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A natural flavonoid lawsonaringenin induces cell cycle arrest and apoptosis in HT-29 colorectal cancer cells by targeting multiple signalling pathways

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Abstract

Colorectal cancer is the third most common malignancy in the world having a high mortality rate. Flavonoids possess many biological activities including anti-cancer activity. lawsonaringenin (LSG) is a flavonoid isolated from leaves of Lawsonia alba Lam. The objective of this study was to demonstrate the anti-cancer potential of LSG in colorectal cancer for the first time. The HT-29 cells were treated with LSG or 5-fluoruracil, as a positive control, to determine its effect on cell cytotoxicity by a MTT cell proliferation assay, and cell cycle progression and apoptosis using flowcytometry. We also determined the mechanisms underlying LSG-mediated growth inhibition of HT-29 cells by by investigating the expression of key oncogenes and apoptosis genes using q-RT PCR and immunocytochemical analysis. The cell cytotoxicity data showed that the IC50 value of LSG was significantly less than the IC50 value of 5-FU (50 µM). The anti-proliferative effect of LSG was mediated by arresting cells in the S phase of the cell cycle which then led to the induction of apoptosis the q-RT PCR and immunocytochemical analysis showed that LSG reduced the expression of ß-catenin (non-phosphorylated) and its downstream signalling target c-Myc, whereas it increased the phosphorylation of ß-catenin. Furthermore, LSG also downregulated the expression of oncogene K-Ras and anti-apoptotic proteins, Bcl-2, and Bcl-xL. In conclusion, our data demonstrates that LSG exerted its anti-tumor activity by arresting the cell cycle in S phase, and by downregulating the expression of oncogenes including ß-catenin, c-Myc, K-Ras and anti-apoptosis proteins Bcl-2 and Bcl-xL. This study suggests a potential use of natural flavonoid, lawsonaringenin, to attenuate colorectal cancer growth; however, further pre-clinical/clinical studies are required to establish its role as a therapeutic agent.

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Abbreviations

ß-Cat-NP:

Beta catenin (non-phosphorylated)

ß-Cat-P:

Beta catenin (phosphorylated)

5-FU:

5-Fluorouracil

Bcl-2:

B-cell lymphoma 2 protein

Bcl-Xl:

B-cell lymphoma extra-large protein

c-Myc:

Avian myelocytomatosis virus oncogene cellular homolog

EGCG:

Epigallocatechin-3gallate

EGFR:

Epidermal growth factor receptor

G2/M phase:

Gap2 mitosis phase

HT-29:

Human colonic adenocarcinoma cell line

K-Ras:

Kirsten rat sarcoma

PI3K:

Phosphatidylinositol 3-kinase

S phase:

Synthesis phase of the cell cycle

Wnt:

Wingless-type MMTV integration site family member

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Acknowledgements

M.I.C. acknowledges the enabling role of the Higher Education Commission, Islamabad, Pakistan, through a financial support under, “Studies on the chemoprevention of mammary carcinogenesis by dietary agents” (Project No. 20-3790). A.A. thanks Ms. Narjis Fatima and Dr. Waqar Hameed for the kind gift of Bcl-2 primer.

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Authors and Affiliations

  1. Dr. Panjwani Center for Molecular Medicine and Drug Research, International Center for Chemical and Biological Sciences, University of Karachi, Karachi, 75270, Pakistan

    Areeba Anwar & Muhammad Iqbal Choudhary

  2. H.E.J. Research Institute of Chemistry, International Center for Chemical and Biological Sciences, University of Karachi, Karachi, 75270, Pakistan

    Nizam Uddin, Bina Shaheen Siddiqui, Sabira Begum & Muhammad Iqbal Choudhary

  3. Nutrition Science and Food Chemistry Laboratory, Agriculture Research Station, Virginia State University, Petersburg, VA, 23806, USA

    Rafat Ali Siddiqui

  4. Department of Biochemistry, Faculty of Sciences, King Abdulaziz University, Jeddah, 21412, Saudi Arabia

    Muhammad Iqbal Choudhary

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  1. Areeba Anwar
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Anwar, A., Uddin, N., Siddiqui, B.S. et al. A natural flavonoid lawsonaringenin induces cell cycle arrest and apoptosis in HT-29 colorectal cancer cells by targeting multiple signalling pathways. Mol Biol Rep 45, 1339–1348 (2018). https://doi.org/10.1007/s11033-018-4294-5

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