DOCTOR FAX

ISSUE 28 1ST August 1997

Contents

• HIV DRUG RESISTANCE AND TREATMENT STRATEGIES PART 2

  A Report from the International Workshop on HIV Drug Resistance, Treatment Strategies and Eradication. 25-28 June 1997, St Petersburg, Florida, USA.

  

• Validation of Viral Load During Treatment as Prognostic Indicator

  Validation of HIV-1 RNA and CD4 count as surrogate markers in the CAESAR trial: preliminary results

  HIV-1 RNA quantification in 1280 patients in the Delta trial and relationship to clinical outcome

  Clinical, immunological and virological outcomes in ACTG 320

  

• The durability of response to protease inhibitor therapy is predicted by viral load
  

  Detection of HIV-1 RNA in the plasma of patients in whom HIV-1 RNA is undetectable using commercial assays

  Continued evolution of HIV-1 during combination therapy despite levels of HIV-1 <500 copies/ml

  Evidence for prevention of new HIV-1 infection cycles in patients treated with indinavir plus zidovudine plus lamivudine

  

• Eradication, immune restoration and long-term suppression strategies
  

  T cell immune reconstitution in HIV-negative hosts: inherent limitations and therapeutic prospects

  Partial immunologic recovery after 13 months of potent antiretroviral therapy

  Duration of HIV-1 load suppression in patients treated with indinavir

  

• OTHER NEWS

  Excess Renal Cell Carcinoma Prevalence Seen In HIV-Infected Patients

  Undetectable Antibody Reported in a Patient Infected With Typical HIV

  Sourced, Compiled and Edited by Paul Blanchard
  Medical Advisor - Dr Graeme Moyle , Chelsea & Westminster Hospital.

  

  HIV DRUG RESISTANCE AND TREATMENT STRATEGIES: PART 2

  A Report from the International Workshop on HIV Drug Resistance, Treatment Strategies and Eradication. 25-28 June 1997, St Petersburg, Florida, USA.

  This is the second of a two part report from the International Workshop. Delegates from the UK included AIDS Treatment Project Executive Director Raffi Babakhanian and leading clinicians Graeme Moyle, Mike Youle, Ian Weller and Ian Williams. Part 1 appeared in ATPs Doctor Fax issue 27 and focused on presentations at the conference on antiretroviral drug resistance and cross-resistance. Part 2 in this issue looks at presentations validating viral load as a surrogate marker of treatment response, durability of viral suppression and implications for treatment strategies.
  
  References: All abstract numbers refer to the: Program and Abstracts of the International Workshop on HIV Resistance, Treatment Strategies, and Eradication; St. Petersburg FL, USA, June 25-28, 1997.
  
  Source: Meeting reports by Graeme Moyle (Chelsea & Westminster Hospital), Raffi Babakhanian (AIDS Treatment Project), Steven G Deeks (HIV InSite) and William OBrien (healthcg.com).
  

  

  Validation of Viral Load During Treatment as Prognostic Indicator

  

  Validation of HIV-1 RNA and CD4 count as surrogate markers in the CAESAR trial: preliminary results

  A study presented by Julio Montaner on behalf of the CAESAR Coordinating Committee indicates the predictive value of reductions in viral load and increases in CD4 lymphocyte count for the clinical benefit of treatment (abstract 61). Validation of viral load changes as an outcome measure requires a large trial with a significant clinical benefit. This has previously only been demonstrated for a few clinical trials, although the practice of viral load monitoring is widely accepted. Such validation has also been suggested by virology substudies of ACTG 175 (see abstract 59 below). The CAESAR trial enrolled 1,895 patients with having a median baseline CD4 lymphocyte count of 134 cells/mm3 to receive placebo or lamivudine (3TC) or 3TC+ loviride in addition to their current antiretroviral regimen. The risk of progression to AIDS was reduced 60% in patients in whom 3TC was added. A subset of 794 patients had plasma HIV RNA levels measured to determine the proportion of the effect of adding 3TC to patients receiving ZDV therapy. The mean baseline viral load was nearly 100,000 copies/ml. Cox proportional hazard models were applied to calculate the proportion of treatment effect explained. Although either CD4 lymphocyte count increases or plasma HIV RNA decreases explained a substantial amount of the treatment effect (76% and 67%, respectively), using both of these markers explains essentially all of the treatment effect. The 95% confidence intervals were quite broad, but the lower limit explains the minimum effect seen, and further demonstrates that these markers are valid for assessing response to therapy. As use of these markers is validated in more and more trials, we will move farther away from the requirement for new antiretroviral treatments to demonstrate a clinical benefit. Because of the large number of potential treatments available, it will be impossible to maintain study patients in assigned, blinded treatment arms for very long when the effects of treatment on these markers are apparent to both patients and treatment providers.

