Anticancer Properties of Plectranthus ornatus-Derived Phytochemicals Inducing Apoptosis via Mitochondrial Pathway

Abstract

Since cancer treatment by radio- and chemotherapy has been linked to safety concerns, there is a need for new and alternative anticancer drugs; as such, compounds isolated from plants represent promising candidates. The current study investigates the anticancer features of halimane (11R*,13E)-11-acetoxyhalima-5,13-dien-15-oic acid (HAL) and the labdane diterpenes 1α,6β-diacetoxy-8α,13R*-epoxy-14-labden-11-one (PLEC) and forskolin-like 1:1 mixture of 1,6-di-O-acetylforskolin and 1,6-di-O-acetyl-9-deoxyforskolin (MRC) isolated from Plectranthus ornatus in MCF7 and FaDu cancer cell lines. Cytotoxicity was assessed by MTT assay, ROS production by Di-chloro-dihydro-fluorescein diacetate assay (DCFH) or Red Mitochondrial Superoxide Indicator (MitoSOX) and Mitochondrial Membrane Potential (MMP) by fluorescent probe JC-1 (5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolylcarbocyanine iodide). In addition, the relative amounts of mitochondrial DNA (mtDNA) were determined using quantitative Real-Time-PCR (qRT-PCR) and damage to mitochondrial DNA (mtDNA) and nuclear DNA (nDNA) by semi-long run quantitative Real-Time-PCR (SLR-qRT-PCR). Gene expression was determined using Reverse-Transcription-qPCR. Caspase-3/7 activity by fluorescence was assessed. Assessment of General In Vivo Toxicity has been determined by Brine Shrimp Lethality Bioassay. The studied HAL and PLEC were found to have a cytotoxic effect in MCF7 with IC50 = 13.61 µg/mL and IC50 = 17.49 µg/mL and in FaDu with IC50 = 15.12 µg/mL and IC50 = 32.66 µg/mL cancer cell lines. In the two tested cancer cell lines, the phytochemicals increased ROS production and mitochondrial damage in the ND1 and ND5 gene regions and reduced MMP (ΔΨm) and mitochondrial copy numbers. They also changed the expression of pro- and anti-apoptotic genes (Bax, Bcl-2, TP53, Cas-3, Cas-8, Cas-9, Apaf-1 and MCL-1). Studies demonstrated increase in caspase 3/7 activity in tested cancer cell lines. In addition, we showed no toxic effect in in vivo test for the compounds tested. The potential mechanism of action may have been associated with the induction of apoptosis in MCF7 and FaDu cancer cells via the mitochondrial pathway; however, further in vivo research is needed to understand the mechanisms of action and potential of these compounds.