Establishment Of Tumor Microenvironment Research Articles

Steroidal alkaloid solanidine impedes hypoxia-driven ATM phosphorylation to switch on anti-angiogenesis in lung adenocarcinoma

PurposeThe declined oxygen tension in the cancer cell leads to the hypoxic adaptive response and favors establishment of tumor micro environment [TEM]. The complex TME consists of interwoven hypoxic HIF-1α and DNA damage repair ATM signaling. The ATM/HIF-1α phosphorylation switch on angiogenesis and abort apoptosis. Targeting this signaling nexus would be a novel therapeutic strategy for the treatment of cancer. BackgroundSteroidal alkaloid solanidine is known for varied pharmacological role but with less molecular evidences. Our earlier findings on solanidine proven its anti-neoplastic activity by inducing apoptosis in lung cancer. In continued research, efforts have been made to establish the underlying molecular signaling in induction of DNA damage in prevailing hypoxic TME. MethodsThe solanidine induced DNA damage was assessed trough alkali COMET assay; signaling nexus and gene expression profile analysis through IB, qRT-PCR, Gelatin Zymography, IHC, IF and ELISA. Pathophysiological modulations assessed through tube formation, migration, invasion assays. Anti-angiogenic studies through CAM, rat aorta, matrigel assays and corneal neovascularization assay. Anti-tumor activity through in-vivo DLA ascites tumor model and LLC model. ResultsThe results postulates, inhibition of hypoxia driven DDR proteins pATMser1981/pHIF-1αser696 by solanidine induces anti-angiogenesis. Systematic study of both non-tumorigenic and tumorigenic models in-vitro as well as in-vivo experimental system revealed the angio-regression mediated anticancer effect in lung cancer. These effects are due to the impeded expression of angiogenic mediators such as VEGF, MMP2&9 and inflammatory cytokines IL6 and TNFα to induce pathophysiological changes ConclusionThe study establishes new role of solanidine by targeting ATM/HIF-1α signaling to induce anti-angiogenesis for the first time. The study highlights the potentiality of plant based phytomedicine solanidine which can targets the multiple hallmarks of cancer by targeting interwoven signaling crosstalk. Such an approach through solanidine necessary to counteract heterogeneous complexity of cancer which could be nearly translated into drug.

Abstract 3153: Prediction of metabolites from cancer-associated fibroblasts that can have profound effects on cancer cells

Abstract Cancer-associated fibroblasts (CAFs) are the most abundant component of tumor microenvironment (TME) that is an ensemble of non-cancerous cells surrounding cancer cells. In particular, CAFs are known to have tumor-assisting functions, which may promote active growth and survival of the interacting cancer cells through their metabolic reprogramming despite being in a harsh environment (e.g., hypoxia or low pH). In this regard, CAF-derived metabolites are expect to have profound effects on metabolism of the cancer cells. To identify novel CAF-derived metabolites that may heavily influence the cancer cells, CAF-specific genome-scale metabolic models (GEMs) were reconstructed and simulated. GEM is a computational model that can predict a metabolic state in a cell-specific manner by integrating omics data (e.g., RNA-seq data). In this study, a total of six GEMs were reconstructed by integrating RNA-seq data from one normal fibroblast, originating from pancreas, and five CAFs associated with breast, colon, skin, lung, and pancreatic tumors. Simulation of the GEMs predicted 20 metabolites that appeared to be heavily used by all the five CAF GEMs in comparison with the one normal fibroblast GEM. Such metabolites were expected to be significantly associated with establishment of TME that is favorable for the cancer cells’ growth. Some of these metabolites are associated with pyrimidine metabolism, fatty acid metabolism, and primary bile acid biosynthesis. Upon experimental validation, the predicted metabolites are expected to help better understand CAFs and TME in terms of metabolites, and also provide clues for identifying effective drug targets for cancer treatment. Citation Format: Hae Deok Jung, Yoo Jin Sung, Jae Yong Ryu, Hyun Uk Kim. Prediction of metabolites from cancer-associated fibroblasts that can have profound effects on cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3153.

Phospholipase A2 Drives Tumorigenesis and Cancer Aggressiveness through Its Interaction with Annexin A1

Phospholipids are suggested to drive tumorigenesis through their essential role in inflammation. Phospholipase A2 (PLA2) is a phospholipid metabolizing enzyme that releases free fatty acids, mostly arachidonic acid, and lysophospholipids, which contribute to the development of the tumor microenvironment (TME), promoting immune evasion, angiogenesis, tumor growth, and invasiveness. The mechanisms mediated by PLA2 are not fully understood, especially because an important inhibitory molecule, Annexin A1, is present in the TME but does not exert its action. Here, we will discuss how Annexin A1 in cancer does not inhibit PLA2 leading to both pro-inflammatory and pro-tumoral signaling pathways. Moreover, Annexin A1 promotes the release of cancer-derived exosomes, which also lead to the enrichment of PLA2 and COX-1 and COX-2 enzymes, contributing to TME formation. In this review, we aim to describe the role of PLA2 in the establishment of TME, focusing on cancer-derived exosomes, and modulatory activities of Annexin A1. Unraveling how these proteins interact in the cancer context can reveal new strategies for the treatment of different tumors. We will also describe the possible strategies to inhibit PLA2 and the approaches that could be used in order to resume the anti-PLA2 function of Annexin A1.

Proteinaceous Hydrogels for Bioengineering Advanced 3D Tumor Models.

