Androgen-dependent apoptosis in male germ cells is regulated through the proto-oncoprotein Cbl

The proto-oncoprotein Cbl is known to control several signaling processes. It is highly expressed in the testis, and because spermatogenesis is androgen dependent, we investigated the androgen dependency expression of Cbl through its testicular sublocalization and its expression levels in rats that were exposed to the antiandrogen flutamide or were hypophysectomized. We report the androgen dependency of Cbl as it localizes in pachytene spermatocytes during androgen-dependent stages, is down-regulated upon flutamide exposure, and is up-regulated with testosterone in hypophysectomized rats. Coculture experiments showed the key control exerted by the Sertoli cell on Cbl activity. As flutamide induces germ cell apoptosis, we investigate members of the Bcl-2 family upon flutamide exposure. We show that the proapoptotic Bcl-2 family member Bim mirrored Cbl expression through a posttranscriptional process. We also show that in Cbl knockout mouse testes, the imbalance between the high expression of Bim and Smac/Diablo and antiapoptotic factors such as cellular inhibitor of apoptosis 2 favors a survival process, which makes these mice unresponsive to androgen withdrawal and could explain their hypofertility

A protective role against oxidative stress for the E3-ubiquitin ligase c-Cbl : usefulness as a prognostic marker for carcinomas

Le travail présenté a porté sur l’analyse in vivo du proto-oncogène c-cbl, dont la forme connue est c-Cbl (p120cbl). Il s’agit d’une E3-Ubiquitine ligase et un poly-adaptateur moléculaire. Nous avons montré l’androgéno-dépendance de l’expression de c-Cbl dans les cellules germinales testiculaires et les cellules épithéliales de la prostate de rats et de souris. Nous avons montré la régulation anti-apoptotique exercée in vivo par la c-Cbl dans la prostate par comparaison des souris c-cbl invalidées ou non pour c-cbl (KO ou WT). L’effet exercé par c-cbl dans le testicule est pro-apoptotique (J.Cell Biol, 2005), que nous avons ultérieurement attribué à une nouvelle isoforme testiculaire de c-Cbl (Δ-c-Cbl). La comparaison des MEF KO et WT après induction d’apoptose par l’étoposide, a conforté l’effet anti-apoptotique exercée in vivo par c-Cbl dans la prostate. Elle a aussi montré la forte apoptose des MEF KO au peroxyde d’hydrogène : c-Cbl peut être considérée comme un protecteur du stress oxydant. L’intensité du stress oxydant associé aux cancers et leur forte résistance à l’apoptose sont des propriétés qui pourraient être reliées à c-Cbl. L’analyse in situ effectuée à partir de tumeurs congelées et de Tissue Microarrays (TMA) a montré une expression élevée de c-Cbl dans certains cancers, dont l’intensité pourrait correspondre à la gravité de l’atteinte anatomo-pathologique. La protéine c-Cbl est apparue être un marqueur d’agressivité du cancer de la prostate, probablement de l’ovaire, de l’utérus, du cerveau, du poumon, du colon et du rectum. Nous la considérons aussi comme une cible thérapeutique car, protecteur du stress oxydant, elle prendrait part à la résistance à l’apoptose des cellules tumorales. Un brevet a été déposé (2009, co-inventeurs : S.Yakoub et al). Un article rapportant ces résultats est en cours de soumission (S. Yakoub et al)This work has focused on the in vivo analysis of the proto-oncogene c-cbl, coding for c-Cbl (p120cbl). We demonstrated the androgen-dependency of c-Cbl in the testicular germ cells and the prostatic epithelial cells of rats and mice. We then identified the anti-apoptotic regulation exerted by p120cbl in the prostate, comparing mouse c-cbl KO and WT, unlike this exerted in the testis (J.Cell Biol, 2005). We reported this difference to the high expression in testis of a new c-Cbl isoform, Δc-Cbl. The comparison of MEF KO and WT allowed confirming the anti-apoptotic regulation to etoposide exerted by c-Cbl. A very high apoptotic effect was observed in MEF KO with H2O2: c-Cbl is a strong stress oxidative protector. Knowing the intensity of oxidative stress in several cancers and their particular resistance to apoptosis as well, the in situ analysis of these malignancies was made from frozen tumours and tissue microassays (TMA). c-Cbl was indeed highly expressed and its intensity appears to reflect the aggressiveness of the pathology. c-Cbl could then be considered as a marker of severity of prostate cancer but probably also ovary, uterus, brain, lung, colon and rectum. It can also be considered as a therapeutic target involved in resistance to apoptosis as a stress oxidative protector. A patent was filed in the United States (2009, co-inventors: S. Yakoub et al

Down-regulation of DcR2 sensitizes androgen-dependent prostate cancer LNCaP cells to TRAIL-induced apoptosis.

International audienceUNLABELLED: ABSTRACT: BACKGROUND: Dysregulation of many apoptotic related genes and androgens are critical in the development, progression, and treatment of prostate cancer. The differential sensitivity of tumour cells to TRAIL-induced apoptosis can be mediated by the modulation of surface TRAIL receptor expression related to androgen concentration. Our previous results led to the hypothesis that downregulation of TRAIL-decoy receptor DcR2 expression following androgen deprivation would leave hormone sensitive normal prostate cells vulnerable to the cell death signal generated by TRAIL via its pro-apoptotic receptors. We tested this hypothesis under pathological conditions by exploring the regulation of TRAIL-induced apoptosis related to their death and decoy receptor expression, as also to hormonal concentrations in androgen-sensitive human prostate cancer, LNCaP, cells. RESULTS: In contrast to androgen-insensitive PC3 cells, decoy (DcR2) and death (DR5) receptor protein expression was correlated with hormone concentrations and TRAIL-induced apoptosis in LNCaP cells. Silencing of androgen-sensitive DcR2 protein expression by siRNA led to a significant increase in TRAIL-mediated apoptosis related to androgen concentration in LNCaP cells. CONCLUSIONS: The data support the hypothesis that hormone modulation of DcR2 expression regulates TRAIL-induced apoptosis in LNCaP cells, giving insight into cell death induction in apoptosis-resistant hormone-sensitive tumour cells from prostate cancer. TRAIL action and DcR2 expression modulation are potentially of clinical value in advanced tumour treatment