that we really need to make that decision.

Yes.

DR. MONTO: I would feel quite comfortable with the reverse genetics procedure, given certain caveats in terms of what you got out being what you put in. And we can do that in terms of sequence ‑‑ sequences, etcetera.

I think that would be ‑‑ if you follow Dr. Karron's suggestion and come up with a hierarchy, that would be my ‑‑ my preference in terms of developing. And I think we need to develop protocols to ‑‑ to be able to handle these different options.

I think the option two is an attractive option, but there I think there would be many questions, certainly relative to what Dr. Minor said, in terms of how do you prove that it's ‑‑ that nothing adventitious has gotten in there? How many plaque purifications would you require? Would there be additional safety testing required, especially ‑‑ do you still see that the egg ‑‑ going back to the egg ‑‑ is going to filter out any human or primate viruses that could have gotten into the mix?

So it would be useful to see more detailed protocols developed for some of these ‑‑ some of these options.

CHAIRMAN OVERTURF: Dr. LaRussa.

DR. LaRUSSA: Is there a sense that in the coming months there is going to be more detailed information available, so that the committee could actually evaluate the different rescue options and make recommendations? Or is it just really too early?

DR. EGAN: It's probably a bit early for that. But, you know, this is an issue that certainly arose last year, and we'd like to have a fuller discussion of it. And that's why we're bringing the ‑‑ you know, the question back to this committee. And, you know, maybe some of the virologists here can, you know, give some additional comments.

CHAIRMAN OVERTURF: Dr. Karron.

DR. KARRON: I would just say that there's probably ‑‑ to follow up on what Dr. Overturf said ‑‑ there's probably diversity of opinion among the committee members about whether a virus that, say, under scheme two had been isolated and was then either recovered by reverse genetics or plaque purified and put through eggs ‑‑ you know, how safe that would be.

And I'm not ‑‑ I'm not sure that there's a unanimity of opinion and that that would be less safe. I think ‑‑ my guess is that there's probably a diversity of opinion on that issue.

CHAIRMAN OVERTURF: I think the diversity is partially based upon an awful lot of suppositions and guessing that we really can't substantiate in any way, shape, or form. So I'm thinking there's a lot of uncertainty.

I think one of the things that favors the current methodologies is a long record of ‑‑ of, we think, safety. But even there there are some ‑‑ there are some ‑‑ some loopholes. But I think unless you have strong evidence to deviate from that in safety and other systems, that's probably ‑‑ probably ‑‑ I think most of us probably would be uncomfortable with that.

I think I also would agree that the reverse genetics approach probably has the best assurances ultimately for proceeding. But I think we're ‑‑ we're not there yet at this point. I don't know how others feel.

DR. DECKER: I would agree with that.

CHAIRMAN OVERTURF: Dr. Decker.

DR. DECKER: Yes, just I agree. And following up on Dr. Karron's comment, I mean, I think that was helpful, and I think it's been implicit, but I'll make it explicit. A, isolation remains ‑‑ if we had our choice, that's the way we'd prefer to see it come.

And if it can't come that way, I would say that the reverse genetics is probably second choice, because in my mind having an allegedly certified cell line out of somebody else's hands for some period of time really drops us right into option two. So that those two go together with me.

If you're going to go with that certified cell line in a non-central, non-FDA level, CDC-level lab, or something else, then you're already in option two. So those merge together for me to become the third choice.

CHAIRMAN OVERTURF: Dr. Karron.

DR. KARRON: Maybe Nancy can comment on this. A non-FDA, non-CDC level laboratory ‑‑ I'm not familiar enough with the WHO influenza laboratories to know if they are ‑‑ this may be a loaded question ‑‑ but comparable to yours in terms of abilities to, for example, handle a qualified cell line, maintain it, do the kind of quality assurance that's necessary.

DR. COX: I would have to say that the majority of the national influenza centers would not have the resources to do this. You need to have a separate room which you can devote for a specific period of time. It has to be completely cleaned up.

You have to have the cell ‑‑ the certified cells locked and in a, you know, very clearly delineated area of the freezer. You have to have very extensive documentation of all of the media components, and so on and so forth, just to meet GLP requirements. And so for ‑‑ I doubt that most would have the resources, really, to devote to this kind of an activity.

DR. KARRON: Right. I was actually speaking about the WHO collaborating centers. I was differentiating ‑‑

DR. COX: Oh, the collaborating centers.

DR. KARRON: ‑‑ between ‑‑ between option one and option three in the ‑‑ in the range that we were given.

DR. COX: There are four WHO collaborating centers, and I would have to speak with each of the directors individually to find out if they would have those resources.

CHAIRMAN OVERTURF: Dr. Markovitz.

DR. MARKOVITZ: Yes. It seems like what Nancy said a while ago seems to be when we're looking for other options other than standard options, I mean, the idea of, you know, getting the original clinical isolate to a lab such as yourself ‑‑ yours or perhaps a comparable lab here, that seems to make infinite good sense as an alternative.

And I'm a bit perplexed. In spite of being a molecular geneticist for about 80 percent of my professional life, I'm not quite sure where we actually stand on these reverse genetics in practical use right now with flu, you know, for vaccine. I mean, it looks ‑‑ I mean, I've heard the term bandied about, but I'm not completely sure where we stand.

Could somebody clarify that for me in five easy sentences?