  HIV-1 RNA quantification in 1280 patients in the Delta trial and relationship to clinical outcome

  The virologic sub-study from the Delta trial was presented by J. P. Alboulker (abstract 59). In this study of ZDV vs. ZDV/ddI vs. ZDV/ddC, approximately one third of the patients (1,280) had measurement of plasma samples using the NASBA assay which has an 800 copies/ml sensitivity. This is an excellent study for virologic analysis, since it was large, with a three year median clinical follow-up, and achievement of many clinical endpoints. The baseline viral load was 4.65 log10 copies/ml and the mean CD4 lymphocyte count was 205. The response to treatment in the combination arms approached 1.0 log, and was less than 0.5 log in the ZDV group. The proportion of patients with undetectable plasma HIV RNA (< 800 copies/ml) was higher in the combination groups with 61% of 299 ZDV/ddI patients, 40% of 304 ZDV/ddC patients, and only 9% of 295 ZDV patients in Delta I. Time to virological failure (defined as HIV RNA level of 0.5 log of both combination arms) detection was significantly longer in those that had a greater viral load reduction initially, and in those with a baseline HIV RNA of < 20,000 copies/ml. The conclusions reached in this virology substudy was that the virological response in the first 16 weeks was highly predictive of clinical outcome. CD4 cell counts were also highly predictive.

  Comment: Until recently, "body count" studies, in which survival was the only relevant outcome, was the gold standard in large scale clinical trials. The results from this analysis indicate that studies designed to analyse surrogate endpoints only are probably sufficient, and that large, expensive clinical endpoint studies may no longer be necessary.

  Clinical, immunological and virological outcomes in ACTG 320

  
  Despite the benefits seen with two drug RT inhibitor therapy, most HIV specialists now opt for more potent regimens which also include a protease inhibitor. Studies validating the predictive value of viral load response with protease inhibitor containing triple combinations were also presented in addition to those of the double NA combinations outlined above.
  
  Scott Hammer, Harvard Deaconess Medical Center, presented data from the ACTG 320 trial, in which 1,156 3TC and protease inhibitor-naive subjects were randomised to ZDV/3TC/indinavir or ZDV/3TC/placebo (abstract 67). Stavudine (d4T) substitution for ZDV was permitted. Despite a fairly advanced stage of disease with mean CD4 lymphocyte count less than 90 and mean viral load approximately 100,000 copies/ml, good treatment responses were achieved. CD4 lymphocyte count increases of greater than 120 cells/mm3 were seen at week 40, and viral load reductions below 500 copies/ml were seen in 40% of patients at 40 weeks. Importantly, there was clinical confirmation of the benefit of this therapy with a significant reduction in appearance of AIDS or death (63 or 11% vs. 33 or 6%; HR0.50, 95% CI 0.33, 0.76; P = 0.001). Thus, even in patients with advanced disease and prior ZDV therapy, addition of two new drugs can still result in a substantial clinical and virologic benefit. This presentation concluded that the HIV-1 disease marker response of viral load and CD4 count parallel the clinical results seen in this study.
  

  Comment: The ACTG 320 experience suggests that about one half of patients who initiate AZT, 3TC and indinavir late in the natural history of the disease process can expect to obtained long-term, complete viral suppression. This is a much lower number than previously reported (Merck 035).

  

  The durability of response to protease inhibitor therapy is predicted by viral load
  

  

  Clinicians clearly need tests which will allow them to predict who will fail a protease inhibitor, so modifications can be made before high level resistance develops. Kempf and his colleagues from Abbott reviewed clinical data from several ritonavir studies to determine which virological markers predict a durable response to protease inhibitor therapy.
  
  To be eligible for their initial analysis, patients must have reached a documented viral load "nadir" (i.e., their lowest viral load), and then rebounded with a virus containing at least two resistance mutations. Of the 29 patients studied, 6 received ritonavir monotherapy, 16 received zidovudine (AZT) plus ritonavir, and seven received the combination of ritonavir and saquinavir. Baseline RNA did not predict how long the patient would respond to a given therapy. The absolute value of the viral load nadir was highly predictive of the durability of viral suppression (R2=0.885): the lower the nadir, the longer the response. Patients who achieved levels below the level of quantification (< 200 copies/ml) had the longest response.
  
