Multicomponent phytotherapeutic approach gaining momentum: is the "one drug to fit all" model breaking down?
Received 6 May 2013
Received in revised form 22 June 2013
Accepted 26 July 2013
Natural product based drugs constitute a substantial proportion of the pharmaceutical market particularly in the therapeutic areas of infectious diseases and oncology. The primary focus of any drug development program so far has been to design selective ligands (drugs) that act on single selective disease targets to obtain highly efficacious and safe drugs with minimal side effects. Although this approach has been successful for many diseases, yet there is a significant decline in the number of new drug candidates being introduced into clinical practice over the past few decades. This serious innovation deficit that the pharmaceutical industries are facing is due primarily to the post-marketing failures of blockbuster drugs. Many analysts believe that the current capital-intensive model-"the one drug to fit all" approach will be unsustainable in future and that a new "less investment, more drugs" model is necessary for further scientific growth. It is now well established that many diseases are multi-factorial in nature and that cellular pathways operate more like webs than highways. There are often multiple ways or alternate routes that may be switched on in response to the inhibition of a specific target. This gives rise to the resistant cells or resistant organisms under the specific pressure of a targeted agent, resulting in drug resistance and clinical failure of the drug. Drugs designed to act against individual molecular targets cannot usually combat multifactorial diseases like cancer, or diseases that affect multiple tissues or cell types such as diabetes and immunoinflammatory diseases. Combination drugs that affect multiple targets simultaneously are better at controlling complex disease systems and are less prone to drug resistance. This multicomponent therapy forms the basis of phytotherapy or phytomedicine where the holistic therapeutic effect arises as a result of complex positive (synergistic) or negative (antagonistic) interactions between different components of a cocktail. In this approach, multicomponent therapy is considered to be advantageous for multifactorial diseases, instead of a "magic bullet" the metaphor of a "herbal shotgun" might better explain the state of affairs. The different interactions between various components might involve the protection of an active substance from decomposition by enzymes, modification of transport across membranes of cells or organelles, evasion of multidrug resistance mechanisms among others.
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Is the drug combination strategy the future of drug discovery?
Switching over from single drug to multi-drug therapy
Traditional medicine unceasingly continues to provide frontline medicinal products for millions of people all across the globe. Although their usage is often considered with skepticism by the western pharmaceutical establishments, but these medicinal extracts which have been used in ancient system of medicine like Ayurveda in the Indian subcontinent and traditional Chinese medicine (TCM) are a very rich bioresource of therapeutic leads for pharmaceutical companies. In these systems of medicine, parent extracts have been "clinically" tested in their traditional background, in some cases over millennia. The transformation of traditional medicines into modern western drugs has its origins in the examples of the antimalarial quinine and the anti-pyretic analgesic aspirin. The success of these two early "blockbuster" drugs set the stage for ongoing drug discovery efforts from traditional medicines. Compounds obtained from medicinal extracts are more advantageous and appealing than synthetics for several reasons. Firstly, they are often stereochemically very complicated, multi-or macrocyclic molecules with limited likelihood of prior chemical synthesis and secondly they are embellished with interesting biological properties (Schmidt et al., 2007). Despite these advantages, the route from traditional medicine to western pharmaceutical is full of hurdles and challenges such as discovering the traditional medicine, isolation and synthesis of the bioactive compound, establishing the molecular mechanism of action and finally development as a pharmaceutical agent (Fig. 1). This has resulted in the introduction of fewer New Chemical Entities (NCE's) into the drug discovery process. Many analysts believe that the current model--"the one drug to fit all", approach will be unsustainable in future and that a new model is necessary for further scientific growth. It is believed that the blockbuster model is dying away.
A new chemical entity (NCE) travels a path from laboratories to clinics, involving target identification, lead identification, lead optimization, preclinical studies, and then four phases of the clinical trials: a 12-14 years odyssey is usual! The extremely complex and capital-intensive process makes companies 'target rich' but 'lead poor'. The pharmaceutical industries are facing a serious innovation deficit. Although we have become high throughput in technology, yet we have remained low throughput in thinking.
The pharmaceutical industry has historically seen incredible growth due primarily to the industry's strategy of focusing efforts toward development of "blockbuster" drugs with the potential to generate over $1 billion in sales. However, recent trends indicate that this model may no longer ensure high growth rates (Frantz, 2005). The average cost of discovering, developing and launching a new drug in June 2008 was inordinately high and represented a dramatic increase over the average cost from 1995. R&D expenses have risen from $2 billion in 1980 to over $40 billion in 2007. Surprisingly, these increases have not led to a corresponding increase in the number and efficacy of new drugs. From 1995 to 2000 (as compared to the previous five years), the number of New Molecular Entities (NMEs) approved dropped by nearly 50%, to about 40, and the number of New Chemical Entities (NCEs) produced per company declined by 41%. Moreover, the number of approvals for New Molecular Entities (NMEs) has steadily declined reaching a low of 17 in 2002 and even lower to less than 10 in 2007 and 2008 (Frantz, 2007; Thayer, 2004).
Post marketing failures of bestseller drugs have become major cause of concern for pharma-industries, leading to a paradigm shift in favor of single to multi targeted drugs and affording greater respect to traditional knowledge. Multi-target approaches are coming into the main stream (Zimmermann et al., 2007). Drugs designed to act against individual molecular targets cannot usually combat multigenic diseases such as cancer, or diseases that affect multiple tissues or cell types such as diabetes and immunoinflammatory disorders. Combination drugs that impact multiple targets simultaneously are better at controlling complex disease systems, are less prone to drug resistance and are the standard of care in many important therapeutic areas. The combination drugs currently employed are primarily of rational design, but the increased efficacy they provide justifies in vitro discovery efforts for identifying novel multi-target mechanisms. Alternative and Complementary approaches in therapeutics which are becoming popular options are based on the principle of multi-component drugs acting synergistically in a holistic fashion. Typical reductionist approach of modern science is being revisited over the background of systems biology and holistic approaches of traditional practices. Scientifically validated and technologically standardized botanical products may be explored on a fast track using innovative approaches like reverse pharmacology and systems biology, which are based on traditional medicine knowledge. Traditional medicine constitutes an evolutionary process as communities and individuals continue to discover practices transforming techniques. Many modern drugs have origin in ethnopharmacology and traditional medicine. Traditional system of medicine is based on the principle of multi-component therapy which involves synergistic interactions giving rise to a therapeutic effect. Synergism plays a key role in the traditional system of medicine like Ayurveda, traditional Chinese medicine, etc. (Corson and Crews, 2007; Patwardhan et al., 2008; Zimmermann et al., 2007).
