3.3 Douglas Richman: Antiviral Drug Resistance and Combination ART

Doug Richman: [00:00:00] I guess to follow on a couple of things that John [Mellors] said, in terms of preparation, I did my training before HIV was appreciated. I think that my years of training with Tom Merigan (b. 1934), my assignment, he had just discovered human interferon and assigned me as a medical student to figure out the mechanism of action of human interferon. It was a very [00:00:30] educational opportunity to find out that you couldn't possibly achieve your assignment.

Then I went to the NIH. Sam Broder and I were house officers together and we became good friends, and we went out to the NIH together for the major purpose of avoiding our Vietnam obligations as yellow berets. But it turned out to be an incredible education and medical virology with Bob (Robert M.) Chanock (1924–2010), where I worked on influenza. [00:01:00] On a related note, the two influenza fellows on the campus at the time were myself and Bob Chanock’s lab and Dave Barry (1944–2002), who was at the [FDA] Bureau of Biologics where we became friends. That's sort of how I got into AZT between those connections.

With that background, I should just—One other thing I should mention about this AZT story is Dave asked Margaret [Fischl] and I to put together the trials as investigators [00:01:30] together with the BW (Burroughs Wellcome) folks and to conduct them. To show how times were different when it was time to write them up, he had the statisticians put together two cardboard boxes with about two feet of tables and figures and data lines and shipped them to each of us. I bought my box over the Christmas vacation in my mother in law's [00:02:00] kitchen, put together one of the two papers. That was the way things were done back in that time. Things have changed quite a bit.

With the background on AZT, I will just move right on and not describe any more about the studies. Having participated, San Diego [VA Medical Center] participated in the phase II study. We collected the lymphocytes, and I tried to isolate virus from these [00:02:30] individuals. This was all done on the run, but we had a number of patients in whom we had sequential isolates, in whom you can see the progressive reduction in drug susceptibility over a period of months. Subsequently, we cloned and population sequenced and then Brendan Larder and Sharon [D. Kemp], his [00:03:00] wife, published the results showing that the progressive increase in resistance was due to the progressive accumulation of mutations in the virus. (1)

Now, sort of related to what John [Coffin] was talking about with Howard Temin may be on a shorter scale, and maybe a little less imposing, is all truth passes through three stages. The first is: It is ridiculed. [00:03:30] Before I conducted these studies, when I found out that Sam told me about the results with his blinded studies with “compound S” as it was called, which turned out to be AZT, I put together a proposal to look at the drug resistance.

My first grant proposal was not even scored. At that time to be not be scored, put you in the bottom 5%, 10%. [00:04:00] Both of the reviewers said that this is an absurd proposal because it's inconceivable that such a highly functional enzyme would be sufficiently plastic to tolerate mutations. I had to find some additional funds, as well as get a bit of support from Burroughs Wellcome to do the studies on the run with the phase II study. [00:04:30] When we put the paper together, it was violently opposed by several people saying it wasn't important, it was just an observation. I got calls from the boards, the director at the Burroughs Wellcome from people who were going to lose the opportunity to sell their [stock] options. It was a very difficult thing once we had published it with a lot of criticism that was just an observation with no clinical significance.

The third step in the [00:05:00] truth passing through three stages, [drug resistance] is now accepted as being self-evident. It's a standard of practice for drug development, and clinical management even it's being incorporated in Sub-Saharan Africa as just part of standard clinical care. After the initial observations with the nuclear sides, compounds with other mechanisms of action were starting to be identified [00:05:30] by the various pharmaceutical companies and the folks at Boehringer [Ingelheim]—many of these companies, as Burroughs Wellcome initially, were uncomfortable or didn't work with BL3 (biosafety level 3) agents. I was contracted by Boehringer Ingelheim to look at a compound that ended up being nevirapine (NVP, Viramune) and to assess its behavior against HIV in [00:06:00] cell culture.

What we did is a preclinical selection for drug resistance. (2, 3) With this compound and then with all compounds subsequently, this is the approach to assessing the relationship of resistance to a new drug. We did in vitro selection. You can see one log increments here with various passages between passage 12 and 20, [00:06:30] a log increase in drug resistance with passage in the presence of drug.

