Nitric Oxide (NO) is a soluble endogenous gas with various biological functions like signaling, and working as an effector molecule or metabolic regulator. In response to inflammatory signals, immune myeloid cells, like macrophages, increase production of cytokines and NO, which is important for pathogen killing. Under these proinflammatory circumstances, called “M1”, macrophages undergo a series of metabolic changes including rewiring of their tricarboxylic acid (TCA) cycle. Here, we review findings indicating that NO, through its interaction with heme and non-heme metal containing proteins, together with components of the electron transport chain, functions not only as a regulator of cell respiration, but also a modulator of intracellular cell metabolism. Moreover, diverse effects of NO and NO-derived reactive nitrogen species (RNS) involve precise interactions with different targets depending on concentration, temporal, and spatial restrictions. Although the role of NO in macrophage reprogramming has been in evidence for some time, current models have largely minimized its importance. It has, therefore, been hiding in plain sight. A review of the chemical properties of NO, past biochemical studies, and recent publications, necessitates that mechanisms of macrophage TCA reprogramming during stimulation must be re-imagined and re-interpreted as mechanistic results of NO exposure. The revised model of metabolic rewiring we describe here incorporates many early findings regarding NO biochemistry and brings NO out of hiding and to the forefront of macrophages immunometabolism.

Breast cancer is the second leading cause of cancer death among U.S. women although the mortality is on a steadily decreasing trend since early 90s. We previously demonstrated a strong correlation between elevated NOS2 and COX2 expressions suggesting these genes can serve as markers of poor prognosis in estrogen receptor negative (ER-) breast cancer. In other words, NOS2 and COX2 is generally associated with the immune response. Induction of NOS2 and COX2 has also been shown to orchestrate multiple pro-oncogenic pathways that may contribute to the immunosuppressive nature of cancer cells. To better understand the interactions between cytokines and breast cancer cells and develop effective therapeutics for breast cancer, we examined the effects of selected cytokines on MDA-MB-231, a human (ER-) breast adenocarcinoma cell line. We found that in combination with cytokines, such as IFNG plus TNF, IL1B, or IL17, significantly induced NOS2 and COX2. IFNG plus TNF or IL1B most strongly induced NOS2 and COX2 where IFNG or IL17 to a lesser extent. It indicates that expressions of NOS2 and COX2 required IFNG but at a low dose in presence of other cytokines. Single-cell RNAseq analysis shows that cells treated with IFNG plus TNF or IL1B clustered with those expressed NOS2 and COX2 as revealed in tSNE plots. Expression of other factors such as IL8 and IL6 (suggested markers for prognosis of breast cancer) or PDL1 and IDO1 (suggested cancer therapeutic targets) also co-clustered with NOS2 and COX2 expressions. In addition, we found increased IL1A and IL1B expressions provide fortification to NOS2 and COX2 expressions. Taken together, our current findings indicate the interactions between breast cancer cells and cytokines. We believe further spatial transcriptomic and proteomic studies may be beneficial for clarifying the complex networks of the immunosuppressive tumor microenvironment. Citation Format: Yuk Sing Robert Cheng, Lisa Ridnour, Veena Somasundaram, Dana Bhattacharyya, Christopher McGinity, Daniel McVicar, Stephen Anderson, Stephen Lockett, David Wink. Interferon gamma induces NOS2 and COX2 expression in estrogen receptor negative breast cancer leading to immunosuppressive tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3804.

The metabolic requirements and functions of cancer and normal tissues are vastly different. Due to the rapid growth of cancer cells in the tumor microenvironment, distorted vasculature is commonly observed, which creates harsh environments that require rigorous and constantly evolving cellular adaption. A common hallmark of aggressive and therapeutically resistant tumors is hypoxia and hypoxia-induced stress markers. However, recent studies have identified alterations in a wide spectrum of metabolic pathways that dictate tumor behavior and response to therapy. Accordingly, it is becoming clear that metabolic processes are not uniform throughout the tumor microenvironment. Metabolic processes differ and are cell type specific where various factors promote metabolic heterogeneity within the tumor microenvironment. Furthermore, within the tumor, these metabolically distinct cell types can organize to form cellular neighborhoods that serve to establish a pro-tumor milieu in which distant and spatially distinct cellular neighborhoods can communicate via signaling metabolites from stroma, immune and tumor cells. In this review, we will discuss how biochemical interactions of various metabolic pathways influence cancer and immune microenvironments, as well as associated mechanisms that lead to good or poor clinical outcomes.

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