The datasets analyzed during the current study are available from the corresponding author on reasonable request. Clinacanthus nutans Burm. However, scientific evidence is generally lacking to support this traditional claim. This study aims to investigate the in vitro, ex-vivo and in vivo effects of C.
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Metrics details. Clinacanthus nutans extracts have been consumed by the cancer patients with the hope that the extracts can kill cancers more effectively than conventional chemotherapies. Our previous study reported its anti-inflammatory effects were caused by inhibiting Toll-like Receptor-4 TLR-4 activation.
However, we are unsure of its anticancer effect, and its interaction with existing chemotherapy. We investigated the anti-proliferative efficacy of polar leaf extracts LP , non-polar leaf extracts LN , polar stem extract SP and non-polar stem extracts SN in human breast, colorectal, lung, endometrial, nasopharyngeal, and pancreatic cancer cells using 3- 4,5-dimethylthiazolyl -2,5-diphenyltetrazolium bromide, MTT assay.
The most potent extracts was tested along with gemcitabine using our established drug combination analysis. The effect of the combinatory treatment in apoptosis were quantified using enzyme-linked immunosorbent assay ELISA , Annexin V assay, antibody array and immunoblotting. A p -value of less than 0. All extracts tested were not able to induce potent anti-proliferative effects. This is the first report of C.
In the presence of SN extracts, we can reduce the dose of gemcitabine 2. The synergism was independent of TLR-4 expression in pancreatic cancer cells. These results provide strong evidence of C. Hence, it should not be used as a total substitution for any chemotherapy agents. However, SN extracts may synergise with gemcitabine in the anti-tumor mechanism. Peer Review reports. Many cancer patients use therapies promoted as viable alternatives to conventional cancer treatment with questionable outcomes.
Such unproven modalities can be potentially harmful. Furthermore, an even greater proportion of cancer patients use complementary therapies such as herbs and supplements along with conventional cancer treatment such as chemotherapy and radiation therapy.
Some of these may have been proven to be adjunctive approaches that control symptoms and enhance quality of life. There is much controversy as to whether these natural health products should be taken during conventional cancer treatments and both sides of the divide provide valid arguments. More importantly, the drug-herb interaction effects of such complementary therapies with chemotherapy agents are often not studied during clinical trials or even receive post-marketing surveillance [ 1 , 2 ].
Cancer development and progression is usually not driven by single cells. The tumor microenvironment drives the drug resistance and tumor survival [ 3 , 4 , 5 ]. It is hard to believe any single agent may effectively suppress cancer development and progression. Researchers have been actively targeting the Mother Nature to explore any potential regimen for cancer.
Despite dietary or plant-isolated compounds [ 6 , 7 , 8 , 9 , 10 , 11 , 12 ] exhibiting a potent anticancer effect, thorough scientific investigation should be conducted in order to validate their effects on cancer treatment.
Clinacanthus nutans , or Sabah Snake Grass SSG as it is locally known in Malaysia, is a plant with indigenous origins in Southeast Asia, although its actual origin is unknown.
It was originally isolated from Sabah, West Malaysia and hence, it is named after the location. The genus Clinacanthus consists of two species, C. Among cancer patients in Malaysia, SSG has been known to cure the latter stages of liver cancer; however, its consumption is advised to be carried out only following conventional treatments of chemotherapy and radiotherapy due to possible adverse effects that could arise. To the best of our knowledge, this claim has no scientific evidence to support it, and is made purely due to the cautioning of concomitant use of chemotherapy agents with other unproven agents.
Several isolated studies have therefore investigated the claims. It was suggested the methanolic extracts of C. In another study, however, C.
Furthermore, C. Thus, it is rather challenging for clinicians to recommend or to discourage the use of C. The need to standardise the experimental procedures, including using the standardised extracts, and to use a standardised in vitro anticancer procedure, is of the utmost importance to mitigating the anticancer potential of C.
In our previous study, we have prepared the standardised polar and non-polar fractions of C. These extracts were found to exhibit anti-inflammatory properties through inhibiting Toll-like Receptor 4 TLR-4 activation and nitric oxide production, one of the key inflammatory mediators.
