What Will It Take to Cure HIV?

IAS–USA Topics in Antiviral Medicine

Perspective What Will It Take to Cure HIV?

Investigational strategies to attempt HIV cure or remission include very detectable virus during 3 years of treat- early initiation of antiretroviral therapy to limit the latent HIV reservoir and ment but exhibited viral rebound 2 preinfection vaccination. In the setting of viral suppression, strategies include weeks to 3 weeks after stopping treat- reactivation of latently infected cells (eg, through “shock” therapy with ment. Similarly, an HIV-infected baby histone deacetylase inhibitors or other agents); use of broadly neutralizing in Milan began antiretroviral treatment antibodies, therapeutic vaccines, immunotoxins, or other immune-based 12 hours after birth and had no detect- therapies to kill latently infected cells; and gene editing to induce target able virus for 3 years while receiving cell resistance (eg, by eliminating the CC chemokine receptor 5 [CCR5] treatment but exhibited viral rebound 2 coreceptor). Improved ability to detect and quantify very low levels of virus weeks to 3 weeks after stopping treat- is needed. This article summarizes a presentation by Jintanat Ananworanich, ment.5-9 MD, PhD, at the IAS–USA continuing education program held in New York, In general, it is believed that earlier New York, in October 2014. initiation of antiretroviral therapy and the longer it is maintained improve the Keywords: HIV, cure, remission, viral reservoir, latent infection chance of limiting the viral reservoir and achieving HIV remission. How- ever, among the infants who eventually HIV cure research currently focuses 7 months, respectively, after receiv- exhibited rebound after stopping anti- on 2 goals: complete eradication of ing bone marrow transplantation with retroviral therapy, time to viral rebound HIV from the body and HIV remission. cells that included the CCR5 corecep- was longest for the Mississippi baby During remission, virus can still be de- tor. The HIV-infected Mississippi baby, despite a later start of therapy (30 tected at low levels in cells or through born to an HIV-infected mother who hours) than the Canadian baby (<24 ultrasensitive testing in plasma, but did not receive antiretroviral therapy, hours) or the Milan baby (12 hours) antiretroviral therapy is not necessary began antiretroviral therapy at 30 hours and a shorter duration of therapy (18 unless continued monitoring reveals after birth. HIV infection was confirmed months, 3 years, and 3 years, respec- increased viremia. and the infant achieved viral suppres- tively). The baseline HIV RNA level of sion 1 month after starting treatment. the Mississippi baby (19,812 copies/ Cases of Remission and Effects Treatment was stopped after 18 months mL) was between that of the Canadian of Antiretroviral Therapy in because the child was lost to follow- baby (808 copies/mL) and the Milan Infants With Early Acute HIV up care, and no virus was detected baby (152,560 copies/mL). The Missis- Infection for a subsequent 27 months. In mid- sippi baby’s longer remission period 2014, the child’s plasma HIV RNA level might be partially explained by the Timothy R. Brown, also referred to as rebounded to 10,000 copies/mL to shorter duration of antiretroviral ther- the Berlin patient, remains the only 16,000 copies/mL, and it was subse- apy before an HIV RNA level of less known person in whom HIV appears quently determined that HIV RNA level than 50 copies/mL was achieved (1 to be eradicated. Brown stopped taking had increased to 9 copies/mL 2 weeks month of therapy for the Mississippi 1-4 antiretroviral therapy approximately 7 before the rebound. baby vs 6 months for the Canadian years ago and has had no detectable There have been several other cases baby and 3 months for the Milan baby). virus capable of replicating since. The of infants who received antiretroviral Testing performed while the Missis- mechanism of the possible eradica- therapy during early acute HIV infec- sippi baby was in remission and was tion in this case included bone marrow tion, resulting in the absence of de- not taking antiretroviral therapy and the transplantation with cells lacking the tectable virus. An HIV-infected baby in Milan baby and Canadian baby were CC chemokine receptor 5 (CCR5) core- California who received antiretroviral taking antiretroviral therapy showed ceptor, rendering the cells resistant treatment at 4 hours after birth contin- that whereas all 3 had negative test to HIV infection. Two other patients, ued therapy for 14 months with no de- results for HIV DNA in peripheral blood, referred to as the Boston patients, ex- tectable cellular or plasma virus. Four replication-competent virus, and anti- perienced HIV remission 3 months and HIV-infected Canadian babies who re- HIV antibody, the Canadian baby and ceived antiretroviral treatment within the Milan baby displayed evidence sug- their first 24 hours have remained on gestive of ongoing viral replication. The Dr Ananworanich is Associate Director for HIV Therapeutics Research at the US Military treatment for 2.5 years to 7 years with Milan baby had detectable HIV-specific HIV Research Program and the Henry M. no detectable virus. Another Canadian T cells and a high percentage of acti- Jackson Foundation for the Advancement baby who received antiretroviral treat- vated T cells, and the Canadian baby of Military Medicine in Bethesda, Maryland. ment within 24 hours of birth had no had detectable cell-associated HIV RNA 80 HIV Cure Volume 23 Issue 2 May/June 2015

