Senescence-associated tumor growth is promoted by 12-Lipoxygenase.

Radiation therapy is a commonly used treatment modality for cancer. Although effective in providing local tumor control, radiation causes oxidative stress, inflammation, immunomodulatory and mitogenic cytokine production, extracellular matrix production, and premature senescence in lung parenchyma. The senescence associated secretory phenotype (SASP) can promote inflammation and stimulate alterations in the surrounding tissue. Therefore, we hypothesized that radiation-induced senescent parenchymal cells in irradiated lung would enhance tumor growth. Using a murine syngeneic tumor model of melanoma and non-small cell lung cancer lung metastasis, we demonstrate that radiation causes a significant increase in markers of premature senescence in lung parenchyma within 4 to 8 weeks. Further, injection of B16F0 (melanoma) or Lewis Lung carcinoma (epidermoid lung cancer) cells at these time points after radiation results in an increase in the number and size of pulmonary tumor nodules relative to unirradiated mice. Treatment of irradiated mice with a senolytic agent (ABT-737) or agents that prevent senescence (rapamycin, INK-128) was sufficient to reduce radiation-induced lung parenchymal senescence and to mitigate radiation-enhanced tumor growth. These agents abrogated radiation-induced expression of 12-Lipoxygenase (12-LOX), a molecule implicated in several deleterious effects of senescence. Deficiency of 12-LOX prevented radiation-enhanced tumor growth. Together, these data demonstrate the pro-tumorigenic role of radiation-induced senescence, introduces the dual TORC inhibitor INK-128 as an effective agent for prevention of radiation-induced normal tissue senescence, and identifies senescence-associated 12-LOX activity as an important component of the pro-tumorigenic irradiated tissue microenvironment. These studies suggest that combining senotherapeutic agents with radiotherapy may decrease post-therapy tumor growth.

MAL2-Induced Actin-Based Protrusion Formation is Anti-Oncogenic in Hepatocellular Carcinoma.

Recent studies report that the polarity gene myelin and lymphocyte protein 2 (MAL2), is overexpressed in multiple human carcinomas largely at the transcript level. Because chromosome 8q24 amplification (where MAL2 resides) is associated with hepatocellular- and cholangio-carcinomas, we examined MAL2 protein expression in these human carcinoma lesions and adjacent benign tissue using immunohistochemistry. For comparison, we analyzed renal cell carcinomas that are not associated with chromosome 8q24 amplification. Surprisingly, we found that MAL2 protein levels were decreased in the malignant tissues compared to benign in all three carcinomas, suggesting MAL2 expression may be anti-oncogenic. Consistent with this conclusion, we determined that endogenously overexpressed MAL2 in HCC-derived Hep3B cells or exogenously expressed MAL2 in hepatoma-derived Clone 9 cells (that lack endogenous MAL2) promoted actin-based protrusion formation with a reciprocal decrease in invadopodia. MAL2 overexpression also led to decreased cell migration, invasion and proliferation (to a more modest extent) while loss of MAL2 expression reversed the phenotypes. Mutational analysis revealed that a putative Ena/VASP homology 1 recognition site confers the MAL2-phenotype suggesting its role in tumor suppression involves actin remodeling. To reconcile decreased MAL2 protein expression in human carcinomas and its anti-oncogenic phenotypes with increased transcript levels, we propose a transcriptional regulatory model for MAL2 transient overexpression.

BRAF Inhibition Stimulates Melanoma-Associated Macrophages to Drive Tumor Growth.

PURPOSE: To investigate the roles of melanoma-associated macrophages in melanoma resistance to BRAF inhibitors (BRAFi). EXPERIMENTAL DESIGN: An in vitro macrophage and melanoma cell coculture system was used to investigate whether macrophages play a role in melanoma resistance to BRAFi. The effects of macrophages in tumor resistance were examined by proliferation assay, cell death assay, and Western blot analyses. Furthermore, two mouse preclinical models were used to validate whether targeting macrophages can increase the antitumor activity of BRAFi. Finally, the number of macrophages in melanoma tissues was examined by immunohistochemistry. RESULTS: We demonstrate that in BRAF-mutant melanomas, BRAFi paradoxically activate the mitogen-activated protein kinase (MAPK) pathway in macrophages to produce VEGF, which reactivates the MAPK pathway and stimulates cell growth in melanoma cells. Blocking the MAPK pathway or VEGF signaling then reverses macrophage-mediated resistance. Targeting macrophages increases the antitumor activity of BRAFi in mouse and human tumor models. The presence of macrophages in melanomas predicts early relapse after therapy. CONCLUSIONS: Our findings demonstrate that macrophages play a critical role in melanoma resistance to BRAFi, suggesting that targeting macrophages will benefit patients with BRAF-mutant melanoma.

SECTM1 produced by tumor cells attracts human monocytes via CD7-mediated activation of the PI3K pathway.

Tumor-associated macrophages (TAMs) have essential roles in tumor progression and metastasis. Tumor cells recruit myeloid progenitors and monocytes to the tumor site, where they differentiate into TAMs; however, this process is not well studied in humans. Here we show that human CD7, a T-cell and NK cell receptor, is highly expressed by monocytes and macrophages. Expression of CD7 decreases in M-CSF-differentiated macrophages and in melanoma-conditioned medium-induced macrophages (MCMI/Mφ) in comparison to monocytes. A ligand for CD7, SECTM1 (secreted and transmembrane protein 1), is highly expressed in many tumors, including melanoma cells. We show that SECTM1 binds to CD7 and significantly increases monocyte migration by activation of the PI3K (phosphatidylinositol 3'-kinase) pathway. In human melanoma tissues, tumor-infiltrating macrophages expressing CD7 are present. These melanomas, with CD7-positive inflammatory cell infiltrations, frequently highly express SECTM1, including an N-terminal, soluble form, which can be detected in the sera of metastatic melanoma patients but not in normal sera. Taken together, our data demonstrate that CD7 is present on monocytes and tumor macrophages and that its ligand, SECTM1, is frequently expressed in corresponding melanoma tissues, possibly acting as a chemoattractant for monocytes to modulate the melanoma microenvironment.

Melanoma-derived conditioned media efficiently induce the differentiation of monocytes to macrophages that display a highly invasive gene signature.

The presence of tumor-associated macrophages (TAMs) in melanomas is correlated with a poor clinical prognosis. However, there is limited information on the characteristics and biological activities of human TAMs in melanomas. In this study, we developed an in vitro method to differentiate human monocytes to macrophages using modified melanoma-conditioned medium (MCM). We demonstrate that factors from MCM-induced macrophages (MCMI-Mφ) express both M1-Mφ and M2-Mφ markers and inhibit melanoma-specific T-cell proliferation. Furthermore, microarray analyses reveal that the majority of genes up-regulated in MCMI-Mφ are associated with tumor invasion. The most strikingly up-regulated genes are CCL2 and MMP-9. Consistent with this, blockade of both CCL-2 and MMPs diminish MCMI-Mφ-induced melanoma invasion. Finally, we demonstrated that both MCMI-Mφ and in vivo TAMs express the pro-invasive, melanoma-associated gene, glycoprotein non-metastatic melanoma protein B. Our study provides a framework for understanding the mechanisms of cross-talk between TAMs and melanoma cells within the tumor microenvironment.