We had just incorporated PCR and [gene] sequencing in the late 80s. What we looked at was the appearance of mutations in the target enzyme with this non-nucleoside RT (reverse transcriptase) inhibitor (NNRTI). What we showed is that—we [00:07:00] used two different viruses. What's interesting here we call it HIV-1 Bru, because Jean-Claude Chermann and Françoise Barré-Sinoussi were gracious enough to ship the virus to San Diego to let us look at it and that's the name that came with it. As you can see that with one virus from the phase II study patient and another from the Paris isolate, [00:07:30] either one or three passages were sufficient to start to see mixtures of wild-type and the Y181C mutation associated with drug resistance. Then it was only five passages and one had already, by purifying selection, come up with a pure population of drug-resistant virus. [00:08:00]

We looked at cross-resistance with the nevirapine resistant virus. You can see it the wild type virus has a susceptibility of 0.04μM with 12 passages that went up to 10μM, and you can see there is no cross-resistanceto either nucleoside (AZT) triphosphate or foscarnet (phosphonomethanoic acid, Foscavir). There is cross-resistance against another non-nucleoside RT inhibitor. [00:08:30] This is a demonstration of cross-resistance and this has become standard, looking at the cross-resistance patterns of new mutations and new drug-resistant isolates.

With the Boehringer Ingelheim folks, we tried to assess what the mechanism of action of the drug resistance was and they had a photoaffinity labeled nevirapine. As you can see on the left, [00:09:00] the B is the drug-resistant mutant and A is the wild type, and the photoaffinity labeled pro binds to the P66 subunit of RT and doesn't bind to the one that has the cytosine mutation at 181.

We then went through the process of confirming that that mutation did, in fact, confer the drug resistance by creating [00:09:30] site-directed mutants and showing that the same phenotype was generated by the site-directed mutant. That approach to drug-resistance has pretty much been followed with more sophisticated techniques subsequently. I should mention that exactly in parallel with what we were doing, Jack Nunberg and Emilio [A. Emini] were looking at a non-nucleoside RT inhibitor at Merck & Co., Inc. (Merck Sharp & Dohme) that didn't end up getting approved but basically selected [00:10:00] for the same mutation.

Once we were able to initiate phase I and phase II studies through the ACTG (AIDS Clinical Trials Group) with nevirapine, we made a couple of observations regarding drug resistance in vivo. (4) Let me see if—[00:10:30] Here we go. A couple of things are interesting. One is: at one week you can see this dramatic reduction and I think this is the first documentation of the dynamics, the rapid turnover, and the impact of drug on the clearance of virus with the introduction of antiviral drug. [00:11:00]

This is all by p24 [antigen level measurement], the PCR assay wasn't quite available back at that time. What you can also see is within two weeks, a dramatic increase in CD4 cell count, all very encouraging but within a month, you can see the complete loss of activity and the loss of CD4 cell counts with basically the neutralization of both [00:11:30] the antiviral effect and the CD4 recovery effect attributable to the appearance or in association with the appearance of this Y181C mutation which we could see by week two. In association with that, the isolate had become two logs more resistant.

We also did population sequencing [00:12:00] of the isolates from these patients and we observed something that's now been seen with almost all agents that if you look for the selection in vitro of mutations, you'll see some mutations. But in vivo which has many orders of magnitude higher amounts of virus replication, the virus explores [00:12:30] a lot more sequence space and fitness space.

What we observed here is [that] quite a few more mutations were observed in vivo than we had selected for in vitro. Secondly, in patients who were given to nevirapine plus AZT, the mutation that's frequent with monotherapy [00:13:00] doesn't appear. This was a consequence of the observation that the Y181C mutation hypersensitized to AZT and so the virus is always searching for a better fitness landscape and it chooses an alternative set of mutations if there's AZT present.

One other thing we observed that I'm not showing you slide for [00:13:30] is that if you do population sequencing in the first week or two the virus has explored all sorts of different variants, but it is exploring this fitness landscape and it finally decides based on the level of resistance, the fitness of the virus, perhaps some immune selective pressures as well, purifying selection ends up resulting in usually a predominant strain after [00:14:00] a period of several months that is most fit and most resistant.

That was the early period of the RT inhibitors of drug resistance. It was clear to us from the very beginning that we were going to want to look at combination therapy. This is actually the first study that we did through the ACTG of combination therapy. (5) The decision to use these two drugs (AZT and ddC) was based on a very logical [00:14:30] principle and that is they were the only two available.

