Molecular Understanding and Modern Application of Traditional Medicines: Triumphs and Trials

Leading Edge Essay

Timothy W. Corson1 and Craig M. Crews1,2,* 1Department of Molecular, Cellular & Developmental Biology, Yale University, New Haven, CT 06511, USA 2Departments of Chemistry and Pharmacology, Yale University, New Haven, CT 06511, USA *Correspondence: [email protected] DOI 10.1016/j.cell.2007.08.021

Traditional medicines provide fertile ground for modern drug development, but first they must pass along a pathway of discovery, isolation, and mechanistic studies before eventual deployment in the clinic. Here, we highlight the challenges along this route, focusing on the compounds artemisinin, triptolide, celastrol, capsaicin, and curcumin.

Traditional medicines continue to Peruvian Indians to suppress shiver- interesting biological properties. But provide front-line pharmacotherapy ing and used since the 17th century perhaps most importantly, parent for many millions of people world- in the treatment of malarial fevers extracts have been “clinically” tested wide. Although their application is (Greenwood, 1992). Similarly, aspirin in their traditional milieu, in some often viewed with skepticism by the was derived from salicylic acid in the cases over millennia. Western medical establishment, bark of the willow tree (Salix species), Despite these advantages, the path medicinal extracts used in ancient used traditionally to treat fever and from traditional medicine to Western medical traditions such as Ayurveda inflammation in many cultures world- pharmaceutical is fraught with chal- on the Indian subcontinent and tra- wide for at least four millennia (Mahdi lenges. Here, we discuss the chal- ditional Chinese medicine (TCM) are et al., 2006). The successes of these lenges of each of the four steps in this a rich source of therapeutic leads for two early “blockbuster” drugs set pipeline (see Figure 1): Western “dis- the pharmaceutical industry. the stage for ongoing drug discovery covery” of a traditional medicine, iso- The transformation of traditional efforts from traditional medicines. lation and/or synthesis of the active medicines into modern drugs has its Compounds derived from medici- component, elucidation of the molec- origins in the archetypal examples of nal extracts are appealing for several ular mechanism, and development the antimalarial quinine and the anti- reasons (Schmidt et al., 2007). They as a pharmaceutical. We focus on pyretic analgesic aspirin. The alkaloid are often stereochemically complex, five interesting and timely examples quinine was isolated in 1820 from multi- or macrocyclic molecules with derived from traditional medicines in the bark of several species of Cin- limited likelihood of prior chemi- varied therapeutic classes, each at chona, thought to have been used by cal synthesis, and they tend to have a different stage in the development

Figure 1. The Route from Traditional Medicine to Modern Drug Shown are five traditional medicines—artemisinin, triptolide, celastrol, capsaicin, and curcumin—and the points in the pathway from ancient remedy to modern drug where they face the biggest hurdles.