The establishment of tumor microenvironment using biomimetic in vitro models that recapitulate key tumor hallmarks including the tumor supporting extracellular matrix (ECM) is in high demand for accelerating the discovery and preclinical validation of more effective anticancer therapeutics. To date, ECM‐mimetic hydrogels have been widely explored for 3D in vitro disease modeling owing to their bioactive properties that can be further adapted to the biochemical and biophysical properties of native tumors. Gathering on this momentum, herein the current landscape of intrinsically bioactive protein and peptide hydrogels that have been employed for 3D tumor modeling are discussed. Initially, the importance of recreating such microenvironment and the main considerations for generating ECM‐mimetic 3D hydrogel in vitro tumor models are showcased. A comprehensive discussion focusing protein, peptide, or hybrid ECM‐mimetic platforms employed for modeling cancer cells/stroma cross‐talk and for the preclinical evaluation of candidate anticancer therapies is also provided. Further development of tumor‐tunable, proteinaceous or peptide 3D microtesting platforms with microenvironment‐specific biophysical and biomolecular cues will contribute to better mimic the in vivo scenario, and improve the predictability of preclinical screening of generalized or personalized therapeutics.

Extracellular vesicles isolated from human renal cell carcinoma tissues disrupt vascular endothelial cell morphology via azurocidin.

Cancer-associated extracellular vesicles (EVs) are intimately involved in establishment of tumor microenvironment and occurrence of metastasis. However, previous studies have mainly relied on experiments with cultured cell lines or mouse models, making it difficult to gain a full understanding of EV functions in human body. Hence, we extracted EVs directly from surgically resected viable clear cell renal cell carcinoma (ccRCC) tissues and adjacent normal renal tissues (n = 20). Quantitative LC/MS analysis identified 3,871 tissue-exudative EV (Te-EV) proteins, among which azurocidin (AZU1) was highly enriched in tumor Te-EVs (p = 2.85 × 10-3 , fold-change = 31.59). Importantly, AZU1 content was also significantly higher in serum EVs from ccRCC patients compared to those from healthy donors. We further found that ccRCC-derived EVs had AZU1-dependent membrane permeabilizing activity for the vascular endothelial cell layer. Thus Te-EVs should be ideal resource for investigation of physiological EV functions.

Abstract TMEM-032: MULTIDISCIPLINARY CHARACTERIZATION OF OVARIAN CANCER SPHERES

Abstract Sphere-forming cultures have been widely used in stem cell biology. Ovarian cancer sphere culture is also a good model to study ovarian cancer ascites spheroids, which propagate without attachment to a substratum. We studied non-adherent ovarian cancer sphere cultures derived from ovarian cancer cell lines and epithelial cancer cells isolated from clinical samples. Western blot analysis and immunofluorescence showed that ovarian cancer spheres expressed elevated levels of stem cell markers CD133, c-Myc, Nanog, and Oct4. Metabolically, ovarian cancer sphere cells showed decreased ATP levels and were under constant oxidative stress, with significant depletion of the endogenous antioxidant glutathione. Accordingly, supplementation with antioxidant supplement N-acetylcysteine (NAC) in cancer sphere cultures relieved oxidative stress and improved growth of the sphere cultures in a dose-dependent manner. In contrast, adherent cancer cell cultures did not show any significant growth enhancement. Furthermore, microarray gene expression profiling and activity-based proteomic profiling using a sulfonate ester chemical probe revealed that the ovarian cancer spheres demonstrated increased level of FOS/JUN expression, which activated the expression and activity of the omega class of glutathione-S transferase GSTO1, an enzyme involved in the metabolism of xenobiotics and cisplatin resistance. Knockdown of FOS expression using siRNA reduced the levels of GSTO1 and abrogated chemoresistance of the sphere cultures. Separately, gene set enrichment analysis of the cancer sphere expression profiles revealed elevated expression of endosomal pathway genes for exosome secretion in the sphere cultures. Knockdown of one elevated gene, Rab27B, significantly reduced the exosome number in the spent medium of ovarian cancer sphere cultures. Clinically, measurement of the exosome number in patient body fluids using the NanoSight instrument and a flow cytometry method indicated that plasma and urine samples from a panel of ovarian cancer patients had increased number of exosomes compared with samples derived from benign patients. CONCLUSIONS: Our multidisciplinary studies have discovered a plethora of properties of ovarian cancer spheres that may promote ovarian cancer growth. Elevated expression and activity of GSTO1 in ovarian cancer spheres may represent a novel mechanism by which ovarian cancer spheroids respond to heightened oxidative stress and chemotherapeutic agents. Antioxidant supplements may not have the intended benefits to cancer survivors. Instead, they may promote cancer growth by relieving the oxidative stress of ovarian cancer spheroids. In contrast, inhibitors that target GSTO1 may have better clinical value in prolonging survivorship of ovarian cancer patients with malignant ascites. Ovarian cancer spheres also have increased expression of endosomal pathway genes for exosome secretion, which may promote the establishment of tumor microenvironment for cancer propagation, and is consistent with the increased exosomes in the plasma and urine samples of ovarian cancer patients. Citation Format: Shubai Liu, Junzheng Yang, Pui-Wah Choi, Jamie Sui-Lam Kwok, Kathleen Hasselblatt, Daniel Nomura, Wing Ping Fong, Stephen KW Tsui, Allison Vitonis, Daniel Cramer, William R. Welch, Benjamin Cravatt, Ross Berkowitz, and Shu-Wing Ng. MULTIDISCIPLINARY CHARACTERIZATION OF OVARIAN CANCER SPHERES [abstract]. In: Proceedings of the 11th Biennial Ovarian Cancer Research Symposium; Sep 12-13, 2016; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(11 Suppl):Abstract nr TMEM-032.