Fig 2: NKTR-255 induces proliferation and activation of BM-derived NK cells, reducing viability of autologous MM cells. (A) Whole BM samples (n = 6) from patients newly diagnosed with MM without prior treatment were incubated for 120 hours with and without 4 increasing doses of NKTR-255. The effect of the compound on the NK (effector) and MM (target) cell populations in whole BM was assessed through an automated flow cytometry platform that allows for absolute cell counting. Based on this data, T:E ratio for each sample was calculated at baseline and it was not modified since the whole BM sample was employed unaltered for this assay. Absolute numbers of NK cells within whole BM samples were measured at baseline and after 120 hours of incubation and results are presented as mean ± standard deviation fold increase in NK cell absolute number in the 6 patients according to concentration level of NKTR-255. (B-C) CD69 and NKG2D expression was measured as a marker of NK cell activation. Percentages of NK cells expressing NKG2D (B) and CD69 (C) over total NK cell population after 120 hours of incubation at different NKTR-255 concentrations are shown as mean and range (minimum to maximum). (D-E) Percentage of live CD138+ MM cells in BM samples of each patient (D) was calculated at baseline and after 120 hours of culture with and without different doses of NKTR-255. Mean of CD138+ cell viability compared with untreated is shown (E). ∗∗P < .01, ∗∗∗P < .001, nsP ≥ .05. BM, bone marrow; E:T, effector-to-target; ns, not significant; NK, natural killer; MM, multiple myeloma; SD, standard deviation.

Fig 3: Interactome analyses of SIRP-ß2 identified DAP12 as signaling component and a strong association of SIRP-ß2 with MHC-class I and potentiate HPV-E7-specific T cell responses. (A) HLA-ABC expression of THP-1.EV, THP-1.SIRP-ß2 and THP-1.SIRP-ß2.K202L determined by flow cytometry. (B) mRNA expression of HLA-A2 (CT value 30 (EV), 26 (SIRP-ß2), 27 (SIRP-ß2.K202L)). (C) HLA-B (CT value 24 (EV), 21 (SIRP-ß2), 22 (SIRP-ß2.K202L)). (D) HLA-C (CT value 25 (EV), 25 (SIRP-ß2), 26 (SIRP-ß2.K202L)) n=3. (E) RNAseq analysis from THP-1.EV, THP-1.SIRP-ß2 and THP-1.SIRP-ß2.K202L, ~300 differently regulated genes. (F, G) Major upregulation of MHC-II from RNAseq analysis. (H) Quantitative mRNA HLA-DR expression on THP-1.EV, THP-1.SIRP-ß2 and THP-1.SIRP-ß2.K202L. (I) Activation of Jurkat Nuclear Factor of Activated T-cells (NFAT)-E7 with THP-1.EV, THP-1.SIRP-ß2 and THP-1.SIRP-ß2.K202L, including different Effector to Target (E:T) ratio’s. (J) Expression of CD69 and CD25 on CD4/CD8 T cells co-cultured with THP-1.EV, THP-1.SIRP-ß2 and THP-1.SIRP-ß2.K202L and E7 peptide for 6 h (n=4). (K) Quantitative IFN-γ secretion by primary E7-TCR specific T cells in co-culture with THP-1.EV, THP-1.SIRP-ß2 and THP-1.SIRP-ß2.K202L and E7 peptide for 48 h (n=5). p values are indicated as: *** p < 0.001, ** p < 0.01, and * p < 0.05. ns (not significant).