Towards TRAIL to silencing of SMURF and NEDD4: FLIP is flopped.

Farooqi, Ammad Ahmad; Fayyaz, Sundas; Bhatti, Shahzad; Ismail, Muhammad; Mansoor, Qaisar

Objective: Prostate cancer is a multifactorial anomaly that arises and exacerbates because of miscellaneous key mediators instrumental in disease aggressiveness. Heterogeneity of the disease offers stumbling blocks in standardization of therapeutic interventions. TRAIL and cFLIP are two diametrically opposed key mediators that antagonize each others function. TRAIL has central role in killing neoplastic cells and sparing innocent cells. However this ligand also undergoes a downregulation in prostate carcinogenesis. In this particular study we hypothesized whether abrogation of negative regulators of TGF signaling potentiates and abolishes the expression of TRAIL and cFLIP respectively or not. Methods: In this particular study we have used androgen sensitive prostate cancer cell line (LNCaP) and treated it with TGF and etoposide. RNA interference technique was used to unfold the correlation betweeen TGF signaling and TRAIL and cFLIP expression in the LNCaP cell line. Enforced expression of SMURF and NEDD4 was done by transfecting cell line with respective constructs. The results were analyzed by RT-PCR and western blot. Results: We have treated the TGF and etoposide treated cell line with siRNA of NEDD4 and SMURF. There was a successful blockade of both genes at transcriptional level as evidenced by RT-PCR study. Simultaneously there was remarkable upregulation in the expression of TRAIL. Another interesting observation was that TGF treatment triggered expression of TRAIL and ablated cFLIP and this activity was pronounced after abrogation of negative regulators of TGF signal transduction. Further investigation into the molecular mechanism was done by reconstitution of SMURF and NEDD4 depleted LNCaP cells which resulted in an enhanced expression of cFLIP. Conclusion: In this study, we show that TRAIL expression is upregulated and cFLIP is downregulated upon exposure of LNCaP cell lines to TGF-β and etoposide and that TRAIL is a major contributor to apoptosis mediated by TGF-β. Consistent with the interpretations, it is obvious that NEDD4 and SMURF are the major proteins involved in the deviation of core biological systems. Combinatorial blockade of these genes by rational drug design, or alternatively, drugging negative regulators of TGF pathway might offer exciting avenues in key aspects of the science of therapeutic individualization.

PROSTATE cancer; DISEASE exacerbation; STANDARDIZATION; HETEROGENEITY; CANCER cells

Journal of Experimental & Integrative Medicine, 2011, Vol 1, Issue 2, p111

1309-4572

Academic Journal

10.5455/jeim.200311.or.004