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Supplementary Table S3 - PDF file 54K, Binding constant between CH5126766 and non-phosphorylated MEK1. Non-phosphorylated His6-MEK1 (WT) was imobilized and affinities between His6-MEK1 and MEK inhibitors were analyzed with SPR

Tumors with mutant RAS are often dependent on extracellular signal–regulated kinase (ERK) signaling for growth; however, MEK inhibitors have only marginal antitumor activity in these tumors. MEK inhibitors relieve ERK-dependent feedback inhibition of RAF and cause induction of MEK phosphorylation. We have now identified a MEK inhibitor, CH5126766 (RO5126766), that has the unique property of inhibiting RAF kinase as well. CH5126766 binding causes MEK to adopt a conformation in which it cannot be phosphorylated by and released from RAF. This results in formation of a stable MEK/RAF complex and inhibition of RAF kinase. Consistent with this mechanism, this drug does not induce MEK phosphorylation. CH5126766 inhibits ERK signaling output more effectively than a standard MEK inhibitor that induces MEK phosphorylation and has potent antitumor activity as well. These results suggest that relief of RAF feedback limits pathway inhibition by standard MEK inhibitors. CH5126766 represents a new type of MEK inhibitor that causes MEK to become a dominant-negative inhibitor of RAF and that, in doing so, may have enhanced therapeutic activity in ERK-dependent tumors with mutant RAS. Cancer Res; 73(13); 4050–60. ©2013 AACR.

Supplementary Figure S1 - PDF file 70K, Induction of p27Kip1 protein and cell cycle arrest by CH5126766

Supplementary Materials and Methods - PDF file 65K, Antibodies and kinase panel assays

Phosphoinositide 3-kinase (PI3K) is a promising anti-cancer target, because various mutations and amplifications are observed in human tumors isolated from cancer patients. Our dihydropyrrolopyrimidine derivative with a phenylurea moiety showed strong PI3K enzyme inhibitory activity, but its pharmacokinetic property was poor because of lack of solubility. Herein, we report how we improved the solubility of our PI3K inhibitors by introducing a solubilizing group and ortho substituents to break molecular planarity.

Substituting a carbon atom with a nitrogen atom (nitrogen substitution) on an aromatic ring in our leads 11a and 13g by applying nitrogen scanning afforded a set of compounds that improved not only the solubility but also the metabolic stability. The impact after nitrogen substitution on interactions between a derivative and its on- and off-target proteins (Raf/MEK, CYPs, and hERG channel) was also detected, most of them contributing to weaker interactions. After identifying the positions that kept inhibitory activity on HCT116 cell growth and Raf/MEK, compound 1 (CH5126766/RO5126766) was selected as a clinical compound. A phase I clinical trial is ongoing for solid cancers.

A novel series of 2-amino-1,3,5-triazines bearing a tricyclic moiety as heat shock protein 90 (Hsp90) inhibitors is described. Molecular design was performed using X-ray cocrystal structures of the lead compound CH5015765 and natural Hsp90 inhibitor geldanamycin with Hsp90. We optimized affinity to Hsp90, in vitro cell growth inhibitory activity, water solubility, and liver microsomal stability of inhibitors and identified CH5138303. This compound showed high binding affinity for N-terminal Hsp90α (Kd=0.52nM) and strong in vitro cell growth inhibition against human cancer cell lines (HCT116 IC50=0.098μM, NCI-N87 IC50=0.066μM) and also displayed high oral bioavailability in mice (F=44.0%) and potent antitumor efficacy in a human NCI-N87 gastric cancer xenograft model (tumor growth inhibition=136%).

A facile methodology effective in obtaining a set of compounds monofluorinated at various positions (fluorine scan) by chemical synthesis is reported. Direct and nonselective fluorination reactions of our lead compound 1a and key intermediate 2a worked efficiently to afford a total of six monofluorinated derivatives. All of the derivatives kept their physicochemical properties compared with the lead 1a and one of them had enhanced Raf/MEK inhibitory activity. Keeping physicochemical properties could be considered a benefit of monofluorinated derivatives compared with chlorinated derivatives, iodinated derivatives, methylated derivatives, etc. This key finding led to the identification of compound 14d, which had potent tumor growth inhibition in a xenograft model, excellent PK profiles in three animal species, and no critical toxicity.

