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6064 Background: Certain histopathologic features may be risk factors for recurrent disease after resection of papillary thyroid cancer (PTC). Age is also an important prognostic factor, with highe...

291 Background: Triple negative breast cancer is often chemoresistant creating the need for novel improved drugs for this disease. Naturally derived withanolides such as withaferin A (WA) have shown potent preclinical efficacy in breast cancers. The purpose of this study is to better define the anticancer activity of WA and novel analogs(X001 and X003) containing a 19-OH substitution, in triple negative MDA-MB-231cells where BRCA1 is important for cell survival. Methods: Cells were treated with increasing doses of WA, X001, or X003 for 24 h. MTS and trypan blue exclusion assays were used to examine cell proliferation and viability. Cell cycle was examined using propidium iodide (PI) staining on flow cytometry (FC). Annexin-V/PI co-staining on FC was used to detect apoptosis. Western analysis was used to evaluate BRCA1 protein levels as well as to confirm apoptosis by PARP cleavage and caspase-3 activation. Results: X003, WA, and X001 inhibited MDA-MB-231 cell viability (IC50=125, 500 and 1300nM respectively). IC50 level of WA induced G2/M phase cell cycle arrest while 2.5μM also induced apoptosis at 24 h as well as a significant decrease in both total and phospho-Akt protein levels. In addition, WA decreased BRCA1, heat shock factor 1, and mutant p53 protein levels in a dose-dependent manner. 2.5 μM of WA decreased BRCA1 protein levels by 75% compared to controls at 24 h (p50) and 6.5μM (5xIC50) and a G2/M cell cycle arrest at only 50nM (50%IC50) and 2μM (1.5xIC50) respectively after 24 hours. Conclusions: Natural withanolides such as WA and its potent analog X003 demonstrate anticancer effects on triple negative breast cancer cells through cell cycle arrest, apoptosis and down-regulation of BRCA1 protein expression. Targeting BRCA1 in triple negative breast cancer with natural withanolides represents a novel approach for future translational studies in this difficult disease.

279 Background: Systemic doxorubicin (DOX) and platinum agents in breast cancer can be limited by their toxicity and have low penetration into lymphatics contributing to locoregional (LR) recurrence. Subcutaneous (s.c.) delivery of single agent nanoconjugated (HA-DOX) or cisplatin (HA-CIS) can significantly boost LR drug delivery with lower toxicity in vivo. The purpose of this study is to evaluate histologic response and toxicity of combination s.c. delivery of HA-DOX/HA-CIS as a significant improvement over IV delivery with standard drugs reduces dose levels and expands the therapeutic dosing range for these drugs. Methods: 1.5x106 MDA-MB-468LN human breast cancer cells (metastatic line) were injected into the mammary fat pad of female Nu/Nu mice. When tumors reached 6mm, mice (n=8 per group) were then treated with standard DOX/CIS IV combination therapy or s.c. injections of HA-DOX/HA-CIS at 25%,50%,75%,100%,125%, and 150% MTD for these drugs. Drugs were dosed weekly x 3 wks and tissues were collected for histology at 4 and 12wk timepoints. Results: Only the nanoconjugate groups demonstrated a complete clinical response (CR) to therapy and a complete pathologic response (CPR) histologically [CR =62.5% of mice and CPR=12.5% at 50% MTD;CR=87.5% and CPR=25% at 75%MTD; p Conclusions: Subcutaneous nanocarrier-delivery of combination DOX/CIS demonstrated significantly improved tumor response with CPR and lower toxicity on histology compared to standard drugs. This delivery platform successfully treats LR and distant metastatic disease at lower doses and improves the dosing range of these drugs compared to standard agents.

5582 Background: 17-allylaminogeldanamycin (17-AAG) is a benzoquinone ansamycin antibiotic that destabilizes heat shock protein 90 (HSP-90) complexes, including RET and multiple cancer-involved pro...

e17003 Background: Epithelial cancers, particularly lung cancer and head and neck squamous cell carcinoma (HNSCC), continue to pose formidable challenges in clinical practice. Novel chemotherapeutic agents have been developed, but even those have limited long-term benefits in the treatment of these tumors, illustrating the need to continue to improve systemic therapy for affected patients. The objective of the present study was to investigate the effect of withaferin A (WA), a plant-derived small molecule, on cancer cell growth, heat shock protein expression and induction of apoptosis in human HNSCCs. Methods: MDA1986 and JMAR HNSCC cells were used in all experiments. The effect of WA on cell viability was determined by MTS assay. Apoptosis and mitochondrial membrane potential changes were assessed by annexin V/propidium iodide and JC-1 staining respectively using standard flow cytometry methods. Effect of WA on modulation of heat shock proteins was determined by Western blot analysis. Results: Withaferin A reduces cell viability in both MDA1986 and JMAR cells with IC50 levels of 265 ± 5 nM by MTS assay, which is 5 fold higher than its reported activity in breast cancer cells. WA completely down-regulates HSP90beta, GRP94, and TRAP-1 expression at 250 nM concentration in HNSCC cells at 24 hours treatment. In addition, WA markedly increased HSP70 levels and mildly increased HSP27 levels in a dose-dependent manner at 24 hours treatment. Flow cytometry with Annexin V/PI staining shows that 5 μM WA treatment for 24 hours induced apoptosis in 63% of MDA1986 and 60% of JMAR cells. WA at 5 μM also reduced mitochondrial membrane potential by JC-1 staining with flow cytometry to less than 10% of controls in both JMAR and MDA1986 cells at 24 hours treatment. Conclusions: Withaferin A is a potent novel inhibitor of HSP90 in human HNSCCs. In addition to HSP modulation, its anticancer mechanistic effects involve apoptotic cell death through the mitochondrial pathway. This molecule shows promise for further in vivo studies to establish preclinical proof of concept as a novel anticancer therapy in this disease. No significant financial relationships to disclose.