Cell 130, September 7, 2007 ©2007 Elsevier Inc. 769 Table 1. Five Traditional Medicines in Clinical Trials Clinical Trials Compound Disease Number Principal Sponsors Artemisinin Malaria 81 31 charities, institutes, universities, and companies based in Austra- lia, Austria, Belgium, Colombia, Ethiopia, France, Gambia, Germany, Ghana, Guinea-Bissau, the Netherlands, Papua New Guinea, South Africa, Sudan, Sweden, Switzerland, the United Kingdom, and the USA; and working in numerous Asian, African, and South American locations Cytomegalovirus 1 Hadassah Medical Organization, Israel infection Schistosomiasis 1 Dafra Pharma, Belgium Triptolide & Celastrol Rheumatoid 1 National Institute of Arthritis and Musculoskeletal and Skin Diseases, (T. wilfordii extract) arthritis MD, USA Capsaicin Chronic pain 13 NeurogesX, CA, USA; AlgoRx Pharmaceuticals, NJ, USA Postoperative pain 5 National Institute of Dental and Craniofacial Research, MD, USA; AlgoRx Pharmaceuticals, NJ, USA Radiation-induced 1 North Central Cancer Treatment Group, MN, USA mucositis Alopecia areata 1 University of Minnesota, USA Morton’s neuroma 1 AlgoRx Pharmaceuticals, NJ, USA Osteoarthritis 1 Khon Kaen University, Thailand Interstitial cystitis 1 National Institute of Diabetes and Digestive and Kidney Diseases, MD, USA Curcumin Colon cancer 6 Chao Family Comprehensive Cancer Center, CA, USA; Tel-Aviv Sourasky Medical Center, Israel; Johns Hopkins University, MD, USA; University of Michigan Comprehensive Cancer Center, USA; University of Pennsylvania, USA; University of Medicine and Dentistry, NJ, USA Pancreatic cancer 3 Rambam Medical Center, Israel; M.D. Anderson Cancer Center, TX, USA; Tel-Aviv Sourasky Medical Center, Israel Alzheimer’s disease 2 John Douglas French Foundation, CA, USA; Chinese University of Hong Kong Chemotherapy- 1 Hadassah Medical Organization, Israel induced mucositis Multiple myeloma 1 M.D. Anderson Cancer Center, TX, USA Psoriasis 1 University of Pennsylvania, USA Cystic fibrosis 1 Seer Pharmaceuticals, CT, USA Includes registered, open, closed, terminated, and completed trials of these compounds, parent extracts, or derivatives. For details see www.clinicaltrials.gov. Data current as of August 21, 2007. process, highlighting successes and least the fourth century CE, it has radicals are currently thought to be roadblocks on the path to status as a been used in the treatment of fever responsible for artemisinin’s antima- Western drug. attributed to malaria. This long history larial activity. The classic method of of use prompted Chinese researchers cell fractionation after treatment with Artemisinin: Production Problems to seek the active antimalarial prin- radiolabeled artemisinin has identi- The antimalarial artemisinin (and ciple; artemisinin was isolated and fied numerous cellular constituents derivatives) represents one of the its structure determined in the mid alkylated by artemisinin (Asawamaha- greatest recent clinical success sto- 1970s (Liu et al., 2006). sakda et al., 1994); the strongest vali- ries arising from a traditional medi- Artemisinin, an endoperoxide ses- dated target for artemisinin is PfATP6, cine, echoing the success of quinine quiterpene lactone with a complex the Plasmodium sarco-endoplas- two centuries earlier. Artemisinin polycyclic ring structure, is modified mic reticulum Ca2+ ATPase (SERCA), (see Figure 1) is derived from Arte- by Fe2+ ions to structures containing which is inhibited by artemisinin (Eck- misia annua L., the sweet wormwood carbon-centered free radicals. Given stein-Ludwig et al., 2003). (qinghao), a shrub first documented that the intracellular environment Clinical studies, initiated in the in TCM in 168 BCE as a hemorrhoid of the Plasmodium malaria parasite 1970s prior to any mechanistic treatment (Liu et al., 2006). Since at is rich in this ion from heme, these insights into artemisinin function,

