Oral cancers although preventable, possess a low five-year survival rate which has remained unchanged over the past three decades. In an attempt to find a more safe, affordable and effective treatment option, we describe here the use of 1'S-1'-acetoxychavicol acetate (ACA), a component of Malaysian ginger traditionally used for various medicinal purposes.
Geranylated 4-phenylcoumarins, DMDP-1 & -2 isolated from Mesua elegans were investigated for anticancer potential against human prostate cancer cells. Treatment with DMDP-1 & -2 resulted in cell death in a time and dose dependent manner in an MTT assay on all cancer cell lines tested with the exception of lung adenocarcinoma cells. DMDP-1 showed highest cytotoxic efficacy in PC-3 cells while DMDP-2 was most potent in DU 145 cells. Flow cytometry indicated that both coumarins were successful to induce programmed cell death after 24 h treatment. Elucidation on the mode-of-action via protein arrays and western blotting demonstrated death induced without any significant expressions of caspases, Bcl-2 family proteins and cleaved PARP, thus suggesting the involvement of caspase-independent pathways. In identifying autophagy, analysis of GFP-LC3 showed increased punctate in PC-3 cells pre-treated with CQ and treated with DMDP-1. In these cells decreased expression of autophagosome protein, p62 and cathepsin B further confirmed autophagy. In contrary, the DU 145 cells pre-treated with CQ and treated with DMDP-2 has reduced GFP-LC3 punctate although the number of cells with obvious GFP-LC3 puncta was significantly increased in the inhibitor-treated cells. The increase level of p62 suggested leakage of cathepsin B into the cytosol to trigger potential downstream death mediators. This correlated with increased expression of cathepsin B and reduced expression after treatment with its inhibitor, CA074. Also auto-degradation of calpain-2 upon treatment with DMDP-1 &-2 and its inhibitor alone, calpeptin compared with the combination treatment, further confirmed involvement of calpain-2 in PC-3 and DU 145 cells. Treatment with DMDP-1 & -2 also showed up-regulation of total and phosphorylated p53 levels in a time dependent manner. Hence, DMDP-1 & -2 showed ability to activate multiple death pathways involving autophagy, lysosomal and endoplasmic reticulum death proteins which could potentially be manipulated to develop anti-cancer therapy in apoptosis resistant cells.
Geranylated 4-phenylcoumarins DMDP-1 and DMDP-2 isolated from Mesua elegans were elucidated for their role in inducing caspase-independent programmed cell death (CI-PCD) in prostate cancer cell lines, PC-3 and DU 145, respectively. Cell homeostasis disruption was demonstrated upon treatment, as shown by the increase in calcium ion through colourimetric assay and endoplasmic reticulum (ER) stress markers GRP 78 and p-eIF2α through western blot. Subsequently, cytoplasmic death protease calpain-2 also showed increased activity during DMDP-1 & -2 treatments, while lysosomic death protease cathepsin B activity was significantly increased in PC-3 treated with DMDP-1. Flow cytometry showed a reduction in mitochondrial membrane potential in both cell lines, while western blotting showed translocation of mitochondrial death protease AIF into the cytoplasm in its truncated form. Furthermore, DMDP-1 & -2 treatments caused significant increase in superoxide level and oxidative DNA damage. Concurrent inhibition of calpain-2 and cathepsin B during the treatment showed an attenuation of cell death in both cell lines. Hence, DMDP-1 & -2 induce CI-PCD in prostate cancer cell lines through calpain-2 and cathepsin B.
1'-acetoxychavicol acetate (ACA) extracted from the rhizomes of Alpinia conchigera Griff (Zingiberaceae) has been shown to deregulate the NF-ĸB signaling pathway and induce apoptosis-mediated cell death in many cancer types. However, ACA is a hydrophobic ester, with poor solubility in an aqueous medium, limited bioavailability, and nonspecific targeting in vivo. To address these problems, ACA was encapsulated in a nanostructured lipid carrier (NLC) anchored with plerixafor octahydrochloride (AMD3100) to promote targeted delivery towards C-X-C chemokine receptor type 4 (CXCR4)-expressing prostate cancer cells. The NLC was prepared using the melt and high sheer homogenization method, and it exhibited ideal physico-chemical properties, successful encapsulation and modification, and sustained rate of drug release. Furthermore, it demonstrated time-based and improved cellular uptake, and improved cytotoxic and anti-metastatic properties on PC-3 cells in vitro. Additionally, the in vivo animal tumor model revealed significant anti-tumor efficacy and reduction in pro-tumorigenic markers in comparison to the placebo, without affecting the weight and physiological states of the nude mice. Overall, ACA-loaded NLC with AMD3100 surface modification was successfully prepared with evidence of substantial anti-cancer efficacy. These results suggest the potential use of AMD3100-modified NLCs as a targeting carrier for cytotoxic drugs towards CXCR4-expressing cancer cells.
Plants are rich in phytoconstituent biomolecules that served as a good source of medicine. More recently, they have been employed in synthesizing metal/metal oxide nanoparticles (NPs) due to their capping and reducing properties. This green synthesis approach is environmentally friendly and allows the production of the desired NPs in different sizes and shapes by manipulating parameters during the synthesis process. The most commonly used metals and oxides are gold (Au), silver (Ag), and copper (Cu). Among these, Cu is a relatively low-cost metal that is more cost-effective than Au and Ag. In this review, we present an overview and current update of plant-mediated Cu/copper oxide (CuO) NPs, including their synthesis, medicinal applications, and mechanisms. Furthermore, the toxic effects of these NPs and their efficacy compared to commercial NPs are reviewed. This review provides an insight into the potential of developing plant-based Cu/CuO NPs as a therapeutic agent for various diseases in the future.
