Lower Than Usual but Detectable Levels of Replication Competent Proviral DNA found in Individuals on HU/ddI/Indinavir
Dr. Franco Lori, who has helped to drive hydroxyurea (HU) research, reported data from this study of 24 individuals who received HU+ddI+Indinavir. Ten of these individuals initiated treatment prior to seroconversion. Using a more senstive test than usual 3/24 individuals were tested and found to have low level replication competent proviral DNA. One person who discontinued treatment and remained off therapy for about 10 months had undetectable plasma viral load; but, using a RNA analysis allowing investigators to view 40 million cells, some cells expressing RNA were found.
The dosing regimen was: HU 300 mg tid (3x/day) if <60kg or 400 mg tid if >60 kg; ddI 200 mg bid; indinavir 800 mg 3x/day. In the USA, the standard HU dose is 500 mg bid but in Europe different mg pills are available. Lori has suggested that tid dosing may be superior to bid dosing. Bristol Myers is planning a study to explore tid vs bid dosing.
Lori said this combination of drugs (protease, NRTI and HU) was selected because they have different targets (protease enzyme, reverse transcriptase enzyme, and a cellualr target by HU of ribonuceotide reductase). They are also hoping two of these drugs (ddI and HU) would efficiently target the cells, such as, macrophages and quiescent lymphocytes which are supposed to be the main reservoir for longer term HIV. A protease inhibitor is added to target activated lymphocytes. See Table 27
The addition of a protease inhibitor with d4T or other drugs will help to increase CD4. See Table 28 for study results: viral loadchanges in semen, blood and lymph tissue.
Among the group who was treated prior to seroconversion, 5 or 6 had Western Blot never became completely positive. One patient remains that way after 18 months. Lori said that is consistent with a strong reduction of virus replication which could be due to fewer antigens presenting itself.
Safety. Lori briefly discussed the concern that absolute neutrophil count can decline from HU treatment. In this study he said it was not a problem. He believes that the HU dosing regimen is important to the effect on ANC. He has suggested that a ANC of 1700 before initiating HU therapy might be a cutoff below which could develop a problem after starting HU therapy. He said, with regards to developing an ANC problem, a persons baseline ANC is more relevant than their CD4. Thrombocytopenia, which is an abnormal lowering of your platelet count, can be a concern from HU. Lori said he saw 1 case in a study of 40 patients receiving HU. If it occurs, he said it should emerge in the first 6-8 weeks.
Low Levels of RNA and Replication Competent DNA are Detectable. Lori discussed one interested patient in the study. This person was infected 57 days prior to starting treatment. Data suggests that by this time there is a steady state viral load (set point). Lori said this patients viral load would not have declined this much without treatment. Their baseline plasma viral load was about 800,000 copies/ml. The viral load went down fast to undetectable. At 39 days after starting treatment, following reaching undetectable, the patient stopped treatment due to an episode of orchitis. Immediately the viral load started to rebound. After starting treatment again at day 42, viral load went back to undetectable. 141 days after starting treatment the patient developed an episode of hepatitis A and the person couldnt take study drugs for 3 weeks, but although concomitant infections can cause viral load rebound, in this case viral load remained undetectable. Then he started treatment again but about a month later, which was about 200 days after starting study treatment, the patient decided to discontinue study drugs. Up til now which is about 460 days after starting treatment the patients plasma HIV RNA remains undetectable. 72 days after stopping treatment investigators analyzed a lymph node. They were unable to detect HIV DNA. They used a nested PCR system that allowed them to go down to one copy. But that testing system is limited to analyzing a maximum of 300,000 cells. Using a RNA analysis which allows them to look at over 40 million cells, and they found cells that were expressing low level RNA.
Lori sent samples from 3 acutely infected individuals in this study to Robert Silicianos lab. In Silicianos lab, it was their experience that they could routinely recover replication competent HIV DNA with a frequency of 0.2 to 16.4 per 105 cells after HAART. Initially, no replication competent HIV was recovered in 2/3 patients. The other patient was the one who interrupted therapy 12 months before testing. The sensitivity of the test was increased 10 times (60 million cells screened), and low levels of replication competent DNA were recovered in these two patients with a frequency of 1 cell/10 million. Why has that one person who discontinued therapy remained undetectable in blood? Lori said, this could be because the virus may not be replication competent in vivo, or because some immunological changes are able to keep the virus under control.
A potentially important mechanism of action for HU, in individuals treated during acute infection as these study participants were, may be to block cell activation and thus limit viral targets and viral production. If you have less activated cells you should have less virus. There are fewer cells available to be infected. As well, using HU with ddI could prevent ddI resistance from being a problem. Data has suggested that even if ddI resistance develops the combination of HU+ddI might allow ddI to be just as effective as if ddI resistance did not occur.
Immunology. For this substudy, a group of 8 treated participants were compared to 8 untreated individuals with a similar time of infection. The treated groups CD4 counts increased, their CD8 counts decreased and therefore their CD4/CD8 ratio increased.
The percentage of CD3-Zeta expression on CD4 and CD8 cells, which is supposed torepresent t-cell functioning, was significantly higher in the treated group than the untreated group. The mean proliferation response to flu (twice as high) and also antigen was higher in the treated than in the untreated groups. The treated group had significantly more naive CD4 and CD8 cells than the untreated group. CD38 and HLA-DR expression on CD8 cells, which represents cell activation, were significantly lower in the treated group compared to the untreated group. CD28 expression, a co-stimulatory t-cell molecule that is essential for t-cell proliferation against antigen, was expressed at higher levels in the treated group than in the untreated group. (ed note: a comparison study might be necessary to detect if these immunological improvements are any more than would be seen with any fully suppressive regimen.
Lori concluded that further studies are required to evaluate these cresults. In fact, HU is getting much attention and many new studies are planned to explore its use in a variety of situations including salvage therapy, acute infection and early infection.
HU+ddI+d4T: Swiss study
In NATAP Reports Jan 98 issue, also available on this web site, the 24 week data from this study was reported. 144 patients were initially randomized to receive ddI+d4T or ddI+d4T+HU. After 24 weeks, 23 of 26 evaluable patients initially randomized to the HU regimen had <200 copies/ml of viral load. At week 48, which was reported at Retrovirus, 16/20 were reported to have <200 copies/ml.
Table 27. Different Mechanisms and Different Targets of HU+ddI+IDV
Table 28. Results - Viral Load Changes in Semen, Blood and Lymph Tissue
* by in situ hybridization
** of these two patients one also had undetectable HIV RNA by in situ hybridization while the other was detectable
*** in the Genital Secretion Report inside this issue, it was reported that even if seminal seminal viral load was <400 copies/ml using a standard test, DNA was detectable and RNA was detectable using a more sensitive test