Podocalyxin enhances breast tumor growth and metastasis and is a target for monoclonal antibody therapy

Abstract

IntroductionPodocalyxin (gene name PODXL) is a CD34-related sialomucin implicated in the regulation of cell adhesion, migration and polarity. Upregulated expression of podocalyxin is linked to poor patient survival in epithelial cancers. However, it is not known if podocalyxin has a functional role in tumor progression.MethodsWe silenced podocalyxin expression in the aggressive basal-like human (MDA-MB-231) and mouse (4T1) breast cancer cell lines and also overexpressed podocalyxin in the more benign human breast cancer cell line, MCF7. We evaluated how podocalyxin affects tumorsphere formation in vitro and compared the ability of podocalyxin-deficient and podocalyxin-replete cell lines to form tumors and metastasize using xenogenic or syngeneic transplant models in mice. Finally, in an effort to develop therapeutic treatments for systemic cancers, we generated a series of antihuman podocalyxin antibodies and screened these for their ability to inhibit tumor progression in xenografted mice.ResultsAlthough deletion of podocalyxin does not alter gross cell morphology and growth under standard (adherent) culture conditions, expression of PODXL is required for efficient formation of tumorspheres in vitro. Correspondingly, silencing podocalyxin resulted in attenuated primary tumor growth and invasiveness in mice and severely impaired the formation of distant metastases. Likewise, in competitive tumor engraftment assays where we injected a 50:50 mixture of control and shPODXL (short-hairpin RNA targeting PODXL)-expressing cells, we found that podocalyxin-deficient cells exhibited a striking decrease in the ability to form clonal tumors in the lung, liver and bone marrow. Finally, to validate podocalyxin as a viable target for immunotherapy, we screened a series of novel antihuman podocalyxin antibodies for their ability to inhibit tumor progression in vivo. One of these antibodies, PODOC1, potently blocked tumor growth and metastasis.ConclusionsWe show that podocalyxin plays a key role in the formation of primary tumors and distant tumor metastasis. In addition, we validate podocalyxin as potential target for monoclonal antibody therapy to inhibit primary tumor growth and systemic dissemination.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-015-0562-7) contains supplementary material, which is available to authorized users.

Emerging Role of Podocalyxin in the Progression of Mature B-Cell Non-Hodgkin Lymphoma

“…Contrasting with these data, in the breast cancer cell lines MDA-MB-231, the highly aggressive MDA-MB-231 clone 4175 and NAMEC8R, all expressing high levels of endogenous PODXL, silencing of PODXL exerted no effect on cell proliferation under monolayer culture conditions [80][81][82]. However, the frequency of tumorsphere-forming cells was markedly decreased in PODXL-silenced MDA-MB-231 breast cancer cell line and, conversely, its overexpression in luminal-like MCF-7 breast cancer cell line, a low metastatic cell line expressing low levels of endogenous PODXL, resulted in increased tumorsphere formation [80]. Consistent with these results, silencing of PODXL in MDA-MB-231 cells reduced primary tumor growth in a mouse model xenograft [80,82].…”

“…However, the frequency of tumorsphere-forming cells was markedly decreased in PODXL-silenced MDA-MB-231 breast cancer cell line and, conversely, its overexpression in luminal-like MCF-7 breast cancer cell line, a low metastatic cell line expressing low levels of endogenous PODXL, resulted in increased tumorsphere formation [80]. Consistent with these results, silencing of PODXL in MDA-MB-231 cells reduced primary tumor growth in a mouse model xenograft [80,82]. Nevertheless, no effect of PODXL silencing on tumor growth was observed when the xenografted cell lines were MDA-MB-231 clone 4175 cells, NAMEC8R, or the pancreatic cancer cell lines SW1990 and Pa03c [81,83].…”

Podocalyxin influences malignant potential by controlling epithelial–mesenchymal transition in lung adenocarcinoma

“…Our results suggest that targeting PODXL might be a novel strategy for the treatment of lung adenocarcinoma. In this regard, there has been some recent progress towards the development of PODXL‐targeted therapies, including PODXL‐targeted monoclonal antibody therapy to inhibit primary tumor growth and systemic dissemination of breast cancer . In addition, Chijiiwa et al .…”

“…In this regard, there has been some recent progress towards the development of PODXL-targeted therapies, including PODXL-targeted monoclonal antibody therapy to inhibit primary tumor growth and systemic dissemination of breast cancer. (33) In addition, Chijiiwa et al (34) suggest that miR-5100, which directly binds to the 3 0 untranslated region of the PODXL transcript and downregulates PODXL expression, could have therapeutic potential for the treatment of metastatic pancreatic cancer.…”

Overexpression of microRNA-5100 decreases the aggressive phenotype of pancreatic cancer cells by targeting PODXL

“…PODXL was previously reported to enhance tumor aggressiveness in breast cancer, prostate cancer, oral squamous cell carcinoma and astrocytoma (56)(57)(58)(59). hsu et al reported PODXL-ebP50-ezrin molecular complex enhances the metastatic potential of renal cancer through recruiting RAC1 guanine nucleotide exchange factor, ARhgeF7 (60).…”