P-182: Natural killer cells are active against myeloma cells with loss of tumor suppressor genes

Introduction: In multiple myeloma (MM) and other neoplasias, it its challenging to target therapeutically tumor cells with loss of function (LOF) for tumor suppressor genes (TSGs), because pharmacological mechanisms to restore the function(s) of such genes are not considered to be readily feasible, in contrast to e.g., inhibition of oncogenic drivers. We reasoned, however, that, LOF for TSGs in MM cells leads to de-repressed cell growth, it may not necessarily protect tumor cells from immune attack. We thus explored the hypothesis that CRISPR-based loss of function of TSGs in MM cell lines may still be associated with substantial response to immune effector cells such as NK cells, which have the advantage to kill tumor cells across HLA barriers.

Methods: We examined our genome-scale or focused CRISPR screens for LOF (CRISPR-based gene editing) or gain of function (GOF, CRISPR activation) in MM lines (KMS11, LP1 and MM1S) exposed to allogeneic donor-derived NK cells (vs. control cultures without NK cells) and specificallt evaluated the performance of genes known to represent recurrent TSGs based on genomic data of MM patient samples or cell lines; as well as candidate TSGs, based on results from genome-scale CRISPR gene editing screens (e.g., CERES or CHRONOS scores >0.4 in multiple DepMap releases and TPM>1 [RNA-seq]) in the same cell lines as the NK cell resistance screens. These analyses sought to identify any TSGs whose LOF may potentially alter the response of MM cells to NK cells.

Results: Overall, 34 genes were identified as top recurrent TSGs in MM patient samples (e.g., based on prior genomic studies) or based on in vitro CRISPR knockout screens. These genes were evaluated for their metrics of sgRNA enrichment or depletion in genome-scale CRISPR gene editing or activation screens in the context of exposure to NK cells. These genes included known and recurrent TSGs (e.g., PTEN, FAM46C, TP53, RB1) as well as other, previously underappreciated candidate TSGs (e.g., HIF1A, ASXL1, ATRX). Perturbation of none of these genes was identified to meet criteria for association with significant resistance to allogeneic donor-derived NK cells (log2FC>1.0, at least 3-4 sgRNAs with enrichment upon CRISPR KO or depletion with CRISPR activation, p-value < 0.05, depletion [or enrichment] rank < 100) in any of the 3 MM cell lines examined in LOF or GOF CRISPR screens for NK cell resistance. In fact, for a limited set of cases (e.g., PTEN in KMS11 cells), KO of a TSG was associated with sensitization to NK cell treatment. These in-house results with MM cells exposed to NK cells are concordant with results for these TSGs in CRISPR screens of non-MM cells treated with cytotoxic T-cells (e.g., 4T1 or RENCA cells; GSE149933).

Conclusions: MM cells deficient for diverse TSGs are equally responsive to NK cells as their TSG-proficient counterparts. NK cell-based therapies may thus be a promising approach to target TSG-deficient MM cells for which specific pharmacological therapies are not currently available.