CD73 Inhibition Reverses Immunosuppression and Has Potential As an Immunomodulatory Therapy in Patients with Multiple Myeloma

Introduction: Adenosine accumulation within the tumor microenvironment can severely limit antitumor immunity by promoting the expansion of immunosuppressive cell types and impairing immune cell function, including T cells, natural killer cells and dendritic cells [Allard et al., Immunol Lett 2019]. Generation of adenosine from adenosine monophosphate (AMP) requires the activity of a cell surface ecto-5'-nucleotidase, CD73, expressed on the surface of various cell types including immune cells. Overexpression of CD73 is observed in many tumor types and correlates with unfavorable clinical outcomes. In patients with multiple myeloma (MM), elevated adenosine levels in the bone marrow (BM) correlate with progression of MM through a CD73-mediated pathway [Horenstein et al. Mol Med. 2016, Yang et al., J Immunother Cancer 2020]. We recently reported a functional role of CD73 signaling in BM of patients with MM, indicating that CD73 inhibition is a unique vulnerability and treatment strategy for MM [Ray et al., Blood Cancer J 2022; Ray et al., Blood 2021]. The essentiality of CD73 in generating adenosine makes CD73 inhibition a promising immunomodulatory strategy in oncology. Our previous work established that CD73-mediated adenosine activity suppresses the cytolytic antitumor immune function in the MM BM milieu and ORIC CD73 inhibitors can overcome immune suppression and restore lysis of MM cells by autologous T cells [Ray et al., Blood 2021]. Methods: To investigate CD73 inhibition in MM, autologous ex vivo cell assays utilizing freshly isolated BM aspirates from patients with MM were used to detect changes in MM cell viability and adenosine generation upon treatment with ORIC-533, a potent, orally bioavailable, AMP-competitive, small molecule inhibitor that is highly selective for CD73. ORIC-533 exhibits picomolar potency in biochemical assays, completely blocking adenosine production from AMP. The majority of BM samples utilized were from patients with relapsed or refractory (r/r) MM after at least three lines of therapy including immunomodulatory drugs, proteasome inhibitors, and CD38 monoclonal antibodies, as well as a patient with relapsed MM post BCMA-CAR-T therapy. Results: In BM aspirates from patients with r/r MM, ORIC-533 significantly reduced MM cell viability at all dose levels tested. Patient-derived bone marrow mononuclear cells (BMNCs) from patients with r/r MM cultured in the presence of ORIC-533 at 0.01, 0.1, or 0.5 μM for 48-72 hours (n=2 patients per dose level) demonstrated significant reduction of viable CD138+ MM cells across all dose-levels. Treatment with ORIC-533 reduced viability by 44% (p=0.008), 51% (p=0.18) and 76% (p=0.014), respectively. Similarly, MM BMNC samples from patients responding to induction therapy or maintenance anti-CD38 monoclonal antibodies were cultured in the presence of ORIC-533 (n=2 patients per dose level). Treatment with ORIC-533 at 1 or 2 μM significantly reduced viability of MM cells by 81% (p<0.05) and 99% (p=0.0001), respectively. To assess ORIC-533 treatment on CD73 activity, CD73-mediated conversion of labeled AMP substrate to adenosine was measured in BM serum from r/r MM samples by mass spectrometry. Treatment with ORIC-533 at 0.01 and 0.1 μM (n=9 r/r MM patients) resulted in a dose-dependent reduction of adenosine on average between 2 to 4-fold, respectively, compared to control. Conclusions: This study, together with our previous reports, confirms that CD73 inhibition can reduce adenosine generation, overcome immune suppression, and restore lysis of MM cells. Based on these results, ORIC-533 is being studied as a single agent in a Phase 1 clinical trial in patients with relapsed or refractory multiple myeloma.