CHAIRMAN OVERTURF: We'll give you six sentences, Dr. Minor. Could you do that in six?

(Laughter.)

DR. MINOR: Okay. I mean, this is purely in technical terms, okay, so it's not in terms of the practicalities of it. Once you've actually got the strain out, what can you do with it, and all that kind of stuff, it's purely technical. Okay?

What you do is you'll extract the nucleic acid, you'll clone it into a plasmid, and then you'll recover your virus from the plasmids that you've cloned. Okay?

To my mind, everything that happens before you actually extract your nucleic acid from your virus is, therefore, cleared out of the way completely, right? Okay. It's completely irrelevant, okay?

Anything that happens after that is your own fault, okay? So if you're ‑‑ okay? So if you're doing things under GNP, or whatever, okay, then I think from that point on you're in the clear. This is why I say I think that if you can it by reverse genetics, I think then you really are clean in terms of you don't really necessarily care where your isolate came from, because you got rid of everything that might have been there in the first place.

DR. MARKOVITZ: Yes, I completely agree with that.

DR. MINOR: Right.

DR. MARKOVITZ: From a virological point of view and molecular. But what I'm curious is ‑‑ the way these isolates are actually arriving at the doorstep prior to being perhaps becoming a vaccine isolate, they're not coming through reverse genetics right now, right? I mean, they're just coming ‑‑ they're growing in cells, right?

DR. MINOR: That's right. Well, I mean, our Vietnam/11/94 thing ‑‑ I mean, this came as an egg isolate from somewhere. I mean, I ‑‑ Hong Kong apparently. Hong Kong? That's okay. So, but I ‑‑ my point is that I don't care where it comes from, if I've gone through reverse genetics. It doesn't even have to come as a virus. I mean, it could come as a ‑‑ it could come as a PCR fragment, provided it's the right PCR fragment. Okay? Provided you can clone it and you can clean it up.

DR. MARKOVITZ: But in terms of what we're doing right now, how big of a percentage of the effort towards a vaccine in a given year is coming from reverse ‑‑ you know, virus derived from reverse genetics? That's what I'm ‑‑

DR. MINOR: Oh, zero. Zero. Zero.

DR. MARKOVITZ: Yes, okay. So that's my impression, but yet I keep hearing reverse genetics, and I ‑‑

DR. MINOR: It's just a good idea, you know?

(Laughter.)

DR. MARKOVITZ: Yes. So, obviously, it's the best idea if it can be up and running. But how close are we to having it up and running in this setting that would impact on the ‑‑ the committee issues, which is year-to-year vaccinology. That's my real question.

DR. COX: There are laboratories that have already produced vaccine candidates for age five, as you heard yesterday. There are intellectual property issues related to this technology, however, that would have to be worked out.

CHAIRMAN OVERTURF: Dr. McInnes.

DR. McINNES: I mean, I think we shouldn't forget there is a product licensed in the United States that starts its origin, its reference virus origin, through reverse genetics. So, I mean, I think the answer that there isn't any, we ‑‑ yes, there is.

DR. COX: Could you clarify?

DR. McINNES: If we're talking about the live attenuated cold-adapted influenza vaccine, what are you saying?

DR. COX: That's classical reassortment, I think, yes.

DR. McINNES: Correct. But, I mean, the concept of using reverse genetics to develop influenza vaccines is not new. I thought that was what you were saying.

DR. COX: No. We were just ‑‑

DR. McINNES: Perhaps I don't understand you.

DR. COX: Yes. We were just saying that there's no license ‑‑ there's no product that's currently used that's derived by reverse genetics.

DR. McINNES: Okay. I'm sorry. I misunderstood you.

CHAIRMAN OVERTURF: Dr. Egan, I think ‑‑ I think you heard a priority here, and I think the feeling of the committee is, as stated by Dr. Karron, probably diverse. But I do think that there is a sense that there is a priority with the first priority really being for reverse genetics.

And then I don't think people are comfortable yet with rescue methods for recovering virus from unqualified cell lines. Did you ‑‑

DR. EGAN: Although there was some comfort I guess through reverse genetics. But ‑‑

CHAIRMAN OVERTURF: Yes.

DR. EGAN: But maybe a little bit less so with several rounds of plaque purification.

CHAIRMAN OVERTURF: Yes.

DR. MARKOVITZ: That sounds right to me, yes.

DR. EGAN: Okay. This has been, you know, very helpful for us. And I think we can certainly take this discussion and the comments that all of the committee members have made and probably ‑‑ and try and put something together, a little more of a directed package for future comment and discussion, both here and with other regulatory agencies.

Thank you.

CHAIRMAN OVERTURF: Was there any further discussion? We actually are about 45 minutes ahead of schedule, mostly because we had no speakers at our open session. So I don't believe there's any other business.

DR. FREAS: No. I would just like to thank the committee members for their discussion, comments, recommendations. We really appreciate it.

Dr. Overturf, you did an excellent job as Chair. I'd like to thank you.

The guest speakers who came from overseas and long distances, we really appreciate it.

And I would like to give special thanks to my friends at CDC. Dr. Cox, we couldn't have done this meeting without you and your colleagues.

And, Dr. Levandowski, I know you had many late hours and many late nights.

We appreciate everybody's input. Thank you.

(Applause.)

It's adjourned.

(Whereupon, at 12:23 p.m., the meeting was adjourned.)