Depletion of intrinsic expression of Interleukin-8 in prostate cancer cells causes cell cycle arrest, spontaneous apoptosis and increases the efficacy of chemotherapeutic drugs.
BACKGROUND
The progression of all cancers is characterized by increased-cell proliferation and decreased-apoptosis. The androgen-independent prostate cancer (AIPC) is the terminal stage of the disease. Many chemokines and cytokines are suspects to cause this increased tumor cell survival that ultimately leads to resistance to therapy and demise of the host. The AIPC cells, but not androgen-responsive cells, constitutively express abundant amount of the pro-inflammatory chemokine, Interleukin-8 (IL-8). The mechanism of IL-8 mediated survival and therapeutic resistance in AIPC cells is unclear at present. The purpose of this report is to show the pervasive role of IL-8 in malignant progression of androgen-independent prostate cancer (AIPC) and to provide a potential new therapeutic avenue, using RNA interference.


RESULTS
The functional consequence of IL-8 depletion in AIPC cells was investigated by RNA interference in two IL-8 secreting AIPC cell lines, PC-3 and DU145. The non-IL-8 secreting LNCaP and LAPC-4 cells served as controls. Cells were transfected with RISC-free siRNA (control) or validated-pool of IL-8 siRNA. Transfection with 50 nM IL-8 siRNA caused >95% depletion of IL-8 mRNA and >92% decrease in IL-8 protein. This reduction in IL-8 led to cell cycle arrest at G1/S boundary and decreases in cell cycle-regulated proteins: Cyclin D1 and Cyclin B1 (both decreased >50%) and inhibition of ERK1/2 activity by >50%. Further, the spontaneous apoptosis was increased by >43% in IL-8 depleted cells, evidenced by increases in caspase-9 activation and cleaved-PARP. IL-8 depletion caused significant decreases in anti-apoptotic proteins, BCL-2, BCL-xL due to decrease in both mRNA and post-translational stability, and increased levels of pro-apoptotic BAX and BAD proteins. More significantly, depletion of intracellular IL-8 increased the cytotoxic activity of multiple chemotherapeutic drugs. Specifically, the cytotoxicity of Docetaxel, Staurosporine and Rapamycin increased significantly (>40% at IC50 dose) in IL-8 depleted cells as compared to that in C-siRNA transfected cells.


CONCLUSIONS
These results show the pervasive role of IL-8 in promoting tumor cell survival, and resistance to cytotoxic drugs, regardless of the cytotoxic mechanism of antiproliferative drugs, and point to potential therapeutic significance of IL-8 depletion in men with AIPC.
Singh
Rajendra K
RK
Department of Urology and Sylvester Comprehensive Cancer Center, Miller School of Medicine, University of Miami, Miami, Florida, USA. Singhrkbio@rediffmail.com
Lokeshwar
Bal L
BL
D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects*; D048708:Cell Growth Processes / Q000187:drug effects; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002633:Chemotaxis / Q000187:drug effects; D020868:Gene Silencing; D006801:Humans; D016209:Interleukin-8 / Q000096:biosynthesis / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism; D008297:Male; D009361:Neoplasm Invasiveness; D009362:Neoplasm Metastasis; D011471:Prostatic Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D034741:RNA, Small Interfering / Q000378:metabolism / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism
D000970:Antineoplastic Agents; D051017:Apoptosis Regulatory Proteins; D016209:Interleukin-8; D034741:RNA, Small Interfering; D011994:Recombinant Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A