Carbazole alkaloids from Murraya koenigii trigger apoptosis and autophagic flux inhibition in human oral squamous cell carcinoma cells

Abstract

Carbazole alkaloids, a major constituent of Murraya koenigii (L.) Sprengel (Rutaceae), exhibit biological effects such as anticancer activity via the induction of apoptosis, and they represent candidate chemotherapeutic agents. Oral squamous cell carcinoma (OSCC) is the most prevalent cancer of the oral cavity and a growing and serious health problem worldwide. In this study, we investigated the anticancer properties and mechanisms of action of two carbazole alkaloids derived from M. koenigii leaves, mahanine and isomahanine, in the OSCC cell line CLS-354. At 15 μM, mahanine and isomahanine were cytotoxic to CLS-354 cells, triggering apoptosis via caspase-dependent and -independent mechanisms. Autophagosomes, visualised using monodansylcadaverine (MDC) labelling, were numerous in carbazole alkaloid-treated cells. Mahanine and isomahanine markedly induced the expression of the autophagosome marker microtubule-associated protein 1 light chain 3, type II (LC3B-II). Genetic and chemical inhibition of autophagy via silencing of the Autophagy protein 5 gene and exposure to bafilomycin A1 (BafA1), respectively, did not arrest carbazole alkaloid-induced apoptosis, indicating that it occurs independently of autophagic activation. Surprisingly, both carbazole alkaloids caused increased accumulation of p62/sequestosome1 (p62/SQSTM1), with coordinated expression of LC3B-II and cleaved caspase-3, suggesting inhibition of autophagic flux. Our results suggest that inhibition of autophagic flux is associated with carbazole alkaloid-induced apoptosis. Our findings provide evidence of a novel cytotoxic action of natural carbazole alkaloids and support their use as candidate chemotherapeutic agents for the treatment of OSCC.

Dysregulated Expression of MITF in Subsets of Hepatocellular Carcinoma and Cholangiocarcinoma

“…4A). Mahanine was shown to induce cell death in human cancer cells through inhibiting the autophagic degradation activity, a mechanism different from cisplatin (Utaipan et al 2017). Treatment for 24 h with mahanine at 25 µM significantly decreased the viability of HepG2 hepatocellular carcinoma cells and HuCCT1 and KKU-100 cholangiocarcinoma cells (Fig.…”

“…Moreover, mahanine was reported to inhibit the autophagic degradation activity in human oral squamous cell carcinoma cells (Utaipan et al 2017). Here we have shown that mahanine exerted the potent cell toxicity in the hepatic cancer cell lines, including HuCCT1 cells that were resistant to cisplatin (Samatiwat et al 2016); namely, mahanine may kill hepatic cancer cells through the mechanism different from cisplatin.…”

“…Subsequently, we found the increase in MITF expression in human hepatic cancer cells treated with mahanine, a carbazole alkaloid identified in the leaves of Murraya koenigii (Linn.) Spreng (curry leaf) (Roy et al 2004;Utaipan et al 2017). This plant is used as folk medicine in Thailand, where the liver flukeassociated cholangiocarcinoma is common (Shaib and El-Serag 2004).…”

Medicinal Profile, Phytochemistry, and Pharmacological Activities of Murraya koenigii and its Primary Bioactive Compounds

“…Dexter (DLD-1) colon cancer cells; interestingly, this inhibition of the Akt/mTOR pathway could possibly activate the intrinsic apoptotic program [86]. Mahanine and isomahanine derived from M. koenigii leaves exert anticancer effects on oral squamous cell carcinoma cells via the induction of microtubule-associated protein 1 light chain 3, type II (LC3B-II), and cleaved caspase-3, suggesting the inhibition of autophagic flux [96]. In human leukemic cells, Mahanine was reported to induce apoptosis by interrupting signal transfer between Apo-1/Fas signaling and the Bid protein and via mitochondrial pathways in humans [59].…”

Autophagy modulators for the treatment of oral and esophageal squamous cell carcinomas

“…It was also observed that 71 and 72 might lead to the generation of ROS, causing mitochondrial dysfunction and triggering apoptosis. Microscopic analysis showed an increased formation of autophagic vacuoles and WB showed increased levels of LC3‐II and p62, suggesting that the two compounds were autophagy inducers able to stop the autophagic flux resulting in autophagic cell death . In another paper by Irimie et al, the activity of epigallocatechin‐3‐gallate ( 73 ), one of the most important bioactive compounds of green tea, was evaluated against OSCC.…”