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Page 18 of 38 J Cancer Metastasis Treat 2020;6:5 I http://dx.doi.org/10.20517/2394-4722.2020.13

Cyr61 stimulates active angiogenesis in various tumors, although the mechanism is unknown. Here,
we report that Cyr61 enhances the activity of tip cells during angiogenesis by regulating VEGFR2-
Hippo pathway. Microvessel networks and directional vascular cell migration patterns were deformed in
Cyr61-knockdown zebrafish embryos. Moreover, Cyr61 promoted the endothelial sprouting activity in
angiogenesis. Cyr61 induced the interaction of integrin αvβ3 with VEGFR2, which activated downstream
MAPK/PI3K signaling pathways, YAP/TAZ, and Rho effector mDia1 to enhance tip cell activity and Cyr61
itself. Integrin αvβ3 inhibitor repressed tip cell number and sprouting in postnatal retinas from endothelial
cell-specific Cyr61 transgenic mice (VE-Cadherin:Cyr61), and allograft tumors in Cyr61 transgenic mice
also showed hyperactive vascular sprouting. Cancer patients with high Cyr61 expression have poor survival
outcomes and positive correlation with integrin αvβ3 and high YAP/TAZ. Thus, our data underscore the
positive feedback regulation of tip cells by Cyr61 through integrin αvβ3/VEGFR2 and YAP/TAZ activity,
suggesting a promising therapeutic intervention for pathological angiogenesis.

25. Silibinin targets bone morphogenic protein 2 in its efficacy against ultraviolet B radiation-
induced promotion/progression of microscopic basal cell carcinoma formation

Rajesh Agarwal, Chapla Agarwal

Department of Pharmaceutical Sciences Skaggs School of Pharmacy and Pharmaceutical Sciences, University
of Colorado Anschutz Medical Campus, Aurora, CO 80045, USA.

Background and aim: Non-melanoma skin cancers (NMSCs) account for about half of all malignancies
diagnosed annually in the United States. Around 80% of NMSCs are basal cell carcinoma (BCC) and 20%
are squamous cell carcinoma (SCC). Whereas the efficacy of several chemopreventive agents has been
examined and reported against both BCC and SCC, a majority of these studies have focused on the test
agent’s activity in a long-term setting to determine the number of tumors formed. Notably, the studies
evaluating the efficacy of chemopreventive agents during early stage(s) of BCC development are lacking.
Accordingly, utilizing the well-established patched (Ptch)+/− mouse model of ultraviolet b (UVB)-induced
BCC formation, we excised skin samples from UVB exposed mice prior to tumor formation to study the
promotion/progression of BCC and to determine the target(s) of silibinin, a well-known skin cancer (SCC)
chemopreventive agent, in BCC tumor growth inhibition.

Experimental procedure: We used a multifactor approach: (1) long-term ultraviolet B radiation-induced
mouse skin tumorigenesis in Ptch heterozygous mice focusing on BCC; (2) investigating and quantifying
expression of molecular regulators and cyclobutane pyrimidine dimers by immunohistochemistry and/or
immunoblotting; and (3) real-time PCR with mouse signal transduction pathway finder PCR array.

Results: At as early as one month, we found that UVB exposure significantly increased the number of
mast cells in Ptch+/− mice by about 48% (P < 0.05), which was completely inhibited (to control levels)
by silibinin topical treatments. In Ptch+/+ mice, which do not develop BCC tumors, we did not observe
any increase in mast cells following UVB exposure, suggesting this could be a specific pathway in the
development of BCC. To decipher the molecular mechanism of these findings, we performed a PCR
profiler array analysis of several genes involved in signal transduction pathways which showed strong
differences between Ptch+/+ and Ptch+/− mice that were unexposed, UVB irradiated, and silibinin treated.
Most notably, following UVB exposure for one month, in Ptch+/− mice, the expression of Bone Morphogenetic
Protein 2 (BMP-2), Hairy/enhancer-of-split related with YRPW motif 1 (Hey1), and Inhibitor of DNA
binding 1 (Id1) was significantly upregulated when compared to Ptch+/+ mice. Additional studies focusing
on BMP-2 found that silibinin strongly inhibits UVB-induced expression of BMP-2 in Ptch+/− mouse