6 (A) Blood DNA 6 (B) Lymph node DNA central memory cells are longer lived 5 5 and constitute a latently infected cel- cells cells 6 6 lular reservoir during HIV infection. 4 4 During chronic HIV infection, the la- 3 3 tent reservoir is large and central 2 2 memory CD4+ cell infections still con-

DNA copies/10 DNA copies/10 stitute a major part of the latently 1 1 10 10 infected cell pool even after years of Log Log 0 0 0 20 40 60 80 100 120 140 160 180 0 20 40 60 80 100 120 140 160 180 antiretroviral therapy. The reservoir size is much smaller during acute HIV 6 (C) Gut DNA 8 (D) Blood RNA infection, with years of antiretroviral 7 5 cells therapy resulting in a marked decrease 6 6 4 in latently infected cells. 5 Levels of integrated HIV DNA were 3 4 ART interruption examined among HIV-infected patients 3 2 in Thailand who started antiretroviral 2 DNA copies/10

10 1 therapy within 2 weeks of infection, 1

Log 0 0 within 3 weeks to 4 weeks of infec- 0 20 40 60 80 100 120 140 160 180 HIV RNA in Blood (copies/mL) 0 50 100 150 200 250 300 tion, or during chronic infection, and Time After SIV Infection (Days) had plasma viral loads below detection limits at 2 years.12 Starting treatment Figure 1. Simian immunodeficiency virus (SIV) proviral DNA was detected in the lymph nodes within the first 2 weeks of infection (B) and gut (C) but not in peripheral blood (A) after antiretroviral therapy (ART) was initiated in resulted in much lower levels of inte- macaques 3 days after infection. All macaques had detectable blood HIV RNA after interrup- grated HIV DNA in all CD4+ cell sub- 11 tion of ART (D). Adapted from Whitney et al. sets than when antiretroviral therapy was initiated during chronic HIV infec- levels; these findings were not present Is Early Antiretroviral Therapy tion, and persistence of the viral res- in the Mississippi baby.2,5,8 Crucial to Limiting HIV ervoir was intermediate among pa- These cases of transient HIV remis- Persistence? tients starting antiretroviral therapy at sion also show how current tools are 3 weeks to 4 weeks after infection. limited in their ability to detect low Initiation of antiretroviral therapy in Studies in the VISCONTI (Virological numbers of HIV-infected cells. Acute macaques 3 days after establishment and Immunological Studies in Con- HIV infection is associated with low of simian immunodeficiency virus (SIV) trollers After Treatment Interruption) levels of HIV DNA, which are further infection resulted in undetectable pro- cohort of posttreatment controllers— reduced by antiretroviral therapy dur- viral DNA levels in peripheral blood a group of 14 patients in France who ing early acute infection to levels that but not in the lymph nodes or gut, started antiretroviral therapy early may be undetectable. Currently, whether with DNA levels at these sites declining and exhibited control of viremia after virus is present or in what amount it during 6 months of treatment (Figure interrupting treatment for 6 years or is present cannot be determined below 1).11 Viral rebound was observed when more—showed that early treatment the detection limits of current assays. antiretroviral treatment was stopped, could skew the distribution of latently Potential methods for measuring on- suggesting that seeding of the viral HIV-infected cells to shorter-lived tran- going viral replication in the reser- reservoir begins very early and that sitional memory cells that may be voir include measurement of cell- initiation of treatment at 3 days in this more rapidly cleared by the immune associated HIV RNA, single-copy HIV animal model is not early enough to system (Figure 2).13 RNA, and replication-competent vi- prevent it. rus using viral outgrowth assays. Re- The size and composition of the la- Strategies to Eliminate HIV cently, it has been shown that the tent HIV reservoir are affected by early Persistence replication-competent HIV reservoir antiretroviral therapy. In basic CD4+ may be 60 times greater than what cell differentiation, stimulation of naive One HIV remission and cure strategy is currently measured by viral out- CD4+ cells by antigens causes them to currently under investigation is pre- growth assay, as there are viruses differentiate into memory CD4+ cells infection vaccination. In a study of 16 that are intact but not induced by this that consist of stem cell, central, tran- macaques, a cytomegalovirus (CMV)- method.10 Investigators are examining sitional, and effector memory CD4+ vector SIV vaccine given prior to SIV the use of inducible HIV RNA assays cells. The shorter-lived transitional and infection did not prevent infection but to activate HIV-infected cells, in an effector memory cells are more likely did result in control of viremia in 9 ma- attempt to measure reservoirs capable to differentiate into terminally differ- caques and in SIV eradication in 8 of replicating. entiated cells and then die, whereas macaques, with no evidence of virus in

