CROI 2020 Abstract eBook

Abstract eBook

Poster Abstracts

the Duesseldorf patient (IciStem#19). This HIV-infected male patient (50y, heterozygous CCR5∆32 allele) received unmodified stem-cell transplantation (SCT) from a 10/10 matched CCR5Δ32/Δ32 donor in Feb/13 for acute myeloid leukemia. At time of SCT complete western blot pattern was detected, proviral load was 1.45 log 10 cop/Mio PBMCs with R5-coreceptor-tropism. In Jun/13 complete remission was achieved by 5-Azacytidine and donor lymphocyte infusions (DLI) after a 2nd relapse. PBMC were negative for HIV-DNA by qPCR/ ddPCR during relapse and thereafter. However, in T-cell subsets few positive signals were observed. qVOA/mVOA were negative. Biopsies: CSF Jul/14, rectum Apr/15+Mar/16, ileumMar/16 and bone marrow Aug/15, and lymph nodes (LN) May/17 were HIV-DNA negative by PCR. In situ hybridization assays (RNAscope, DNAscope) detected few positive signals in LN. Moderate acute and mild chronic GvHD occurred after DLI but Tacrolimus could be finally stopped in Oct/17. He remained on ART with undetectable plasma VL until analytic therapy interruption (ATI) in Nov/18. Methods: PBMC/tissues analysed by ddPCR/qPCR and in situ hybridization. T-cell responses with peptide stimulation assays. qVOA analysed on CD4+T- cells. Drug level assessment by liquid chromatography mass spectrometry. Results: After ATI no antiretrovirals could be detected in multiple plasma samples. In Jul/19 no HIV DNA was detected in CD45+ cells extracted from biopsies (duodenum/ileum/rectum). Neutrophils and IFN-1 responses in the GI tract were very low. CD4 T cells were abundant within GI tract follicular aggregates, RNAscope was negative, DNAscope showed few positive signals, but not clearly above the false detection rate. In Nov/19, 12 mo after ATI, HIV DNA was negative in naive, central memory, transitional memory, and effector/ effector memory CD4 T-cells. qVOA in total CD4 T cells was also negative. Peptide stimulation assays showed CCR5-negative HIV-specific CTL with loss of recognition of RTYV9-specific and decrease of Gag-specific CTL after stopping immunosuppression. The absence of HIV-antigen is confirmed by fading humoral reactivity. Conclusion: No viral rebound was observed for 14 months following ATI, 83 months after allogeneic CCR5Δ32 SCT. In depth analyses of the viral reservoir still showed traces of HIV DNA in LN and GI tract, not clearly representing infectious virus though, since all functional assays were negative. These results are compatible with sustained remission of HIV. 349LB EDITING OF SIV IN NONHUMAN PRIMATES BY CRISPR-CAS9 IN VIRAL RESERVOIRS Jennifer Gordon 1 , Tricia H. Burdo 1 , Pietro Mancuso 1 , Chen Chen 1 , Rafal Kaminski 1 , Mark G. Lewis 2 , Kamel Khalili 1 1 Temple University, Philadelphia, PA, USA, 2 BIOQUAL, Inc, Rockville, MD, USA Background: Antiretroviral therapy (ART) suppresses but does not eliminate replication competent HIV proviral DNA from latently infected cells, thus resulting in viral reactivation upon ART cessation. Therefore, removal of HIV proviral DNA from infected individuals is needed. We have assessed a CRISPR- Cas9 based gene editing strategy for the elimination of the SIV proviral DNA in the rhesus macaque model. Methods: An all-in-one AAV9 gene therapy vector was constructed to deliver CRISPR-Cas9 plus two gRNAs targeting sequences within the 5' and 3' viral LTRs and the Gag gene to excise the intervening proviral DNA fragment. Ten adult Indian rhesus macaques were i.v. infected with SIVmac239 then treated daily with a drug regimen of tenofovir, emtricitabine and dolutegravir (5.1/50/2.5mg/ kg daily s.q.). Animals were randomized into groups to receive low versus high dose of AAV9-CRISPR-Cas9 in a single i.v. infusion (low dose: 1.4x10 12 GC/kg n=4; high dose: 1.4x10 13 GC/kg n=3) as well as control SIV infected animals (n=3). Longitudinal blood samples and lymph node biopsies were collected, and animals were necropsied at 3 (n=8) or 6 months (n=2) after CRISPR treatment. Results: SIV-infected animals treated with AAV9-CRISPR-Cas9 at both high and low doses showed vivo excision of viral DNA from serial blood and lymph node samples. Results from Sanger sequencing confirmed the precise breakpoint of the viral DNA in samples in which excision was detected. Biodistribution of the AAV9-CRISPR-Cas9 vector was assessed by PCR to detect the presence of the Cas9 gene sequence. DNA and RNA scope were performed on lymph nodes in parallel to detect the AAV9-CRISPR-Cas9 viral vector and expression of the Cas9 gene. Broad excision of SIV proviral DNA was observed in lymph nodes and other tissues known to be viral reservoirs including spleen, gut, and brain. A dose response between low and high doses, as well as temporal distribution between 3 and 6 months, was observed for AAV9-CRISPR-Cas9 viral DNA in the blood.