  To confirm this observation, Kempf and colleagues looked at 202 subjects who received at least 30 days of ritonavir or ritonavir plus AZT, and who had at least a 0.5 log decrease in viral load. Subjects who discontinued therapy were excluded form the analysis. Of the 152 patients eligible for analysis, baseline viral load at the time ritonavir was initiated did not predict duration of maximal response. Once again, subjects who achieved a viral RNA < 200 copies/ml had a durable response, while those who never went below 1000 copies/ml rebounded back toward baseline quickly. Most subjects reached their nadir by week 12.

  Nadir RNA (copies/ml)	> 1000	200-1000	200
  Time to failure (days)	60	128	199

  Notably, many patients who went below 200 copies did eventually rebound. It is unclear if more sensitive assays (lower limit of detection 20 copies/mL) could more accurately predict outcome [abstract 62]. These data underline the importance of achieving maximal treatment response with the initial therapy but also the concern that even patients achieving this response will ultimately fail.
  

  Comment: This data suggest that the viral load at week 12 of therapy with a protease inhibitor is highly predictive of outcome. Therefore, clinicians may want to modify or change therapy at week 12 if the viral load remains greater than 200 copies/ml and there is no evidence that it is still falling towards a nadir.

  Detection of HIV-1 RNA in the plasma of patients in whom HIV-1 RNA is undetectable using commercial assays
  Current standard of care is to treat HIV aggressively, with an explicit goal of attaining "undetectable" levels of plasma HIV-1 RNA (typically defined as a viral load less than 200 to 500 copies/ml). The natural history of patients who achieved undetectable levels remains unclear.
  
  In a study reported by Natarjan and colleagues, an ultrasensitive viral load assay (RT-PCR, lower level of detection 50 copies/ml) was used to better characterised patients with 5 months of documented viral load levels less than 500 copies/ml (Chiron, bDNA assay) or 400 copies/ml (Roche, RT-PCR assay). Most patients were receiving aggressive combination therapy. Of the 72 patients studied with the ultrasensitive assay, 50 had detectable levels (>50 copies/ml) [Natrajan et al, abstract 51].
  

  Comment: Plasma HIV-1 RNA can be readily detected in the majority of patients with so-called "undetectable levels" using current assays. This suggests that many patients have persistent low level viral replication, despite the use of combination therapy. Whether these patients are more likely to eventually fail a given regimen remains to be determined. Biologically, one can assume that a patient with a viral load < 50 copies/ml is more likely to achieve long-term viral suppression than a patient with a viral load of 50 to 500 copies/mL. Once these ultrasensitive assays are commercially available, clinicians should be able to identify patients at high risk for failing a given regimen, and therefore be able to intervene before high level resistance develops.

  
  

  Continued evolution of HIV-1 during combination therapy despite levels of HIV-1 < 500 copies/ml

  A group from the NIH performed intensive virological studies on four patients who attained long-term viral suppression on indinavir and IL-2. All four patients achieved undetectable levels of HIV-1 RNA, and remained there for at least two years. The investigators looked at viral genotype, both at baseline and 1.5 years into therapy. Surprisingly, in all 4 patients, the protease gene appeared to evolve over time. This observation, limited by technical difficulties in obtaining sufficient PCR products from patients with low levels of plasma HIV-1 RNA, indicates that viral evolution can occur in the presence of effective protease inhibitor therapy. If true, viral replication must be ongoing [Imanici et al, abstract 63].
  

  Comment: This provocative study may not be generalisable, since patients also received IL-2. However, it does raise concerns about the ability of current regimens to suppress viral replication indefinitely. Other studies presented at the meeting suggest that HIV does not evolve in the presence of potent therapies [Emini et al, abstract 128; Gunthard et al, abstract 66. See below].

  Evidence for prevention of new HIV-1 infection cycles in patients treated with indinavir plus zidovudine plus lamivudine

  
  Merck 035 is an important study of 90 patients treated with AZT + 3TC, indinavir monotherapy or AZT + 3TC + indinavir. Approximately 86% of patients receiving the three-drug combination had viral loads of < 500 copies/mL through 68 weeks of therapy. This impressive data set has established AZT + 3TC + indinavir as the standard of care in most clinical studies.
  
  Two groups performed intensive studies on these patients in an attempt to determine if viral replication is completely suppressed. Gunthard and colleagues in San Diego studied the emergence of genotypic resistance to 3TC in 10 patients. Since the M184V mutation typically occurs rapidly in the presence of 3TC, this group assumed that the lack of M184V after prolonged therapy indicates complete, or near complete, viral suppression.
  
