Measurement of proliferative index in squamous cell carcinoma by flow cytometry

Squamous cell carcinoma of the upper aerodigestive tract presents significant problems in determining appropriate treatment regimens. Staging aided by sophisticated investigations allows for planning of treatment, but there is a definite need for a specific and reproducible marker to quantify biological aggressiveness. For some classes of tumors the DNA ploidy of cells, as determined by flow cytometry, has shown good correlation to pathologic grading and prognosis. Using a mathematical model, it is possible to calculate the S-phase fraction, which is indicative of proliferative activity and may reflect tumor aggressiveness. However, this parameter is often difficult to determine reliably in squamous cell carcinoma. An optimal marker would be a measureable protein related to proliferation. An attempt was made to use flow cytometry to measure the nuclear enzyme topoisomerase II to assess proliferation in cultured cell lines. Although the antibody was specific to the extracted protein, constituents of the rabbit serum bound non-specifically throughout fixed cells. Further purification of this antibody preparation could realize its diagnostic potential. Antibody against proliferating cell nuclear antigen was more specific, resulting in good correlation flow cytometrically with the S-phase fraction of the cultured cell lines. The results obtained with these protein markers deems further investigation into their use as prognostic indicators. The protocol has been established to apply this technology to tumor samples and establish a meaningful parameter of biological behavior of tumors, upon which treatment regimens can be confidently based.