We put them together, we were restricted to a very small study so we could really only observe tolerability and it took a very large randomized phase three study to show that actually adding a second nucleoside to AZT had some incremental benefit but still not sufficient to [00:15:00] have a sustained benefit without the selection for resistance. 

Based on the very simple principles that had been clearly elaborated in the 1950s with tuberculosis, mutants preexist in the population to any single drug. The combination of drugs basically has [00:15:30] the probability of selecting for the resistance probability of being collinear, as the product of the two probabilities. So that what we proposed theoretically—and this is a slide that I made in a little review—that the more drug you gave, the higher the selective pressure, the more rapidly you would select for resistance. (6) We actually documented this in 1990 with the AZT studies. Mark [Harrington] [00:16:00] raised the question, why did we go into the phase II right away?

The issue was urgency. Patients were just dying in our clinics right and left, in contrast to what Jim Curran said, some of us, ID (infectious disease) doctors, decided to go into HIV. It was, as Paul [Volberding] and Sam [Broder] mentioned, quite an experience. We wanted to get drug into [00:16:30] people as rapidly as we could, but we realized there was toxicity. In the late '80s, a number of studies through the ACTG (AIDS Clinical Trials Group) then looked at a more tolerable dosing and we ultimately went to that. In a study that we published in 1990, we showed that the higher dose AZT selected for resistance more rapidly than lower dose AZT and that's because of the higher selective pressure. (7) [00:17:00]

Back in the mid-'70s when a lot of people were proposing antiviral drugs because they put them into culture and they prevented replication, it was observed that most of those things that worked with detergent, soaps and other things that killed the cells, which actually do prevent virus replication. The proposal was made by [Paul E.] Hermans in the mid-'70s that the definition of an antiviral drug is one that can select for drug resistance. (8) [00:17:30] That is a pretty reliable one that rules out nonspecific effects.

Anyway, we did document that the more drug you had, the higher the selective pressure but also proposed that once you slow down replication, you would start reducing the probability of resistance, and if you could completely suppress replication there would be no more evolution. [00:18:00]

When we finally had classes of drugs other than the nucleosides, this could be tested. (9) The standard of practice by the early '90s was combination to dual nucleoside therapy. In the early '90s, I think it may have been '93 or so, John Mellors and I and John Ryan, who was at the time head of ID at Merck, [00:18:30] had a breakfast meeting at [unintelligible 00:18:31] and discussed the design of combination study with Merck. This was the evaluation of indinavir (IDV, Crixivan, a protease inhibitor) compared to what was then the standard of practice, AZT and 3TC. What was observed is AZT did what you've already seen, AZT [00:19:00] and 3TC, rapid activity and then rapid loss of the activity with the emergence of resistance. Indinavir actually had some activity in patients that was a little bit greater than the combination, but it began to get lost in most patients.

It was much slower and what we subsequently learned is protease inhibitor [00:19:30] resistance requires the accumulation of multiple mutations. The combination was greater with all three drugs, and it was sustained, and this plateau was due to the limit of detection of the RNA. The results from that original paper show that driving virus below the limit of detection using the viral load assays, the dual nucleoside combination couldn't do it. (9) [00:20:00] Indinavir actually did it in about 30% of people. Those were the very lowest viral loads and highest CD4 cell counts but the combination gave 80% reduction that was pretty much sustained and some of those failures were people who not just were poorly adherent but dropped out and other things. This was the new paradigm, the one that was announced at [the 11^th International Conference on AIDS] Vancouver (1996) [00:20:30] and really changed the approach to antiretroviral therapy.

I should mention that, done in parallel, the Boehringer Ingelheim people also put together a combination study with the Canadians and European group. (10) I think it's important to mention the senior author (Joep M. A. Lange, 1954–2014) should have been here to participate in this [00:21:00] meeting but Vladimir Putin's henchman shot him and his lovely partner and collaborator Jacqueline [van Tongeren] (1949–2014) out of the sky in the Ukraine on the Malaysian flight and, unfortunately, he isn't able to share his brilliant experience with us.

These are the results of that study, showing [00:21:30] a similar pattern, although the percentage is lower which I think is attributable to the fact that ddI and nevirapine are not quite as good as 3TC and idinivir. 

