School of Medicine and Health Sciences Poster Presentations

Title

HIV-Specific T Cells can be Expanded from Virus-Naive Donors to Target a Range of Epitopes: Implications for a Cure Strategy

Poster Number

23

Document Type

Poster

Publication Date

3-2016

Abstract

Background: The Berlin patient remains the only case of functional HIV cure and long-term independence from anti-retroviral drugs, despite multiple attempts to eradicate infection with allogeneic hematopoietic stem cell transplant (HSCT). HIV+ individuals with hematologic malignancies who receive allogeneic HSCT may face viral rebound during the immune reconstitution period following transplant. One potential approach to prevent this is to administer virus-specific T cells, a strategy that has proven success in preventing other viral reactivation post-transplant, such as CMV and EBV.

Objective: We have previously expanded broadly-HIV-specific T cells from HIV+ patients; however allogeneic transplants only contain virus-naïve T cells. We thus sought to develop a robust, reproducible platform that can expand HIV-specific cells from the naïve pool in the allogeneic setting. We hypothesize that HIV-specific T cells can also be primed ex vivo from seronegative individuals to effectively target and suppress HIV replication in vitro.

Design/Methods: Peripheral blood mononuclear cells isolated from virus-naïve donors are used to generate dendritic cells and T cells. T cells are stimulated three times with DCs pulsed with HIV-pepmix and a combination of cytokines that promote proliferation and differentiation. We tested T cells for (1) specificity against HIV antigens and individual peptides, (2) pro-inflammatory cytokine secretion in response to stimulation with HIV peptides, and (3) ability to suppress HIV replication.

Results/Discussion: We successfully expanded (75.705 mean fold expansion) HXTCs recognizing HIV antigens from virus naïve donors. IFNg ELISPOT showed HXTCs (n=8) were specific against Gag (mean=331.25 SFC/1e5 cells) and Nef (mean=242.63 SFC/1e5 cells) vs Irrelevant (mean=13 SFC/1e5 cells). HXTCs produced significantly pro-inflammatory responses (p<0.05) to stimulation by gag/nef, as determined by levels of TNF-alpha, IL-2, IL-6, IL-8, and perforin (n=3). Importantly, HXTCs (n=4) were able to suppress HIV replication more than non-specific CD8+ T cells when co-cultured with autologous CD4+ T cells infected with HIV SF162 (HXTC 78.62% viral suppression compared to CD8+ T cell 34.19% viral suppression). HXTCs showed both HLA Class I or II specificity for individual HIV epitopes, as determined by HLA blocking and IFNg ELISPOT. This is the first report demonstrating generation of functional, multi-HIV antigen specific T-cells from HIV-negative donors, which has implications for using allogeneic HSCT as a functional HIV cure. The low frequency of circulating HXTCs post-infusion suggests these HXTCs could have a significant effect on prevent

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Presented at: GW Research Days 2016

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HIV-Specific T Cells can be Expanded from Virus-Naive Donors to Target a Range of Epitopes: Implications for a Cure Strategy

Background: The Berlin patient remains the only case of functional HIV cure and long-term independence from anti-retroviral drugs, despite multiple attempts to eradicate infection with allogeneic hematopoietic stem cell transplant (HSCT). HIV+ individuals with hematologic malignancies who receive allogeneic HSCT may face viral rebound during the immune reconstitution period following transplant. One potential approach to prevent this is to administer virus-specific T cells, a strategy that has proven success in preventing other viral reactivation post-transplant, such as CMV and EBV.

Objective: We have previously expanded broadly-HIV-specific T cells from HIV+ patients; however allogeneic transplants only contain virus-naïve T cells. We thus sought to develop a robust, reproducible platform that can expand HIV-specific cells from the naïve pool in the allogeneic setting. We hypothesize that HIV-specific T cells can also be primed ex vivo from seronegative individuals to effectively target and suppress HIV replication in vitro.

Design/Methods: Peripheral blood mononuclear cells isolated from virus-naïve donors are used to generate dendritic cells and T cells. T cells are stimulated three times with DCs pulsed with HIV-pepmix and a combination of cytokines that promote proliferation and differentiation. We tested T cells for (1) specificity against HIV antigens and individual peptides, (2) pro-inflammatory cytokine secretion in response to stimulation with HIV peptides, and (3) ability to suppress HIV replication.

Results/Discussion: We successfully expanded (75.705 mean fold expansion) HXTCs recognizing HIV antigens from virus naïve donors. IFNg ELISPOT showed HXTCs (n=8) were specific against Gag (mean=331.25 SFC/1e5 cells) and Nef (mean=242.63 SFC/1e5 cells) vs Irrelevant (mean=13 SFC/1e5 cells). HXTCs produced significantly pro-inflammatory responses (p<0.05) to stimulation by gag/nef, as determined by levels of TNF-alpha, IL-2, IL-6, IL-8, and perforin (n=3). Importantly, HXTCs (n=4) were able to suppress HIV replication more than non-specific CD8+ T cells when co-cultured with autologous CD4+ T cells infected with HIV SF162 (HXTC 78.62% viral suppression compared to CD8+ T cell 34.19% viral suppression). HXTCs showed both HLA Class I or II specificity for individual HIV epitopes, as determined by HLA blocking and IFNg ELISPOT. This is the first report demonstrating generation of functional, multi-HIV antigen specific T-cells from HIV-negative donors, which has implications for using allogeneic HSCT as a functional HIV cure. The low frequency of circulating HXTCs post-infusion suggests these HXTCs could have a significant effect on prevent