Nuray Erin

Schwann cells are glial cells responsible for the protection and repair of axons. Schwann cells activated by factors released from cancer cells resemble reactive Schwann cells and may enhance cancer growth and metastasis. Breast carcinoma is a heterogeneous disease and our previous studies demonstrated further heterogeneity in site-dependent metastasis. Specifically cells isolated from liver metastasis of 4T1 mammary carcinoma cells (4TLM) demonstrate distinct phenotypic features leading to diverse tumor microenvironment compared to 4T1 mammary carcinoma cells metastasized to brain (4TBM). In accordance, non-metastatic/hardly metastatic cells obtained from 4T1 mammary carcinoma (67NR cells) interacts with their microenvironment differently. Hence, we examined the effects of secretomes obtained from 67NR, 4TBM and 4TLM cells on primary Schwann cells obtained from mouse sciatic nerve, specifically focusing on inflammatory cytokine release. Secretomes of liver metastatic subset of mammary carcinoma cells (4TLM) induced striking increases in IL-6 secretion, which was markedly higher than IL-6 secretion induced by secretomes of 4TBM and 67NR cells. Similarly, only 4TLM secretomes induced remarkable increases in inflammatory chemokines KC and MIP-2. Monocyte Chemoattractant Protein-1 (MCP-1) (also called CCL2), involved in migration of monocyte, is highly secreted by 4TLM cells but not by 4TBM and 67NR cells. Secretomes of 4TLM cells also induced robust increases in MCP-1 secretion from Schwann cells. Interestingly metastatic cells decreased IL-1b secretion from Schwann cells. Next we determined how co-culturing of Schwann cells with carcinoma cells alter cytokine/chemokine secretion in order to determine contact-dependent changes. Schwann cells were co-cultured with cancer cells at 1/5, ½.5 and 1/1 ratio and secretomes were analyzed for cytokine secretion. Interestingly, co-culturing markedly suppressed IL-6 secretion from Schwann cells. Secretion of chemokines such as KC and MCP-1, however increased following co-culturing. Lastly we have performed proteomic analysis of Schwann cell secretomes in the presence and absence of secretomes from cancer cells. Peptides were analyzed using the Orbitrap Fusion mass spectrometry instrument (Thermo Scientific) coupled with the Dionex Ultimate 3000 RSLCnano system. For bioinformatics analysis, the Proteome Discoverer (PD) program was used. As the Venn diagram shows, 446 proteins were identified in secretomes of unstimulated Schwann cells. Interestingly 363 of these proteins were also found in metastatic and non-metastatic breast cancer cells. This was surprising given the differences in cell lineages. Analysis of the 363 common proteins by KEGG pathways revealed several important carcinoma-associated pathways, such as the HIF-1 signaling pathway. The 37 proteins unique to Schwann cells pointed out two reactome pathways: Regulation of Insulin-like Growth Factor transport and post-translational protein phosphorylation. Protein content of secretomes of Schwann cells stimulated with secretomes of cancer cells differed from unstimulated Schwann cells such that 13 proteins were markedly downregulated while over 200 proteins were upregulated. We are in process of analyzing these results. This study is supported by TÜBİTAK 122Z333

Purpose: Mammary carcinoma is comprised heterogeneous groups of cells with different metastatic potential. 4T1 mammary carcinoma cells metastasized to heart (4THM), liver (4TLM) and brain (4TBM) and demonstrate cancer-stem cell phenotype. Using these cancer cells we found thatTGF-β is the top upstream regulator of metastatic process. In addition, secretion of bone morphogenetic protein 1 (BMP-1), which is crucial for the proteolytic release of TGF-β, was markedly high in metastatic mammary cancer cells compared to non-metastatic cells. Although TGF-β inhibitors are in clinical trials, systemic inhibition of TGF-β may produce heavy side effects. We here hypothesize that inhibition of BMP-1 proteolytic activity inhibits TGF-β activity and induces anti-tumoral effects. Method and Results: Effects of specific BMP-1 inhibitor on liver and brain metastatic murine mammary cancer cells (4TLM and 4TBM), as well as on human mammary cancer MDA-MB-231 and MCF-7 cells, were examined and compared with the results of TGF-β inhibition. Inhibition of BMP-1 activity markedly suppressed proliferation of cancer cells and enhanced anti-tumoral effects of doxorubicin. Inhibition of BMP-1 activity but not of TGF-β activity decreased colony and spheroid formation. Differential effects of BMP-1 and TGF-β inhibitors on TGF-β secretion was also observed. Conclusions: These results demonstrated for the first time that the inhibition of BMP-1 activity has therapeutic potential for treatment of metastatic mammary cancer and enhances the anti-tumoral effects of doxorubicin.