I think the principles of combination therapy are there. Getting back to the drug resistance question, we did a study of a weighted [00:22:00] sample from the United States that represented the 200,000 people under care back in '97 and by that time because of all the prior nucleoside therapy, 78% of people under care had a drug-resistant isolate. (11) Even only, it was a couple of years after the approval of the PI’s (protease inhibitors) and NNRTI’s, because of the use of these drugs as add-ons rather than combination, we already had multiple [00:22:30] drug resistance in half of the population in the country that were under care.

The consequence of that also was that these people were transmitting their virus—and this is a combination study led by Susan Little in our group, of 10 North American cities showing the progressive transmission of drug resistance to new patients and in some sites still on the West coast, there was rates are approaching 20% [00:23:00] although they have plateaued and, in fact, they've improved. (12)

I should just mention that the current drugs, and you'll hear more from John, are getting up in the 90% range and we were talking about 8% or 10% failure. (13) That is not drug failure with drug resistance. Drug failure with drug resistance with the current regimens is 1% or less often. These 90% is attributable to the fact that people who drop off or lost to [00:23:30] follow up are considered as non-successes. The true success rate in as-treated is just absolutely remarkable with the current regimens and John [Martin] will tell you more about it. 

What's happening with resistance in the resource-rich countries, I think is best exemplified by this study from the Swiss cohort which is the most densely observed [00:24:00] cohort, and they're following almost two-thirds of the patients in Switzerland. (14) You can see there have been 8,000 patients under care as of 2013 but back in the late '90s, it was always 5,000. You look at the rates of resistance and those who started before '99, it was approaching that 80% rate that I showed you in the US. Those who started between '99 and 2006, it dropped to below 50% and those who started since [00:24:30] 2007, the rates of emergence of resistance to people who start combination therapy is a tiny fraction. So the rates of development of resistance in the whole Swiss cohort are in the single digits or teens annually and the people who have multiple class resistance with no treatment options has dropped to almost zero. We can treat almost everybody, even those who fail. It's absolutely remarkable [00:25:00].

I'll just close mentioning resource-limited countries. What has happened as a result of the success in resource-rich countries and the leadership of a number of people including Joep Lange, the development of the PEPFAR program, there are now over 17 million people under care. The majority in sub-Saharan Africa, [00:25:30] they're getting combination therapy. However, the rates of resistance are recapitulating to a large extent. (15) What we had seen in the United States are not quite as great because they are getting a combination regimen of two nucleosides and a non-nucleoside. The longer under care, the more resistance. The resistance is restricted to nucleosides and non-nucleoside because that's the drugs available and in [00:26:00] a limited number of patients, PIs are available second-line therapy and the rates of transmitted resistance are really a function of how long the rollout has been in the country.

The reason there is still resistance in these countries is because suboptimal regimens, until very recently, were being used, thymidine analogs were only being replaced in the last few years with a tenofovir based regimen [00:26:30] and there were insufficient second line and salvage regimens. Failure of methods and resources to monitor viral loads only recently being introduced. Nonadherence in some areas was more due to stock-outs rather than patient nonadherence and the perinatal trans resistance was due to the fact that just limited perinatal treatment was being used rather than fully [00:27:00] treating the mother with fully suppressive combination therapy. It's called option B.

All of those are things that are being addressed but contributed to the fact that while resistance was disappearing in resource-rich areas, it was expanding where the treatment was expanding. The availability of dolutegravir (DTG, Tivicay) and the [00:27:30] Gilead integrase inhibitor you'll hear about, plus the availability of TAF (tenofovir alafenamide fumarate, Vemlidy) as you'll hear about should provide dramatic improvements in ART in low and middle-income countries and, hopefully, address what was really a frightening impediment to effective therapy when we started.

I will stop here and I am glad to answer any questions.

[appause] [00:28:00].

John Mellors (moderator): Thank you, Doug. I don't remember what we had for breakfast, do you?

Doug: Boy, no. I had coffee. I know that much.

David Gludish: Thanks, Doug. That's a great talk. Just the last slide. You paint a nice picture or at least a hopeful picture [00:28:30] about the possibility combination therapy actually working in places like Sub-Saharan Africa. How do you see the epidemic playing out there in the next several years? Are you cautiously optimistic?

