You are here
ANALYTIC TREATMENT INTERRUPTION (ATI) AFTER ALLOGENEIC CCR5-D32 HSCT FOR AML IN 2013
Björn-Erik O. Jensen1, Elena Knops2, Nadine Lübke1, Annemarie Wensing3, Javier Martinez-Picado4, Rolf Kaiser2, Monique Nijhuis3, Maria Salgado4, Thomas Harrer5, Eva Heger2, Johanna M. Eberhard6, Ilona Hauber7, Carsten Münk1, Dieter Häussinger1, Guido Kobbe1
1Heinrich Heine University Hospital, Düsseldorf, Germany,2University of Cologne, Cologne, Germany,3Utrecht University, Utrecht, Netherlands,4IrsiCaixa Institute for AIDS Research, Badalona, Spain,5University Hospital Erlangen, Erlangen, Germany,6University Hospital Hamburg–Eppendorf, Hamburg, Germany,7Heinrich Pette Institute, Hamburg, Germany
A 49y-old HIV-infected male patient received unmodified HSCT from a 10/10 CCR5-d32 donor in Feb 2013 because of acute myeloid leukemia (AML) while in 2nd complete remission (CR). At time of HSCT proviral HIV DNA load was 1.45 log10cop/Mio PBMCs. All anticipated antibodies were detected by western blot. HIV coreceptor-usage was predicted R5 (Sanger: FPR 44.5%; NGS: 0.14% X4 at 3.5% FPR, geno2pheno), confirmed by phenotypic testing (TropChase). He had a 2nd relapse of AML in Jun/13 but after 8 courses of 5-azaC and 4 donor lymphocyte infusions CR was achieved. Immunosuppression was stopped in Oct/17. During HSCT the patient remained on ART until Nov/18 with undetectable plasma viral load.
PBMC and tissues were analysed by ddPCR, qPCR and in situ hybridization in several laboratories as well as humoral and T-cell responses. Infectious virus was analysed on CD4+T-cells (qVOA, MVOA). Patient was registered to IciStem as #19.
Almost all PBMC samples were negative for proviral HIV-DNA by qPCR/ddPCR at multiple time points. CSF (Jul/14), rectum (Apr/15, Mar/16), ileum (Mar/16) and bone marrow (Aug/15) were negative. Further testing with 0.1 Mio cells from ileum showed in 1/4 replicates one positive droplet with LTR-, but none with gag-primers. There was also a signal in TCM 0.2 Mio cells (ddPCR 1 positive droplet, qPCR neg) and in TEM 0.36 Mio cells (qPCR 5cop/Mio cells, ddPCR neg) while all other T-cell subsets were negative in ddPCR & qPCR. No HIV-DNA could be detected by PCR in lymph nodes of May/17, but in situ hybridization assays (RNAscope, DNAscope) detected few positive signals. Viral outgrowth assays (qVOA) in Feb/16, Mar/16 and May/16 were negative (23 Mio CD4+T-cells, IUPM<0.031/Mio cells CD4+T-cells). Mouse VOA (Apr/16: Rag2-/-γc-/-, Apr/17: NOD-SCID IL2gR-/-) were also negative. Gp160 was the only remaining band on the blot. Peptide stimulation assays revealed the presence of CCR5-negative HIV-specific CTL recognizing HLA-A2-restricted RT-epitope YV9 and HLA-B7-restricted Gag-p6 epitope YL9.
Despite low signals in ultrasensitive assays no virus could be detected in qVOA/mVOA in the Duesseldorf patient. Taking into account the homozygous CCR5-d32 status we consider a viral rebound to be unlikely. An ATI is the only way to find out whether HIV has been eradicated by allogeneic CCR5-d32 HSCT. Therefore ART was stopped in Nov/18 after thorough discussion with the patient. Despite all plasma samples being negative after ATI longer surveillance is essential.