Chemopreventive potential of hydroethanolic Murraya koenigii leaves extract against DMBA induced breast carcinogenesis: In-silico and in-vivo study

Abstract

Ethnopharmacological relevance: Murraya koenigii (MK), a member of the Rutaceae family and widely known as the curry-leaf tree, is indigenous to India, Sri Lanka, and other south Asian nations. It is a renowned medicinal herb because of the wide range of bioactive components found in its leaves, such as girinimbine, koenimbine, mahanimbine and mahanine among others. All these bioactive components make this plant beneficial for treating a variety of ailments and diseases. Biological and pharmacological activities of MK include anti-oxidant, anti-microbial, anti-ulcer, anti-helminthic, anti-malarial, anti-trichomonal, hepatoprotective, anti-diabetic, etc. AIM OF THE STUDY: The present study aimed to evaluate the possible protective effect of hydroethanolic Murraya koenigii leaves extract (HEMKLE) against 7,12-Dimethylbenz[a]anthracene (DMBA)-induced breast cancer in rats, which further paves the way for future breast cancer treatment.

Materials and methods: For the preparation of hydroethanolic Murraya koenigii leaves extract (HEMKLE), Murraya koenigii (MK) leaves were taken from the botanical garden of the Panjab University campus, Chandigarh, and authenticated from the Department of Botany, Panjab University (accession number 22417). The phytochemical characterization of HEMKLE was performed using liquid chromatography-mass spectrometry (LC-MS). Following this, an in-silico molecular docking analysis was performed using Maestro Schrodinger software, and an in-vivo study was conducted. For the in-vivo study, female SD rats were divided into four different groups. Group I (C), Group II (DMBA), Group III (HEMKLE), and Group IV (HEMKLE + DMBA). Histopathogy, oxidative and antioxidant status, immunohistochemistry of estrogen receptor-α, TUNEL assays, mRNA and protein expression of apoptotic pathway genes were conducted in in-vivo study.

Results: In LC-MS, major phytochemical constituents including flavonoids and carbazole alkaloids were identified. In-silico docking study revealed the strong binding affinity between the identified compounds with caspase-3. Additionally, koenine displayed the highest binding affinity/minimum energy of -9.21 kcal/mol with 6BDV as compared to other phytochemicals. Furthermore, in-vivo experimentation revealed that HEMKLE administration in Group IV(HEMKLE + DMBA) significantly inhibits the tumor incidence and volume as compared to alone DMBA treated group. The antioxidant action of HEMKLE was proven from the in-vivo analysis of antioxidant marker enzymes, histopathology, immunohistochemistry of ER-α studies. Further, increase number of TUNEL positive cells was observed in co-treated animals as compared to alone DMBA treated animals. In Group IV (HEMKLE + DMBA), upregulated expression of pro-apoptotic genes and downregulated expression of anti-apoptotic gene were observed when compared to Group II(DMBA) suggested the apoptotic effect of HEMKLE.

Conclusion: The results of the present study provide clear evidence of the chemopreventive capabilities of HEMKLE in rats with DMBA-induced breast cancer. The observed outcomes could potentially be attributed to the existence of diverse phytochemicals within the HEMKLE.

Keywords: Apoptosis; Breast cancer; Estrogen receptor-α; In-silico study; LC-MS; Murraya koenigii; Oxidative stress; TUNEL assay.