  Viral sequences were analysed using automated dideoxynucleotide (ABI) and high-density oligonucleotide (Affymetrix) sequencing techniques. Plasma HIV-1 RNA and peripheral blood mononuclear cells (RNA, DNA) were obtained at baseline and after one year of treatment. Samples were also obtained from lymph node biopsies. In subjects with undetectable levels of HIV-1 RNA after one year of therapy, M184V did not evolve in each of the compartments evaluated [Gunthard et al, abstract
  66].
  
  The clinical scientists at Merck performed a similar analysis. During the course of Merck 035, several patients receiving AZT, 3TC and indinavir had transient elevations in viral RNA (to > 500 copies/mL) or persistent low levels of viral RNA (using ultrasensitive viral load assays). Genetic analysis from eight such patients revealed no new mutations in the reverse transcriptase or protease genes. One patient did have a new M184V, suggesting loss of viral control [Emini, abstract 128].
  

  Comment: These studies provide evidence that potent combination therapy can fully suppress viral replication. If true, resistance mutations theoretically can not evolve. Long term viral suppression, and possibly viral eradication, is therefore feasible.

  

  Eradication, immune restoration and long-term suppression strategies
  

  

  Updates on studies of recent seroconverter patients or chronically infected patients with high CD4 cell counts treated with 2 nucleoside analogues plus one or two PIs were presented. Follow-up has now reached a mean of 17.6 months. Presentation focused on the patients with viral load persistently below assay detection (100 copies/ml). Additional tested sites include CSF, semen, tonsil or lymph node and gastrointestinal lymphoid tissue obtained at sigmoidoscopy. Tissue co-culture and PCR for mRNA is consistently undetectable in all seroconverter patients. Similarly RNA PCR is undetectable in semen but DNA remains positive in all sites in all but one seroconverter patient. Decisions to stop treatment in any patients are not expected for > 1 year. (abstracts 126 & 127).
  
  Data from other areas of medicine such as oncology suggest that in adults (>16 years) it is unclear if CD4 repertoire will fully restore after substantial depletion possibly due to limitation or loss of thymic function. Immune restoration must therefore occur from peripheral expansion of remaining cells.
  
  

  T cell immune reconstitution in HIV-negative hosts: inherent limitations and therapeutic prospects

  
  The potential for immune reconstitution was reviewed in a plenary session by Crystal Mackall, National Cancer Institute (abstract 119). She reviewed the experience with immune reconstitution in patients treated for chemotherapy, as well as in many animal models. Using a murine bone marrow transplantation model, thymic-dependent T cell regeneration restores T cell numbers including na ve T cells in immune competent hosts within 12 weeks. In mice with insufficient thymic function, however, antigen-driven peripheral expansion of mature T cell populations predominates, and lymphopoiesis is significantly impaired. This results in a restricted restoration with limited T cell repertoire diversity, and impaired functional responses. The thymic-dependent pathways give rise to "naive" cells while the peripheral expansion pathways generate "memory" phenotype. The oncology experience with immune restoration indicates that younger children, having a functional thymus, recover immune competence much better than older patients.
  
  

  Partial immunologic recovery after 13 months of potent antiretroviral therapy

  
  The same phenomenon is now reported for HIV infection. In a 13 month interim report of 11 HIV-infected patients receiving ZDV/3TC/nelfinavir (abstract 123), recovery of naive CD4 lymphocytes was strongly related to patient age. Total mean CD4 lymphocyte count increased from 249 cells/mm3 baseline to 380 cells/mm3 at 13 months. Incomplete immune restoration in older individuals was demonstrated by report of a significant, inverse correlation between age and the naive/memory CD4 + lymphocyte ratio. This fell from approximately 0.5 - 0.1 for patients under age 30 to 0.1 for patients 50 or older. This suggests that a functional thymus is important for immune restoration in patients whose CD4 lymphocyte population has been depleted by chronic HIV infection. This may be encouraging, however, for the potential benefit of children treated with intensive therapy regimens.
  
  Restoration of CD8+ cells appears less impaired (abstract 119). Treatment at seroconversion with ZDV/3TC/IDV appeared to result in normalising of memory (RO) vs. na ve (RA) CD8+ lymphocytes compared with the loss of RA cells which typically occurs at and after seroconversion, suggesting possible maintenance of the immune repertoire (abstract 120). In 33 patients receiving triple combination therapy with ZDV/3TC/ritonavir initial rises in CD4 cell count were related to redistribution or expansion of CD4 memory cells over the first 3 weeks. A slower rise in these cells and eventually in na ve CD4 cells (abstract 122). In a similar cohort of 11 ZDV/3TC/nelfinavir treated patients followed for 13 months with viral load sustained below detection, the CD4+ na ve cell population increased from a mean 62 to 119/mm3 with similar normalisation of the CD8+ na ve/memory cell ratio to that reported in abstract 120 (abstract 123).
  
