PI3Kδ activation, IL6 over-expression, and CD37 loss cause resistance to the targeting of CD37-positive lymphomas with the antibody-drug conjugate naratuximab emtansine

Abstract

PurposeThe transmembrane protein CD37 is expressed almost exclusively in lymphoid tissues, with the highest abundance in mature B cells. CD37-directed antibody- and, more recently, cellular-based approaches have shown preclinical and promising early clinical activity. Naratuximab emtansine (Debio 1562, IMGN529) is an antibodydrug conjugate (ADC) that incorporates an anti-CD37 monoclonal antibody conjugated to the maytansinoid DM1 as payload. Naratuximab emtansine has shown activity as a single agent and in combination with the anti-CD20 monoclonal antibody rituximab in B cell lymphoma patients.Experimental DesignWe assessed the activity of naratuximab emtansine usingin vitromodels of lymphomas, correlated its activity with CD37 expression levels, characterized two resistance mechanisms to the ADC, and identified combination partners providing synergy.ResultsThe anti-tumor activity of naratuximab emtansine was tested in 54 lymphoma cell lines alongside its free payload. The median IC50of naratuximab emtansine was 780 pM, and the activity, primarily cytotoxic, was more potent in B than in T cell lymphoma cell lines. In the subgroup of cell lines derived from B cell lymphoma, there was some correlation between sensitivity to DM1 and sensitivity to naratuximab emtansine (r=0.28, P = 0.06). After prolonged exposure to the ADC, one diffuse large B cell lymphoma (DLBCL) cell line developed resistance to the ADC due to the biallelic loss of theCD37gene. After CD37 loss, we also observed upregulation of IL6 (IL-6) and other transcripts from MYD88/IL6-signaling. Recombinant IL6 led to resistance to naratuximab emtansine, while the anti-IL6 antibody tocilizumab improved the cytotoxic activity of the ADC in CD37-positive cells. In a second model, resistance was sustained by an activating mutation in thePIK3CDgene, associated with increased sensitivity to PI3Kδinhibition and a switch from functional dependence on the anti-apoptotic protein MCL1 to reliance on BCL2. The addition of idelalisib or venetoclax to naratuximab emtansine overcame resistance to the ADC in the resistant derivative while also improving the cytotoxic activity of the ADC in the parental cells.ConclusionsTargeting B cell lymphoma with the CD37 targeting ADC naratuximab emtansine showed vigorous anti-tumor activity as a single agent, which was also observed in models bearing genetic lesions associated with inferior outcomes, such as MYC translocations and TP53 inactivation or resistance to R-CHOP. Resistance DLBCL models identified active combinations of naratuximab emtansine with drugs targeting IL6, PI3Kδ, and BCL2.Despite notable progress in recent decades, we still face challenges in achieving a cure for a substantial number of lymphoma patients (1,2). A pertinent example is diffuse large B cell lymphoma (DLBCL), the most prevalent type of lymphoma (3). More than half of DLBCL patients can achieve remission, but around 40% of them experience refractory disease or relapse following an initial positive response (3). Regrettably, the prognosis for many of these cases remains unsatisfactory despite introducing the most recent antibody-based or cellular therapies (3,4), underscoring the importance of innovating new therapeutic strategies and gaining insights into the mechanisms of therapy resistance.CD37 is a transmembrane glycoprotein belonging to the tetraspanin family, primarily expressed on the surface of immune cells, principally in mature B cells but also, at lower levels, in T cells, macrophages/monocytes, granulocytes and dendritic cells (5) (6-8). CD37 plays a crucial role in various immune functions, including B cell activation, proliferation, and signaling, although its precise role still needs to be fully elucidated. CD37 interacts with multiple molecules, including SYK, LYN, CD19, CD22, PI3Kδ, PI3Kγ, and different integrins, among others (6-8). In mice, the lack of CD37 is paired with reduced T cell-dependent antibody-secreting cells and memory B cells, apparently due to the loss of CD37-mediated clustering of α4β1integrins (VLA-4) on germinal center B cells and decreased downstream activation of PI3K/AKT signaling and cell survival (5). Reflecting the expression pattern observed in normal lymphocytes, CD37 exhibits elevated expression in all mature B-cell lymphoid neoplasms, including most lymphoma subtypes, and absence in early progenitor cells or terminally differentiated plasma cells (6,8-14). In DLBCL, CD37 expression has been reported between 40% and 90% of cases across multiple studies performed using different antibodies (10,14-16).CD37-directed antibody- and, more recently, cellular-based approaches have shown preclinical (7,10-14,17-23) and early promising clinical activity (24-32). Among the CD37-targeting agents, naratuximab emtansine (Debio 1562, IMGN529) is an antibody-drug conjugate (ADC) that incorporates the anti-CD37 humanized IgG1 monoclonal antibody K7153A conjugated to the maytansinoid DM1, as payload, via the thioether linker, N-succinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate (SMCC) (10).Based on the initialin vitroandin vivoevidence of anti-tumor activity in lymphoma and chronic lymphocytic leukemia (CLL) (7,10), naratuximab emtansine entered the clinical evaluation as a single agent. The phase 1 study exploring naratuximab emtansine enrolled 39 patients with relapsed/refractory B cell lymphoma (27). The overall response rate (ORR) was 13% across all patients and 22% in DLBCL patients, including the only observed complete remission (CR) (27). In preliminary results of a phase 2 trial exploring the combination of naratuximab emtansine with the anti-CD20 monoclonal antibody rituximab (18), based on positive preclinical data (18), the ORR was 45% in 76 patients with DLBCL with 24 CRs (32%), 57% in 14 patients with follicular lymphoma (five CR), 50% in four MCL patients (2 CR) (31).Here, we studied the pattern of activity of naratuximab emtansine across a large panel of cell lines derived from DLBCL and other lymphoma subtypes and characterized two resistance mechanisms to the ADC.

