Acharya, Abhinav P

Abstract Background: Despite the success of chimeric antigen receptor (CAR) T cell therapy in hematological malignancies, its efficacy in solid tumors remains limited. Macrophages, as key players in the innate immune response, offer unique advantages in the solid tumor context due to their ability to infiltrate tissues and phagocytose tumor cells. Preclinical studies have demonstrated that CAR-engineered macrophages (CAR-Ms) can penetrate solid tumors, initiate phagocytosis, and promote adaptive immunity. Early-phase clinical trials infusing CAR-Ms into patients have shown a favorable safety profile. However, within the immunosuppressive tumor microenvironment (TME), macrophages are often reprogrammed into tumor-promoting, M2-like tumor-associated macrophages (TAMs), which limit their therapeutic efficacy. Metabolic reprogramming is a critical driver of this polarization and TCA derived succinate has been shown to promote M1 phenotype in macrophages. In this study, we investigate whether succinate-loaded nanoparticles (PES NPs) can metabolically rewire CAR-Ms to maintain a pro-inflammatory (M1-like) phenotype and enhance antitumor activity in ovarian cancer (OC). Results: We synthesized polyethylene succinate (PES) nanoparticles (NPs) and analyzed their size, polydispersity, and zeta (ζ) potential using dynamic light scattering (DLS). The results showed a negative potential, indicative of stability, and suggested that succinate could be effectively encapsulated in nanoparticles. OC cells abundantly express mesothelin protein; therefore, mesothelin-targeting CARs were chosen to transfect mouse bone marrow-derived macrophages (BMDMs) or human monocyte-derived macrophages (HMDMs) to generate CAR-Ms. Our results showed that mouse and human CAR-Ms treated with PES NPs exhibited increased mitochondrial respiration and glycolysis, seen as elevated ECAR and OCR relative to vehicle control or cell-permeable dimethyl succinate ester (DMSA). Flow cytometry analysis revealed that PES treatment resulted in increased expression of markers of the classical M1 phenotype in both mouse and human CAR-Ms. The CAR-Ms also had elevated mRNA levels of proinflammatory cytokines like IL-6, TNFα, and IL1β, while anti-inflammatory cytokines like IL-10 remained low. Furthermore, IL-4-induced M2-like CAR-Ms could be reversed to an M1-like phenotype by PES NPs. Consistent with these results, PES NP-treated CAR-Ms exhibited increased phagocytic activity toward OC cells expressing mesothelin. Building on these in vitro results, the effect of PES NP-treated CAR-Ms on the reduction of tumor growth in OC tumor models will be further investigated. Conclusions: Our findings demonstrate that PES NPs effectively reprogram CAR-Ms to maintain a pro-inflammatory M1 phenotype. This reprogramming enhances their phagocytic activity and target cell killing in vitro. Collectively, these results support intracellular succinate delivery as a promising strategy to improve the efficacy of CAR-M-based therapies in OC. Citation Format: Remya Raja, Alexis Hand, Madhan Mohan Chandra Sekhar Jaggarapu, Jason Tonne, Richard Vile, Abhinav Acharya, Marion Curtis. Succinate based metabolic reprogramming of CAR-macrophages for ovarian cancer immunotherapy [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Ovarian Cancer Research; 2025 Sep 19-21; Denver, CO. Philadelphia (PA): AACR; Cancer Res 2025;85(18_Suppl):Abstract nr B063.

Abstract Rheumatoid arthritis (RA) causes immunological and metabolic imbalances in tissue, exacerbating inflammation in affected joints. Changes in immunological and metabolic tissue homeostasis at different stages of RA are not well understood. Herein, the changes in the immunological and metabolic profiles in different stages in collagen induced arthritis (CIA), namely, early, intermediate, and late stage is examined. Moreover, the efficacy of the inverse‐vaccine, paKG(PFK15+bc2) microparticle, to restore tissue homeostasis at different stages is also investigated. Immunological analyses of inverse‐vaccine‐treated group revealed a significant decrease in the activation of pro‐inflammatory immune cells and remarkable increase in regulatory T‐cell populations in the intermediate and late stages compared to no treatment. Also, glycolysis in the spleen is normalized in the late stages of CIA in inverse‐vaccine‐treated mice, which is similar to no‐disease tissues. Metabolomics analyses revealed that metabolites UDP‐glucuronic acid and L‐Glutathione oxidized are significantly altered between treatment groups, and thus might provide new druggable targets for RA treatment. Flux metabolic modeling identified amino acid and carnitine pathways as the central pathways affected in arthritic tissue with CIA progression. Overall, this study shows that the inverse‐vaccines initiate early re‐establishment of homeostasis, which persists through the disease span.

Abstract Adoptive Cell Transfer (ACT) therapies have recently fostered excitement for the treatment of hematogenous cancers. However, efficacy has largely been stunted in the treatment of solid tumors, especially for ovarian cancer, in part due to nutrient restriction within the tumor microenvironment. We recently demonstrated that intracellular delivery of biodegradable microparticles (MPs) made from polymers of alpha-ketoglutarate modulated dendritic cell energy metabolism and immunophenotype, altering regulatory and helper T cell frequencies. To determine if continuous metabolite release from MPs would rescue T cell dysfunction in nutrient-poor, immunosuppressive conditions, we generated MPs of polymerized glycolytic and tricarboxylic acid cycle central-carbon metabolites. Flow cytometric analysis and confocal microscopy indicated that FITC-conjugated MPs were efficiently loaded into the cytoplasm of activated primary murine T cells via membrane adsorption or electroporation. The MPs were retained intracellularly and sustained proliferation of mouse T cells for over five days in vitro. Glycolysis and mitochondrial respiration stress tests using an Agilent Seahorse XFe96 Analyzer demonstrated that primary mouse CD8 T cells utilized the MPs as a metabolic substrate. In particular, MPs increased metabolic function, as indicated by increased spare respiratory capacity, glycolysis, and glycolytic capacity. The results of this study provide preliminary evidence in support of utilizing sustained-releasing particles of metabolic substrates for the metabolic reprogramming of immune cells used in ACT therapies. This strategy has potential to improve or expand the efficacy of existing ACT therapies for solid tumors.

Disruption of metabolic homeostasis at the organismal level can cause metabolic syndrome associated with obesity. The role of adipose tissue in cancer has been investigated over the last several decades with many studies implicating obesity as a risk factor for the development of cancer. Adipose tissue contains a diverse array of immune cell populations that promote metabolic homeostasis through a tightly controlled balance of pro- and anti-inflammatory signals. During obesity, pro-inflammatory cell types infiltrate and expand within the adipose tissue, exacerbating metabolic dysfunction. Some studies have now shown that the intracellular metabolism of immune cells is also deregulated by the lipid-rich environment in obesity. What is not fully understood, is how this may influence cancer progression, metastasis, and anti-tumor immunity. This review seeks to highlight our current understanding of the effect of adipose tissue on immune cell function and discuss how recent results offer new insight into the role that adipose tissue plays in cancer progression and anti-tumor immunity.

Metabolites control immune cell functions, and delivery of these metabolites in a sustained manner modulate the function of the immune cells.