81 IAS–USA Topics in Antiviral Medicine

P = .001 Strategies to kill latently a pseudomonas endotoxin and antiret- 18 P = .003 HIV-infected cells include roviral therapy was used (Figure 3).

P = .007 use of broadly neutralizing Inducing resistance to HIV infection 100 antibodies and vaccines. in cells has been examined by using Transitional memory P = .003 Broadly neutralizing anti- gene therapy to eliminate CCR5 core- 80 CD4+ T cells NS bodies bind cell-free virus ceptor. In one study, HIV-infected pa- P = .032 and might clear infected tients underwent leukapheresis and Central 60 memory Effector cells. In one study, adminis- their cells underwent gene editing with CD4+ T cells memory tration of 1 or 2 doses of the zinc finger nucleases to remove CCR5 CD4+ T cells 40 broadly neutralizing mono- genes. Modified cells that no longer ex- clonal antibody PGT121 to pressed CCR5 were then proliferated HIV Reservior (%) 20 Naive macaques with low base- and reinfused into the same patient. Contribution to Resting CD4+ T cells line chimeric SIV/HIV (SHIV) HIV DNA levels were reduced but in- 0 viral load resulted in remis- terruption of antiretroviral therapy re- Posttreatment Controllers sion of virus for more than sulted in viral rebound in all patients. 1 year, long after PGT121 However, viral rebound was followed Figure 2. The HIV reservoir skewed to shorter-lived tran- was no longer detectable by spontaneous control of virus in 1 pa- sitional memory cell subsets in the VISCONTI (Virological in the blood.17 Numerous tient who was heterozygous for the and Immunological Studies in Controllers After Treatment Interruption) cohort of posttreatment controllers. NS in- such monoclonal antibod- CCR5 gene and who exhibited the high- dicates not a statistically significant difference. Adapted ies are being investigated for est level of engraftment of the modified 19 from Saez-Cirion et al.13 use in humans. A study of cells (see Tebas et al, 2014 ). Studies are monoclonal antibody VRC01 currently examining additional doses organs or blood in the latter group.14 administered during acute HIV infec- of the modified cells and use of che- This response likely reflects the ability tion is planned, and the AIDS Clinical motherapy preconditioning to improve of the replicating CMV vector to gen- Trials Group (ACTG) network is plan- engraftment. erate ongoing immune responses. In ning a study of this antibody in chroni- responding animals, the vaccine was cally HIV-infected patients. Summary able to generate a very rapid early Immunotoxins may also be used mucosal immune response that con- to kill HIV-infected cells, particularly HIV cure strategies currently being ex- tained the virus before drastic systemic in tissue. When this strategy was pre- amined in human studies (Figure 420) spread of infection. Further, immune viously assessed as monotherapy it begin with minimizing the HIV reser- response was not limited to immune- was ineffective, but it has since pro- voir through early antiretroviral ther- dominant epitopes, with the broadness duced promising results when used apy and use of broadly neutralizing of response preventing viral escape. in combination with antiretroviral antibodies. Once viral load is sup- A prime-boost adenovirus 26 and therapy. A study in humanized mice pressed, latently infected cells can be modified vaccinia virus Ankara (MVA) showed a marked reduction in HIV reactivated (eg, with HDAC inhibitors vaccine resulted in control of SIV vir- RNA-positive cells in tissue when a or activation of toll-like receptors or emia in 3 of 8 macaques. Control of combination of an immunotoxin with protein kinase C) and immune-based viremia was associated with a broad CD8+ cell response and neutralizing 10 8 No antiretroviral therapy Antiretroviral therapy only Antiretroviral therapy and antibody response.15 This vaccine for immunotoxin 3B3 - PE38 HIV will be investigated in patients 10 7 starting antiretroviral therapy during early acute HIV infection, to determine 10 6 whether it may help them achieve HIV remission. 10 5 Another remission strategy, in the setting of suppressed HIV viral load, is 4