347LB SUSTAINED REMISSION IN A 4-YEAR-OLD HIV-INFECTED CHILD TREATED IN FIRST YEAR OF LIFE Gloria P. Heresi 1 , Douglas D. Richman 2 , Roukaya Al Hammoud 1 , Gilhen Rodriguez 1 , Norma Perez 1 , James R. Murphy 1 1 University of Texas at Houston, Houston, TX, USA, 2 University of California San Diego, La Jolla, CA, USA Background: Very rarely children with vertically acquired HIV and given antiretroviral therapy (ART) soon after birth, then stop ART, have extended periods without detectable HIV in peripheral blood by routine testing. We report a child with intrauterine-acquired HIV, who started on combined antiretroviral therapy at 33 hours of life and remains undetectable over 3 years after discontinuing ART. Methods: In addition to routine clinical assays, HIV DNA was assayed using droplet digital PCR (ddPCR) for gag and pol using DNA extracted from available CD4 lymphocytes purified by negative selection. Results: A healthy newborn was born to a mother with no prenatal care and a 6-year history of diagnosed, but untreated HIV infection, with 14,400 HIV RNA copies/ml and 27% CD4 at delivery. The child was started on ART at 33 hours of life. A blood sample submitted for HIV DNA on day of life (DOL) 1 and another for HIV RNA on DOL 2 failed due to technical issues. A DOL 14 sample tested positive for HIV DNA. Because of this finding dried blood spots from DOL 1 from routine newborn screening were tested for HIV DNA with a positive result (CDC). The mother discontinued the child’s ART after 1 year. From birth through 4 years old the child remained clinically well with undetectable HIV RNA (<20) by routine laboratory testing, and HIV specific antibodies becoming and remaining negative from 15 months. Testing by HIV ddPCR-DNA was performed at intervals beginning at DOL 114 and were intermittently detected with the most recent one showing <1 copy of gag and pol DNA/ million CD4 cells. Conclusion: We present a child with intrauterine- acquired HIV infection, initiation of ART at 33 hour of life who was maintained on ART for 1 year and has remained clinically well through 4 years of age including 3 years without ART. Whether viral control was affected by ART, characteristics of the child or virus are being investigated.

Poster Abstracts

348LB CCR5Δ32 SCT-INDUCED HIV REMISSION: TRACES OF HIV DNA BUT FADING IMMUNE REACTIVITY Bjoern-Erik O. Jensen 1 , Dieter Häussinger 1 , Elena Knops 2 , Annemarie Wensing 3 , Javier Martinez-Picado 4 , Monique Nijhuis 3 , Maria Salgado 4 , Jacob D. Estes 5 , Nadine Lübke 1 , Rolf Kaiser 2 , Thomas Harrer 6 , Johannes Fischer 1 , Julian Schulze zur Wiesch 7 , Johanna M. Eberhard 8 , Guido Kobbe 1 1 Heinrich Heine University Düsseldorf, Düsseldorf, Germany, 2 University of Cologne, Cologne, Germany, 3 Utrecht University, Utrecht, Netherlands, 4 IrsiCaixa Institute for AIDS Research, Badalona, Spain, 5 Oregon Health and Sciences University, Portland, OR, USA, 6 University Hospital Erlangen, Erlangen, Germany, 7 University Medical Center Hamburg–Eppendorf, Hamburg, Germany, 8 University Hospital Hamburg– Eppendorf, Hamburg, Germany Background: To date only 3 patients have achieved long-term HIV-remission after analytic therapy interruption (ATI). Here we provide an update of

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