Existing multi-target therapeutics
Increasingly, drug combinations are the standard of care for the treatment of diseases including cancer, type 2 diabetes mellitus (T2DM), viral and bacterial infection, and asthma. Often, these combinations are applied as co-therapy regimens, but in many cases the individual components of the combination are co-formulated as a single pill or injection. A new generation of multi-target drugs is currently emerging from clinical development: single chemical entities that act simultaneously at multiple molecular targets. There are several categories of multi-target therapeutics that can be defined on the basis of target relationship. In the first class, the therapeutic effect occurs at separate molecular targets that can reside within individual signaling pathways, between pathways within a cell or at separate tissues in the body. In the second category, modulation of one target facilitates action at a second target, for example by altering compound metabolism, inhibiting efflux pumps or blocking other resistance mechanisms (Table 1). Third, a coordinated action at multiple sites on a single target or macromolecular complex (e.g. prokaryotic ribosome) yields the therapeutic effect.
Note that the set of targets in each of these three cases can be modulated either by a mixture of separate chemical entities or by a single compound designed to have multiple actions. Although multi-target action can be achieved in several ways, it is the coordinated effect at the set of targets that results in the biological and, hopefully, therapeutic effect (Kubinyi, 2003). Table l lists some examples of multi-target therapeutics from various indications.
The benefits of multi-target action are well established in cancer. Traditional chemotherapeutic agents have been routinely applied as co-therapies. For example, adjunctive agents can sensitize cancer cells to DNA-damaging drugs, and newer co-therapy protocols including 5-FU, leucovorin and oxaliplatin (i.e. FOLFOX) are now applied in colorectal cancer. Currently, the molecularly targeted agents such as Herceptinl (trastuzumab) and Erbituxl (cetuximab) are being developed in combination protocols with traditional antineoplastics, estrogen blockade and other targeted agents (Dancey and Chen, 2006; Johnston, 2005). For example, Herceptinl, which targets ErbB2 (HER-2/neu), is being applied in combination with the anti-VEGF (vascular endothelial growth factor) antibody Avasti n1 to treat breast cancer, and Erbi tux1 (which targets ErbB1) is applied in combination with irinotecan for the treatment of colorectal cancer. A novel class of receptor tyrosine kinase (RTK) inhibitors that possess multi-target action in a single chemical entity are currently in clinical development. Lapatinib and canertinib are examples of a new class of pan-ErbB inhibitors. These new agents with multi-target action will almost certainly be applied in combination with other molecularly targeted or traditional chemotherapeutic agents once they reach the market (Britten, 2004; Hynes and Lane, 2005).
Table 1 Some of the examples of drug-combination products. Trade name Indication Compound 1 Compound 2 Vytorin Hyperlipidemia Ezetimibe Simvastatin Caduet Coronary heart Amlodipine Atorvastatin disease Lotrel Hypertension Amlodipine Benezapril Glucovance Type 2 diabetes Metformin Clyburide mellitus Avandamet Type 2 diabetes Metformin Rosiglitasone mellitus Truvada Anti-HlV Emtricitabine Tenofovir Kaletra Anti-HIV Lopinavir Ritonavir Rebetron Anti-Hepatitis PEC-interferon Ribavirin C Bactrim Antibacterial Trimethoprim Sulfamethoxazole Trade name Target or Target or mechanism of mechanism of action 1 action 2 Vytorin Dietary HMC-CoA cholesterol reductase Caduet Calcium-channel HMC-CoA antagonist reductase Lotrel Calcium-channel ACE inhibitor antagonist Glucovance Gluconeogenesis Insulin secretagogue Avandamet Cluconeogenesis PPAR-y agonist Truvada RT inhibitor RT inhibitor Kaletra Protease Protease inhibitor inhibitor Rebetron Interferons B Antimetabolite Bactrim Dihydrofolate Dihydropteroate reductase synthase
What synergy means in phytomedicine?
It is very difficult at the first instance to provide an unequivocal universal definition for the term synergism or synergy effect, because synergy has a precise mathematical definition according to the method used to prove it. The term synergy comes from the Greek word "Synergos" meaning "working together". Synergy means working together of two or more substances to produce an effect greater than the sum of their individual effects. In nature, synergy phenomena are ubiquitous ranging from physical science (e.g. the different combinations of quarks to form protons or neutrons) to chemistry (hydrogen and oxygen combine to produce water) to the cooperative combination among the genes to produce genomes (Wikipedia). The concept of synergy is based on the view that a cohesive group is more than the sum of its parts; synergy is the ability of a group to outperform even its best individual member. However, synergy is not always positive, in various cases synergy produces negative results.
Conventional medicine follows a reductionist approach. Specific compounds are assigned for defined biological functions or target molecules. Chemotherapy of infectious diseases represents a successful example for this "one target-one drug" concept. This is what Paul Ehrlich had in mind with his idea of "magic bullets" to specifically target diseases. This approach explains the reluctance of western medicine toward multi-component therapies with broad spectrum activities such as phytotherapy. Observable treatment successes by phytopoharmaceuticals are frequently neglected or classified as placebo effects. On the other side, the majority of diseases are multi-factorial and targeting a single cause of a disease by a single drug may not deliver satisfactory treatment results. Traditional medicinal systems including European phytotherapy, traditional Chinese medicine (TCM) or Ayurveda have a holistic approach. Instead of mono-substances, mixtures of medicinal plants with complex interactions of dozens to hundreds of compounds are used. Here, multicomponent recipes are considered as advantageous for pleiotropic diseases compared to single compounds. Instead of a "magic bullet", the metaphor of an "herbal shotgun" might better describe this situation (Wagner and Ulrich-Merzenich, 2009; Williamson, 2001).
As for as drug discovery is concerned, the therapeutic value of synergistic interactions among the different components has been known since early time and many traditional healing systems still rely on this principle of synergy. Aromatherapy is fundamentally based on the principle of combination of highly complex mixtures of essential oils to produce a therapeutic effect. The use of polyherbals has been carried down the ages and today allopathic medicine is based on the principle of synergy in which various compounds are mixed in single or isolated dosage forms which are then administered concomitantly. The combination therapy has recently got tremendous response and is now widely accepted particularly in the treatment of infectious diseases (Efferth and Koch, 2011). The world Health Organization (WHO) has advised pharmaceutical companies to use artemisinin combination therapy not only because of its 95% cure rate against Plasmodium falciparum - the causative malarial parasite, but this combination therapy may also decrease the incidence of resistance. The WHO has also urged to stop artemisinin derivatives monotherapy which not only has a lower cure rate than combination therapy but it has also higher chances of developing resistance (Douglas et al., 2010). In the fight against the multidrug resistant microbes, the combination therapy is becoming more and more of utmost importance. The current treatment regimen for tuberculosis constitutes a cocktail of five drugs viz., pyrazinamide, rifampicin isoniazid, streptomycin and ethambutol (PRISE). Another renowned antimicrobial agent having a synergistic effect in combination is amoxicillin (a 13-lactam antibiotic) and clavulanic acid. Clavulanic acid binds to 13-lactamase producing microorganisms, which protects amoxicillin from 13-lactamase attack, which in turn results in an extended spectrum of activity for amoxicillin (Chan and lseman, 2002; KaIan and Wright, 2011; Matsuura et al., 1980). Multi-drug therapy is also being practiced worldwide in the treatment of AIDS and other infectious diseases, hypertension numerous types of cancer and rheumatic diseases. The multi-drug concept in current cancer therapy has been designated as biomodulatory-metronomic chemotherapy. The idea is to fight the tumor via a process of concerted and concomitant action not through direct destruction of the tumor but rather by suppression or activation of different processes which are essential for the tumor's survival (e.g. by angiogenesis, induction of apoptosis, activation of the immune system, etc.). Multi-target therapy is more effective and less vulnerable to adaptive resistance because the biological systems are less able to compensate the action of two or more substances simultaneously. As a result, mono-target drugs are incapable of effectively combating complex pathological conditions like cancer and infectious diseases (Berenbaum, 1989).