Both sensory neurons and immune cells, albeit at markedly different levels, express the vanilloid (capsaicin) receptor, Transient Receptor Potential, Vanilloid-1 (TRPV1). Activation of TRPV1 channels in sensory afferent nerve fibers induces local effector functions by releasing neuropeptides (most notably, substance P) which, in turn, trigger neurogenic inflammation. There is good evidence that chronic activation or inactivation of this inflammatory pathway can modify tumor growth and metastasis. TRPV1 expression was also demonstrated in a variety of mammalian immune cells, including lymphocytes, dendritic cells, macrophages and neutrophils. Therefore, the effects of TRPV1 agonists and antagonists may vary depending on the prominent cell type(s) activated and/or inhibited. Therefore, a comprehensive understanding of TRPV1 activity on immune cells and nerve endings in distinct locations is necessary to predict the outcome of therapies targeting TRPV1 channels. Here, we review the neuro-immune modulation of cancer growth and metastasis, with focus on the consequences of TRPV1 activation in nerve fibers and immune cells. Lastly, the potential use of TRPV1 modulators in cancer therapy is discussed.

Pterygium, a disease of the ocular surface, is characterized by the proliferation and invasion of fibrovascular tissue. Chronic inflammation contributes to pterygium occurrence. Sensory neuropeptides of TRPV1-positive nerve fibers are involved in inflammation and corneal wound healing. The possible association between TRPV1 in nerve fibers and neuropeptides such as Substance P (SP) and Vasoactive Intestinal Peptide (VIP) in the recurrence of pterygium has not been examined before. The pterygia from 64 patients were used to determine changes in SP and VIP levels using 10 min acetic-acid extraction that yielded mainly neuronal peptides. There was a sufficient amount of pterygium tissues from the 35 patients for further immunohistochemical analysis of TRPV1 and S100, which is a glial marker to visualize nerve fibers. SP and VIP levels increased markedly in cases with primary and secondary recurrences, and there was a close correlation between SP and VIP levels. TRPV1 expression increased in the epithelium, while stromal expression decreased in recurrences. Nerve fibers were demonstrated mainly in the stroma, and serial sections confirmed the localization of TRPV1 with the nerve fibers. These results together with previous findings demonstrated that the increased epithelial expression of TRPV1 in recurrent pterygia might be involved in the pathogenesis, and the inhibition of epithelial TRPV1 activity may prevent recurrence.

Interactions between the immune system and the nervous system are crucial in maintaining homeostasis, and disturbances of these neuro-immune interactions may participate in carcinogenesis and metastasis. Nerve endings have been identified within solid tumors in humans and experimental animals. Although the involvement of the efferent sympathetic and parasympathetic innervation in carcinogenesis has been extensively investigated, the role of the afferent sensory neurons and the neuropeptides in tumor development, growth, and progression is recently appreciated. Similarly, current findings point to the significant role of Schwann cells as part of neuro-immune interactions. Hence, in this review, we mainly focus on local and systemic effects of sensory nerve activity as well as Schwann cells in carcinogenesis and metastasis. Specific denervation of vagal sensory nerve fibers, or vagotomy, in animal models, has been reported to markedly increase lung metastases of breast carcinoma as well as pancreatic and gastric tumor growth, with the formation of liver metastases demonstrating the protective role of vagal sensory fibers against cancer. Clinical studies have revealed that patients with gastric ulcers who have undergone a vagotomy have a greater risk of stomach, colorectal, biliary tract, and lung cancers. Protective effects of vagal activity have also been documented by epidemiological studies demonstrating that high vagal activity predicts longer survival rates in patients with colon, non-small cell lung, prostate, and breast cancers. However, several studies have reported that inhibition of sensory neuronal activity reduces the development of solid tumors, including prostate, gastric, pancreatic, head and neck, cervical, ovarian, and skin cancers. These contradictory findings are likely to be due to the post-nerve injury-induced activation of systemic sensory fibers, the level of aggressiveness of the tumor model used, and the local heterogeneity of sensory fibers. As the aggressiveness of the tumor model and the level of the inflammatory response increase, the protective role of sensory nerve fibers is apparent and might be mostly due to systemic alterations in the neuro-immune response. Hence, more insights into inductive and permissive mechanisms, such as systemic, cellular neuro-immunological mechanisms of carcinogenesis and metastasis formation, are needed to understand the role of sensory neurons in tumor growth and spread.

The ability of tumor cells to evade apoptosis is established as one of the hallmarks of cancer. The deregulation of apoptotic pathways conveys a survival advantage enabling cancer cells to develop multi-drug resistance (MDR), a complex tumor phenotype referring to concurrent resistance toward agents with different function and/or structure. Proteins implicated in the intrinsic pathway of apoptosis, including the Bcl-2 superfamily and Inhibitors of Apoptosis (IAP) family members, as well as their regulator, tumor suppressor p53, have been implicated in the development of MDR in many cancer types. The PI₃K/AKT pathway is pivotal in promoting survival and proliferation and is often overactive in MDR tumors. In addition, the tumor microenvironment, particularly factors secreted by cancer-associated fibroblasts, can inhibit apoptosis in cancer cells and reduce the effectiveness of different anti-cancer drugs. In this review, we describe the main alterations that occur in apoptosis-and related pathways to promote MDR. We also summarize the main therapeutic approaches against resistant tumors, including agents targeting Bcl-2 family members, small molecule inhibitors against IAPs or AKT and agents of natural origin that may be used as monotherapy or in combination with conventional therapeutics. Finally, we highlight the potential of therapeutic exploitation of epigenetic modifications to reverse the MDR phenotype.