Human colorectal cancers with an intact p16/cyclin D1/pRb pathway have up-regulated p16 expression and decreased proliferation in small invasive tumor clusters

A systematic spatial heterogeneity with high proliferative activity at the luminal border and low activity at the invasive margin is an unexpected beh avior that has been observed in colorectal cancer (CRC). To clarify this ph enomenon and possible underlying regulatory mechanisms, we have by immunohi stochemistry elucidated the proliferative activity and the expression of G1 /S regulatory proteins in small and large tumor cell clusters at the invasi ve margin in 97 CRCs. By identifying small tumor clusters at the tumor fron t, actually invading cancer cells could be characterized. and analyzed sepa rately. These cells could then be compared. with the main tumor mass repres ented by the larger tumor clusters. The proliferation was significantly low er in small tumor clusters compared with larger clusters (P < 0.001) and th e decrease in proliferation was correlated with a p16 up-regulation (r(s =) -0.41, P < 0.001), Interestingly, CRCs lacking p16 expression (18%) or tum ors with other aberrations in the p16/cyclin D2/pRb pathway had a less pron ounced decrease in proliferation between large and small clusters (P < 0.00 1), further strengthening the association between p16 and ceased proliferat ion at the invasive margin. This contrasts to tumors with low p27 or abnorm al p53 levels showing sustained proliferation in small tumor clusters. Our findings imply that invading CRC cells generally have low proliferative act ivity, and this phenomenon seems to be mediated through p16 and the p16/cyc lin D1/pRb pathway.