Potential Of Withaferin A Research Articles

Computational Insights into the Potential of Withaferin-A, Withanone and Caffeic Acid Phenethyl Ester for Treatment of Aberrant-EGFR Driven Lung Cancers.

The anticancer activities of Withaferin-A (Wi-A) and Withanone (Wi-N) from Ashwagandha and Caffeic Acid Phenethyl Ester (CAPE) from honeybee propolis have been well documented. Here, we examined the binding potential of these natural compounds to inhibit the constitutive phosphorylation of epidermal growth factor receptors (EGFRs). Exon 20 insertion mutants of EGFR, which show resistance to various FDA approved drugs and are linked to poor prognosis of lung cancer patients, were the primary focus of this study. Apart from exon 20 insertion mutants, the potential of natural compounds to serve as ATP competitive inhibitors of wildtype protein and other common mutants of EGFR, namely L858R and exon19del, were also examined. The potential of natural compounds was compared to the positive controls such as erlotinib, TAS6417 and poziotinib. Similar to known inhibitors, Wi-A and Wi-N could displace and binds at the ATP orthosteric site of exon19del, L858R and exon20, while CAPE was limited to wildtype EGFR and exon 20 insertion mutants only. Moreover, the binding free energy of the natural drugs against EGFRs was also comparable to the positive controls. This computational study suggests that Wi-A and Wi-N have potential against multiple mutated EGFRs, warranting further in vitro and in vivo experiments.

Potential of Withaferin-A, Withanone and Caffeic Acid Phenethyl ester as ATP-competitive inhibitors of BRAF: A bioinformatics study.

Serine/threonine-protein kinase B-raf (BRAF) plays a significant role in regulating cell division and proliferation through MAPK/ERK pathway. The constitutive expression of wild-type BRAF (BRAFWT) and its mutant forms, especially V600E (BRAFV600E), has been linked to multiple cancers. Various synthetic drugs have been approved and are in clinical trials, but most of them are reported to become ineffective within a short duration. Therefore, combinational therapy involving multiple drugs are often recruited for cancer treatment. However, they lead to toxicity and adverse side effects. In this computational study, we have investigated three natural compounds, namely Withaferin-A (Wi-A), Withanone (Wi-N) and Caffeic Acid Phenethyl ester (CAPE) for anti-BRAFWT and anti-BRAFV600E activity. We found that these compounds could bind stably at ATP-binding site in both BRAFWT and BRAFV600E proteins. In-depth analysis revealed that these compounds maintained the active conformation of wild-type BRAF protein by inducing αC-helix-In, DFG-In, extended activation segment and well-aligned R-spine residues similar to already known drugs Vemurafenib (VEM), BGB283 and Ponatinib. In terms of binding energy, among the natural compounds, CAPE showed better affinity towards both wild-type and V600E mutant proteins than the other two compounds. These data suggested that CAPE, Wi-A and Wi-N have potential to block constitutive autophosphorylation of BRAF and hence warrant in vitro and in vivo experimental validation.

Abstract 3678: Enhanced activity of chemotherapeutic drugs by blueberry anthocyanidins and withaferin A against human lung cancer cells.

Abstract Lung cancer is the leading cause of all cancer deaths in the U.S., Europe and the World. With only an expected 16% relative 5-year survival rate in the U.S., treatment of this aggressive and progressive disease continues to remain a fundamental problem. Moreover, resistance to chemotherapy is a major problem in cancer treatment. In an attempt to overcome drug resistance and improve treatment efficacy, new molecular targeted drugs are being developed and tested in clinical studies. Over the recent years various epidemiological studies have provided compelling evidence for the chemopreventive/therapeutic efficacies of various functional foods and natural products. We hypothesize that non-toxic natural compounds such as a native mixture of blueberry anthocyanidins (BB anthos) and withaferin A (WFA), alone and in combination with a suboptimal dose of the chemo drug paclitaxel (PAC) will have therapeutic response and also chemosensitize the lung cancer cells. We evaluated whether BB anthos could enhance the sensitivity of H1299 cells to PAC and reduce its effective dose to inhibit cell proliferation. Data show significantly enhanced cell death by PAC at a suboptimal concentration (1 nM) (IC50 ≈ 40 nM) when combined with 5 μM (25% vs. 5% inhibition) and 10 μM (49% vs. 5% inhibition) BB anthos (1:1 mixture of Dp, Cy, Mv, Pe and Pt). The significant inhibition of H1299 lung cancer xenografts in nude mice was also evident from the same combination, while no significant growth inhibition was observed with the individual entities. Since BB is a known antioxidant, we determined if BB anthos would attenuate oxidative stress-induced by PAC in H1299 lung cancer cells by measuring intracellular ROS production. Our data showed that addition of BB anthos (10 μM each) did not affect the ROS levels produced by PAC (10 nM), as determined by cell permeable fluorescence probe DCFH-DA. Similarly, we also determined the antiproliferative potential of WFA alone and in combination with chemo drugs, PAC and docetaxel (DOC) against non-small-carcinoma lung cancer cells. WFA demonstrated high antiproliferative activity against the growth of both H1299 and A549 cells as determined by MTT assay. When suboptimal concentrations of WFA were tested in combination with suboptimal concentration of DOC and PAC, significantly enhanced antiproliferative activities were observed against H1299 and A549 compared with individual entities. The combination effect was found to be synergistic as analyzed using the CalcuSyn software. Together, our results suggest that both BB anthos and WFA enhanced therapeutic response of chemo drugs against lung cancer and reduce their required dose thus minimizing their toxicity. (Supported from Duggan endowment fund.) Citation Format: Farrukh Aqil, Radha Munagala, Hina Kausar, Jeyaprakash Jeyabalan, Ramesh Gupta. Enhanced activity of chemotherapeutic drugs by blueberry anthocyanidins and withaferin A against human lung cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3678. doi:10.1158/1538-7445.AM2013-3678

