The bone marrow microenvironment is a key source of ARQ197 drug resistance for myeloma cells. It makes cytokines these as HGF and IL-6, which encourage the ARQ197 growth of myeloma cells and add to their resistance to cure . Myeloma cells were seeded on NKtert at a concentration 20 × that of NKtert cells. Forty-eight hours of treatment with 3 μM ARQ 197 resulted in cell death of 54% in U266 cells (Figure 2E) and 69% in OPM-2 cells (Figure 2F) when co-cultured with NKtert cells these values were similar to that of U266 cells cultured alone (55% annexin V +/PI + cells Figure 2E) and OPM-2 cells cultured alone (82% annexin V +/PI + cells Figure 2F). Importantly, ARQ 197 did not induce a notable increase in NKtert cell death (22% at 3 μM), suggesting that ARQ 197 spares normal stromal cells.
Activity of ARQ 197 in Melphalan- and Dexamethasone-Resistant Cell Lines
Melphalan is an alkylating agent that was one of the earliest treatments introduced for myeloma. We tested ARQ 197 on paired cell lines that were melphalan resistant (8226/LR-5) and melphalan sensitive (8226/S). ARQ 197 inhibited cell growth by 54% at 3 μM in 8226/S cells (Figure 3A). This was associated with 77% cell death (Figure 3B). Despite the melphalan-resistant 8226/LR-5 cells having a high percentage of endogenous cell death (44%), 3 μM ARQ 197 was still able to inhibit cell growth by 54% and induced 98% cell death (Figure 3, A and B). The correlation in cell death was high between 8226/S and 8226/LR-5 cells in their response to ARQ 197 (Pearson correlation 0.8718, P = .0105).
Dexamethasone is another approved agent for myeloma treatment. Hence, we tested both MM.1R cells, which are resistant to the synthetic steroid dexamethasone, and MM.1S cells, which are sensitive, to determine whether ARQ 197 can induce cell death in these cell lines. At 1 and 3 μM ARQ 197, cell growth was inhibited by 33% and 49%, respectively, in sensitive cells and by 28% and 44%, respectively, in resistant cells (Figure 3C). Similar to the growth inhibition data, an increase in cell death was detectable with 3 μM ARQ 197 and was similar in both cell lines (Figure 3D). The correlation in cell death was high between MM.1R and MM.1S cells in their response to ARQ 197 (Pearson correlation 0.9741, P = .0010).
Activity of ARQ 197 in Cells Resistant to Bortezomib or Lenalidomide
Newly approved drugs for myeloma include proteasome inhibitors such as bortezomib and immunomodulatory drugs such as thalidomide or lenalidomide. However, patients acquire resistance to these drugs. We examined two pairs of cell lines that were either sensitive or resistant to bortezomib (ANBL-6 vs ANBL-6/V10R and KAS-6 vs KAS-6/V10R). In these cell lines, growth inhibition was observed starting with 0.3 μM, and by 3 and 10 μM, 50% inhibition was achieved in both sensitive and resistant cell lines (Figure 4, A and C). This finding was consistent with that of induction of cell death (Figure 4, B and D), which was also similar in sensitive and resistant cell lines. Both paired cell lines exhibited high correlation in their response to ARQ 197 (Pearson correlation 0.9844, P < .0001 between ANBL-6 and ANBL-6/V10R and Pearson correlation 0.9898, P < .0001 between KAS-6 and KAS-6/V10R).