Gram of Tissue 10 the use of “shock” therapy to activate

latently infected cells to produce virus. Cells/HIV RNA-Producing 3 In a study using the histone deacety- 10 lase (HDAC) inhibitor romidepsin as shock therapy, reactivation of HIV was 10 2 observed but was not accompanied by Tissue Samples From Mice a reduction in HIV DNA level, indicat- ing that few, if any, HIV-infected cells Figure 3. Effects of immunotoxin 3B3-PE38 combined with antiretroviral therapy on HIV RNA were killed after reactivation.16 in tissue in mice. Adapted from Denton et al.18

82 HIV Cure Volume 23 Issue 2 May/June 2015

References 1. Hutter G, Nowak D, Mossner M, et al. Long-term control of HIV by CCR5 Del- ta32/Delta32 stem-cell transplantation. N Engl J Med. 2009;360(7):692-698. 2. Persaud D, Gay H, Ziemniak C, et al. Ab- sence of detectable HIV-1 viremia after treatment cessation in an infant. N Engl J Med. 2013;369(19):1828-1835. 3. Henrich TJ, Hanhauser E, Marty FM, et al. Antiretroviral-free HIV-1 remission and viral rebound after allogeneic stem cell transplantation: report of 2 cases. Ann Intern Med. 2014;161(5):319-327. 4. Luzuriaga K, Gay H, Ziemniak C, et al. Vi- remic relapse after HIV-1 remission in a perinatally infected child. N Engl J Med. 2015;372(8):786-788. 5. Giacomet V, Trabattoni D, Zanchetta N, et al. No cure of HIV infection in a child despite early treatment and apparent viral clearance. Lancet. 2014;384(9950):1320. Figure 4. Different HIV cure strategies currently used in human studies that may need to 6. Rainwater-Lovett K, Luzuriaga K, Persaud be combined in order to achieve HIV remission. CCR5 incidates CC chemokine receptor 5; D. Very early combination antiretroviral PD, programmed cell death.20 therapy in infants: prospects for cure. Curr Opin HIV AIDS. 2015;10(1):4-11. 7. Bitnun A, Samson L, Chun TW, et al. Early therapies (eg, broadly neutralizing an- the tools used to measure the HIV res- initiation of combination antiretroviral therapy in HIV-1-infected newborns can tibodies, therapeutic vaccines, or anti– ervoir in tissue and blood. achieve sustained virologic suppression programmed cell death [PD] 1 and In conclusion, SIV and HIV eradica- with low frequency of CD4+ T cells car- anti–PD ligand 1 agents) can be used tion has thus far been achieved via a rying HIV in peripheral blood. Clin Infect Dis. 2014;59(7):1012-1019. to kill HIV-infected cells, or cells can be CMV vector vaccine that maintained 8. Brophy J, Chun TW, Samson L et al. 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In Special Issue: sion or cure has been achieved is chal- HIV will be difficult or impossible to Abstracts From the 2014 Conference on lenging. Treatment cessation is the ulti- achieve in the near future. HIV remis- Retroviruses and Opportunistic Infections. mate test of HIV remission, and en- sion is a more attainable goal that will Top Antivir Med. 2014;(e-1):37-38. 10. Ho YC, Shan L, Hosmane NN, et al. Repli- suring safety requires frequent moni- require testing of combination therapies cation-competent noninduced proviruses toring of viral load and a low threshold to reduce the size of the HIV reservoir in the latent reservoir increase barrier to for restarting treatment.21 In some plan- and boost HIV-specific immunity. In HIV-1 cure. Cell. 2013;155(3):540-551. 