Wang, Enfeng

Abstract Metastasis, the leading cause of cancer death, requires tumor cell intravasation, migration through the bloodstream, arrest within capillaries, and extravasation to invade distant tissues. Few mechanistic details have been reported thus far regarding the extravasation process or re-entry of circulating tumor cells at metastatic sites. Here, we show that neuropilin-2 (NRP-2), a multifunctional nonkinase receptor for semaphorins, vascular endothelial growth factor (VEGF), and other growth factors, expressed on cancer cells interacts with α5 integrin on endothelial cells to mediate vascular extravasation and metastasis in zebrafish and murine xenograft models of clear cell renal cell carcinoma (RCC) and pancreatic adenocarcinoma. In tissue from patients with RCC, NRP-2 expression is positively correlated with tumor grade and is highest in metastatic tumors. In a prospectively acquired cohort of patients with pancreatic cancer, high NRP-2 expression cosegregated with poor prognosis. Through biochemical approaches as well as Atomic Force Microscopy (AFM), we describe a unique mechanism through which NRP-2 expressed on cancer cells interacts with α5 integrin on endothelial cells to mediate vascular adhesion and extravasation. Taken together, our studies reveal a clinically significant role of NRP-2 in cancer cell extravasation and promotion of metastasis. Cancer Res; 73(14); 4579–90. ©2013 AACR.

Although biomedical applications of nanotechnology, which typically involve functionalized nanoparticles, have taken significant strides, biological characterization of unmodified nanoparticles remains underinvestigated. Herein we demonstrate that unmodified gold nanoparticles (AuNPs) inhibit the proliferation of cancer cells in a size- and concentration-dependent manner by abrogating MAPK-signaling. In addition, these AuNPs reverse epithelial-mesenchymal transition (EMT) in cancer cells by reducing secretion of a number of proteins involved in EMT, up-regulating E-Cadherin, and down-regulating Snail, N-Cadherin, and Vimentin. Inhibition of MAPK signaling and reversal of EMT upon AuNP treatment inhibits tumor growth and metastasis in two separate orthotopic models of ovarian cancer. Western blot analyses of tumor tissues reveal up-regulation of E-Cadherin and down-regulation of Snail and phospho-MAPK, confirming the reversal of EMT and inhibition of MAPK signaling upon AuNP treatment. The ability of a single self-therapeutic nanoparticle to abrogate signaling cascades of multiple growth factors is distinctive and purports possible medical applications as potential antitumor and antimetastatic agent.

Abstract VEGF/vascular permeability factor (VEGF/VPF or VEGF-A) is a pivotal driver of cancer angiogenesis that is a central therapeutic target in the treatment of malignancy. However, little work has been devoted to investigating functions of VEGF that are independent of its proangiogenic activity. Here, we report that VEGF produced by tumor cells acts in an autocrine manner to promote cell growth through interaction with the VEGF receptor neuropilin-1 (NRP-1). Reducing VEGF expression by tumor cells induced a differentiated phenotype in vitro and inhibited tumor forming capacity in vivo, independent of effects on angiogenesis. Autocrine activation of tumor cell growth was dependent on signaling through NRP-1, and Ras was determined to be a critical effector signaling molecule downstream of NRP-1. Our findings define a novel function for VEGF in dedifferentiation of tumor cells expanding its role in cancer beyond its known proangiogenic function. Cancer Res; 72(16); 3912–8. ©2012 AACR.

AbstractThe Na+/H+ exchanger regulatory factor-2 (NHERF-2) is an integral component of almost all endothelial cells (ECs), yet its endothelial function is not known. Here, we found that NHERF-2, is a key regulator of endothelial homeostasis because NHERF-2–silenced ECs proliferate at a much higher rate even in the absence of mitogens such as VEGF compared with control ECs. We further show that the hyperproliferation phenotype of NHERF-2–silenced EC is because of an accelerated cell cycle that is probably caused by a combination of the following factors: increased cytoplasmic calcium, increased expression of c-Myc, increased expression of cyclin D1, and reduced expression of p27. Using an experimental mouse model of human hemangioma, we found that the endothelial neoplasms derived from NHERF-2–silenced cells were much larger in volume than those derived from control cells. Thus, NHERF-2 is a negative regulator of endothelial proliferation and may have important roles in endothelial homeostasis and vascular modeling.

Abstract Prostate cancer is the most common cancer and the second leading cause of cancer death in men in the United States. Definitive treatment consists of either radical prostatectomy or radiotherapy followed by androgen deprivation. Although majority of the prostate cancers usually responds to these therapies and can be cured, some of them will eventually relapse. It is believed that the relapsed cancers obtained new ability that facilitates their resistance to these therapeutic stresses. Increased expression of VEGF-C and its receptor Neuropilin-2 (NRP-2) in hormone-refractory and metastatic prostate cancers indicates that the VEGF-C/NRP-2 axis could play an important role in the progression of prostate cancer. Indeed, we observed VEGF-C/Neuropilin-2 axis promotes survival of prostate cancer cells during stress induced by hydrogen peroxide or radiation. Detail signaling events downstream of neuropilin-2 for this stress-resistant function have also been determined. Our findings therefore demonstrated a novel and distinct function of NRP-2 in protecting cancer cells from stress-induced cell death, thereby facilitating cancer recurrence and metastasis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3950.