Tumors with mutant RAS are often dependent on extracellular signal–regulated kinase (ERK) signaling for growth; however, MEK inhibitors have only marginal antitumor activity in these tumors. MEK inhibitors relieve ERK-dependent feedback inhibition of RAF and cause induction of MEK phosphorylation. We have now identified a MEK inhibitor, CH5126766 (RO5126766), that has the unique property of inhibiting RAF kinase as well. CH5126766 binding causes MEK to adopt a conformation in which it cannot be phosphorylated by and released from RAF. This results in formation of a stable MEK/RAF complex and inhibition of RAF kinase. Consistent with this mechanism, this drug does not induce MEK phosphorylation. CH5126766 inhibits ERK signaling output more effectively than a standard MEK inhibitor that induces MEK phosphorylation and has potent antitumor activity as well. These results suggest that relief of RAF feedback limits pathway inhibition by standard MEK inhibitors. CH5126766 represents a new type of MEK inhibitor that causes MEK to become a dominant-negative inhibitor of RAF and that, in doing so, may have enhanced therapeutic activity in ERK-dependent tumors with mutant RAS. Cancer Res; 73(13); 4050–60. ©2013 AACR.

Our lead compound for a phosphoinositide 3-kinase (PI3K) inhibitor (1) was metabolically unstable because of rapid glucuronidation of the phenol moiety. Based on structure-activity relationship (SAR) information and a FlexSIS docking simulation score, aminopyrimidine was identified as a bioisostere of phenol. An X-ray structure study revealed a hydrogen bonding pattern of aminopyrimidine derivatives. Finally, aminopyrimidine derivatives 33 showed strong tumor growth inhibition against a KPL-4 breast cancer xenograft model in vivo.

Macrocyclic compounds bearing a 2-amino-6-arylpyrimidine moiety were identified as potent heat shock protein 90 (Hsp90) inhibitors by modification of 2-amino-6-aryltriazine derivative (CH5015765). We employed a macrocyclic structure as a skeleton of new inhibitors to mimic the geldanamycin-Hsp90 interactions. Among the identified inhibitors, CH5164840 showed high binding affinity for N-terminal Hsp90α (K(d)=0.52nM) and strong anti-proliferative activity against human cancer cell lines (HCT116 IC(50)=0.15μM, NCI-N87 IC(50)=0.066μM). CH5164840 displayed high oral bioavailability in mice (F=70.8%) and potent antitumor efficacy in a HCT116 human colorectal cancer xenograft model (tumor growth inhibition=83%).

In two series of small-molecule ligands, one inhibiting human cathepsin L (hcatL) and the other MEK1 kinase, biological affinities were found to strongly increase when an aryl ring of the inhibitors is substituted with the larger halogens Cl, Br, and I, but to decrease upon F substitution. X-ray co-crystal structure analyses revealed that the higher halides engage in halogen bonding (XB) with a backbone C=O in the S3 pocket of hcatL and in a back pocket of MEK1. While the S3 pocket is located at the surface of the enzyme, which provides a polar environment, the back pocket in MEK1 is deeply buried in the protein and is of pronounced apolar character. This study analyzes environmental effects on XB in protein-ligand complexes. It is hypothesized that energetic gains by XB are predominantly not due to water replacements but originate from direct interactions between the XB donor (Caryl-X) and the XB acceptor (C=O) in the correct geometry. New X-ray co-crystal structures in the same crystal form (space group P2(1)2(1)2(1)) were obtained for aryl chloride, bromide, and iodide ligands bound to hcatL. These high-resolution structures reveal that the backbone C=O group of Gly61 in most hcatL co-crystal structures maintains water solvation while engaging in XB. An aryl-CF3-substituted ligand of hcatL with an unexpectedly high affinity was found to adopt the same binding geometry as the aryl halides, with the CF3 group pointing to the C=O group of Gly61 in the S3 pocket. In this case, a repulsive F2C-F⋅⋅⋅O=C contact apparently is energetically overcompensated by other favorable protein-ligand contacts established by the CF3 group.

5a-Carba-β-D-glucopyranose derivatives were synthesized and identified as novel SGLT2-selective inhibitors. These inhibitors exhibited potent SGLT2 inhibition with high selectivity over SGLT1. Among the tested compounds, 6f indicated the most potent hSGLT2 inhibition and the highest selectivity over hSGLT1. Moreover, the pharmacokinetics data also showed that 6h, which had the same aglycon structure as sergliflozin-active (3-active), had a threefold longer half-life time (T(1/2)) than sergliflozin (3) with a high distribution volume in db/db mice. Subsequently, 6h lowered blood glucose levels as much as 3 and showed longer hypoglycemic action than 3 in db/db mice.

