The PDGF-B/Rβ signaling pathway has been characterized in invasive breast cancer. PDGFRα and β are commonly overexpressed both in tumor and stromal cells (27
), and PDGF receptor β has been shown to be upregulated in breast cancer endothelial cells (28
). Autocrine PDGFR signaling has been shown to promote breast cancer metastasis (29
). PDGF-D is the most recent addition to the PDGF family (10
). Whether PDGF-D is expressed in human breast cancer tissues and whether it affects tumor growth and metastasis in vivo
are not known. In this study, we observed for the first time that PDGF-D is highly expressed in human breast cancers. We found that overexpression of PDGF-D activates the PDGFRβ signaling and enhances tumor growth and metastasis. We then, blocked PDGF-D signaling using both genetic (shRNA) and pharmacological (imatinib) approaches. We observed that PDGF-D knockdown decreased tumor growth, but inhibition of PDGFRβ by imatinib has only minimal effect on parental MDA-MB-231 tumor growth. The efficacy of the pharmacological approach may be reduced due to the abnormal tumor microenvironment resulting in decreased drug delivery. However, it is also possible that PDGF-D exerts its pro-tumor effects via other signaling pathways. Indeed, recent studies reported PDGF-D crosstalk with PI3K/Akt, Notch and NF-κB signaling pathways and increased tumor angiogenesis and invasion (7
). In summary, we conclude that although PDGF-D pathway alone does not provide a sufficient autocrine signal to accelerate MDA-MB-231 cell growth in vitro
, it potentiates tumor growth and metastasis in vivo
via: (i) stimulation of cancer cell proliferation through ERK1/2 and p38 MAPK activation, (ii) protection of cancer cells from apoptosis via Akt activation, and (iii) stimulation of tumor cell metastasis via activation of SDF-1α/CXCR4 pathway.
Others and we have previously shown that overexpression of PDGF-D increased tumor metastasis in renal cell carcinoma (8
) and PDGF-D contributes to epithelial-mesenchymal transition (EMT) phenotype in prostate cancer cells (30
). Here, we show a novel mechanism by which PDGF-D enhances metastasis. We found that overexpression of PDGF-D increase nodal metastasis of orthotopic MDA-MB-231 breast carcinomas via CXCR4 signaling. Blocking CXCR4 with AMD3100, a specific inhibitor of CXCR4, abolished PDGF-D-induced metastasis, suggesting that CXCR4 signaling directly mediates PDGF-D induced metastasis. A growing body of literature has indicated that SDF-1α/CXCR4 pathway plays a critical role in cancer cell metastasis (31
). CXCR4 expression level in human breast cancers is positively correlated with lymph node metastasis and inversely correlated with patient prognosis (32
). In our study, we observed PDGF-D increased CXCR4 in tumor tissues but not in tumor cells in culture, suggesting that PDGF-D in tumor microenvironment either indirectly upregulates CXCR4 expression or increases recruitment of CXCR4 expressing stromal cells. Further study is needed to dissect the mechanism of PDGF-D induced CXCR4 expression and pinpoint the cell type in which CXCR4 is upregulated.
Another important finding of this study is that overexpression of PDGF-D improves the delivery and efficacy of cytotoxic therapies via normalization of tumor vasculature. In breast cancer, VEGF and PDGF families have been identified as angiogenic factors (34
). PDGF-D has been shown to upregulate VEGF and to stimulate angiogenesis (36
). In our study, overexpression of PDGF-D did not change VEGF expression in tumor cells or in host tissues (data not shown). As a result, we did not observe differences in tumor angiogenesis between parental and PDGF-D overexpressing tumors. However, we found that PDGF-D overexpression normalized tumor blood vessel morphology and increased perivascular cell coverage. This vessel maturation can attenuate vascular hyperpermeability, lower tumor interstitial fluid pressure (IFP), and restore pressure gradients across vessel walls as well as into tissues (20
). As a result, tissue penetration of chemotherapeutic agents is improved (20
). In some cases, blocking PDGFR signaling by imatinib has been shown to lower IFP (37
), presumably via different mechanisms, such as fibroblast contraction. In this study, we showed that PDGF-D could normalize vessel function and increase the penetration of chemotherapeutic agents and ultimately improve the efficacy of anti-tumor therapy in an orthotopic breast cancer model, suggesting a new strategy for enhancing the effects of chemotherapy.
In summary, PDGF-D is highly expressed in human breast carcinoma tissues. On one hand, overexpression of PDGF-D leads to increased tumor growth and metastasis; on the other hand, overexpression of PDGF-D normalizes tumor blood vessels and improves delivery and efficacy of chemotherapeutic drugs. Our preclinical study suggests that PDGFR-β blockers should be used cautiously, especially in combination with chemotherapy in the clinic. For example, in patients with high levels of PDGF-D in tumors, imatinib treatment should be avoided at the time of cytotoxic therapy. Future studies are thus necessary to explore the risks and benefits of PDGFR-β blockade for anticancer treatment.