The total phenolic contents and total flavonoid contents were correlated with its anti-inflammatory potency.
More importantly, we have established that these standardised bioactive extracts of C. In this study, we aimed to expand our knowledge by investigating the anticancer effects of these standardised C.
Since most patients are likely also to take both chemotherapy agents and C. The current investigation was also designed to determine the possible cell death behind the interaction between C.
As established in the previous study [ 18 ], the C. Clinacanthus nutans polar leaf extracts LP , non-polar leaf extracts LN , polar stem extract SP and non-polar stem extracts SN were prepared based on the previous study without modification [ 18 ]. The leaves and stem bark were dried and turned to powder separately before extraction. The dried powders were immersed in polar solvents methanol and dichloromethane or non-polar solvents hexane and diethyl ether for 3 days at room temperature.
The preparation was in accordance to the way the extracts were normally consumed by the public and from methods described in published literature. We routinely determined the total phenolic content TPC and total flavonoid content TFC of extracts using standardized assays as per described in previous studies.
Cancer cells derived from cancers that had high prevalence or high mortality were selected for this study. Human nasopharyngeal cancer cells were shared with us by Professor Sam C. K, University of Malaya, Malaysia. As a comparison, we also tested vitexin and isovitexin Sigma Aldrich, USA , the pure compounds that were commonly isolated Clinacanthus nutans extracts [ 13 ]. IC 50 values were calculated based on the dose-response curve generated.
The drug combination analysis was performed using the method developed by Chou and Talalay [ 24 ]. Multiple fixed-ratio dose—effect curves and the calculations of combination index CI and drug reduction index DRI were generated using Calcusyn 2. DRI values were used to describe the dose reduction potential of the combination by predicting the dose of gemcitabine and SSG needed when used in combination to achieve a defined effect level fraction affected, Fa in comparison with the single-agent dose required for such effect.
Since we could not exclude the possibility that SN extracts and gemcitabine might act simultaneously through different mechanisms, we also included the Bliss independence model to define the combinatorial effects. The volume of Bliss interaction for dose-response combination matrix was statistically evaluated and graded accordingly by MacSynergy II program version 1. Cancer cells treated with 0. The enrichment factors of apoptosis were calculated based on the absorbance of cells treated with compound or combination over absorbance of cells treated with vehicle control 0.
The fold changes were calculated after being normalised with cells treated with 0. Fifty micrograms of total protein was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis followed by immunoblotting. To evaluate whether C.
SN possess the most potent IC 50 : Vitexin and isovitexin, the most commonly isolated C. To investigate whether there was synergism between SN extracts and gemcitabine, via enhancement of gemcitabine sensitivity in the pancreatic cancer cells, we evaluated the anti-proliferative effects of SN extracts in combination with gemcitabine in the squamous-like SW and BxPC3 pancreatic cancer cells, and the progenitor-like AsPC1 pancreatic cancer cells.
As shown in Fig. The CI values and the combination analyses based on the HSA and Bliss independence models indicated that the combination of SN extracts and gemcitabine exerted significant synergism to inhibit the squamous subtype pancreatic cancer cells SW and BxPC3 being tested Figs. The treatment concentration ratio of SN to gemcitabine shown was Combenefit software was used to generate and assess the dose-response surface curves and drug interaction effects following the HSA model and Bliss independence model as implemented in the software.
The level of synergy blue or antagonism red at each combination was indicated by the color scale shown. The net Bliss interaction volumes of the matrix drug combination were calculated based on MacSynergy II program and graded accordingly. All the experiments were conducted in triplicate and performed independently. Together, our results implied that synergistic interaction exists between SN extracts and gemcitabine, with SN extracts strongly enhancing gemcitabine sensitivity in squamous subtype of pancreatic cancer cells, and the favourable DRI trend obtained might be exploited as an added benefit from combining SN with gemcitabine in pancreatic cancer cells.
