Bryan Manning’s research while affiliated with Wistar Institute and other places

Patients with metastatic ovarian cancer (OvCa) have a 5-year survival rate of <30% due to the persisting dissemination of chemoresistant cells in the peritoneal fluid and the immunosuppressive microenvironment in the peritoneal cavity. Here, we report that intraperitoneal administration of β-glucan and IFNγ (BI) induced robust tumor regression in clinically relevant models of metastatic OvCa. BI induced tumor regression by controlling fluid tumor burden and activating localized antitumor immunity. β-glucan alone cleared ascites and eliminated fluid tumor cells by inducing intraperitoneal clotting in the fluid and Dectin-1-Syk–dependent NETosis in the omentum. In omentum tumors, BI expanded a novel subset of immunostimulatory IL27+ macrophages and neutralizing IL27 impaired BI efficacy in vivo. Moreover, BI directly induced IL27 secretion in macrophages where single agent treatment did not. Finally, BI extended mouse survival in a chemoresistant model and significantly improved chemotherapy response in a chemo-sensitive model. In summary, we propose a new therapeutic strategy for the treatment of metastatic OvCa.

Patients with metastatic ovarian cancer (OvCa) have a 5-year survival rate of less than 30% due to persisting dissemination of chemoresistant cells in the peritoneal fluid and the immunosuppressive microenvironment in the peritoneal cavity. Here, we report that intraperitoneal administration of β-glucan and IFNγ (BI) induced robust tumor regression in clinically relevant models of metastatic OvCa. BI induced tumor regression by controlling fluid tumor burden and activating localized antitumor immunity. β-glucan alone cleared ascites and eliminated fluid tumor cells by inducing intraperitoneal clotting in the fluid and Dectin-1-Syk-dependent NETosis in the omentum. In omentum tumors, BI expanded a novel subset of immunostimulatory IL27+ macrophages and neutralizing IL27 impaired BI efficacy in vivo. Moreover, BI directly induced IL27 secretion in macrophages where single agent treatment did not. Finally, BI extended mouse survival in a chemoresistant model and significantly improved chemotherapy response in a chemo-sensitive model. In summary, we propose a new therapeutic strategy for the treatment of metastatic OvCa.

Significance: Most ovarian cancer (OC) patients ultimately succumb to drug-resistant intraperitoneal metastases. There is an unmet need to develop novel therapies against metastatic OC. Whether activating myeloid cells can promote OC regression has not been explored extensively. The purpose of this study is to develop a novel immunotherapy that activates myeloid cells with the goal of eradicating metastatic OC. Methods: In this study, we mainly relied on syngeneic mouse models of metastatic OC. To model metastatic OC in mice, we performed intraperitoneal injection of clinically relevant murine OC cell lines that were recently established by Dr. Robert Weinberg’s group, including homologous recombination (HR)-proficient KPCA cells (KRASG12VTrp53R172HCcne1OEAkt2OE ) and HR-deficient BPPNM cells (Brca1−/−Trp53R172HPten−/− Nf1−/−MycOE ). To activate myeloid cells, we used b-glucan and interferon (IFN) γ. b-glucan is a yeast cell wall polysaccharide that is currently in clinical trials to treat multiple cancers (NCT05159778). It canonically activates myeloid cells through the Dectin-1-spleen tyrosine kinase (SYK) pathway and induces infiltration of monocytes and neutrophils. IFNγ activates macrophages to an anti-tumor status. We investigated whether and how b-glucan affects metastatic OC with or without IFNγ using bioluminescence imaging, confocal imaging, flow cytometry, and single-cell RNA sequencing (scRNA seq). Finally, we tested whether combining myeloid cell activation with carboplatin could eradicate metastatic OC.Results: First, we found that b-glucan alone was highly efficient in clearing ascites. b-glucan captured free-floating OC cells into solid nodules within 5 hours after its injection through two non-redundant and equally important pathways: (1) an unexpected Dectin-1-SYK-independent, heparin-sensitive pathway that was mediated by resident macrophages in the peritoneal fluid; (2) a Dectin-1-SYK-PAD4-dependent pathway that was mediated by neutrophil extracellular traps in the omentum. Second, we found that combining b-glucan with IFNγ (b-glucan/IFNγ) not only cleared ascites but also reduced total metastases compared to PBS-, b-glucan-, or IFNγ-treated mice. This anti-tumor immunity required T cells and non-tumor IFNγ signaling in the host. Third, using scRNA seq, we found that b-glucan/IFNγ induced the enrichment of unique subsets of tumoricidal neutrophils and macrophages compared to other groups. Interestingly, the novel macrophage subset selectively expressed an interleukin (IL)-12 family cytokine, IL-27. We found that blocking IL-27 significantly impaired the anti-tumor response. Finally, combining b-glucan/IFNγ with carboplatin nearly eradicated chemoresistant KPCA tumors and chemosensitive BPPNM tumors. Patients with high expression of the genes identified in the novel myeloid cell subsets correlate with better overall survival. Conclusion: We have identified a novel combination therapy of myeloid activation and chemotherapy that could potentially transform treatments against metastatic OC. Citation Format: Brennah Murphy, Taito Miyamoto, Bryan Manning, Toshitha Kannan, Anastasia Ghilkovsky, Yulia Nefedova, Daniel Claiborne, Rugang Zhang, Andrew Kossenkov, Nan Zhang. Novel myeloid activation therapy promotes regression of metastatic ovarian cancer [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr B086.