770 Cell 130, September 7, 2007 ©2007 Elsevier Inc. demonstrated that artemisinin and A. annua itself. The plant has proven since then (Yang et al., 1998 and ref- its derivatives are powerful antima- genetically tractable: Several of the erences therein). Like artemisinin, larials. They have proved particularly isoprenoid biosynthetic enzymes however, triptolide is currently derived effective for treating severe malaria necessary for artemisinin produc- from its plant of origin with low yield: and, in combination with traditional tion have been cloned, and A. annua 6–16 ng/g in one study (Brinker and antimalarials, for combatting Plas- can be successfully transformed with Raskin, 2005). Little work has been modium drug resistance. Combina- Agrobacterium tumefaciens to over- done to investigate biotechnological tion therapies containing artemisinin express key biosynthetic genes (Liu routes to triptolide production, which are now considered the treatment et al., 2006). are important to reduce reliance on of choice for malaria in Asia, with Perhaps the most promising strat- the natural source. Moreover, contin- growing adoption in Africa (see Table egy is the use of microbes to produce ued development of derivatives of trip- 1 for information on clinical trials). artemisinin. In a triumph of genetic tolide such as the succinyl sodium salt Artemisinin may also have efficacy engineering, Ro et al. combined PG490-88 will be valuable to improv- against other parasites and as an genetic activation of the endoge- ing the solubility and side-effect pro- anticancer compound, possibly act- nous mevalonate isoprenoid synthe- file of this compound (Tao and Lipsky, ing via antiangiogenic and proapop- sis pathway with introduction of A. 2000). totic mechanisms in the latter case annua genes to produce artemisinic Determination of triptolide’s cellu- (Efferth, 2007). acid in the budding yeast Saccha- lar target has proven to be an even Despite these dramatic findings, romyces cerevisiae (Ro et al., 2006). greater challenge. This is not unusual: widespread deployment of artemis- This precursor compound, which can Many a promising therapeutic natural inin has been hindered by produc- be readily converted to artemisinin product has faltered when no clear- tion difficulties. Although a dozen in the laboratory, is secreted in large cut mechanism of action could be synthetic routes to artemisinin have quantities from the yeast. Such cre- identified. Although progression into been described, all are complex and ative strategies, leveraging the power the clinic without such knowledge is low yielding, rendering them eco- of genetics and in vivo biochemistry, possible, as was the case with arte- nomically unfeasible (Liu et al., 2006). can provide a valuable counterpart misinin, a solid knowledge of molec- Synthetic chemistry has, however, to synthetic chemistry and natural ular mechanism (ideally at the struc- offered semi-synthetic artemisinin sources in the production of natural tural, not just the molecular, level) derivatives with improved solubil- product medicines. allows medicinal chemists to perform ity (such as sodium artesunate) and rational derivatization to improve stability (such as artemether) (Efferth, Triptolide and Celastrol: affinity, specificity, pharmacoki- 2007). Even a totally synthetic trioxo- Harnessing the Power of the netics, and stability. Knowledge of lane compound RBX11160 (OZ277), Thunder God Vine mechanism can also potentially lead inspired by the trioxane endoperox- Trypterygium wilfordii Hook F., the to more specific clinical trials and, in ide moiety of artemisinin, has shown “thunder god vine” (lei gong teng), cases like triptolide, completely new promise as an antimalarial (Venner- is another TCM. This vine has been insights. strom et al., 2004). used traditionally for the treatment A large body of work describes Artemisinin for clinical use is pre- of arthritis and other diseases, and triptolide’s inhibitory effects on tran- dominantly produced naturally in it is the source of several biologically scription mediated through NF-κB A. annua plants. Despite efforts to active secondary metabolites (Tao and NFAT (Qiu and Kao, 2003), but maximize agricultural production, the and Lipsky, 2000). Some of its TCM until recently, direct cellular targets artemisinin content in plant extracts uses might rely on the presence of were elusive. Nonetheless, careful varies widely due to environmental multiple active components, and clin- cell fractionation with [3H]-triptolide conditions: 0.01%–0.8% dry weight ical studies have been performed on enabled identification of the Ca2+ (Efferth, 2007). This in turn makes the extracts of the plant (Table 1), rather channel polycystin-2 (encoded by the drug itself expensive—particularly than on a single compound (Tao and PKD2 gene) as a possible triptolide- problematic for an antimalarial, which Lipsky, 2000). However, substantial binding protein (others also likely is needed in large quantities in many work has focused on two major bio- exist) (Leuenroth et al., 2007). PKD2 poorer countries. Cell and plantlet active