Saturday, May 17, 2025
5/17/2025
Delivering miR-29a to enhance anti-viral and anti-tumor responses in vivo - Exhausted T cells (TEX) differentiate in cancer and chronic infections. TEX do not function optimally and they do not provide protection. Immunotherapy reinvigorates TEX by antagonizing surface inhibitory receptors, such as PD-1. However, the epigenetic stability of TEX inhibits complete reversal of exhaustion, therefore significantly limiting the clinical benefits of immunotherapy. We recently proposed the use of microRNAs to attenuate exhaustion. MicroRNAs regulate cell differentiation by targeting multiple mRNAs. Indeed, microRNA-29a (miR- 29a) regulates CD8 T cell differentiation, enhances CD8 T cell persistence and function and promotes memory- like responses in chronic infection by epigenetically altering TEX differentiation. Therefore, we suggest the use of miR-29a to enhance TEX reinvigoration. However, delivering microRNAs to TEX in vivo is a major challenge. The goal of this proposal is to reinvigorate TEX and improve anti-viral and anti-tumor responses using a novel method for delivery of miR-29a in vivo. Our preliminary data suggest that miR-29a synergizes with αPD-1 to enhance CD8 T cell responses and promote durable, functional CD8 T cell reinvigoration. However, our results are based on retroviral overexpression in CD8 T cells, which does not allow for delivery of miR-29a to TEX later when exhaustion has progressed. Therefore, we developed a novel method to deliver miR-29a to already differentiated TEX in vivo. We conjugated miR-29a to αPD-1 (miR-29a/αPD-1). MiR-29a/αPD-1 increased miR- 29a expression and decreased miR-29a target expression in TEX, suggesting that miR-29a delivery is efficient and functional. Therefore, our overall hypothesis is that miR-29a/αPD-1 alters T cell differentiation, reinvigorates TEX, and enhances anti-viral and anti-tumor T cell responses. Aim 1. Reinvigorate TEX with miR-29a/αPD-1 in vivo. MiR-29a regulates key transcriptional circuits and synergizes with αPD-1 to promote TEX reinvigoration. Therefore, we hypothesize that miR-29a delivery by miR- 29a/αPD-1 can promote TEX reinvigoration both early and late during the differentiation of exhaustion. We will administer miR-29a/αPD-1 to mice with chronic viral infection (LCMV) or tumors (B16 melanoma or KPC-PDAC). We will elucidate the role of miR-29a in TEX reinvigoration during the early and late exhaustion phase. Aim 2. Delineate targeting of immune cells and tumor cells by miR-29a/αPD-1. PD-1 is highly expressed in TEX, but also in other immune cells, and tumor cells. Interestingly, miR-29a can suppress tumorigenesis and inhibit tumor cell proliferation. Therefore, we hypothesize that miR-29a/αPD-1 can provide significant clinical benefits by targeting both immune and tumor cells. To test this, we will use a fluorescence labelled miR-29a/αPD- 1 to elucidate miR-29a delivery and identify other immune cells and tumor cells targeted by miR-29a/αPD-1. At the completion of these studies we will a) demonstrate the ability of miR-29a to prevent and/or reverse exhaustion, b) develop a novel immunotherapeutic to provide immune reinvigoration with a dual effect on immune cells and tumor cells, and c) develop a novel platform for in vivo delivery of miRNAs to TEX.
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