Targeting Intrinsically-Resistant Breast Cancer Stem Cells with Gamma-Secretase Inhibitors.

Abstract

Abstract Background: We showed previously that tumorigenic, mammosphere-forming human breast cancer cells characterized by high CD44 and low or undetectable CD24 levels (CD44+/CD24-/low) are intrinsically resistant to conventional chemotherapy, and therefore may be responsible for cancer relapse. Our goal is identify novel drugs that selectively target these chemotherapy-resistant, tumor-initiating cells. Gene expression analysis of CD44+/CD24-/low cells vs. non-tumorigenic cells implicated the Notch, PI3K, and Hedgehog signaling pathways in regulating CD44+/CD24-/low cells. Thus, Notch, PI3K-AKT, and/or Hedgehog inhibitors may eliminate this unique subpopulation of cancer cells, either alone or in combination with chemotherapy, and could improve patient outcome. To test this hypothesis, we are carrying out a series of preclinical and clinical studies using a gamma-secretase inhibitor (GSI) to target the Notch pathway. Methods: For preclinical studies, stable xenograft lines were generated by transplantation of human tumor biopsy fragments into immunocompromised mice. Mice with tumors (n = 32, 150-300 mm3) were randomized to four treatment groups: 1) vehicle control, 2) chemotherapy: docetaxel 3) drug: GSI (MRK-003, Merck) or 4) combination: docetaxel + MRK-003. During treatment, mice were monitored for tumor volume and body weight. At the end of the treatment cycle, residual tumors were characterized by FACS for the percentage of CD44+/CD24-/low cells, as well as for mammosphere-forming efficiency (MSFE) and tumor-initiating capacity. In a complementary clinical trial, breast cancer biopsies taken before and after treatment with GSI (MK-0752, Merck) were characterized for expression of CD44, CD24, and ALDH by FACS and for MSFE. Results and Conclusions: In preclinical studies using two independent triple negative xenograft lines, Notch pathway inhibition reduced mammosphere formation but did not affect tumor volume, with no consistent change in marker expression by FACS. In patient samples, MSFE also declined after the first cycle of GSI/chemotherapy and remained low after subsequent cycles. This response corresponded with a stasis of metastatic growth during five cycles of treatment, but metastatic burden began to increase coincident with the sixth cycle of treatment. Marker analysis suggests that GSI treatment chemo-sensitizes a significant proportion of the otherwise chemo-resistant CD44+/CD24-/low cell population indicating that they are dependent on the Notch pathway for survival. The decrease of MSFE in both preclinical and clinical studies suggests that inhibition of the Notch pathway by GSI may reduce the number of tumorigenic cancer cells that would otherwise remain after chemotherapy. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 48.