Definition and proof of synergy
If two drugs A and B are combined, and if drug A has an effect and drug B has no effect and if in combination they have an effect that is greater than that of drug A, then it is enhancement or potentiation. We can describe the effect simply as percent enhancement or - fold of potentiation. If A and B alone each has an effect, then in combination they may produce a synergistic, an additive, or an antagonistic effect. By definition, synergism is an effect that is more than additive, whereas the definition for antagonism is an effect that is less than additive. Synergism occurs when two or more herbal ingredients mutually enhance each other's effect more significantly than the simple sum of these ingredients. Synergy represents a form of interaction as opposed to a simple addition response. Among all the methods proposed for the proof of synergy effects, the "isobole method" of Berenbaum (1989) appears to be one of the most experimentally practicable and also the most demonstrative method. An isobole is an "iso-effect" curve, in which a combination of constituents (da, db) is represented on a graph, the axes of which are the dose-axes of the individual agents (Da and Db). If the agents do not interact, the isobole (the line joining the points representing the combination to those on the dose axes representing the individual doses with the same effect as the combination) will be a straight line. If synergy is occurring, i.e. the effect of the combination is greater than expected from their individual dose-response curves, the dose of the combination needed to produce the same effect will be less than for the sum of the individual components and the curve is said to be 'concave..The opposite applies for antagonism, in which the dose of the combination is greater than expected, and produces a 'convex' isobole (Figs. 2 and 3). It is quite possible to have synergy at one dose combination and antagonism at another, with the same substances and this would give a complicated isobole with a wave-like or even elliptical appearance. To demonstrate synergy effect, in vitro or animal models are utilized for the demonstration of the isoboles of a mixture of two substances (Berenbaum, 1989: Chou, 2006, 2010).
Combination index ((CI)
A combination index (Cl) method was put forth by Chou and Talalay (1983) for quantifying the synergism or antagonism of two drugs and assesses the nature of the interaction (synergy, additivity, or antagonism) (Chou, 2006). The combination index is calculated on the basis of their concentration and biological activity (like cell growth inhibition, [IC.sub.50]). Cl analysis provides quantitative data and the numerical value is calculated as:
CI = [C.sub.A, X]/[IC.sub.X, A] + [C.sub.B, X]/[IC.sub.X, B]
where [C.sub.A, X] and [C.sub.B, X] are the concentrations of the drug A and drug B used in combination to produce a mean effect X ([IC.sub.50]). [IC.sub.X, A] and [IC.sub.X, B] are the median effect values ([IC.sub.50]) for single drug A and B. Combination index (CI) is used to quantitatively depict synergism (CI < I ), additive effect (CI = 1) and antagonism (CI > 1) (Table 2). The combination index or the interaction index is a quantitative measure of the degree of synergism or sub-additivity that occurs when two drugs are mixed together. Doses of drugs that give the same effect are called isoboles and the method of analysis is called isobolar method. An isobologram is a cartesian plot of doses that in combination yield a specified level of effect. It is a convenient and currently popular way of graphically exhibiting results of drug-combination studies, because paired values of experimental points that lie below or above the line joining the axial points ([IC.sub.50] or [ED.sub.50] values) represent supra- and sub-additive combinations, respectively.
Table 2 Symbols and description of synergism or antagonism in drug combination models analyzed with the combination index (Cl) method. This method is based on those described by Chou and Talalay (1984). CI < 1, CI = 1, and CI > 1 indicate synergism, additive effect and antagonism, respectively. Range of combination Description Graded symbols index (Cl) <0.1 Very strong synergism +++++ 0.1-0.3 Strong synergism ++++ 0.3-0.7 Synergism +++ 0.7-0.85 Moderate synergism ++ 0.85-0.90 Slight synergism + 0.90-1.10 Nearly additive [+ or -] 1.10-1.20 Slight antagonism - 1.20-1.45 Moderate antagonism -- 1.45-3.3 Antagonism --- 3.3-10 Strong antagonism ---- >10 Very strong antagonism -----
Experimental evidence in favor of synergism
It is a routine practice for phytochemists to examine and prepare extracts from medicinal plants keeping in view isolating the single chemical entity responsible for the therapeutic effect. But this approach may lead to inconclusive findings. If a mixture of substances is required for the effect, then the bioassay-led isolation, which narrows activity down initially to a fraction and finally to a compound, is doomed to failure and this has led to the result that the plants are in fact lacking the desired therapeutic effect. An excellent example is that of Kigelia pinnata, in which the previously reported cytotoxic activity was destroyed after fractionation. These and other related misconceptions regarding synergy can be dispelled by clinical trials. Most of the time when activity is thought to be lost during purification, synergy should be suspected. If synergism is known or suspected to be present, the mixture is necessary for the therapeutic effect. Sometimes the presence of whole plant material, which may contain for instance antioxidants, may protect the active secondary metabolites from decomposition and degradation. Sometimes the active compound may be a minor unidentified compound.
The following examples investigated through in vitro and in vivo experiments are forwarded in support of synergism:
Example 1. Marihuana (Cannabis sativa)
The well-known cannabis and tetra hydrocannabinol (THC = 6,9--THC) possess antispastic action, hallucinogenic, antiemetic, anxiolytic and analgesic effects. Baker et al. (2000) have proved it in an immunogenic animal model of multiple sclerosis. Because there were some indications that showed a stronger muscle-antispastic effect of the extract than of pure THC, a comparative i.v. test of 1 mg/THC and 5 mg/kg Cannabis extract, the latter standardized on a concentration of 20% of THC, was carried out. It was shown that the whole cannabis extract with equimolar THC concentration was much more effective antispastic agent than THC alone (Baker et al., 2000). Since in a preliminary investigation, THC free extract did not exhibit strong antispastic effect, concomitant chemical constituents of the Cannabis extract, most probably cannabidiol, may be responsible for the synergy effects. It has been reported that cannabidiol which is also a constituent of the extract promotes an increase in the transport of ananclamide through the brain membrane not evident with THC. This could explain the stronger antispastic effect of the Cannabis extract (Wilkinson et al., 2003; Williamson and Evans, 2000; Zuardi et al., 1982).
