Cancer stem cells related marker CD44 and Notch activation in metastatic colon cancer patients

Background: Notch pathway has been suggested to play an important role in regulating cancer stem cells (CSCs) in colon cancer. CSCs are responsible for colon cancer initiation, progression and may be involved in resistance to conventional therapies. Moreover, CSCs promote angiogenesis throughout several factors such as MDK, EGF, IL-8. CD44 is a transmembrane glycoprotein reported as a putative marker for isolating CSCs. Preclinical studies and a clinical study from our group (Negri et al, 2015) showed that strong Notch intracellular cleaved domain (NICD) is associated with resistance to anti-vascular endothelial growth factor (VEGF) therapy. Based on these reports, we assessed NICD and CD44 expression in a series of metastatic colon cancer patients treated with anti-VEGF monoclonal antibody bevacizumab within first-line clinical trials. Methods: NICD and CD44 expression were assessed by immunohistochemistry (IHC) on paraffin-embedded samples from 42 pretreatment primary tumors. Samples were stained by antibody against NICD (1:50, Cell Signaling Technology, MA, USA) and CD44 (1:50, Novocastra Laboratories, Newcastle upon Tyne, UK), processed using the streptavidin-biotin-peroxidase complex (Dako Corp.,CA, USA) and counterstained with hematoxylin eosin solution. Level of positivity on scale from 0 (negative) to 3 (strong positivity) was assessed for NICD and CD44 staining. Results: Table 1 shows a positive association between NICD and CD44 expression in colon cancer patients. Cases with strong NICD reactivity and high CD44 staining experienced a shorter progression-free (PFS) and overall survival (OS) as compared with cases with strong NICD and low CD44 expression (PFS: 4.7 vs 14.7 months, p = 0.029; OS: 14.5 vs 33.4 months, p = 0.073). Conclusions: Our hypothesis is that Notch activation could maintain tissue stem cells and that in this scenario the anti-VEGF therapy could be of reduced relevance.