The in vitro susceptibilities of 140 laboratory reference strains of fungi, including type strains, and 165 clinical yeast isolates from Japan towards isavuconazole compared with fluconazole (FLC), itraconazole (ITC), voriconazole and amphotericin B were measured. Broth microdilution methods based on Clinical and Laboratory Standards Institute (CLSI) methods were used for yeasts, and RPMI-MOPS medium semi-solidified with 0.2% low-melting-point agarose based on CLSI guidelines was used for moulds. The range of isavuconazole minimum inhibitory concentrations (MICs) was 0.0004-0.21 mg/L for Candida albicans, 0.0036-0.4 mg/L for Candida glabrata, 0.023-0.058 mg/L for Candida krusei, 0.0026-0.032 mg/L for Cryptococcus neoformans, 0.1-0.39mg/L for Aspergillus fumigatus and 0.2-0.39 mg/L for Aspergillus terreus. Isavuconazole was as active as ITC against the dimorphic true pathogenic fungi, with a range of MICs from

A series of benzofuran-based farnesyltransferase inhibitors have been designed and synthesized as antitumor agents. Among them, 11f showed the most potent enzyme inhibitory activity (IC 50 = 1.1 nM) and antitumor activity in human cancer xenografts in mice.

An estimated 170 million individuals worldwide are infected with hepatitis C virus (HCV), a serious cause of chronic liver disease. Current interferon-based therapy for treating HCV infection has an unsatisfactory cure rate, and the development of more efficient drugs is needed. During the early stages of HCV infections, various host genes are differentially regulated, and it is possible that inhibition of host proteins affords a therapeutic strategy for treatment of HCV infection. Using an HCV subgenomic replicon cell culture system, here we have identified, from a secondary fungal metabolite, a lipophilic long-chain base compound, NA255 (1), a previously unknown small-molecule HCV replication inhibitor. NA255 prevents the de novo synthesis of sphingolipids, major lipid raft components, thereby inhibiting serine palmitoyltransferase, and it disrupts the association among HCV nonstructural (NS) viral proteins on the lipid rafts. Furthermore, we found that NS5B protein has a sphingolipid-binding motif in its molecular structure and that the domain was able to directly interact with sphingomyelin. Thus, NA255 is a new anti-HCV replication inhibitor that targets host lipid rafts, suggesting that inhibition of sphingolipid metabolism may provide a new therapeutic strategy for treatment of HCV infection.

1,3-beta-D-Glucan synthase, which synthesizes a main component of fungal cell wall, is one of the promising targets for antifungal agents. In order to identify novel chemical classes of 1,3-beta-D-glucan synthase inhibitors, we screened a chemical library monitoring inhibition of the Candida albicans 1,3-beta-D-glucan synthase activity. The piperazine propanol derivative GSI578 [(2,6-difluoro-phenyl)-carbamic acid 3-(4-benzothiazol-2-yl-piperazine-1-yl)-propyl ester] was identified as a potent inhibitor against 1,3-beta-D-glucan synthase with an IC50 value of 0.16 microM. GSI578 exhibited in vitro antifungal activity against pathogenic fungi including C. albicans and Aspergillus fumigatus. Temperature-sensitive mutations of the FKS1 gene in the Deltafks2 background of Saccharomyces cerevisiae, where FKS1 and FKS2 encode putative catalytic subunits of 1,3-beta-D-glucan synthase, altered sensitivity to GSI578. This suggests that the antifungal activity of the piperazine propanol derivative has an effect on 1,3-beta-D-glucan synthase inhibition. Results of our initial evaluation suggest that the piperazine propanol derivative is a novel chemical structure of the class of antifungals which inhibit fungal cell growth by inhibiting fungal 1,3-beta-D-glucan synthase.

The correlation between the molecular electrostatic potential (MEP) on benzofuran inhibitors and the N-myristoyltransferase (Nmt) inhibitory activity is investigated by a three-dimensional quantitative structure–activity relationship (3D-QSAR) approach. To each compound, a Kohonen neural network (KNN) is used to project the MEP values on a Connolly surface into a two-dimensional map. Each node in the KNN map is coded by the associated MEP value of the occupying sampling point and all the coding nodes are collected together to define chemical descriptors. The best partial least squares (PLS) model with the selected descriptors is searched among the possible candidate models by genetic algorithm (GA). The resulting best model is significantly more predictive than the one with all chemical descriptors. The model has given additional information about the binding of a benzofuran inhibitor to Nmt.

The C-4 side chain modification of lead compound 1 has resulted in the identification of a potent and selective Candida albicans N-myristoyltransferase (CaNmt) inhibitor RO-09-4609, which exhibits antifungal activity against C. albicans in vitro. Further modification of its C-2 substituent has led to the discovery of RO-09-4879, which exhibits antifungal activity in vivo. The drug design is based on X-ray crystal analysis of a CaNmt complex with benzofuran derivative 4a. The optimization incorporates various biological investigations including a quasi in vivo assay and pharmacokinetic study. The computer aided drug design, synthesis, structure-activity relationships, and biological properties of RO-09-4879 are described in detail.