Example 2. St. John's Wort (Hypericum perforatum)
Standardized Hypericum extracts have been shown by more than 40 placebo-controlled clinical studies to be effective for the treatment of mild, moderate and even severe depression. The pharmacological effects of several of these extracts are comparable with the synthetic psychopharmacological drugs like amitriptyline, imipramin and flumazenil. Evidence for the synergistic pharmacokinetic interactions has been obtained for St. John's Wort extracts (Woelk, 2000). During a bio-assay guided fractionation of a methanolic extract, hypericin and pseudohypericin were identified as components that showed pharmacological effect in the forced swimming test while the purified compounds were found to be inactive after treatment at doses comparable to the total extracts. When a fraction containing procyanidins, which itself was inactive in the test model, was combined with hypericin or pseudohypericin, activity was detected at relatively low oral doses. According to various investigations performed so far on the pharmacological effects of Hypericum, several chemical constituents are believed to be involved in its effectiveness particularly hyper-Ion i n, the hypercines, amentoflavon, rutin, hyperosid, xanthones and proanthocyanidines (Fig. 3). This idea has been hypothesized by various in vitro neurochemical studies with various CNS receptors making the use of radioligand-binding techniques to prove that the antidepressant activity of standardized Hypericum extract might be a result of the cooperative action of several chemical constituents of H. perforatum. As shown in the figure, various targets are believed to be involved like presynaptic and postsynaptic neurons, the pituitary gland and hypothalamus are all involved as possible targets and all the main chemical constituents of St. John's Wort show affinities to any of the above targets (Schulz, 2001, 2003; Simmen et al., 2001).
Example 3. Iberogast[R] (a phytopreperation of nine plant extracts)
This is an example for the multi-target principle which comprises of nine plant extracts and is considered in Germany and other European countries as a leading phytopreperation for the treatment of functional dyspepsia and motility-related intestinal disorders (Fig. 4). Twelve clinical studies, among them two in comparison with the synthetic drugs cisapride and metoclopramide, showed a complete therapeutic equivalence with the two synthetic drugs, with the additional benefit that the phytopreperation showed fewer or no side effects in comparison to the two synthetics. lberogast exhibits a multi-target effect by balancing the disturbed gastrointestinal motility function, by alleviating gastro-intestinal hypersensitivity, by inhibiting the inflammation, suppression of gastric juice secretion and effects on gastro-intestinal autonomic afferent function. On the contrary to this multiphytopreperation, the synthetic monodrugs cisapride and metoclopramide as classical proton pump inhibitors target only one symptom of functional dyspepsia. Each plant extract of the preparation was investigated separately in all relevant pharmacological in vitro and in vivo models with the result that all extracts, some of them multifunctionally or synergistically, are involved in the overall pharmacological effect (Allescher, 2006; Sailer et al., 2002; Wagner, 2006).
Example 4. Kava Kava (Piper methysticum)
Kava Kava (Piper methysticum) is a plant native to the South Pacific islands with anxiolytic and sedative activities. Controlled human clinical trials show it to be superior to placebo for the treatment of anxiety, an equivalent in efficacy to the benzodiazepine oxazepam (Serax[R]) (Pittler and Ernst, 2000). The bioactive chemical constituents from Kava are the kava lactones, particularly kavain, dihyd rokava in, yangonin, dimethoxyyangonin, methysticin and dihydromethysticin (Fig. 5). The kava lactones yangonin and dimethoxyyangonin act in an anticonvulsant manner. Their efficacy is much more in combination with other kava constituents rather than when they are applied separately. One constituent namely di hydromethysticin, seems to be particularly important for the synergy. In some experiments, in mice and dogs it was observed that the oral bioavailability of kavain was greater if it was administered in an extract when compared to an equivalent quantity of the pure constituent. Kava lactones pass the blood--brain barrier and behavioral effects occur at micromolar concentrations. Kava lactones enhance binding to the GABAA receptor in the low micromolar range, through a non-benzodiazepine mechanism. These lactones also block voltage-gated [Na.sup.+] and [Ca.sup.2+] channels in micromolar concentrations (Friese and Gleitz, 1998; Jussofie et al., 1994; Keledjian et al., 1988; Lindenberg and Pitule-Schodel, 1990).
Example 5. Antimicrobial action of berberine potentiated by 5'-methoxyhydnocarpin (5'-MHC)
In a study published in Proceedings of National Academy of Sciences (PNAS) USA, Stermitz et al., 2000 have convincingly demonstrated the occurrence of synergy in some Berberis medicinal plants (Stermitz et al., 2000). Berberine alkaloids are the cationic antimicrobials synthesized by a variety of plants particularly of the family Berberidaceae. These cationic alkaloids are readily extruded by multidrug resistant pumps
(MDRs) present in various bacterial strains. These MDRs protect the microbial cells from both synthetic and natural antimicrobials. Several Berberis medicinal plants producing berberine are also found to synthesize an inhibitor of the Nor A MDR pump of a human pathogen Staphylococcus aureus, namely 5'-methoxyhydnocarpin (5'-MHC), previously known as a minor constituent of Chaulmoogra oil, a traditional therapy for leprosy (Fig. 7). 5'-MHC itself alone does not possess the antimicrobial activity but strongly potentiates the action of berberine and other Nor A substrates against S. aureus (Fig. 6). It has been reported that MDR-dependent efflux of ethidium bromide and berberine from S. aureus cells is completely inhibited by 5'-MHC. The degree of accumulation of berberine in the bacterial cells is increased considerably in the presence of 5'-MHC, proving that this plant compound effectively disabled the bacterial resistance mechanism against the berberine antimicrobial.
The chloroform extracts of the leaves from Berberis repens, Berberis aquifolia, Berberis fremonti had no antimicrobial activity at >500 [micro]g/m1 concentration, but inhibited S. aureus growth completely in the presence of 30[micro]g/m1 berberine, a concentration which is 1/8th the MIC for this compound. This extract that inhibited the cell growth in the presence of berberine but had no activity when added alone is likely to contain an MDR inhibitor namely 5'-MHC.
Objectives of synergistic combinations
The main objectives of the use of synergistic and potentiative drug combinations are the following:
a. The combination therapy is expected to exhibit greater efficacy than the monotherapy.
b. The combination of drugs is expected to reduce the dosage at equal or increased level of efficacy.
c. The combination of drugs may reduce or delay the development of drug resistance due to the inhibition of multiple pathways.
d. The combination therapy is expected to reduce the unwanted side effects but at the same time exhibit enhanced therapeutic action.
Synergistic/antagonistic interactions of natural products with clinically used anticancer and antimicrobial drugs
Cancer with more than 11 million death cases every year is the second leading cause of mortality. It is estimated that there will be 16 million new cases of death due to cancer each year by 2020. Death rate from this dreadful disease in the world is projected to rise, with an estimated 9 million people dying from cancer in 2015 and 11.4 million dying in 2030 (jemal et al., 2005). Given the fact that cancer is a multi-factorial disorder resulting in unlimited division of cells, remedial strategies that target tumor cells while causing fewer side effects on normal cells are desired. In such cases, chemical, biological and clinical informatics methods can be used to discover or design novel treatment strategies for specific tumor cell targeting, overcoming drug resistance and increasing protection of normal cells against antitumor drugs (Krishna and Mayer, 2000). Over 90% of the cancer death cases attributable to failure in chemotherapy are generally related to multidrug resistance (MDR). MDR refers to a series of events characterized by the property of drug resistant tumors exhibiting simultaneous resistance to a number of structurally and functionally unrelated chemotherapeutic agents. Occurrence of multidrug resistance (MDR) mechanisms in cancer has presented serious barrier for successful cancer treatment. Two mechanisms which play a key role in MDR are the amplified activity of efflux pumps, as the multidrug resistance proteins (MRPs) and the detoxification by phase II conjugating evizymes, such as glutathione 5-transferases and UDP-glucuronosyltransferases. A synergistic interaction between these two mechanisms, MRPs and phase II enzymes, in the field of MDR has been reported. Presenting solution to overcome MDR in tumor cells is the main consideration in chemotherapy of cancer, which mainly is focused on combinational strategy (Meijerman et al., 2008; Tsuruo, 2003).