constituent compounds: trip- or the gene encoding its activator, cultures are an appealing alterna- tolide and celastrol (Figure 1). PKD1, causes polycystic kidney dis- tive source of this compound as Triptolide is a diterpenoid epoxide ease (PKD) when mutated because they can be grown under much more with a staggering variety of docu- entry of Ca2+ ions is essential for closely controlled conditions than mented cellular effects. Along with growth arrest of epithelial cells form- whole plants. Indeed, useful yields of anti-inflammatory activity, it shows ing the kidney tubule. Because trip- the compound can be produced by anticancer, immunosuppressive, and tolide activates opening of the poly- feeding cultures artemisinin precur- antifertility effects (Qiu and Kao, 2003). cystin-2 channel, it could potentially sors (Liu et al., 2006). An alternative It was isolated in 1972, and several complement loss of PKD1. This is the approach is the genetic engineering of synthetic routes have been described case in a mouse model of polycys-

Cell 130, September 7, 2007 ©2007 Elsevier Inc. 771 tic kidney disease in which the mice Tarahumara Indians as a remedy for The cloning of TRPV1 kick started lack Pkd1 (Leuenroth et al., 2007). coughs and bronchitis. Similar uses the field of pain receptor pharmacol- Thus, this calcium-dependent activ- plus anti-inflammatory and gastroin- ogy. Numerous pharmaceutical com- ity of triptolide, which is unrelated to testinal applications were adopted in panies are developing both TRPV1 its transcriptional repression activity India after the Portuguese imported antagonists (to block nociception (Leuenroth and Crews, 2005), opens chili peppers in the late 15th century. directly) and agonists (to desensitize a new therapeutic avenue for pursu- In Africa, they are traditionally used nociceptors, as with capsaicin) (Immke ing triptolide, in addition to its effects internally and externally as antisep- and Gavva, 2006). Resiniferatoxin, on the immune and reproductive sys- tics (Dasgupta and Fowler, 1997). another traditional medicine from the tems and in cancer. However, modern usage of capsaicin latex of Euphorbia resinifera, is one Highlighting the complexity of is focused on the treatment of various such agonist with higher potency than plant extracts, the pentacyclic triter- types of pain (see below) and also capsaicin (Immke and Gavva, 2006). pene celastrol (Figure 1) is structur- in the treatment of detrusor hyper- Efforts continue to create TRPV1 ago- ally a very different component of T. reflexia, a form of urinary inconti- nists with better skin permeation and wilfordii with a divergent therapeu- nence (Dasgupta and Fowler, 1997). lacking the distinctive side effect of a tic profile. Celastrol (also known as High-dose oral capsaicin also has burning sensation on application. tripterine) is extracted in small quan- anticancer properties in some animal Capsaicin itself has been used tities from T. wilfordii or other mem- model studies but seems to be a can- clinically with moderate success as a bers of the Celastraceae (bittersweet) cer promoter in others. topical treatment for the pain of rheu- family. To our knowledge, no total Compared with artemisinin, trip- matoid and osteoarthritis, psoriasis, synthesis or alternative production tolide, and celastrol, capsaicin is diabetic neuropathy, and posther- routes have been reported. chemically quite simple (Figure 1). It petic neuralgia (Table 1), but herein Although not yet tested as a single was purified and named in the 19th lies the particular challenge with agent in humans (Table 1), celastrol century and first synthesized in the this molecule: The chronic pain dis- has shown promise as an anti-inflam- 1920s (Dasgupta and Fowler, 1997). orders are notoriously idiosyncratic, matory compound in animal models But the widespread cultivation of and not all patients or all pain syn- of arthritis, lupus, amyotrophic lateral Capsicum makes synthesis unneces- dromes respond to capsaicin (Immke sclerosis, and Alzheimer’s disease sary, as large quantities of capsaicin and Gavva, 2006). The somewhat (Sethi et al., 2007 and references can easily be extracted from readily vague and diffuse traditional uses of therein). It also has antiproliferative available peppers. this compound offer little assistance effects against numerous cancer cell The mechanism of capsaicin in pain here, unlike artemisinin, for instance. lines. Several molecular mechanisms induction has been the topic of much