  In chronically infected patients on long term ZDV/3TC/indinavir treated in the small phase II study (Merck study 035) approximately 86% of patients achieved VL <500 copies/ml maintained at least 68 weeks treatment. Many patients have persistent or transiently detectable levels of viral RNA on ultra-sensitive assays up to 52 weeks, however in only 1 of 7 patients did a resistance associated mutation (an RT mutation 184Val for 3TC resistance) arise over this period. This mutation did not result in loss of virological control. These data, whilst at odds with the continued evolution of viral polymorphism, and appearance of IDV resistance mutation in one patient reported in another similarly treated group of 4 patients (abstract 63), suggest that low levels of detectable RNA in treated patients may reflect production from remaining chronically infected cells rather than continuing rounds of viral replication. Physicians need to be aware that, when more sensitive assays become more widely used, patients may take prolonged periods to achieve viral load counts below the detection of these assays, and that this detectable virus appears, in treatment adherent persons, not to represent a cause for concern. Of course, concern would arise if a nadir is reached which is at an unacceptably high level or drifts back towards baseline.
  
  

  Duration of HIV-1 load suppression in patients treated with indinavir

  
  In a review of 106 optimal responder patients selected from phase II indinavir studies who achieved VL <500 before week 24 and sustained this response for greater than 12 weeks gradual loss of virological control was reported over follow up (abstract 129).
  
  

  Follow-up time	24 weeks	48 weeks	68 weeks
  % <500 (95% CI)	97 (93-100)	87 (80-94)	82 (74-91)

  
  These data suggest that even in selected groups of optimal responders, triple combination therapy including a PI is associated with loss of virological response over prolonged follow-up. Thus while optimal responses may forestall resistance they will not necessarily prevent resistance. This suggests that strategic planning for management of both inadequate responders and of failure after initial optimal response is required at the initiation of therapy if HIV infection is to become a chronically manageable infection.
  
  Maintaining options may be achieved in some patients by subtraction therapy approaches starting with a multi-drug initial regimen to achieve a low viral load then withdrawing one or more agents to leave fewer agents as maintenance therapy. In 5 treatment na ve patients commencing ZDV/ddI/nevirapine who interrupted ddI therapy after achieving VL <20 copies/ml for >6 weeks were found to sustain response on 2 drugs, with follow-up in subjects beyond 1 year, suggesting the feasibility of this approach (abstract 130).
  
  
  

  

  OTHER NEWS

  

  Excess Renal Cell Carcinoma Prevalence Seen In HIV-Infected Patients
  

  
  New research from the Medical Center of Central Georgia indicates that the prevalence of renal cell carcinoma is over 8.5 times higher in HIV-positive hospitalized patients than in HIV-negative patients. Lead author Dr. Stephen Baynham and colleagues report in the June issue of AIDS Patient Care and STDs that 308 of 66,715 adults admitted to the medical center between 1990 and 1994 were HIV-positive. Two of the HIV- infected patients and 48 of the non-HIV patients had renal cell carcinoma. The researchers explained: "In our general adult hospital population, the probability of a patient having [renal cell carcinoma ... was estimated to 75/100,000. In our HIV adult hospital population, the estimated prevalence of [renal cell carcinoma] was 649/100,000."
  Source: AIDS Daily Summaries
  
  

  Undetectable Antibody Reported in a Patient Infected With Typical HIV
  

  
  In the July issue of Clinical Infectious Diseases, researchers present the case of a patient infected with a typical strain of HIV who repeatedly tested HIV seronegative. Reports show that over a four-year period of treatment for symptoms of immunosuppression, the patient tested negative for HIV antibodies 35 times. However, polymerase chain reaction and p24 assays proved the patient to be infected. Dr. Larry Reimer of the VA Medical Center in Salt Lake City and others said the case was unusual because the patient "had advance infection while he was still HIV seronegative." The researchers noted that "the most important implication of false-negative test results is the effect it may have on the blood supply." Moreover, Reimer added: "Physicians who treat patients with AIDS-defining conditions, but for whom standard HIV antibody testing is negative, should consider the possibility that HIV infection is present and may be identified by additional testing procedures."
  Source: AIDS Daily Summaries