Combinational drug therapy has a long history and roots in traditional Chinese medicines. Today, parallel to new advances in cancer chemotherapy, cancer combinational drug therapy has been well developed and wide ranges of scientific efforts are focused on that. More and more medicinal chemists are turning their attention to natural medicines in order to overcome 'more investment, less drugs' challenge in drug discovery. Natural medicines are thought to be a hidden treasure of unexplored new-entity drugs. Although, some promising drug candidates have been derived from natural medicines, but because of the fact that the efficacy of most natural medicines lies in the synergy of diverse components rather than a single component, it is a great challenge to find single-component new entity drugs from the natural medicine. Even if the drug molecule is isolated from the source, either it is not as active as expected or it can be toxic. In many other cases, even if the drug candidate is developed, but with the passage of time it develops resistance and becomes therapeutically inefficient. Therefore, understanding the synergistic mechanisms of natural medicines is of great significance to drug discovery (Gertsch, 2011; Junio et al., 2011; Li and Vederas, 2009).
The fact that synergy does exist in natural medicines can be detected, at least, at two levels. First, the various components present in a single medicinal agent might work synergistically. For example, the high antimicrobial potential of some medicinal plants (e.g. Berberis sp.) results from the synergy of antimicrobial agents (e.g. berberine) and multidrug resistance (MDR) inhibitors (e.g. 5'-methoxyhydnocarpin). Some mostly used anticancer natural medicines, such as Rhizoma Polygoni Cuspidati, Fructus Schisandrae Chinensis and Rhizoma Zingiberis Recens, also comprise anticancer agents (e.g. apigenin and limonene) and MDR inhibitors for cancer cells (e.g. quercetin and [beta]-elemene). Of course most of the MDR inhibitors have no biological activities (such as microbicidal or anticancer activities) on their own, they effectively potentiate the antimicrobial or anticancer effects of bioactive agents by avoiding them from being pumped out of the cells by the MDR pumps. Secondly, synergism can also be observed by combining various individual natural agents in a formula to enhance the therapeutic efficacy and/or reduce toxicity. Traditional Chinese Medicine (TCM), which has collected more than 100 000 formulae in the past 1500-2000 years best reveals the above fact. The synergistic mechanisms of some TCM formulae have been understood on at least preliminary level. For instance, the combination of Realgar (tetraarsenic tetrasulfide), Indigo Naturalis, Radix Salviae Miltiorrhizae and Radix Pseudostellariae has been proved to be effective in the treatment of human acute promyelocytic leukemia (APL). Through pinpointing the roles of active ingredients derived from Realgar, Indigo Naturalis and Radix Salviae Miltior-rhizae, the synergy of this formula basically has been elucidated. That is, tetraarsenic tetrasulfide (Fig. 7) directly attacks promyelocytic leukemia retinoic acid receptor a (PML-RARa) oncoprotein and promotes APL cell differentiation. The principal components of Indigo Naturalis and Radix Salviae Miltiorrhizae, which is indirubin and tanshinone 11A (Fig. 7), potentiate tetraarsenictetrasulfide-induced ubiquitination and degradation of PML-RARa. In addition, indirubin and tanshinone IIA enhance the expression of aquaglyc-eroporin 9, which helps transport tetraarsenic tetrasulfide into APL cells and, thus, augments its efficacy (Qiu, 2007; Wang et al., 2008).
Although the rational design of targeted drugs has made lot of progress
due to advances in genomics and cell biology, but the effects of these targeted drugs are not durable when they are used alone. This problem arises partially because agents directed at an individual target often exhibit limited efficacies, poor safety and resistance profiles. It is now well established that cellular pathways operate more like webs than highways. There are often multiple ways or alternate routes that may be switched on in response to the inhibition of a specific target. This facilitates the emergence of resistant cells or resistant organisms under the specific pressure of a targeted agent, resulting in drug resistance and clinical failure of the drug. To overcome this limitation of monotherapy, combination therapies involving more than one agents, are often required to effectively cure many malignant tumors and infectious diseases. This combinatorial approach to drug design has been supported by clinical successes with combination therapies and multi-target agents and attempts have been made in the direction of the discovery of new multicomponent therapies. Understanding the molecular mechanism of action of drug combinations involving synergistic and antagonistic interactions could pave the way for the discovery of novel effective combination therapies.
Natural products have always played a significant role in the drug discovery process. In particular, natural product combinations have been extensively investigated, clinically tested, and widely used in traditional, folk and alternative medicines. The novel multi-target mechanisms of natural product combinations may be valuable sources of information for developing the multi-target therapeutics. Various natural product combinations produce significantly better effects than equivalent doses of their components. Synergism effect could not only occur among natural compounds, but also between natural compounds and marketed chemotherapeutic agents. A list of recently used natural products in cancer combination therapy is given in Table 3. List of patents pertaining to use of natural products in cancer combination therapy is given in Table 4.
Anticancer enhancing effects of isoprenoid natural products from essential oils
The toxic effects of anticancer drugs during the treatment is a growing concern. The use of non-toxic potentiating agent or synergistic compound in combination with anticancer drugs may considerably potentiate their effectiveness while minimizing their toxicity. Most of the currently used cancer treatments employ a cocktail of various anticancer drugs acting in synergism. For example, MVAC protocol comprises of four drugs including methotrexate, vinblastine, cloxorubicin and cis-platin. But, each component of these types of cocktails is toxic as well as their combination (Blick et al., 2012). If a compound which itself is non-toxic but potentiates the efficacy of other anticancer drugs is identified, it can solve many problems of the clinical cancer treatment.