11. Whitney JB, Hill AL, Sanisetty S, et al. ned studies, viral load will be measured this early stage of HIV cure research, Rapid seeding of the viral reservoir prior to every 3 days to 7 days and treatment there will be many disappointments, SIV viraemia in rhesus monkeys. Nature. will be restarted with viral recrudes- but it is important to iteratively learn 2014;512(7512):74-77. 12. Ananworanich J, Vandergeeten C, Chom- cence. Many of the drugs being tested from these and steadily move the field chey N, et al. Early ART intervention re- for HIV cure are cancer drugs with po- forward. stricts the seeding of the HIV reservoir in long-lived central memory CD4 T cells tential toxic efftects and many require [Abstract 47]. 20th Conference on Ret- intravenous infusion. Studies of the var- Presented by Dr Ananworanich in October roviruses and Opportunistic Infections ious HIV cure strategies will be a major 2014. First draft prepared from transcripts (CROI). March 3-6, 2013; Atlanta, Georgia. by Matthew Stenger. Reviewed and edited by 13. Saez-Cirion A, Bacchus C, Hocqueloux burden to patients, requiring frequent L, et al. Post-treatment HIV-1 controllers follow-up, blood draws, and tissue sam- Dr Ananworanich in March 2015. The views with a long-term virological remission af- expressed are those of the authors and should pling. Because there are many pro- ter the interruption of early initiated anti- not be construed to represent the positions of retroviral therapy ANRS VISCONTI Study. posed treatments in the pipeline and the US Army or the Department of Defense. PLoS Pathog. 2013;9(3):e1003211. because combination treatments will be 14. Hansen SG, Piatak M, Jr., Ventura AB, et al. Immune clearance of highly patho- needed, novel study designs to quickly Financial affiliations in the past 12 months: genic SIV infection. 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83 IAS–USA Topics in Antiviral Medicine

16. Kjaer A, Sogaard OS, Graverson ME, et al. 18. Denton PW, Long JM, Wietgrefe SW, et al. 21. Rothenberger MK, Keele BF, Wietgrefe The HDAC inhibitor romidepsin is safe Targeted cytotoxic therapy kills persisting SW, et al. Large number of rebounding/ and effectively reverses HIV-1 latency in HIV infected cells during ART. PLoS Pathog. founder HIV variants emerge from multi- vivo as measured by standard clinical as- 2014;10(1):e1003872. focal infection in lymphatic tissues after says [TUAA0106LB]. 20th International treatment interruption. Proc Natl Acad Sci AIDS Conference. July 20-25, 2014; Mel- 19. Tebas P, Stein D, Tang WW, et al. Gene USA. 2015;112(10):E1126-E1134. bourne, Australia. editing of CCR5 in autologous CD4 T cells of persons infected with HIV. N Engl J 17. Barouch DH, Whitney JB, Moldt B, et al. Med. 2014;370(10):901-910. Therapeutic efficacy of potent neutralizing HIV-1-specific monoclonal antibodies 20. Ananworanich J, Fauci AS. HIV cure re- in SHIV-infected rhesus monkeys. Nature. search: a formidable challenge. Journal of Top Antivir Med. 2015;23(2):80-84. 2013;503(7475):224-228. Virus Education. 2015;1:1-3. ©2015, IAS–USA. All rights reserved

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