Shweta Kaushik1, Hari Shyam2, Satish Agarwal3, Ramesh Sharma2, Tapas C Nag4, Anil K Dwivedi3, Anil K Balapure5. 1. Division of Biochemistry, CSIR-Central Drug Research Institute, Lucknow 226031, India; Academy of Scientific and Innovative Research, Taramani, Chennai 600113, India. 2. Division of Biochemistry, CSIR-Central Drug Research Institute, Lucknow 226031, India. 3. Division of Pharmaceutics, & Pharmacokinetics, CSIR-Central Drug Research Institute, Lucknow 226031, India. 4. Department of Anatomy, All India Institute of Medical Sciences, New Delhi 110029, India. 5. Division of Biochemistry, CSIR-Central Drug Research Institute, Lucknow 226031, India; Academy of Scientific and Innovative Research, Taramani, Chennai 600113, India. Electronic address: anilbalapure58@gmail.com.
Abstract
AIMS: Recently, strategies of cancer treatment using combination of agents with distinct molecular mechanism(s) of action are considered more promising due to its high efficacy and reduced systemic toxicity. The study is aimed to improve the efficacy of selective estrogen receptor modulator, Centchroman (CC) by combination with the phytoestrogen Genistein (GN). METHODS: Cytotoxicity was evaluated by Sulforhodamine B assay. Cell cycle analysis was done through flow cytometry. Further, Apoptosis was analyzed using Annexin V/PI staining, tunel assay and electron microscopic examination and verified using western blot analysis. In order to validate the in vitro results, in vivo analysis was performed using 4T1-syngeneic mouse model. KEY FINDINGS: In this study, we report that the dietary isoflavone genistein (GN) synergistically improved antineoplasticity of CC in breast cancer by arresting cells at G2/M phase culminating in ROS dependent apoptosis. The combination of CC plus GN caused dysregulation of Bax and Bcl-2 ratio inducing mitochondrial dysfunction, activation of Caspase-3/7, -9 and PARP cleavage. Further, combination significantly suppresses phosphorylation of PI3K/Akt/NF-κB, enhancing apoptosis. Additionally, combination markedly reduced tumor growth compared to CC and GN alone in mouse 4T1 breast tumor model. SIGNIFICANCE: Together, these studies suggest that GN represents a potential adjunct molecule whose role in CC induced apoptosis deserves attention.
AIMS: Recently, strategies of cancer treatment using combination of agents with distinct molecular mechanism(s) of action are considered more promising due to its high efficacy and reduced systemic toxicity. The study is aimed to improve the efficacy of selective estrogen receptor modulator, Centchroman (CC) by combination with the phytoestrogen Genistein (GN). METHODS:Cytotoxicity was evaluated by Sulforhodamine B assay. Cell cycle analysis was done through flow cytometry. Further, Apoptosis was analyzed using Annexin V/PI staining, tunel assay and electron microscopic examination and verified using western blot analysis. In order to validate the in vitro results, in vivo analysis was performed using 4T1-syngeneic mouse model. KEY FINDINGS: In this study, we report that the dietary isoflavone genistein (GN) synergistically improved antineoplasticity of CC in breast cancer by arresting cells at G2/M phase culminating in ROS dependent apoptosis. The combination of CC plus GN caused dysregulation of Bax and Bcl-2 ratio inducing mitochondrial dysfunction, activation of Caspase-3/7, -9 and PARP cleavage. Further, combination significantly suppresses phosphorylation of PI3K/Akt/NF-κB, enhancing apoptosis. Additionally, combination markedly reduced tumor growth compared to CC and GN alone in mouse4T1breast tumor model. SIGNIFICANCE: Together, these studies suggest that GN represents a potential adjunct molecule whose role in CC induced apoptosis deserves attention.