The Role of Darpp-32 and t-Darpp in Breast Cancer Tumorigenesis and the Development of Acquired Drug Resistance

Each year approximately 233,000 women in the United States are diagnosed with breast cancer. Survival depends greatly not only on the grade and stage of the cancer at diagnosis, but also on the molecular type of breast cancer. Around 20-25% of women with breast cancer overexpress the human epidermal growth factor receptor 2 (HER2) and are diagnosed with HER2-positive breast cancer. Overexpression of HER2 stimulates malignant growth and survival. Currently there are four FDA-approved drugs targeted to HER2; however, women with this type of cancer continue to have a poor prognosis and low disease-free survival rate, largely because acquired resistance to targeted therapies develops so rapidly. In order to improve cancer patient outcomes it is imperative that we identify novel therapeutic targets.;Upregulation of the protein t-Darpp (truncated variant of Darpp-32) has been linked directly to resistance to trastuzumab (HerceptinRTM), a well-established treatment for HER2-positive breast cancer. The full-length protein Darpp-32 (dopamine and cAMP-regulated phosphoprotein of 32 kDa) reverses the trastuzumab resistance phenotype induced by t-Darpp. This suggests that these two proteins have antagonistic functions in determining a cell's sensitivity to trastuzumab and possibly in breast cancer in general. Our research goal was to better understand the role of Darpp-32 and t-Darpp in tumor development and progression.;In order to determine the role of Darpp-32 and t-Darpp in tumor development, Darpp-32 and t-Darpp expression in healthy and tumorigenic mouse mammary tissue was examined. We observed a predominance of Darpp-32 in healthy breast tissue and a predominance of t-Darpp in tumor breast tissue. Using Darpp-32/t-Darpp knockout mice, crossed with a mouse mammary tumor model, we showed that these proteins are linked to increased tumor growth. Together, these data suggest that t-Darpp, and to a lesser degree Darpp-32, are directly involved in breast tumor development.;In order to investigate the role of Darpp-32 and t-Darpp in tumor progression, we examined their involvement in resistance to another HER2-targeted therapy, lapatinib (TykerbRTM). Western and RT-PCR analysis showed that lapatinib-resistant cells exhibited a shift in their relative levels of Darpp-32 and t-Darpp toward a predominance of t-Darpp in resistant cells, as compared to parental cells. While overexpression of t-Darpp was insufficient to cause de novo lapatinib resistance in a short-term cell proliferation assay, it was enough to confer a survival and/or growth advantage during prolonged exposure to drug. The mechanism responsible for this survival advantage is not well understood, but it may be linked to failed lapatinib-induced upregulation of the pro-apoptotic protein BIM in cells overexpressing t-Darpp.;Relative levels of Darpp-32 and t-Darpp seem to be important in breast cancer growth and drug resistance, and shifts in expression from Darpp-32 to t-Darpp occur relatively quickly, suggesting a potential epigenetic mechanism of gene regulation. Hypomethylation of DNA in human breast cancer cell lines caused the upregulation of Darpp-32. This evidence suggests that DNA methylation plays a role in regulating gene expression at this locus. A better understanding of the mechanisms regulating Darpp-32 and t-Darpp expression as well as their relative functions in tumorigenesis and drug resistance has the potential to improve patient outcomes.