Such kinds of molecules are terpenes. Terpenes are a class of natural products having wide distribution in nature, mostly found in plants. [beta]-Caryophyllene, a well-known sesquiterpene present as a major component in various essential oils has been reported to enhance the efficacy of paclitaxel synergistically against MCF-7 cell lines (Legault and Pichette, 2007). Cancer cells have been reported to be sensitized to radiation therapy by using one or more monoterpenes or sesquiterpenes particularly perillyl alcohol. The cancer cell that was exposed to the terpenes has been reported to be more sensitive to the irradiation than a control cell that has not been exposed to the above treatment. Geranium oil in combination with paclitaxel slowed down the growth of human breast and lung cancer cells up to 87% (Rajesh et al., 2003). Geraniol, which is an acyclic dietary oxygenated monoterpene found in many essential oils has been reported to exert anti-tumor activity against various cancer cells both in vitro and in vivo. Geraniol inhibited Caco-2 cell growth by reducing DNA synthesis leading to a blockage of the cells in the S-phase of the cell cycle. Geraniol also increased the cytotoxicity of 5-FU and increased its uptake in human colon cancer cell lines. Geraniol caused a 2-fold reduction of thymidylate synthase and thymidine kinase expression in cancer cells (Carnesecchi et al., 2002). In another study, perillyl alcohol, which is a constituent of many essential oils, inhibited the growth of human breast cancer cells (MDA-MB-231, MDA-MB-435, and MCF-7) and induced apoptosis. The novel discovery in the study was that perillyl alcohol (IC20), methyl jasmonate ([IC.sub.20]), and cis-platin (1 [micro]M) in combination exhibited synergistic effects in growth inhibition in MDA-MB-435 and MDA-MB-231 cells. The [IC.sub.50] for cis-platin is 600 [micro]M and in combination with perillyl alcohol and methyl jasmonate, it decreases to 1 [micro]M, which is a 600-fold increase in sensitivity to cis-platin. In case of MDA-MB-231 cells in the presence of perillyl alcohol and methyl jasmonate, a 1200-fold increase in sensitivity to cis-platin was observed. Mechanistic study revealed that the combination treatment increased the TNFR1 expression and decreased mitochondrial membrane potential in MDA-MB-435 and MDA-MB-231 cells (Yeruva et al., 2010). Peril-lyl alcohol has also been reported to inhibit growth of cancer cells and induce apoptosis. Another report indicated that perillyl alcohol sensitized human myeloid U937 cells to pentoxifylline. Combination treatment of perillyl alcohol with pentoxifylline increased Bc1 and Bax expression as well as induced apoptosis. Perillyl alcohol has also been reported to sensitize prostate and malignant glioma cells to cisplatin/radiation via Fas mediated death receptor pathway (Rajesh and Howard, 2003; Rajesh et al., 2003). Malignant glioma cells preincubation with POH exhibited a concentration dependent sensitivity to cisplatin and doxorubicin. Essential oils have also been reported to exhibit synergistic effects with various chemotherapeutic anticancer drugs. For instance geranium oil has been reported to exhibit synergistic effects with some chemotherapeutic anticancer agents used in different cancers including breast, lung, ovary, colon, prostate, liver, kidney, neuroblastoma, leukemia, lymphoma, and other cancers.
Table 3 Natural products and medicinal extracts exhibiting synergism with anticancer agents. Source Natural product Cell line/in vivo model Cotton seeds (--)-Cossypol BxPC-3cell line Ferula Conferone MDCK-MDR1 cells schtschurowskiana Cimcifuga Cycloartane-type triterpenoids Human breast cancer racemosa cell line MCF-7 Solatium Coramsine Murine model of linnaeanum malignant mesothelioma Glycine max Genistein PC-3 human prostate cancer cell line Ailanthus 1 -Methoxy-canthin-6-one Human leukemia altissima (Jurkat), thyroid Swingle carcinoma (ARO and N PA) and hepatocellular carcinoma (HuH7) cell lines Rhizome of Alisma Alisol B 23-acetate(ABA) MDR cell lines orientate HepG2-DR and K562-DR Glycine max Genistein Pancreatic cancer cell lines BxPC-3 cell line Glycine max Genistein Human prostate cancer cells PC-3 cells Glycine max Genistein HeLa (cervical cancer), OAW-42 (ovarian cancer) and L929 (normal fibroblasts) Glycine max Genistein MCF-7 human B RCA cells Rosa sp. Geraniol Colonic cancer cell oil/Cymbopogon lines Caco-2 and martinii SW620 cells oi 1/Cymb op ogon nardus L oil Euphorbia Latilagascenes B Human MDR I gene lagascae transacted mouse lymphoma cells Azadiraclua Neem leaf preparation (NLP) Swiss mice indica diminishes leucopenia And peripheral blood mononuclear cells (PBMC) Panax notoginseng Notoginseng flower extract HCT-116 human (NGF) colorectal cancer cell line Epilohium Oenothein B (OeB) Prostate cancer angustifolium cells PC-3 Olea sp. Oil Oleic acid Breast cancer cells with HER-2/neu oncogene amplification Tanacetum Parthenolide Hs605T. MCF-7 partlienium (L.) Plant products Perillyl alcohol (POH) NSCLC, A549 and H520 cells Viris vinifera Proanthocyanidin K562. A549. CNE seeds cells, experimental transplantation Sarcoma 180 (SI80) and Hepatoma 22 (H22) in vivo Torilb japonica Torilin Multidrug-resistant KB-V1 and MCF7/ADR cells Tripterygiurn Triptolide (TPL) Ovarian cancer and wilfordii Nude mice Hook. Scutellaria Wogonin Jurkat and HL-60 baicalensis cells Georgi Celastnis Dihydro [beta]-agarofuran Human vulcanicola sesquiterpenes MDR1-transfected NIH-3T3 cells A plant alkaloid NSC77037 MDR ovarian cancer cells Camellia sinensis (--)-Epigallocatechin-3-gallate MCF-7Tam cells Source Anticancer agent Refs. Cotton seeds Genistein Mohammad et al. (2005) Ferula Vinblastine Barthomeuf et schtschurowskiana al.(2006) Cimcifuga Tamoxifen Gaube et racemosa al,(2007) Solatium CpG-containing Van der Most linnaeanum oligodeoxy et al.(2006) nucleotides Glycine max SB715992 Davis et al. (2006) Ailanthus Human recombinant Ammirante altissima etal. (2006) Swingle tumor necrosis factor related apoptosis inducing ligand (TRAIL) Rhizome of Alisma Vinblastine. Wang et orientate Puromycin. al.(2004) Paclitaxel, Aciinomycin D,5-Fluorouracil, Cisplatin, Verapamil Doxorubicin Glycine max Erlotinib El-Rayes et al.(2006) Glycine max Docetaxel Li etal. (2006) Glycine max Camptothecins Papazisiset al. [2006) Glycine max Tamoxi fen Mai et al.(2007) Rosa sp. 5-Fluorouracil(5-FU) Carnesecchi et oil/Cymbopogon al. (2002) martinii oi1/Cymbopogon nardus L oil Euphorbia Doxorubicin Duarte et al. lagascae (2007) Azadiraclua Cyclophosphamide Ghosh et al. indica (2006) Panax notoginseng 5-Fluorouracil Wang et al. (2007) Epilohium Arabinosylcytosine Kiss et al. angustifolium (2006) Olea sp. Oil Trasmzumab Menendez et (Herceptine) al. (2005) Tanacetum Parthenolide Wu et al. partlienium (L.) (2006) Plant products Cisplatin Yeruva et al. (2007) Viris vinifera Doxorubicin Zhang et seeds al.(2005) Torilb japonica Adriamycin. Kim et Vinblastine, al.(1998) Taxol and Colchicine Tripterygiurn Carboplatin Westfall et wilfordii al. (2008) Hook. Scutellaria Etoposide Lee et al. baicalensis (2007) Georgi Celastnis Verapamil Torres-Romero vulcanicola et al. (2009) A plant alkaloid Paclitaxel Susa et al. (2010) Camellia sinensis Tamoxifen Farabegoll et al.