To determine the mode of cell deaths induced by the synergistic effects of SN extracts and gemcitabine, we quantified the apoptotic nucleosomes induced by 0. Combination of SN and gemcitabine also induced 3. It was found to be much higher fold of apoptotic cells death as compared to apoptotic induction when pancreatic cancer cells were treated with 0. GAPDH was used as loading control. These results indicated that the synergistic effects of SN extracts and gemcitabine was independent to TLR-4 expression on human pancreatic cancer cells.
Our results showed that SW cells treated with combination of SN extracts and gemcitabine , showed 2. Gemcitabine reduced the expression of anti-apoptotic bodies, namely bcl-2 B-cell lymphoma 2 , cIAP-2 cellular inhibitor of apoptosis 2 , livin, survivin and XIAP X-linked inhibitor of apoptosis protein in SW cells.
However, our results showed that the combination of SN extracts and gemcitabine had a greater reduction of these anti-apoptotic bodies expression in SW cells, compared to cells treated with gemcitabine alone.
As highlighted in Fig. The results were similar to findings observed in antibody array. These results suggest that synergistic apoptotic induction by upregulating bax and downregulating bcl-2, cIAP-2, and XIAP in human pancreatic cancer cells. In recent years, the use of complementary and alternative medicine among cancer patients has been raising.
This exponential increase could be attributed to the enhanced awareness of the general public to social media who are marketing the benefits of complementary medicine and the overall misgivings towards conventional cancer treatments. Furthermore, patients consume complementary and alternative medicine because they believe it will improve their quality of life. Some patients may even think complementary and alternative medicine can prolong life and promote cancer remission.
These common complementary and alternative medicines include herbs, vitamins, minerals, homeopathy, naturopathy and specialized diets [ 33 , 34 , 35 , 36 ]. Despite the popularity of complementary and alternative medicine, there is limited research evaluating the scientific efficacy of complementary and alternative medicine in cancer treatment as well as its interaction with conventional cancer treatment.
In addition, compared to patients receiving conventional cancer treatment, patients who chose complementary and alternative medicine also showed higher refusal rates of surgery, chemotherapy, radiotherapy, and hormone therapy. These factors lower the 5-year overall survival rates, inadvertently leading to greater mortality risks [ 36 ]. Hence, the need for conclusive evidence of the evidence of complementary and alternative medicine in cancer treatment is needed.
Based on folk medicine, C. To the best of our knowledge, only several isolated studies have thus far investigated the claims.
Metrics details. Clinacanthus nutans extracts have been consumed by the cancer patients with the hope that the extracts can kill cancers more effectively than conventional chemotherapies. Our previous study reported its anti-inflammatory effects were caused by inhibiting Toll-like Receptor-4 TLR-4 activation. However, we are unsure of its anticancer effect, and its interaction with existing chemotherapy.
Clinacanthus nutans is a species of plant in the family Acanthaceae. It is also known by the common names belalai gajah Malay , phaya yo Thai , Sabah snake grass , ki tajam Sunda , and dandang gendis Jawa. This plant is used in the traditional herbal medicines of Malaysia, Indonesia, Thailand, and China. Clinacanthus nutans is a herbaceous plant that grows in low shrubs up to 2. Its stems are green, woody, upright and cylindrical.
Context: Medicinal plants have attracted global attention for their hidden therapeutic potential. Clinacanthus nutans Burm. CN contains phytochemicals common to medicinal plants, such as flavonoids. Traditionally, CN has been used for a broad range of human ailments including snake bites and cancer. Objectives: This article compiles the ethnomedicinal uses of CN and its phytochemistry, and thus provides a phytochemical library of CN.
Clinacanthus nutans has had a long history of use in folk medicine in Malaysia and Southeast Asia; mostly in the relief of inflammatory conditions. In this study, we investigated the effects of different extracts of C. Extracts of leaves and stem bark of C. The extracts were standardized by determining its total phenolic and total flavonoid contents. These results provide a basis for understanding the mechanisms underlying the previously demonstrated anti-inflammatory activity of C.