Therapeutic Repurposing of Brincidofovir in Non-Hodgkin Lymphoma – Potential Synergy with Immune Checkpoint Blockade

Introduction: Brincidofovir (BCV) is a novel nucleoside phosphonate analogue with unique dual anti-viral and anti-tumor properties. Notably, BCV elicits potent anti-lymphoproliferative activity, and has demonstrated potential immunogenic effects in pre-clinical studies on T-cell and NK/T-cell lymphoma (T/NK-NHL) cell-line models.

Methods: Activity of BCV was evaluated in forty-four cell-line models, including T/NK-NHL (n=25) and B-cell lymphoma (BCL, n=19), as well as their respective NOD/SCID mice xenograft models (MYC/BCL2-rearranged diffuse large B-cell lymphoma, TP63-rearranged anaplastic large cell lymphoma, JAK3-mutant NK/T-cell lymphoma and peripheral T-cell lymphoma, not otherwise specified; n=1 each). Potential in vivo immunogenic effects were examined in a syngeneic EL4-C57BL/6 murine lymphoma model.

Results: BCV demonstrated potent inhibitory effects in vitro across the majority of cell-lines regardless of EBV positivity, with IC50 values within clinically-achievable human plasma concentrations (2 µg/ml) in 17/25 (68.0%) T/NK-NHL and in 13/19 (68.4%) BCL. In vivo treatment via intraperitoneal BCV (IP, 40mg/kg, 2X per week) significantly inhibited tumor growth in all xenograft models when compared to vehicle control. Notably, RNAseq analysis demonstrated BCV downregulated MYC and MYC-target pathways in T/NK-NHL models. Further mechanistic studies showed that BCV evoked S-phase cell cycle arrest, replication stress, DNA damage and apoptosis, while also triggering STING pathway-mediated interferon responses, PD-L1 expression and immunogenic cell death. In EL4 murine lymphoma, BCV similarly inhibited cell viability in a dose-dependent manner (IC50 516 ng/ml) and increased PD-L1 expression on flow cytometry. In the syngeneic EL4-C57BL/6 model, BCV alone or in combination with anti-PD1 (IP 200 µg 1X per week) significantly inhibited tumor growth at 8 days post-treatment, compared to isotype or anti-PD1 treatment alone. No significant difference in tumor volume or weight was observed with the combination of BCV and anti-PD1, compared with BCV alone. Intriguingly, at this early time point, histologic examination suggested a significantly heavier immune infiltration only in the BCV+anti-PD1 combination group, which was corroborated by the highest scores for adaptive immune response, cytokines/chemokines & receptors, cytotoxic cells, dendritic cells, NK CD56^dim cells and neutrophils (NanoString nCounter, Mouse Immunology Panel). Gene expression of CCL2, CCL12, CXCL9, GZMB, CHIL3 and CTLA4 were also highest in the combination treatment group.

Conclusions: Taken together, these results demonstrate a novel role of BCV in the treatment of lymphoma. The immunogenic effects of BCV suggests potential for combination with PD1/PDL1 and CTLA4 checkpoint immunotherapy.