In the present study, we took advantage of a well-defined β-cell carcinogenesis model to assess the role of COUP-TFII during the angiogenic switch as well as pancreatic tumor progression. Our results highlight the notion that COUP-TFII is critical for the angiogenic switch during tumorigenesis. Notably, COUP-TFII is a member of the nuclear receptor superfamily whose activity can be regulated by small diffusible ligands. Indeed, structural analysis has shown that COUP-TFII contains a potential ligand binding pocket, and its activity could be stimulated by retinoids (25
), thus raising the interesting possibility that antagonists for COUP-TFII can be found, and supporting COUP-TFII as a novel “druggable” target for the treatment of tumors.
Using a xenograft model, we showed previously that COUP-TFII directly regulates Ang1 expression in pericytes to modulate angiogenesis (15
). However, angiogenic profile analysis indicated that there was little change between control and mutant animals with respect to Ang1 expression (). This is not unexpected since Casanovas et al. showed that Ang1 was induced by the islet tumor cells in this cancer model (26
) and the increase of Ang1 in the tumor cells may mask the down-regulation of COUP-TFII in the tumor environment. To assess if COUP-TFII regulates Ang1 expression in the tumor microenvironment in the current model, we isolated adjacent normal pancreatic tissue and pancreatic islet tumor, and compared Ang1 levels in control and mutant mice. Our results indicated that Ang1 expression was indeed decreased in the pancreatic tissue adjacent to tumor of COUP-TFII mutant mice (Supplementary Fig. 4
). These results are consistent with our previous findings (15
). As expected, the expression of Ang1 in islet tumors remains the same in control and mutant tumors due to the high expression of Ang1 elicited by β tumor cells.
Here, we provide compelling evidence that COUP-TFII controls VEGFR-2 signaling by directly suppressing the endothelial expression of VEGFR-1 and the sVEGFR-1 (27
). Consistent with the results from the pancreatic islet tumor model, we found that VEGFR-1 expression is consistently up regulated in the Matrigel Plug model (Supplementary Fig. 5
), indicating that regulation of VEGFR-1 expression by COUP-TFII is a general mechanism in both physiological and pathological neoangiogenesis process. In the RIP-Tag2 tumors, the results from genetic inactivation of VEGF alleles in pancreatic β cells (28
) have documented that VEGFR-2 signaling is a key driver for vascular growth. This notion is supported by the observation that sVEGF-R1 delivered by an adenovirus vector is a potent inhibitor of RIP-Tag2 tumor angiogenesis and tumor progression (30
). Although it has been reported that VEGFR-1 is also expressed in many nonendothelial cell types, including macrophages and various tumor cells (31
), in situ hybridization analysis revealed that VEGFR-1 expression is detectable only in the islet vasculature, at least in this tumor model (32
). Therefore, we conclude that in vivo
up regulation of VEGFR-1 expression in COUP-TFII mutant mice is primarily contributed by endothelial regulation by COUP-TFII.
Our results suggest that COUP-TFII is essential for tumor-induced neo-lymphangiogenesis. Intriguingly, we found that COUP-TFII did not regulate the transcription of VEGFR-1 in lymphatic endothelial cells (Supplementary Fig. 3
). Therefore, lymphangiogenesis defects displayed by COUP-TFII mutant mice is unlikely due to VEGFR-1 regulation by COUP-TFII. However, we recently showed that COUP-TFII enhanced the pro-lymphangiogenic actions of VEGF-C by directly stimulating expression of neuropilin-2, a co-receptor for VEGF-C, to control lymphatic vessel sprouting (22
). Loss of COUP-TFII significantly compromised VEGFR-3 signaling, which is a major receptor mediating VEGF signaling for lymphangiogenesis. Furthermore, COUP-TFII has been shown to physically and functionally interact with Prox1, a key regulator of lymphatic endothelial cell (19
), raising the interesting possibility that COUP-TFII acts jointly with Prox1 to regulate tumor lymphangiogenesis. In this regard, COUP-TFII might exert its effect in tumor lymphangiogenesis through regulation of VEGFR3/Nrp2 signaling and interactions with Prox1.
In summary, our results reveal a central role for COUP-TFII in the pathological angiogenic and lymphangiogenic response, and support COUP-TFII as a promising new target for anti-angiogenic therapy.