Thus, testing for capsaicin efficacy have been identified for these effects, neurophysiological research (Cor- is a matter of clinical trial and error, including gene expression modulation tright et al., 2007). Capsaicin, along largely undermining the “tried and likely mediated through inhibition of with thermal heat, directly activates true” advantage of a traditional medi- NF-κB via TAK1 and IκBα kinase (Sethi nociceptors in the skin, the sensory cine. The major clinical advantage et al., 2007 and references therein), neurons responsible for the sensation that capsaicin holds over other unre- proteasome inhibition, topoisomerase of pain, with the subsequent release lated pain drugs under development II inhibition, and heat shock response of the neurotransmitter substance is its approved status as a foodstuff. activation (Hieronymus et al., 2006 P. Capsaicin’s therapeutic effect on and references therein). Nonethe- pain is due to the desensitization and Curcumin: Awaiting Targets and less, direct targets remain elusive. eventual destruction of nociceptors Outcomes As celastrol and triptolide move into following repeated capsaicin expo- Like capsaicin, the polyphenol cur- human studies, it will be vital not only sure. In a classic example of expres- cumin (Figure 1) is best known as a to better understand their mecha- sion cloning, Caterina et al. identified spice constituent: It is the yellow pig- nisms of action but also to investigate the capsaicin receptor (Caterina et ment component of the curry spice any potential synergistic effects of the al., 1997). Capsaicin was known to turmeric (Curcuma longa, known as two compounds, both at the cellular cause Ca2+ ion influx into nociceptors, haldi in Hindi). It is also, however, a and organismal levels. so these authors transfected a noci- drug used in Ayurveda and TCM in ceptor cDNA library into nonexcit- the treatment of diseases as diverse Capsaicin: Painless for Some? able HEK293 cells and screened for as rheumatism, fever, intestinal dis- Used worldwide, the alkaloid cap- capsaicin-dependent Ca2+ ion influx. orders, trauma, and amenorrhea (see saicin is the main cause of the “hot” The receptor they cloned, now known the Analysis by S. Singh on page 765 sensation associated with chili pep- as TRPV1, is a Ca2+ ion channel that of this issue). Modern research has pers, members of the genus Capsi- also responds to, and integrates, sig- attributed anti-inflammatory, immuno- cum. Beyond their widespread use nals from piperine (the irritant in black modulatory, antimalarial, and antican- as a spice, chili peppers were used pepper), protons, and other noxious cer effects to this multitalented com- in the Americas by the Aztecs and stimuli (Caterina et al., 1997). pound (Aggarwal et al., 2007).

772 Cell 130, September 7, 2007 ©2007 Elsevier Inc. Like capsaicin, synthesis of cur- compared to rigorous clinical trials, or develop alternative processes cumin is trivial and was first reported has slowed the formal validation of such as cell culture or transgenesis in 1910, but sufficient quantities of curcumin. This is confounded by lim- to enable useful-scale production. curcumin for therapeutic use are ited pharmaceutical company inter- Despite continuing advances in syn- available from the spice. This is par- est because curcumin itself is not thetic chemistry, the very complex- ticularly important as low bioavail- patentable (although synthetic meth- ity of many natural products that is ability of the parent compound cou- ods, derivatives, and pharmaceutical responsible for their desirable bio- pled with rapid intestinal metabolism formulations are) and by the percep- logical function can make production dictates large doses for clinical use tion that, as a foodstuff, curcumin difficult. (Sharma et al., 2005); derivatization of is more a nutraceutical (perhaps a With a reliable supply of compound the natural product is actively being dietary cancer preventative) than a available, biologists can then iden- pursued. traditional drug. This perception can tify and validate cellular targets and Given its pleiotropic clinical effects, only be changed by clinical studies mechanisms of action. New tools are it is perhaps not surprising that cur- showing successful disease treat- sorely needed for this particularly cumin has documented effects on ment with curcumin. Phase I studies daunting challenge, such as meth- countless intracellular signaling path- have documented tolerance up to ods that compare the phenotypic