Background Ovarian cancer (OvCa) is the most lethal gynecological cancer and the fifth leading cause of cancer-related deaths in women. Despite an initial positive response to therapy, most patients relapse and present with chemotherapy resistance. The prognosis for recurrent disease is poor, with a 5-year survival rate of < 30%. One contributor to disease recurrence and therapy resistance is the presence of disseminated cancer cells that remain in the peritoneal fluid after treatment. A growing body of evidence suggests that these cells exhibit dormancy characteristics that render them resistant to most therapies. Moreover, cancer cells in fluid are unable to be surgically resected. Because 75% of relapsed patients present with therapy-resistant intraperitoneal disease, developing new strategies to effectively target disseminating OvCa cells in the peritoneal fluid is crucial for the effective treatment of OvCa. Two key players in peritoneal immunity, the omentum and peritoneal resident macrophages (PRMΦs ), are known to sequester pathogens in the peritoneal fluid and coordinate an inflammatory immune response in the peritoneum. One such activator of peritoneal immunity is β-glucan, a sugar found on the cell walls of yeasts. Objective To target disseminating OvCa cells in the peritoneal fluid by activating myeloid cells in the peritoneal cavity via intraperitoneal administration of β-glucan. Methods C57bl/6J mice were injected with GFP-labeled murine OvCa cells ( KPCA : Trp53 −/−R172H Ccne1 OE Akt2 OE KRAS G12V) immediately followed by 500μg β-glucan (i.p.). Five hours later, mice were euthanized and their peritoneal lavage was analyzed for the presence of OvCa cells. To model advanced disease, Luciferase-KPCA cells were seeded in C57bl/6J mice one week prior to biweekly β-glucan treatment and imaged 3 weeks later. Results First, we found that β-glucan was highly efficient in acutely clearing OvCa cells from the peritoneal fluid. Mechanistically, β-glucan captured free-floating OvCa cells into solid nodules through two non-redundant and equally important pathways: (1) an unexpected Dectin-1/SYK-independent, heparin-sensitive pathway that was mediated by PRMΦ aggregation in the peritoneal fluid; and (2) a Dectin-1/SYK/PAD4-dependent pathway that was mediated by neutrophil extracellular traps in the omentum. Second, we found that β-glucan also completely cleared cancer from the peritoneal fluid and prevented ascites accumulation in advanced disease. Combining β-glucan with IFNγ (β-glucan/IFNγ) not only cleared ascites but also regressed omentum tumors and prevented intraperitoneal metastases as compared to PBS-, β-glucan-, or IFNγ-treated mice. Conclusions Intraperitoneal injection of β-glucan clears OvCa cells from the peritoneal fluid and has therapeutic potential to control OvCa metastasis.

Background Eradication of intraperitoneal metastasis of ovarian cancer (OC) remains an unmet challenge. Current T cell-mediating immunotherapies have not been applied to OC due to lack of efficacy in most patients. Immunosuppressive myeloid cells associate with tumor progression and treatment resistance in the metastatic sites of OC. The purpose of this study is to develop a novel immunotherapy that activates myeloid cells with the goal of eradicating metastatic OC. Methods Recently established, clinically relevant murine OC cells, that are homologous recombination proficient, KPCA (KRASG12VTrp53R172HCcne1OEAkt2OE ), were intraperitoneally injected to build the murine metastatic OC model. β-glucan and interferon (IFN) γ were used to activate myeloid cells. β-glucan is a yeast cell wall polysaccharide that is currently in clinical trials to treat multiple cancers. It canonically activates myeloid cells through the Dectin-1 pathway and induces infiltration of monocytes and neutrophils. IFNγ activates macrophages to an anti-tumor status. Tumor burden and tumor microenvironment in the ascites and omentum were evaluated using bioluminescence imaging, confocal imaging, flow cytometry, and single-cell RNA sequencing (scRNA seq) to investigate whether and how β-glucan modulates metastatic OC with or without IFNγ. Furthermore, OC patient survival was analyzed based on gene expression using public dataset. Finally, combination of myeloid cell activation with carboplatin was tested. Results β-glucan alone was efficient in clearing ascites, although it did not affect total metastases. On the other hand, combining β-glucan with IFNγ (β-glucan/IFNγ) not only cleared ascites but also reduced total metastases compared to PBS-, β-glucan-, or IFNγ-treated mice. This anti-tumor immunity required T cells and non-tumor IFNγ signaling in the host. scRNA seq of omental tumors revealed β-glucan/IFNγ induced the enrichment of a unique subset of neutrophils and macrophages compared to other groups. The neutrophil subset upregulated Camp and Ltf, which are granule proteins known to have tumoricidal function. Eukaryotic initiation factor-2 signaling, which is associated with reactive oxygen species (ROS) production, is the most upregulated pathway in the neutrophil subset. The macrophage subset selectively expressed interleukin (IL)-27, which has anti-tumor potential mainly through T cells. Blocking IL-27 significantly impaired the anti-tumor response of β-glucan/IFNγ. OC patients with high expression of these genes identified in the novel myeloid cell subsets correlate with better overall survival. Finally, combining β-glucan/IFNγ with carboplatin nearly eradicated chemoresistant KPCA tumors. Conclusions β-glucan/IFNγ suppressed metastatic OC enriching anti-tumor myeloid cell populations. Combination therapy of this myeloid cell activation and standard-of-care chemotherapy could potentially transform treatments against metastatic OC. Ethics Approval This study is approved by IACUC (#201536).