Targeting survival pathways in cancer stem cells

The discovery of tumor-initiating cells endowed with stem-like features, and therefore referred to as cancer stem cells (CSCs), has added a further level of complexity to the pathobiology of neoplastic diseases. This uncommon cellular subpopulation has been connected with tumor initiation, metastatization and treatment failure. CSCs are protected against standard medical treatments by multiple mechanisms including the abnormal activation of both DNA damage repair signals and canonical survival pathways. Moreover, this cellular subset relies on dedicated signals such as self-renewal-linked molecular circuits. Therefore, the exact definition of the target population and the identification of molecular networks differentially activated in CSCs compared with their differentiated progeny are crucial for an optimal pre-clinical development of molecular targeted agents. We isolated breast CSCs from surgically resected primary breast tumors. A first round of experiments was designed in order to determine whether our clones met the operative criteria to be defined as CSCs. In particular, all these clones possessed: i) the expression of a unique repertoire of markers common to stem and progenitor cells, ii) an unlimited growth in vitro using media optimized for stem cell cultures and iii) the ability to reproduce the parental tumor upon injection in immunocompromised mice. Afterword, the metastatic potential of genetically engineered BCSCs was compared with that of differentiated breast cancer cells (dBCCs) . Upon both intracardiac and orthotopic injection the undifferentiated pool was able to generate distant metastases and to recapitulate the dissemination pattern of the human disease, while dBCCs failed to generate distant lesions. Afterwards, high-throughput assays have been exploited in order to define molecular mechanisms underlying this differential metastatic proclivity. In particular, most up- and down-regulated genes were evaluated for their convergence on canonical signal pathways/biological functions. BCSCs displayed higher levels of DNA repair-linked effectors such as BRCA1, ATR, ATM and Chk1, higher level of the pro-tumorigenic and pro-metastatic protein c-MET and, finally, they were characterized by lower levels of physiological Wnt inhibitors (Dkk family members). Therefore, we identified three different pathways/functions whose hyper-activation seems to be correlated with the metastatic ability of BCSCs. Notably, molecular effectors of the above-mentioned pathways can be pharmacologically antagonized by experimental targeted agents.