or ways. Its anti-inflammatory action 8000 mg/day, allowing a large dose- gene expression profiles induced by can be attributed largely to its inhibi- response range to be tested in phase a small molecule to those induced by tion of NF-κB activity, COX-2 and 5- II studies, several of which are under- known compounds (Hieronymus et LOX expression, and cytokine release way for the treatment of cancer, pso- al., 2006) or chemical enhancer/sup- (Aggarwal et al., 2007). Curcumin riasis, and Alzheimer’s disease (Table pressor screens. Development of in may directly target IκBα kinase to 1) (Hsu and Cheng, 2007). We must silico tools to “dock” small molecules block NF-κB. It also binds to a num- await the outcomes of these studies with protein structures to provide ber of other proteins, including thio- before curcumin can be validated as models for testing in vitro will likely redoxin reductase, several kinases, a pharmaceutical. come into their own with advances and several receptors (Aggarwal et in structural genomics, as sufficient al., 2007). The challenge here, then, Ongoing Challenges computational power becomes avail- as with many other natural products, An effective drug should be facile and able. is deciphering which of these targets economical to produce and deliver, Ideally with a mechanism in hand, is mechanistically valid for which bio- should display favorable absorption, clinicians must then test the com- logical activity. With such a broad distribution, metabolism, excretion, pound in the disease of interest spectrum of potential targets and and toxicity (ADMET) characteristics, (Table 1) while keeping an open mind activities described for curcumin, this and should treat the targeted disease for unexpected therapeutic activities is no easy task. Synthesis of deriva- with specificity and efficacy. Tradi- and working with medicinal chemists tives that selectively ablate certain tional medicines, as with other natu- to produce derivatives with improved cellular and/or therapeutic effects ral products, can offer powerful leads ADMET properties. Finally, regulatory is one possible route to tease apart for therapeutic development because approval must be obtained, as with all this mechanism-function conun- (unlike synthetic libraries) they drugs. This is particularly problem- drum, perhaps in concert with radio- already have documented effects on atic if the active principle is an extract labeled fractionation experiments (as the organism. However, the process or mixture, rather than an isolated described above) or affinity chroma- from plant to product is a slow one. compound; the U.S. Food and Drug tography with immobilized curcumin. Despite the oft-shared limitations Administration has been understand- The very versatility that makes cur- noted here, these five examples of ably reluctant to approve multiple- cumin appealing has also limited its traditional medicines are exceptional agent drugs until recently (Schmidt et rigorous clinical testing: There are in the extent to which they have been al., 2007). Only in 2006 was the first wide-ranging efficacy reports, but studied and the success they have such drug approved: Polyphenon E most are based on preclinical, anec- achieved in the clinic; countless other (MediGene), a topical antiviral pre- dotal, or pilot studies rather than on promising compounds wallow in pared from catechins extracted from randomized, placebo-controlled, obscurity. green tea (Camellia sinensis). double-blind trials (Hsu and Cheng, The challenges are formidable (Fig- Artemisinin, triptolide, celastrol, 2007). Activity has been reported in ure 1): Ethnopharmacologists must capsaicin, and curcumin are “poster several inflammatory and autoim- identify a medicine, its uses, and children” for the power and promise mune diseases and numerous can- active components. These efforts are of turning traditional medicines into cers, both as a preventative agent urgent, as traditional knowledge— modern drugs. However, their stories and treatment, alone or in combina- and traditional plant species—are highlight the ongoing interdisciplinary tion (Hsu and Cheng, 2007). The rela- being lost at an alarming rate. Chem- research efforts that continue to be tive ease and rapid payoff of under- ists must then synthesize the com- necessary to realize the pharmaceuti- taking preclinical or pilot studies, pound using a cost-effective method cal potential of traditional therapeutics.

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