Doug: Anyone who's in AIDS and stays in it has to be an optimist. If you're a pessimist, you got to do something else for a living. I'm involved with cure research. I'm in health. You're either an optimist [00:29:00]or unrealistic in this business.

Emilio Emini: Hey there. Just to address that question, obviously, as you said, Doug, we all have to be the optimist but the challenges in, particularly, in Southern and Eastern Africa are driven by demographic issues. As you know, we're looking at, and you'll see when I talk about this tomorrow. There's an enormous number of young people who are just beginning to enter into that age [00:29:30] range where they become susceptible to HIV infection. If we just keep doing what we're currently doing in Sub-Saharan Africa, the absolute number of infected people 10 years from now is going to be greater than it was 10 years ago. What you said is absolutely correct. We'll talk about prevention tomorrow but from the treatment side, doing better with treatment because the tools are all there. There's some better tools coming along but making sure that they're delivered appropriately and making sure that they come in at a cost that is reasonable [00:30:00]particularly as the number of infected people going on therapy increases substantially, hopefully, over the next few years, is fundamentally critical. If that doesn't happen, then we will be actually in a worse place a few years from now than we are now.

Doug: For the last several years the PEPFAR funding has been flatlined, the European funding has diminished and so there are very practical issues. [00:30:30] I'm optimistic about the United States but I'm nervous as hell.

[laughter]

John Hamilton: John, two points. First, I think in a forum such as this, the goal of which is to create an accurate historical record it would be unfortunate not to mention the contributions of the Concorde Study and the VA Cooperative Study, which separately and independently established the limited time benefits [00:31:00] of early AZT therapy in the face of monotherapy. (16, 17) Secondly, and I see Marty left before I could involve her in this conversation, some derivative benefits of the VA Cooperative Study, at least, was the demonstration in a prospective clinical trial that the development of resistance actually accounted for the time-limited benefit of monotherapy, and secondly, we demonstrated that viral load [00:31:30] in a prospectively collected patient population clinical specimen, was a good surrogate marker for progression. I would hope that that would be included whatever record is created.

John Mellors: Sunday morning, John, I promise.

Wasif Khan: I'm going to bring it back the tolerance to mutations in reverse transcriptase. After all it's an enzyme. Is there a possibility or predictions, [00:32:00] which can be made by modeling or by designing, which can tell where would be the limit. You can't mutate the whole reverse transcriptase and still have function left.

Doug: If I remember correctly—

Wasif: Eventually, you would have, perhaps, a multi-component—

Doug: Maybe John [Coffin] can add to this but as I remember, Esteban Domingo has done a mutational analysis of almost every residue in the damn virus to look at resistance [00:32:30] and fitness and all that. All you have to do is treat patients with drugs and the virus will tell you what the sequence alternatives are going to be. The protease resistance is even more remarkable, where almost a third of the residues can be modified if you exert selective pressure with PI drugs.

John Coffin: Protease [00:33:00] is a very small, very compact, and what looks like a very tightly organized protein, yet 30% as you say, of the residues change almost within a few weeks when you start therapy.

John Mellors: Doug, I thought when you started your talk it was going to be ridiculous and then I violently was opposed to—

Doug: [crosstalk] the talk was going to be ridiculous?

John Mellors: —to some points you made, but in the end, I thought it was all self-evident.

[laughter] [00:33:30]

[clapping]

Thank you, Doug, that was terrific.

[clapping]

[00:33:36] [END OF AUDIO]