CD20 expression regulates CD37 levels in B-cell lymphoma – implications for immunotherapies

“…It has already been reported that rituximab synergizes with naratuximab emtansine by increasing the endocytosis of CD37. 24 While it has been recently demonstrated that the efficacy of naratuximab emtansine relies on the expression levels of CD37, 33 our results suggest that anti-CD37 ADCs may be an option also for r/r RTX-treated patients. Interestingly, the recent work by Arribas et al , who explored the resistance mechanism to naratuximab emtansine, demonstrates that acquired resistance to this anti-CD37 ADC may lead to increased sensitivity to venetoclax.…”

“…Interestingly, the recent work by Arribas et al , who explored the resistance mechanism to naratuximab emtansine, demonstrates that acquired resistance to this anti-CD37 ADC may lead to increased sensitivity to venetoclax. 33 This emphasizes the significance of examining the implications of acquired resistance to identify novel therapeutic pathways with a focus on tailored medical approaches. Finally, we show that decreased CD37 levels (up to 50% in our models) do not influence the efficacy of CD37-targeted CARs in both in vitro and in vivo settings and delineate CD37 CARs as an effective option for CD20-negative RTX-pretreated patients.…”

Comprehensive Analysis of177Lu-lilotomab Satetraxetan in Lymphoma Cell Lines: Implications for Precision Radioimmunotherapy and Combination Schemes

“…Taking advantage of available surface protein expression and RNA data obtained in our laboratory on the same cell lines used in this study 14,23,24 , we analyzed whether the anti-tumor activity of 177 Lulilotomab satetraxetan was affected by the expression levels of its target. 177 Lu-lilotomab satetraxetan IC50 values were negatively correlated with CD37 surface protein expression across 54 cell lines, including both B and T cell lymphoma models (r=-0.48, P=0.003) and also within the group of B-cell lymphomas (r=-0.36, P=0.015) (Figure 2A-B).…”

“…Indeed, one of the first demonstrations of the anti-tumor activity of radioimmunoconjugates in the clinical setting was with the anti-CD37 131 I MB-1 antibody given at high doses with autologous bone marrow support 11 . CD37 has also been used as the target of antibody-drug conjugates (ADCs) 6,7,[12][13][14] . Lutetium-177 ( 177 Lu) lilotomab satetraxetan (Betalutin) is a CD37-targeting radioimmunoconjugate obtained by conjugating the murine anti-CD37 IgG1 lilotomab with the bi-functional chelator p-SCNbenzyl-DOTA (satetraxetan) that chelates 177 Lu 3+ 15 .…”

The Regulation of Exosome Generation and Function in Physiological and Pathological Processes

“…In addition, it controls IL-6 receptor signaling through its interaction with SOCS3. Furthermore, CD37 can mediate the aggregation of α4β1 and subsequent activation of PI3K/AKT signaling and cell survival [ 131 , 132 ]. CD63 may interact with the integrins α4β1, α3β1, α6β1, LFA-1, and β2 [ 133 ], mediating binding to the ECM (extracellular matrix) and promoting tumor cell migration.…”