A series of tumor-activated prodrugs of the inhibitors of dihydropyrimidine dehydrogenase (DPD), an enzyme catabolizing 5-fluorouracil (5-FU: 4g), has been designed and synthesized. RO0094889 (11c) is a prodrug of 5-vinyluracil (4c), a known DPD inhibitor, and was designed to generate 4c selectively in tumor tissues by sequential conversion of 11c by three enzymes: esterase, cytidine deaminase and thymidine phosphorylase, the latter two of which are known to be highly expressed in various tumor tissues. When capecitabine (1), a tumor-activated prodrug of 5-FU, was co-administered orally with 11c, 5-FU in tumor tissues was significantly increased with only a slight increase of 5-FU in plasma as compared with oral capecitabine alone.

Water soluble N-benzyltriazolium or N-benzylimidazolium salt type prodrugs of several highly lipophilic triazole or imidazole antifungals have been synthesized. They were designed to undergo an enzymatic activation followed by a self-cleavage to release a parent drug. The prodrugs such as 16 had enough chemical stability and water solubility for parenteral use and were rapidly and quantitatively converted to the active substance in human plasma.

Modification of the C-2 position of a benzofuran derivative 6 (RO-09-4609), an N-myristoyltransferase (Nmt) inhibitor, has led us to discover antifungal agents that are active in a murine systemic candidiasis model. The drug design is based on the analysis of a crystal structure of a Candida Nmt complex with 2. The optimization has been guided by various biological evaluations including a quasi in vivo assay and pharmacokinetic analysis.

Potent and selective Candida albicans N -myristoyltransferase (CaNmt) inhibitors have been identified through optimization of a lead compound 1 discovered by random screening. The inhibitor design is based on the crystal structure of the CaNmt complex with compound ( S )- 3 and structure–activity relationships (SARs) have been clarified. Modification of the C-4 side chain of 1 has led to the discovery of a potent and selective CaNmt inhibitor 11 (RO-09-4609), which exhibits antifungal activity against C. albicans in vitro.

Highly potent 1,3-β- d -glucan synthase inhibitors, 7b , 10a , 10b and 12 , have been identified by the chemical modification of the ornithine residue of a fungicidal macrocyclic lipopeptidolactone, RO-09-3655 ( 1 ), isolated from the cultured broth of Deuteromycotinia spp. These compounds showed stronger antifungal activity against systemic candidiasis as well as pulmonary aspergillosis in mice, and less hepatotoxicity as compared with 1 .

As in Saccharomyces cerevisiae, the pathogenic fungus Candida albicans harbors three chitin synthases called CaChs1p, CaChs2p, and CaChs3p, which are structurally and functionally analogous to the S. cerevisiae ScChs2p, ScChs1p, and ScChs3p, respectively. In S. cerevisiae,ScCHS1, ScCHS2, and ScCHS3 are all non-essential genes; only the simultaneous disruption ofScCHS2 and ScCHS3 is lethal. The fact that a null mutation of the CaCHS1 is impossible, however, implies that CaCHS1 is required for the viability of C. albicans. To gain more insight into the physiological importance of CaCHS1, we identified and characterized a novel inhibitor that was highly specific to CaChs1p. RO-09-3143 inhibited CaChs1p with a K i value of 0.55 nm in a manner that was non-competitive to the substrate UDP-N-acetylglucosamine. RO-09-3143 also hampered the growth of the C. albicans cells with an MIC50value of 0.27 μm. In the presence of RO-09-3143, theC. albicans cells failed to form septa and displayed an aberrant morphology, confirming the involvement of the C. albicans Chs1p in septum formation. Although the effect of RO-09-3143 on the wild-type C. albicans was fungistatic, it caused cell death in the cachs2Δ null mutants but not in the cachs3Δ null mutants. Thus, it appears that inC. albicans, inhibition of CaChs1p causes cell growth arrest, but simultaneous inhibition of CaChs1p and CaChs2p is lethal.

( 3 H)-Azoxybacilin, a new antifungal agent, was prepared in five steps. The label was introduced by reduction of 2-t-butoxycarbonylamino-succinic-acid-1-t-butylester-carbonic-acid-ethylester anhydride (2) with LiB 3 H 4 generated from tritium gas via Li 3 H.