Citations

1. Larder, Brandon A., and Sharon D. Kemp. “Multiple Mutations in HIV-1 Reverse Transcriptase Confer High-Level Resistance to Zidovudine (AZT).” Science 246, no. 4934 (December 1, 1989): 1155–58. doi:10.1126/science.2479983.
2. Richman, Douglas, Cheng-Kon Shih, Israel Lowy, Janice Rose, Patricia Prodanovich, Stephen Goff, and Johanna Griffin. “Human Immunodeficiency Virus Type 1 Mutants Resistant to Nonnucleoside Inhibitors of Reverse Transcriptase Arise in Tissue Culture.” Proceedings of the National Academy of Sciences 88, no. 24 (December 15, 1991): 11241–45. doi:10.1073/pnas.88.24.11241.
3. Richman, Douglas D., Alan S. Rosenthal, Mark Skoog, Robert J. Eckner, Ting-CHao Chou, John P. Sabo, and Vincent J. Merluzzi. “BI-RG-587 Is Active against Zidovudine-Resistant Human Immunodeficiency Virus Type 1 and Synergistic with Zidovudine.” Antimicrobial Agents and Chemotherapy 35, no. 2 (February 1, 1991): 305–8. doi:10.1128/AAC.35.2.305.
4. Richman, Douglas D., Diane Havlir, Jacques Corbeil, David Looney, Caroline Ignacio, Stephen A. Spector, John Sullivan, et al. “Nevirapine Resistance Mutations of Human Immunodeficiency Virus Type 1 Selected during Therapy.” Journal of Virology 68, no. 3 (March 1, 1994): 1660–66.
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6. Richman, Douglas D. “The Implications of Drug Resistance for Strategies of Combination Antiviral Chemotherapy.” Antiviral Research, Consensus Symposium on Antiviral Therapy, 29, no. 1 (January 1996): 31–33. doi:10.1016/0166-3542(95)00911-6.
7. Richman, Douglas D., J. M. Grimes, and S. W. Lagakos. “Effect of Stage of Disease and Drug Dose on Zidovudine Susceptibilities of Isolates of Human Immunodeficiency Virus.” Journal of Acquired Immune Deficiency Syndromes 3, no. 8 (1990): 743–46.
8. Hermans, Paul E. “Antiviral Agents.” Mayo Clinic Proceedings 52, no. 11 (November 1977): 683–86.
9. Gulick, Roy M., John W. Mellors, Diane Havlir, Joseph J. Eron, Charles Gonzalez, Deborah McMahon, Douglas D. Richman, et al. “Treatment with Indinavir, Zidovudine, and Lamivudine in Adults with Human Immunodeficiency Virus Infection and Prior Antiretroviral Therapy.” New England Journal of Medicine 337, no. 11 (September 11, 1997): 734–39. doi:10.1056/NEJM199709113371102.
10. Montaner, Julio S. G., Peter Reiss, David Cooper, Stefano Vella, Marianne Harris, Brian Conway, Mark A. Wainberg, et al. “A Randomized, Double-Blind Trial Comparing Combinations of Nevirapine, Didanosine, and Zidovudine for HIV-Infected Patients: The INCAS Trial.” JAMA 279, no. 12 (March 25, 1998): 930–37. doi:10.1001/jama.279.12.930.
11. Richman, Douglas D, Sally C Morton, Terri Wrin, Nicholas Hellmann, Sandra Berry, Martin F Shapiro, and Samuel A Bozzette. “The Prevalence of Antiretroviral Drug Resistance in the United States:” AIDS 18, no. 10 (July 2004): 1393–1401. doi:10.1097/01.aids.0000131310.52526.c7.
12. Little, Susan J., Sarah Holte, Jean-Pierre Routy, Eric S. Daar, Marty Markowitz, Ann C. Collier, Richard A. Koup, et al. “Antiretroviral-Drug Resistance among Patients Recently Infected with HIV.” New England Journal of Medicine 347, no. 6 (August 8, 2002): 385–94. doi:10.1056/NEJMoa013552.
13. David Wohl et al., “Tenofovir Alafenamide (TAF) in a Single-Tablet Regimen in Initial HIV-1 Therapy” (Conference on Retroviruses and Opportunistic Infections, Seattle, Washington, February 23–26, 2015), https://www.croiconference.org/croi-2015/.
14. Scherrer, Alexandra U., Viktor von Wyl, Wan-Lin Yang, Roger D. Kouyos, Jürg Böni, Sabine Yerly, Thomas Klimkait, et al. “Emergence of Acquired HIV-1 Drug Resistance Almost Stopped in Switzerland: A 15-Year Prospective Cohort Analysis.” Clinical Infectious Diseases 62, no. 10 (May 15, 2016): 1310–17. doi:10.1093/cid/ciw128.
15. Stadeli, Kathryn M, and Douglas D Richman. “Rates of Emergence of HIV Drug Resistance in Resource-Limited Settings: A Systematic Review.” Antiviral Therapy 18, no. 1 (2012): 115–23. doi:10.3851/IMP2437.
16. Aboulker, Jean-Pierre, and AnnMarie Swart. “Preliminary Analysis of the Concorde Trial.” The Lancet, Originally published as Volume 1, Issue 8849, 341, no. 8849 (April 3, 1993): 889–90. doi:10.1016/0140-6736(93)93096-J.
17. St Clair, M. H., P. M. Hartigan, J. C. Andrews, C. L. Vavro, M. S. Simberkoff, and J. D. Hamilton. “Zidovudine Resistance, Syncytium-Inducing Phenotype, and HIV Disease Progression in a Case-Control Study. The VA Cooperative Study Group.” Journal of Acquired Immune Deficiency Syndromes 6, no. 8 (August 1993): 891–97.

