Treatment of multiple myeloma (MM) cells with sublethal doses of genotoxic drugs leads to senescence and results in increased NK cell recognition and effector functions. Herein, we demonstrated that doxorubicin- and melphalan-treated senescent cells display increased expression of IL15, a cytokine involved in NK cell activation, proliferation, and maturation. IL15 upregulation was evident at the mRNA and protein level, both in MM cell lines and malignant plasma cells from patients' bone marrow (BM) aspirates. However, IL15 was detectable as a soluble cytokine only , thus indicating a functional role of IL15 in the BM tumor microenvironment. The increased IL15 was accompanied by enhanced expression of the IL15/IL15RA complex on the membrane of senescent myeloma cells, allowing the functional trans-presentation of this cytokine to neighboring NK cells, which consequently underwent activation and proliferation. We demonstrated that MM cell-derived exosomes, the release of which was augmented by melphalan treatment in senescent cells, also expressed IL15RA and IL15, and their interaction with NK cells in the presence of exogenous IL15 resulted in increased proliferation. Altogether, our data demonstrated that low doses of chemotherapeutic drugs, by inducing tumor cell senescence and a senescence-associated secretory phenotype, promoted IL15 trans-presentation to NK cells and, in turn, their activation and proliferation, thus enhancing NK cell-tumor immune surveillance and providing new insights for the exploitation of senescence-based cancer therapies..

Natural killer (NK) cells are immune innate effectors playing a pivotal role in the immunosurveillance of multiple myeloma (MM) since they are able to directly recognize and kill MM cells. In this regard, among activating receptors expressed by NK cells, NKG2D represents an important receptor for the recognition of MM cells, being its ligands expressed by tumor cells, and being able to trigger NK cell cytotoxicity. The MHC class I-related molecule A (MICA) is one of the NKG2D ligands; it is encoded by highly polymorphic genes and exists as membrane-bound and soluble isoforms. Soluble MICA (sMICA) is overexpressed in the serum of MM patients, and its levels correlate with tumor progression. Interestingly, a methionine (Met) to valine (Val) substitution at position 129 of the α2 heavy chain domain classifies the MICA alleles into strong (MICA-129Met) and weak (MICA-129Val) binders to NKG2D receptor. We addressed whether the genetic polymorphisms in the MICA-129 alleles could affect MICA release during MM progression. The frequencies of Val/Val, Val/Met, and Met/Met MICA-129 genotypes in a cohort of 137 MM patients were 36, 43, and 22%, respectively. Interestingly, patients characterized by a Val/Val genotype exhibited the highest levels of sMICA in the sera. In addition, analysis of the frequencies of MICA-129 genotypes among different MM disease states revealed that Val/Val patients had a significant higher frequency of relapse. Interestingly, NKG2D was downmodulated in NK cells derived from MICA-129Met/Met MM patients. Results obtained by structural modeling analysis suggested that the Met to Val dimorphism could affect the capacity of MICA to form an optimal template for NKG2D recognition. In conclusion, our findings indicate that the MICA-129Val/Val variant is associated with significantly higher levels of sMICA and the progression of MM, strongly suggesting that the usage of soluble MICA as prognostic marker has to be definitely combined with the patient MICA genotype.

Natural killer (NK) cells are innate cytotoxic lymphocytes that play a key role in cancer immunosurveillance thanks to their ability to recognize and kill cancer cells. NKG2D is an activating receptor that binds to MIC and ULBP molecules typically induced on damaged, transformed or infected cells. The release of NKG2D ligands (NKG2DLs) in the extracellular milieu through protease‐mediated cleavage or by extracellular vesicle (EV) secretion allows cancer cells to evade NKG2D‐mediated immunosurveillance. In this work, we investigated the immunomodulatory properties of the NKG2D ligand MICA*008 associated to distinct populations of EVs (i.e., small extracellular vesicles [sEVs] and medium size extracellular vesicles [mEVs]). By using as model a human MICA*008‐transfected multiple myeloma (MM) cell line, we found that this ligand is present on both vesicle populations. Interestingly, our findings reveal that NKG2D is specifically involved in the uptake of vesicles expressing its cognate ligand. We provide evidence that MICA*008‐expressing sEVs and mEVs are able on one hand to activate NK cells but, following prolonged stimulation induce a sustained NKG2D downmodulation leading to impaired NKG2D‐mediated functions. Moreover, our findings show that MICA*008 can be transferred by vesicles to NK cells causing fratricide. Focusing on MM as a clinically and biologically relevant model of tumour‐NK cell interactions, we found enrichment of EVs expressing MICA in the bone marrow of a cohort of patients. All together our results suggest that the accumulation of NKG2D ligands associated to vesicles in the tumour microenvironment could favour the suppression of NK cell activity either by NKG2D down‐modulation or by fratricide of NK cell dressed with EV‐derived NKG2D ligands.