(2010) Table 4 List of recent patents pertaining to use of natural products in cancer combination therapy. Research area/ A brief synopsis of Year/patent Refs. title the patent number Preparation and This invention 2003/ Wang et al. composition of describes the US6617335 (2003) bis-benzyl- preparation of isoquinoline class various derivatives bis-isoquinoline alkaloids possessing multi- drug resistance (MDR) reversal activities and are used as sensitizing agents (potentiators) in cancer chemotherapy Synergistic This invention 2003/ Lee (2003) compositions and reports a US6537988 methods for pharmaceutical cancer treatment preparation for the synergistic treatment of cancer consisting one agent from antiproliferative cytotoxic class and the other from antiproliferative cytostatic class Methods and use of This invention 2003 / Horwitzetal. combination relates to the US65415Q9 (2003) chemotherapy for synergistic effects the treatment of of Taxol cancer in combination with discodermolide in treatment of cancer. Therapy for B cell This 2005 / Chan et al. disorders using invention describes US20050095243 (2005) combination a combination therapy approach therapy of anti-CD20 antibody with a BLyS antagonist for the treatment of cell based malignancies and B-cell regulated autoimmune diseases. Combination This invention 2007/ Sliwkowski and therapy of HER relates to the use US20070020261 Kelsey expressing of HER2- (2007) cancers dimerization inhibitors (HDIs) and EGER inhibitors for tumors expressing HER2 and EGFR Compounds for This invention 2007/ Ekstrom et al. potentiating and describes methods US20070264241 (2007) augmenting for using Cancer Therapy nucleoside analog prodrugs for augmenting cancer treatment. Composition and This invention is 2008/ Kim et al. methods for related to a US20080268072 (2008) enhancing the composition antiproliferative enriched effect of with 3-0- Pulsatillas radix [O-[alpha]-L- rhamnopyranosyl- (l-2)- [0-[beta]-D] Glucopyranosyl- (1-4)l-[alpha]- L- arabinopyranosyl hederagenin which shows strong antitumor by enzyme -reacting and extracting Pulsatiliac radix Development of This invention 2008/ Scott (2008) methods to relates to methods US20080026400 identify most which uses a potent bioactive specific agents and type of assay synergistic system, the combinations Multi-Pathway High Throughput Assay, in combination with a novel experimental strategy, in which repetitive cycles of experiments result in the identification of the most effective synergistic combinations of potential active agents from a library of substances (compounds). Method of using This invention 2008/ Adimooiam et histone demonstrates US20080153877 al. deacetylase advantageous (2008) inhibitors effects of a and monitoring combinational biomarkers in therapy including a combination histone therapy deacetylase inhibitor and another therapeutic agent as well forecasting the interval administration between them. Hixed drug This invention 2008/ Louie et proportions for describes a US20080I99515 al.(2008) treatment of pharmaceutical hematopoietic composition tumors and cancer comprising a fixed, disorders non-antagonistic molar ratio of cytarabine and an anthracycline for treating hematologic cancers or proliferative disorders. Methods and The invention 2008/ Borisyetal. screening system describes a method L1S20D80194421 (2008) for identifying for screening and Drug-drug identifying drug- interactions drug interactions using combinational arrays Combination cancer This invention 2008/ Xu et al. treatment with a demonstrates the US20080159980 (2008) GST-activated synergistic effects antiproliferative of molecule and effective dose a another anticancer GST-activated therapy anticancer compound and a therapeutically effective dose of another anticancer therapy Combination Cancer This invention 2008/ Arnold therapy relates to the US200802G7957 et al.(2008) evaluation of combinational therapy using an anti-cancer agent and an IGF1R inhibitor compound Phyto- This invention 2008/ Rangel and nutraceurical demonstrates that a US20080260771 Angel synergistic specific (2008) composition combination of for Prostate extracts of plants disorder(s) and nutraceuticals possess synergistic effects, with minimal side effects for treatment of prostate disorders Compositions and This invention is 2008/ Majeed (2008) methods to related to a U520080226571 provide composition enhanced containing photoprotection Labdane- against UV A and diterpenoids that UV induced provides better insult of human photo skin protection against both UV A and UV A radiations in the HaCaT human keratinocyte cell lines. Pharmaceutical This invention 2008/ Chu composition and relates to the US20O80113042 et al.(2008) method for Synergistic effects of cancer therapy geranium essential based on oil or its chemical combinational use constituents and of conventional a chemotherapeuric anticancer agents agent or plant and extract in geranium essential treatment of oil or compounds different cancers thereof Combination This invention 2008/ Moodleyand products relates to a US20080020018 Coulter pharmaceutical (2008) formulation comprising methyxanthine as one active agent, and at least containing a corticosteriod as another active agent Combination This invention 2009/ Johnstone et formulations of demonstrates useful U52G090074848 al. (2009) platinum agents effects of a and cytidine cytidine analogs analog and a platinum agent in augmenting therapeutic effects when are used in combination. Method for This invention 2009/ Lanzara determining drug- describes how to US200900127I7 (2009) molecular combine combinations that pharmaceutical or modulate and biological increase molecules or drugs the therapeutic in safety and order to prepare effectiveness of specific ratio pharmaceutical combinations that drugs are adjusted to increase the overall safety and therapeutic efficacy of the individual molecules or drugs while minimizing side effects and more economical. Fixed ratio drug This invention 2009/ Andrew et al. combination relates to the US20090023680 (2009) therapy for solid methods for tumors treating cancer by administering a pharmaceutical composition containing a fixed, non-antagonistic molar ratio of irinotecan and floxuridine Potentiator of The invention 2009/ Pichette and anticancer agents relates to using US20090286865 Legault in cancer essential oil (2009) treatment terpene or derivative thereof as a potentiator for increasing therapeutic effect of an anticancer agent (paclitaxel) Combinatorial anti This invention is 2009/ Buck et - cancer therapy related to US20090274698 al.(2009) combination therapy comprising an anti -cancer agent that increase pAkt levels in tumor cells and an mTOR inhibitor that binds to and directly inhibits both mTORCI and mTORC2 kinases Anticancer, This invention 2009/ Ricciardiello chemopreventive. relates to using US20090048187 et al. and compositions (2009) Anti- extracted inflammatory from pinoresinol- effects of rich Oka europaea pinoresinol- Caiazzana olives rich in treating cancer olives and its synergic effects with a polyphenols composition isolated from Oka europaea Caiazzana olives. Cucurbitacin 8 and This invention 2009/ Xie et al. its uses demonstrates US20090247495 (2009) methods for preventing or treating various disorders by administering compounds consisting cucurbitacin A Flavopereirine and This invention 2009/ Hail and alstonine presents a method US20090215853 Beljanski cocktails in the for administration (2009) of treatment and an effective dose prevention of of a cocktail of prostate cancer flavopereirine and alstonine for preventing prostate cancer and/or decreasing PSA levels. Cancer therapy This invention 2009/ Shuangand Ci relates to US2D09008240G (2009) presenting a method of regulating the cell cycle and treating cancer with a peroxisome proliterator- activated receptor agonist and a mevalonate pathway inhibitor.
