ASSESSMENT OF HUMAN PROSTATE TUMOR CELL HETEROGENEITY

The intent of this research is to investigate human prostate cancer heterogeneity by employing a multiparametric analysis of data obtained through double beam flow cytometry. Prostate cancer was the third commonest fatal tumor in American males of all ages and the leading cause of cancer death in American males over the age of 55. As high as these incidences are for clinical cancer the situation could be as much as five times worse if all men with incidental or latent prostate cancer were to clinically manifest their disease. To date, there is no sure way of predicting an individual patient's prognosis based upon standard histopathology. However, when histologic techniques that can assess and quantitate tumor cell heterogeneity are employed, differences between the clinically silent and clinically manifest prostatic cancers can be identified. One quantitative technique that was capable of predicting patient prognosis employed manually performed nuclear morphometry. Therefore, the specific aims of this application are to employ double beam flow cytometry to grade human prostate cancer by assessing tumor cell heterogeneity based upon morphologic, genetic, and biochemical parameters. Preliminary studies by the principal investigator have shown that perpendicular light scatter at 448 nm wavelength correlates closely with nuclear shape and nuclear size when assessing living intact enzymatically suspended cells. Forward light scatter is known to correlate with cell size. Also, Hoechst 33342 is known to reversibly and quantitatively measure the DNA content of intact cells. Differentiation between live and dead cells can be accomplished by the addition of trypan blue which, when added to Hoechst-stained cells, will quench fluorescence in dead cells. Lastly, flavone 3-diphosphate wil produce an insoluble fluorescent product when acted upon by acid phosphatase. The multiparametric analysis of nuclear and cellular morphology as determined by light scatter, DNA content determine by Hoechst 33342, and acid phosphatase activity as determined by flavone 3-diphosphate should allow for the quantitative assessment of prostate cancer tumor cell heterogeneity. Following standard histologic diagnosis of prostate cancer, we will clinically follow all patients to determine their ultimate course. The multiparametric data will be used to develop an index to correlate tumor aggressiveness and patient prognosis. (1)