Abstract P052

Emerging role of SAMHD1 in anti-tumor immune responses and sensitivity to chemotherapy in classical Hodgkin lymphoma

Background: The tumor microenvironment (TME) plays a pivotal role in the pathogenesis of classical Hodgkin lymphoma (cHL). SAMHD1, a dNTP hydrolase, is involved in anti-tumor immune responses through DNA damage mechanisms and cGAS-STING pathway. SAMHD1 expression in the neoplastic Hodgkin and Reed-Sternberg cells (HRS) is an independent adverse prognostic factor in cHL, however, the mechanisms underlying its clinical significance are unknown. In this study the biologic impact of SAMHD1 on anti-tumor immune responses and sensitivity to chemotherapy in cHL was investigated.

Methods: The in vitro system included 6 cHL cell lines, in 3 of which (L1236, L428, L540) SAMHD1-deficient clones (KO) were generated using CRISPR-Cas9 gene editing. In addition, the effects of plasmid-mediated forced expression of SAMHD1 were analyzed in 2 cHL cell lines with low (L428) or undetectable (MDAV) endogenous SAMHD1. The cHL cell lines were treated with a STING agonist, STING inhibitor (C-176), or doxorubicin. Protein expression and activation were analyzed by Western blot. Immune responses were evaluated using RT-qPCR for gene expression of critical immunomodulators, and targeted proteomics with Olink Target 48 Cytokine panel on culture supernates.

Results: SAMHD1-KO and -WT cHL cells showed differential gene expression of critical modulators of antitumor immune responses. The effects of SAMHD1 deficiency were associated with changes in cGAS-STING pathway activation, including phosphorylation of TBK1, IRF3 and IRF7. SAMHD1 deficiency resulted in decreased secretion of IL6, CSF2, TNF, CCL2/7. STING stimulation led to significantly increased IFNβ gene expression by HRS cells in SAMHD1-WT but not in SAMHD1-KO cHL cell lines. STING inhibition also resulted in differential changes in immunomodulatory cytokines. Forced expression of SAMHD1 led to upregulation of CXCL10 and IFNβ gene expression in cultured HRS cells. These effects were linked to lower levels of CXCL9, IL10, TNF, IL17a and CXCL8 in MDA-V cells following SAMHD1 forced expression as shown by proteomic profiling. SAMHD1-WT and -KO cHL cells differentially responded to doxorubicin treatment with changes in apoptosis-regulating proteins.

Conclusions: SAMHD1 expression seems to modulate the anti-tumor immune responses partially through the cGAS-STING pathway in cHL. SAMHD1 expression also affects the response to doxorubicin treatment in vitro.