Abstract It is a challenge to develop a universal single drug that can treat breast cancer at single- or multiple-stage complications, yet remains nontoxic to normal cells. The challenge is even greater when breast cancer–specific, estrogen-based drugs are being developed that cannot act against multistaged breast cancer complications owing to the cells differential estrogen receptor (ER) expression status and their possession of drug-resistant and metastatic phenotypes. We report here the development of a first cationic lipid-conjugated estrogenic derivative (ESC8) that kills breast cancer cells independent of their ER expression status. This ESC8 molecule apparently is nontoxic to normal breast epithelial cells, as well as to other noncancer cells. ESC8 induces apoptosis through an intrinsic pathway in ER-negative MDA-MB-231 cells. In addition, ESC8 treatment induces autophagy in these cells by interfering with the mTOR activity. This is the first example of an estrogen structure–based molecule that coinduces apoptosis and autophagy in breast cancer cells. Further in vivo study confirms the role of this molecule in tumor regression. Together, our results open new perspective of breast cancer chemotherapy through a single agent, which could provide the therapeutic benefit across all stages of breast cancer. Mol Cancer Res; 9(3); 364–74. ©2011 AACR.

Abstract Although the importance of RGS-GAIP–interacting protein (GIPC) in the biology of malignant cells is well known, the molecular mechanism of GIPC in the inhibition of tumor progression has not been identified. This study focused on elucidating the molecular role of GIPC in breast cancer progression. By using a human breast tumor specimen, an in vivo mouse model, and breast cancer cell lines, we showed for the first time that GIPC is involved in breast cancer progression through regulation of breast cancer cell proliferation, survival, and invasion. Furthermore, we found that the Akt/Mdm2/p53 axis, insulin-like growth factor-1 receptor, matrix metalloproteinase-9, and Cdc42 were downstream of GIPC signaling in breast cancer cells. Moreover, we showed that wild-type p53 reduced GIPC-induced breast cancer cell survival, whereas mutant p53 inhibited GIPC-induced cell invasion. Finally, we demonstrated that an N-myristoylated GIPC peptide (CR1023, N-myristoyl-PSQSSSEA) capable of blocking the PDZ domain of GIPC successfully inhibited MDA-MB-231 cell proliferation, survival, and further in vivo tumor growth. Taken together, these findings demonstrate the importance of GIPC in breast tumor progression, which has a potentially significant impact on the development of therapies against many common cancers expressing GIPC, including breast and renal cancer. Mol Cancer Res; 8(12); 1591–600. ©2010 AACR.

AbstractEndothelial cell–specific chemotaxis receptor (ECSCR) is a cell surface protein expressed by blood endothelial cells with roles in endothelial cell migration and signal transduction. We investigated the function of ecscr in the development of the zebrafish vasculature. Zebrafish ecscr is expressed in angioblasts and in axial vessels during angioblast migration and vasculogenesis. Morpholino-directed ecscr knockdown resulted in defective angioblast migration in the posterior lateral plate mesoderm, a process known to depend on vascular endothelial-derived growth factor (VEGF). In cultured cells, transfected ECSCR localized to actin-rich membrane protrusions, colocalizing with kinase insert domain protein receptor (KDR)/VEGF receptor 2 in these regions. ECSCR-silenced cells show reduced VEGF-induced phosphorylation of KDR but not of FMS-like tyrosine kinase 1 (FLT1)/VEGF receptor 1. Finally, chemical inhibition of VEGF receptor activity in zebrafish resulted in angioblast deficiencies that partially overlap with those seen in ecscr morphants. We propose that ecscr promotes migration of zebrafish angioblasts by enhancing endothelial kdr sensitivity to VEGF.

Abstract Oncogenic activation of Bmi-1 is found in a wide variety of epithelial malignancies including ovarian cancer, yet a specific mechanism for overexpression of Bmi-1 has not been determined. Thus, realizing the immense pathologic significance of Bmi-1 in cancer, we wanted to investigate if microRNA (miRNA) aberrations played a role in the regulation of Bmi-1 in ovarian cancer. In this report, we identify two miRNAs, miR-15a and miR-16, that are underexpressed in ovarian cell lines and in primary ovarian tissues. We show that these miRNAs directly target the Bmi-1 3′ untranslated region and significantly correlate with Bmi-1 protein levels in ovarian cancer patients and cell lines. Furthermore, Bmi-1 protein levels are downregulated in response to miR-15a or miR-16 expression and lead to significant reduction in ovarian cancer cell proliferation and clonal growth. These findings suggest the development of therapeutic strategies by restoring miR-15a and miR-16 expression in ovarian cancer and in other cancers that involve upregulation of Bmi-1. [Cancer Res 2009;69(23):9090–5]

Abstract Recurrence and subsequent metastatic transformation of cancer develops from a subset of malignant cells, which show the ability to resist stress and to adopt to a changing microenvironment. These tumor cells have distinctly different growth factor pathways and antiapoptotic responses compared with the vast majority of cancer cells. Long-term therapeutic success can only be achieved by identifying and targeting factors and signaling cascades that help these cells survive during stress. Both microarray and immunohistochemical analysis on human prostate cancer tissue samples have shown an increased expression of vascular endothelial growth factor-C (VEGF-C) in metastatic prostate cancer. We have discovered that VEGF-C acts directly on prostate cancer cells to protect them against oxidative stress. VEGF-C increased the survival of prostate cancer cells during hydrogen peroxide stress by the activation of AKT-1/protein kinase Bα. This activation was mediated by mammalian target of rapamycin complex-2 and was not observed in the absence of oxidative stress. Finally, the transmembrane nontyrosine kinase receptor neuropilin-2 was found to be essential for the VEGF-C–mediated AKT-1 activation. Indeed, our findings suggest a novel and distinct function of VEGF-C in protecting cancer cells from stress-induced cell death, thereby facilitating cancer recurrence and metastasis. This is distinctly different from the known function of VEGF-C in inducing lymphangiogenesis. [Cancer Res 2009;69(15):6042–8]