Role of synergism in the antimicrobial properties of essential oils (E0s)
The antimicrobial properties of E0s have been reported in several studies. In many cases the activity results from the complex interactions between the different classes of compounds such as phenols, aldehydes, ketones, alcohols, esters, ethers or hydrocarbons found in E0s. Though in some cases, the bioactivities of E0s are closely related with the activity of the main components of the oils. Several studies have found that a number of these compounds exhibited significant antimicrobial properties when tested separately. It has been reported that E0s containing aldehydes or phenols, such as cinnamaldehyde, citral, carvacrol, eugenol or thymol as major components showed the highest antibacterial activity, followed by E0s containing terpene alcohols (Bakkali et al., 2008: Burt, 2004). Other E0s, containing ketones or esters, such as [beta]-myrcene, [alpha]-thujone or geranyl acetate had much weaker activity. While volatile oils containing terpene hydrocarbons were usually inactive. Different terpenoid components of E0s can interact to either reduce or increase antimicrobial efficacy. The interaction between EO compounds can produce four possible types of effects: indifferent, additive, antagonistic, or synergistic effects. Interestingly, phenolic monoterpenes and phenylprcpanoids (typically showing strong antimicrobial activities) in combination with other components were found to increase the bioactivities of these mixtures. Most of the studies have focused on the interaction of phenolic monoterpenes (thymol, carvacrol) and phenylpropanoids (eugenol) with other groups of components, particularly with other phenols, phenylpropanoids and monoterpene alcohols, while monoterpenes and sesquiterpenes hydrocarbons were used to a lesser extent (Table 5). The combination of phenolics with monoterpene alcohols produced synergistic effects on several microorganisms, in particular, the combination of phenolics (thymol with carvacrol, and both components with eugenol) were synergistically active against E. coli strains. Though other reports have observed additive and antagonism effects (Ben Arfa et al., 2006; Cox et al., 2001; Hammer et al., 1999; Juliani et al., 2002; Lambert et al., 2001) (Table 5).
Table 5 Combination of essential oils and components and their antimicrobial interactions against several microorganisms. Pair combinations Organism Methods Thymol/carvacrol Staphylococcus aureus Half dilution Pseudomonas aeruginosa Escherichia coli Checkerboard S. aureus. Bacillus Checkerboard cereus E coli S. aureus, P. Mixture aeruginosa E. coli Checkerboard Thymol/eugenol E. coli Checkerboard Carvacrol/eugenol E. coli Checkerboard S. aureus. B. cereus, E Checkerboard coli Carvacroi/cymene B. cereus Mixture Carvacrol/linalool Listeria monocytogenes. Checkerboard Eugenol/linalool Enterobacter Eugenol/menthol aerogenes. E. coli. P. aeruginosa Cinnamaldehyde/eugenol Staphylococcus sp., Mixture Micrococcus sp., Bacillus sp., and Enterobacter sp. 1.8-Cineole/aromadendrene Methieillin-resisranr Checkerboard S. aureus (MRSA) and vancomycin-resistant enterococci (VRE) Enterococcus faecalis [alpha]-Pinene/limonene Saccliaromyces Checkerboard cerevisiae [alpha]-Pinene/linalaol L monocytogenes. Mixture Linalool/terpinen-4-ol 0. Yersinia vulgare/Rosmarinus enterocolitica. officinalis 0. Aeromonas hydrophilla. vulgare/T. vulgaris P. fluorescens Lippia multiflora/Mentha E. coli. E. aerogenes, Checkerboard piperita L Enterococcus imiltifiora/O. basiiicum faecalis, L. M. monocytogenes. P. piperita/0, basilicum aeruginosa, Salmonella enterica, S. typhimurium. Shigella, dysenteriae, S. aureus E. coli. E. aerogenes, E. faecalis. L monocytogenes Pair combinations Interaction References Thymol/carvacrol Additive Lambert etal. (2001) Synergism Pei etal. (2009) Antagonism Additive Lambert et al. (2001) Additive Rivas etal. (2010) Thymol/eugenol Synergism Pei et al.(2009) Carvacrol/eugenol Synergism Pei et al. (2009) Antagonism Carvacroi/cymene Synergism Ultee et al,(2000) Carvacrol/linalool Synergism Bassole etal. Eugenol/linalool (2010) Eugenol/menthol Cinnamaldehyde/eugenol Additive Moleyarand Narasimham (1992) 1.8-Cineole/aromadendrene Additive Mulyaningsih et al.(2010) [alpha]-Pinene/limonene Synergism, Tserennadmid additive et al. (2011) [alpha]-Pinene/linalaol Synergism Linalool/terpinen-4-ol 0. vulgare/Rosmarinus officinalis 0. vulgare/T. vulgaris Lippia multiflora/Mentha Synergism, Bassole etal. piperita L additive (2010) imiltifiora/O. basilicum M. piperita/0, basiiicum
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0944-7113/$--see front matter [c] 2013 Elsevier GmbH. All rights reserved.
Manzoor A. Rathera(a), *, Bilal A. Bhata (a), ** Mushtaq A. Qurishi (b)
(a) Medicinal Chemistry Division, Indian Institute of Integrative Medicine, Sanat Nagar, Srinagar, India
(b) Department of Chemistry, University of Kashmir, Srinagar, India
* Corresponding author. Tel.: +91 9622452835.
* Corresponding author.
E-mail addresses: firstname.lastname@example.org, email@example.com (M.A. Rather). firstname.lastname@example.org (B.A. Bhat).
Contents Is the drug combination strategy the future of drug discovery? 2 Switching over from single drug to multi-drug therapy 2 Existing multi-target therapeutics 2 What synergy means in phytomedicine? 3 Definition and proof of synergy 4 Combination index (CI) 4 Experimental evidence in favor of synergism 5 Example 1. Marihuana (Cannabis sativa) 5 Example 2. St. John's Wort (Hypericum perforatum) 6 Example 3. Iberogast[R] (a phytopreperation of 6 nine plant extracts) Example 4. Kava Kava (Piper methysticum) 6 Example 5. Antimicrobial action of berberine 6 potentiated by 5'-methoxyhydnocarpin (5'-MHC) Objectives of synergistic combinations 6 Synergistic/antagonistic interactions of natural products 8 with clinically used anticancer and antimicrobial drugs Anticancer enhancing effects of isoprenoid natural 9 products from essential oils Role of synergism in the antimicrobial properties of 10 essential oils (E0s) References 13
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|Author:||Rather, Manzoor A.; Bhat, Bilal A.; Qurishi, Mushtaq A.|
|Publication:||Phytomedicine: International Journal of Phytotherapy & Phytopharmacology|
|Date:||Dec 15, 2013|
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