Purpose: Increased β-adrenergic receptor (β-AR) signaling has been shown to promote the creation of an immunosuppressive tumor microenvironment (TME). Preclinical studies have shown that abrogation of this signaling pathway, particularly β2-AR, provides a more favorable TME that enhances the activity of anti–PD-1 checkpoint inhibitors. We hypothesize that blocking stress-related immunosuppressive pathways would improve tumor response to immune checkpoint inhibitors in patients. Here, we report the results of dose escalation of a nonselective β-blocker (propranolol) with pembrolizumab in patients with metastatic melanoma. Patients and Methods: A 3 + 3 dose escalation study for propranolol twice a day with pembrolizumab (200 mg every 3 weeks) was completed. The primary objective was to determine the recommended phase II dose (RP2D). Additional objectives included safety, antitumor activity, and biomarker analyses. Responders were defined as patients with complete or partial response per immune-modified RECIST at 6 months. Results: Nine patients with metastatic melanoma received increasing doses of propranolol in cohorts of 10, 20, and 30 mg twice a day. No dose-limiting toxicities were observed. Most common treatment-related adverse events (TRAEs) were rash, fatigue, and vitiligo, observed in 44% patients. One patient developed two grade ≥3 TRAEs. Objective response rate was 78%. While no significant changes in treatment-associated biomarkers were observed, an increase in IFNγ and a decrease in IL6 was noted in responders. Conclusions: Combination of propranolol with pembrolizumab in treatment-naïve metastatic melanoma is safe and shows very promising activity. Propranolol 30 mg twice a day was selected as RP2D in addition to pembrolizumab based on safety, tolerability, and preliminary antitumor activity.

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MDM2 and MDM4 are key regulators of p53 and function as oncogenes when aberrantly expressed. MDM2 and MDM4 partner to suppress p53 transcriptional transactivation and polyubiquitinate p53 for degradation. The importance of MDM2 E3-ligase-mediated p53 regulation remains controversial. To resolve this, we generated mice with an Mdm2 L466A mutation that specifically compromises E2 interaction, abolishing MDM2 E3 ligase activity while preserving its ability to bind MDM4 and suppress p53 transactivation. Mdm2L466A/L466A mice exhibit p53-dependent embryonic lethality, demonstrating MDM2 E3 ligase activity is essential for p53 regulation in vivo. Unexpectedly, cells expressing Mdm2L466A manifest cell cycle G2-M transition defects and increased aneuploidy even in the absence of p53, suggesting MDM2 E3 ligase plays a p53-independent role in cell cycle regulation and genome integrity. Furthermore, cells bearing the E3-dead MDM2 mutant show aberrant cell cycle regulation in response to DNA damage. This study uncovers an uncharacterized role for MDM2’s E3 ligase activity in cell cycle beyond its essential role in regulating p53’s stability in vivo.

BackgroundCIMAvax-EGF is an epidermal growth factor (EGF)-depleting immunotherapy which has shown survival benefit as a switch maintenance treatment after platinum-based chemotherapy in advanced non-small cell lung cancer (NSCLC). The primary objective of this trial is to establish the safety and recommended phase II dose (RP2D) of CIMAvax-EGF in combination with nivolumab as second-line therapy for NSCLC.MethodsPatients with immune checkpoint inhibitor-naive metastatic NSCLC were enrolled using a “3+3” dose-escalation design. Toxicities were graded according to CTCAE V4.03. Thirteen patients (one unevaluable), the majority with PD-L1 0%, were enrolled into two dose levels of CIMAvax-EGF.FindingsThe combination was determined to be safe and tolerable. The recommended phase 2 dose of CIMAvax-EGF was 2.4 mg. Humoral response to CIMAvax-EGF was achieved earlier and in a greater number of patients with the combination compared to historical control. Four out of 12 evaluable patients had an objective response.

Introduction: While combined immunotherapeutic activation of B and T cells is predicted to synergistically enhance both humoral and cell-mediated immunity, this has not been previously tested in human cancer patients. To assess this, we conducted the FIH trial combining B cell-activating IO (CIMAvax-EGF (C)) with T cell-activating IO (PD-1 blockade with nivolumab (N)). C activates B cells to produce endogenous anti-epidermal growth factor (EGF) antibodies (Abs) that deplete EGF from circulation. As a single agent maintenance therapy in aNSCLC, C prolonged overall survival (OS) in a randomized phase III trial. We now present immune correlates of improved OS with this novel combination. Methods: We reported safety and preliminary efficacy data from 13 pts from the phase I study of C+N (AACR 2019). Serial blood samples were analyzed by Luminex for dynamic changes in circulating cytokines and proteins implicated in EGF signaling, and by flow cytometry for peripheral blood mononuclear cell immunophenotype. Cox regression analysis was used to analyze association with OS. Results: 71% of pts achieved anti-EGF Ab titers of >1:4000 after 3 doses of C, compared to historical controls of 39% with C alone. Baseline increase in CD8+ naive T cells, memory B cells, and decrease in MDSC and Th17 T cells all significantly correlated with better OS. Baseline increase of inflammatory cytokines/markers (IL7, IL15, IFNα, IL8, sCD40L and CRP) significantly correlated with worse OS. Despite enrichment for pts with tumor proportion score (TPS) PD-L1 0%, median OS (mOS) of the intent-to-treat population (pts who received at least 1 dose of C+N, n=13) was 13.7 months (mo) vs. 9.9 mo in Checkmate 057 aNSCLC pts with tumor proportion score (TPS) PD-L1 <10%. mOS of per-protocol pts (all 4 loading doses of C+N given over 8 weeks, n=11) was 18.5 mo. In contrast to Checkmate 057 results, where OS benefit of N favored aNSCLC with KRAS mutation (KRASm), C+N showed better mOS of 22.4 mo in EGFR/ALK/KRAS wildtype (wt) pts versus 12.3 mo in KRASm/STKwt pts. Conclusion: This study provides evidence for synergistic effects of combining B and T cell activating IO in aNSCLC pts. C+N generated higher anti-EGF titers in more pts at an earlier time point vs. C alone, which is the first demonstration in pts of enhancement of Ab responses by immune checkpoint blockade. Immune analysis identified new specific immune cell populations and inflammatory markers that significantly correlate with OS with C+N that have not been previously identified with C or N as single agents. C+N improved OS compared to historical controls of aNSCLC pts receiving N in phase III trials. Of particular note, C+N showed better OS in pts with EGFR/ALK/KRAS wt aNSCLC versus KRASm/STKwt pts. The Phase II portion of the study is ongoing. Citation Format: Tania Crombet, Jason Muhitch, Circe Mesa, Rachel Evans, Danay S. Hernandez, Patricia L. Luaces, Zaima Mazorra, Orestes Morales, Carlos Cedeno, Aileen Cinquino, Daniel T. Fisher, Kelvin Lee, Mary Reid, Grace Dy. Evidence for synergistic immune responses in the first-in-human (FIH) combination of B cell-activating immunotherapy (IO) with anti-PD1 immune checkpoint inhibitor nivolumab (N) as 2nd-line therapy in patients (pts) with advanced non-small cell lung cancer (aNSCLC) [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr CT130.