Index

• 1.5 John Coffin — The Origin of Molecular Retrovirology 
• 2.1 Paul Volberding — The First Patients 
• 2.2 James Curran — Deciphering the Epidemiology of AIDS 
• 2.3 Mark Harrington — The Importance of Activism to the US Response 
• 2.5 Françoise Barré-Sinoussi — Discovery of HIV 
• 3.2 Samuel Broder: The First Clinical Trials of Antiretroviral Drugs 
• 3.6 John Martin — Making it Simpler: A Single Pill to Treat HIV 
• 6.5 Emilio Emini — Issues in HIV Vaccine Development: Will the Future be any Easier than the Past? 
• 8.1 John Mellors — MACS and Beyond: Epidemiology, Viremia and Pathogenesis 
• 11th International AIDS Conference, Vancouver, Canada, July 1996
• 3TC (lamivudine)
• Africa, sub-Saharan Africa 
• AIDS Clinical Trials Group (ACTG) 
• AZT (azidothymidine) 
• Barry, David Walter "Dave" (1944–2002) 
• Boehringer Ingelheim 
• Bureau of Biologics (FDA) 
• Burroughs-Wellcome & Company (GlaxoSmithKline) 
• cell culture, tissue culture, immortalized cell line 
• Chanock, Robert M. (1924–2010) 
• Chermann, Jean-Claude (b. 1939) 
• clinical trials (phases of clinical research) 
• cohort study
• cure vs. remission of HIV/AIDS
• ddC (dideoxycytidine, zalcitabine) 
• detergents and soaps 
• Domingo, Esteban 
• drug discovery and development 
• drug resistance 
• DTG (dolutegravir) 
• education and early career 
• Fischl, Margaret 
• fluorescent tag (fluorophore, fluorescent label, fluorescent probe, immunofluorescence) 
• foscarnet (phosphonomethanoic acid, Foscavir) 
• funding and grants 
• Gilead 
• Gludish, David W. 
• highly active antiretroviral therapy (HAART), combination antiretroviral therapy (cART) 
• HIV-1 Bru strain 
• idinivir (IDV, Crixivan) 
• in vitro vs. in vivo 
• infectious disease (medical specialty) 
• influenza 
• integrase inhibitors 
• interferons 
• Kemp, Sharon D. 
• lab safety, biosafety levels, safety protocol 
• Lange, Joep M. A. (1954–2014) 
• Larder, Brendan A. 
• Little, Susan J. 
• Malaysia Airlines Flight 17 (July 24, 2014) 
• mechanism 
• Merck 
• Merigan, Thomas C. (b. 1934) 
• military service and "Yellow Berets" 
• National Institutes of Health (NIH) 
• natural selection, evolutionary selection, evolutionary fitness 
• nevirapine (NVP, Viramune) 
• non-nucleoside reverse-transcriptase inhibitors (NNRTIs) 
• North America
• Nunberg, Jack 
• observation 
• p24 HIV capsid protein 
• PCR (polymerase chain reaction) 
• peer review 
• PEPFAR (President's Emergency Plan For AIDS Relief) 
• prevention of HIV/AIDS
• protease 
• protease inhibitors 
• Putin, Vladimir (b. 1952) 
• reverse transcriptase 
• Ryan, John 
• San Diego Veterans Administration Medical Center 
• Schopenhauer, Arthur (1788–1860) 
• sensitivity and specificity; false positive, false negative; biological specificity 
• specimen exchange 
• stock options 
• Temin, Howard M. (1934–1994) 
• tenofovir (TDF, Viread)
• United States
• van Tongeren, Jacqueline (1949–2014) 
• Vietnam War