Autophagy plays a role in deciding the fate of cells by inducing either survival or death. 1'S-1-acetoxychavicol acetate (ACA) is a phenylpropanoid isolated from rhizomes of Alpinia conchigera and has been reported previously on its apoptotic effects on various cancers. However, the effect of ACA on autophagy remains ambiguous. The aims of this study were to investigate the autophagy-inducing ability of ACA in human non-small cell lung cancer (NSCLC), and to determine its role as pro-survival or pro-death mechanism. Cell viability assay was conducted using MTT. The effect of autophagy was assessed by acridine orange staining, GFP-LC3 punctate formation assay, and protein level were analysed using western blot. Annexin V-FITC/PI staining was performed to detect percentage of cells undergoing apoptosis by using flow cytometry. ACA inhibits the cell viability and induced formation of cytoplasmic vacuoles in NSCLC cells. Acidic vesicular organelles and GFP-LC3 punctate formation were increased in response to ACA exposure in A549 and SK-LU-1 cell lines; implying occurrence of autophagy. In western blot, accumulation of LC3-II accompanied by degradation of p62 was observed, which further confirmed the full flux of autophagy induction by ACA. The reduction of Beclin-1 upon ACA treatment indicated the Beclin-1-independent autophagy pathway. An early autophagy inhibitor, 3-methyaldenine (3-MA), failed to suppress the autophagy triggered by ACA; validating the existence of Beclin-1-independent autophagy. Silencing of LC3-II using short interfering RNA (siRNA) abolished the autophagy effects, enhancing the cytotoxicity of ACA through apoptosis. This proposed ACA triggered a pro-survival autophagy in NSCLC cells. Consistently, co-treatment with lysosomal inhibitor, chloroquine (CQ), exerted a synergistic effect resulting in apoptosis. Our findings suggested ACA induced pro-survival autophagy through Beclin-1-independent pathway in NSCLC. Hence, targeting autophagy pathway using autophagy inhibitor such as CQ represented a novel promising approach to potentiate the cytotoxicity of ACA through apoptosis in NSCLC.
1'-S-1'-acetoxychavicol acetate (ACA) has been previously reported to reduce tumor volume in nude mice, at an effective dose of 1.56 mg/kg body weight. However, the detailed toxicological profile for ACA has not yet been performed. Herein, we investigated the toxicity of intravenous administration of ACA in male and female Sprague-Dawley rats, both acutely (with single doses of 2.00, 4.00 and 6.66 mg/kg body weight, for 14 days), and sub-acutely (with weekly injections of 0.66, 1.33, and 2.22 mg/kg, for 28 days). In both toxicity studies, treatment with ACA did not affect behavior, food/water intake or body weight, nor did it induce any changes in clinically relevant hematological and biochemical parameters or mortality, suggesting that the LD50 of ACA was higher than 6.66 mg/kg body weight, regardless of sex. Sub-acutely, there was however, mild focal inflammation of kidneys and lobular hepatitis, but these were not associated with significant functional adverse effects. Therefore, the no-observed-adverse-effect level (NOAEL) for intravenous administration of ACA in the present 28-day sub-acute study was 2.22 mg/kg body weight, in both male and female rats. These findings provide useful information regarding the safety of ACA use in a healthy, non-tumor-bearing rat model.
Cancer development and progression are extremely complex due to the alteration of various genes and pathways. In most cases, multiple agents are required to control cancer progression. The purpose of this study is to investigate, using a mouse model, the synergistic interactions of anti-cancer agents, 1'-S-1'-acetoxychavicol acetate (ACA), Mycobacterium indicus pranii (MIP), and cisplatin (CDDP) in double and triple combinations to treat chemo-sensitize and immune-sensitize breast cancer. Changes in tumor volume and body weight were monitored. Organs were harvested and stained using hematoxylin-eosin for histopathological assessment. Milliplex enzyme-linked immunosorbent assay (ELISA) was performed to determine cytokine levels, while immunohistochemistry (IHC) was conducted on tumor biopsies to verify systemic drug effects. In vivo mouse models showed tumor regression with maintenance of regular body weight for all the different treatment regimens. IHC results provided conclusive evidence indicating that combination regimens were able to down-regulate nuclear factor kappa-B activation and reduce the expression of its regulated pro-inflammatory proteins. Reduction of pro-inflammatory cytokines (e.g., IL-6, TNF-α, and IFN-ɣ) levels were observed when using the triple combination, which indicated that the synergistic drug combination was able to significantly control cancer progression. In conclusion, ACA, MIP, and CDDP together serve as promising candidates for further development and for subsequent clinical trials against estrogen-sensitive breast cancer.
Previous in vitro and in vivo studies have reported that 1'-S-1'-acetoxychavicol acetate (ACA) isolated from rhizomes of the Malaysian ethno-medicinal plant Alpinia conchigera Griff (Zingiberaceae) induces apoptosis-mediated cell death in tumour cells via dysregulation of the NF-κB pathway. However there were some clinical development drawbacks such as poor in vivo solubility, depreciation of biological activity upon exposure to an aqueous environment and non-specific targeting of tumour cells. In the present study, all the problems above were addressed using the novel drug complex formulation involving recombinant human alpha fetoprotein (rhAFP) and ACA.