Anticancer activity of guggulsterone in head and neck squamous cell carcinoma

B139 Head and neck squamous cell carcinoma (HNSCC) is a one of the most commonly occurring neoplasms worldwide and is associated with a high degree of morbidity and mortality. The severity of this disease largely stems from the very high rates of second primary tumor (SPT) development. HNSCC is thought to be caused by the effects of carcinogens, particularly those in tobacco and alcohol. For these reasons, chemoprevention of HNSCC, aimed at inhibiting the steps leading to cellular transformation and carcinogenesis, represents an appropriate clinical goal. This study of a potential chemopreventive therapy for HNSCC focuses on two molecular targets, nuclear factor kappa B (NFκB) and signal transducer and activator of transcription (STAT)-3. Both of these targets are oncogenic transcription factors known to mediate growth and proliferation of HNSCC and to regulate the expression of target genes that control the cell cycle, apoptosis, tumor invasion and tumor angiogenesis. Guggulsterone, a naturally occurring compound used in traditional Indian medicine, has been shown to have anticancer activity and to inhibit NFκB activity in various cancer cell types, including HNSCC. In the current study of guggulsterone’s anticancer activity, treatment of HNSCC cell lines, 1483 and UM-22b, with a racemic mixture of guggulsterone isomers resulted in a dose-dependent decrease in protein levels of both total and phosphotyrosine STAT3 in vitro , as shown through western blotting. Immunohistochemical staining of xenograft tumors from mice treated with guggulsterone also revealed a decrease in total STAT3 of 87.3% in tumors derived from UM-22b cells and of 44.3% in tumors derived from 1483 cells, compared to a vehicle control. Realtime RT-PCR of mRNA from HNSCC cells treated with guggulsterone suggests, however, that this decrease in STAT3 does not stem from alterations in STAT3 gene transcription. Additionally, as shown through trypan blue dye exclusion assays, guggulsterone treatment decreased numbers of viable HNSCC cells, with EC50’s ranging from ~2-8 µM at 72 hours, depending on cell line and formulation of guggulsterone. Percentages of cell populations in the G1 phase of the cell cycle increased after 24 hours of guggulsterone treatment at 10 and 20 µM by 9.5% and 33.2%, respectively, in UM-22b cells, and by 13.3% and 53.6%, respectively, in 1483 cells, as demonstrated through flow cytometric analysis after propidium iodide staining of HNSCC cells. Guggulsterone treatment of HNSCC cells also resulted in an increase in apoptosis, demonstrated by a 3.6-fold increase in levels of histone-associated DNA fragments present in lysates of UM-22b cells and a decrease in levels of full length caspase 3 in both cells lines. These demonstrations of the anticancer activities of guggulsterone are encouraging in evaluating this compound as a potential chemopreventive agent in HNSCC. Future plans include in vivo studies which will employ a carcinogen-induced model of HNSCC and assess guggulsterone’s chemopreventive potential. Future in vitro studies will focus on investigation of the molecular mechanisms of STAT3 decrease and growth inhibition upon guggulsterone treatment of HNSCC cell lines and will hopefully complement in vivo studies in providing a firm basis for a potential clinical trial of guggulsterone as a chemopreventive therapy for HNSCC.