Abstract 1741: Mitotic catastrophe and inhibition of growth of Castration Resistant Prostate Cancers by a dietary agent Withaferin-A.

Abstract Cell cycle deregulation has been strongly associated with the pathogenesis of prostate cancer. It has been well documented that an increased expression of cell cycle regulatory proteins (cyclin D1-25%, cyclin A-35% and cyclin B-75%) have been reported in castration resistant prostate cancer (CRPC) cells, when compared to normal prostate tissue. There are several cell cycle regulators undergoing clinical trials targeting either G0/G1 or G2/M phase of cell cycle to inhibit the growth of many cancers including prostate cancer. We previously published that Withaferin-A (WA); a natural compound effectively suppresses CRPC growth both in vitro and in vivo. Hence, in this study we intend to determine the cell-cycle regulatory potential of this agent on CRPC cells. WA caused irreversible G2/M arrest in both CRPC cell lines (PC-3 and C4-2B) till 72 hours . Phosphorylation of cdc2 by Wee1 and Myt1 is an important step to maintain an inactive state of cdc2 that lead to G2/M cell cycle arrest. Additionally decreased levels of phosphorylated Wee-1 and Myt1 followed by increase in the phosphorylation status of cdc2 (Tyr15 and Thr14) supported the G2/M phase of cell cycle arrest in both CRPC cell lines. Further checkpoint kinase assay showed increased activity in WA treated CRPC cells suggesting activation of CDC25C that activates cyclinB/cdc2 complex .We also observed down regulation of several cyclin family proteins in both CRPC cells. Altogether, these results conclude d that WA inhibits CRPC cell growth by induction of G2/M cell cycle arrest, highlighting the potential of WA could be a therapeutic agent for CRPC. Currently, we are investigating the effect of WA on cell cycle markers using both xenograft and TRAMP models. The outcome of our study may enable bringing WA to the mainstream of medicine for prevention and/or therapy for CRPC. Citation Format: Aditi Shirish Vadodkar, Suman Suman, Ram Vinod Roy, Chendil Damodaran. Mitotic catastrophe and inhibition of growth of Castration Resistant Prostate Cancers by a dietary agent Withaferin-A. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1741. doi:10.1158/1538-7445.AM2013-1741

Production dynamics of Withaferin A in Withania somnifera (L.) Dunal complex

Withaferin A (WA) is an important withanolide holding promise in cancer treatment and as a relatively safe radiosensitive/chemotherapeutic agent, which is present in traces in all parts of Withania somnifera except the leaves, where as it is reported to be present in only two non-Indian chemotypes (South African chemotype/Israel chemotype 1). The present studies have marked its presence in all Indian populations (wild/cultivated), as well as two identified Indian chemotypes (AGB002 and AGB025). The quantitative dynamics of WA production in Indian populations and interchemotypic hybrids developed at our institute have been studied, and the results were compared with five previously reported chemotypes from Israel, South Africa and India. An analysis on inheritance characteristics based on presence/absence of WA in hybrid plants and their respective parents is given for future studies on the chemogenetics of this complex species in greater detail. Further, the production potential of WA in vitro propagated plants of elite varieties developed at our institute is discussed, in view of maintaining chemotypic fidelity and stability from a production point of view. Also, evidence-based clues suggesting the leaves as the site of the synthesis of WA is provided.