EFFECTS OF INTERMITTENT ANDROGEN SUPPRESSION ON THE STEM-CELL COMPOSITION AND THE EXPRESSION OF THE TRPM-2 (CLUSTERIN) GENE IN THE SHIONOGICARCINOMA

The proportion of tumorigenic stem cells and the expression of the apo ptosis-related gene, TRPM-2 (clusterin), were studied in populations o f Shionogi carcinoma cells subjected to multiple cycles of androgen wi thdrawal and replacement (intermittent androgen suppression). The pare nt androgen-dependent cell line was initially transplanted into a male mouse which was castrated when the estimated weight of the resultant tumour became approximately 3 g. After the tumour had regressed to 40% or less of the original weight, it was transplanted into the next non -castrated male. This was repeated for four cycles of transplantation and castration-induced apoptosis before the tumour progressed to an an drogen-independent state. The proportion of total stem cells in the tu mour, as determined by in vivo limiting dilution assays in male mice, was constant during the first three cycles but increased 15-fold betwe en the third and fourth cycles. In the parent androgen-dependent tumou r before androgen ablation, the androgen-independent stem cell populat ion formed 0.8% of the total stem cell compartment. After the fourth c ycle this population increased to 47%; a population of similar size (3 3%, P=0.8) was found in the androgen-independent recurrent form of the tumour induced by one-time castration. Whether androgen withdrawal th erapy was intermittent or continuous, conversion to androgen independe nce thus occurred when one-third to one-half of the total stem cell co mpartment was populated by androgen-independent stem cells. The androg en-repressed TRPM-2 (clusterin) gene was actively expressed in regress ing tumours after androgen ablation, and also became constitutively ex pressed in non-regressing tumours after the first and subsequent cycle s of androgen withdrawal. Staining of cytoplasm and nuclei with anti-c lusterin antibody was observed in androgen-dependent tumour cells afte r each cycle of intermittent androgen suppression; the nuclear stainin g was more intense in recurrent androgen-independent cells. The anomal ous nuclear localization of clusterin, an anti-cytolytic TRPM-2 encode d protein, may serve to inhibit early events in the apoptotic process and thereby foster the generation and outgrowth of androgen-independen t stem cells in an androgen-depleted environment. Copyright (C) 1996 P ublished by Elsevier Science Ltd.