Hepatitis C virus (HCV) infection represents a serious health-care problem. Previously we reported the identification of NA255 from our natural products library using a HCV sub-genomic replicon cell culture system. Herein, we report how the absolute stereochemistry of NA255 was determined and an enantioselective synthetic method for NA255 derivatives was developed. The structure-activity relationship of the NA255 derivatives and rat pharmacokinetic profiles of the representative compounds are disclosed.

C-Aryl 5a-carba-β-d-glucopyranose derivatives were synthesized and evaluated for inhibition activity against hSGLT1 and hSGLT2. Modifications to the substituents on the two benzene rings resulted in enhanced hSGLT2 inhibition activity and extremely high hSGLT2 selectivity versus SGLT1. Using the created superimposed model, the reason for the high hSGLT2 selectivity was speculated to be that additional substituents occupied a new space, in a different way than known inhibitors. Among the tested compounds, the ethoxy compound 5h with high hSGLT2 selectivity exhibited more potent and longer hypoglycemic action in db/db mice than our O-carbasugar compound (1) and sergliflozin (2), which could be explained by its improved PK profiles relative to those of the two compounds. These results indicated that 5h might be a promising drug candidate for the treatment of type 2 diabetes.

Synthesis of cyclohexyl analogs of restricticin, a novel type of antifungal agent from ⊂-) R-carvone and their in vitro antifungal activity are described.

Hispidospermidin (1) is a novel phospholipase C inhibitor produced by Chaetosphaeronema hispidulum (Cda) Moesz NR 7127. Its structure (C 25 H 47 N 3 O) has been elucidated as a cage compound with a trimethylspermidine side chain based on various NMR studies, including 1 H- 1 H COSY, 13 C- 1 H COSY, HOHAHA, HMBC, COLOC and long range JC-H resolved 2D spectroscopy. The absolute configuration of 1 has been elucidated by modified MOSHER's method on the (R)- and (S)-MTPA amides of a derivative of 1

Proliferation of endothelial cells is critical for angiogenesis. We report orally available, in vivo active antiangiogenic agents which specifically inhibit endothelial cell proliferation. After identifying human umbilical vein endothelial cell (HUVEC) proliferation inhibitors from a cell-based high-throughput screening (HTS), we eliminated those compounds which showed cytotoxicity against HCT116 and vascular endothelial growth factor receptor 2 (VEGFR-2) inhibitory activity. Evaluations in human Calu-6 xenograft model delivered lead compound 1. Following extensive lead optimization and alteration of the scaffold we discovered 32f and 32g, which both inhibited the proliferation and tube formation of HUVEC without showing inhibitory activity against any of 25 kinases or cytotoxicity against either normal fibroblasts or 40 cancer cell lines. Upon oral administration, 32f and 32g had good pharmacokinetic profiles and potent antitumor activity and decreased microvessel density (MVD) in Calu-6 xenograft model. Combination therapy with a VEGFR inhibitor enhanced the in vivo efficacy. These results suggest that 32f and 32g may have potential for use in cancer treatment.

Most acute hepatitis C virus (HCV) infections become chronic and some progress to liver cirrhosis or hepatocellular carcinoma. Standard therapy involves an interferon (IFN)-α-based regimen, and efficacy of therapy has been significantly improved by the development of protease inhibitors. However, several issues remain concerning the injectable form and the side effects of IFN. Here, we report an orally available, small-molecule type I IFN receptor agonist that directly transduces the IFN signal cascade and stimulates antiviral gene expression. Like type I IFN, the small-molecule compound induces IFN-stimulated gene (ISG) expression for antiviral activity in vitro and in vivo in mice, and the ISG induction mechanism is attributed to a direct interaction between the compound and IFN-α receptor 2, a key molecule of IFN-signaling on the cell surface. Our study highlights the importance of an orally active IFN-like agent, both as a therapy for antiviral infections and as a potential IFN substitute.

Heat shock protein 90 (Hsp90) is a molecular chaperone which regulates maturation and stabilization of its substrate proteins, known as client proteins. Many client proteins of Hsp90 are involved in tumor progression and survival and therefore Hsp90 can be a good target for developing anticancer drugs. With the aim of efficiently identifying a new class of orally available inhibitors of the ATP binding site of this protein, we conducted fragment screening and virtual screening in parallel against Hsp90. This approach quickly identified 2-aminotriazine and 2-aminopyrimidine derivatives as specific ligands to Hsp90 with high ligand efficiency. In silico evaluation of the 3D X-ray Hsp90 complex structures of the identified hits allowed us to promptly design CH5015765, which showed high affinity for Hsp90 and antitumor activity in human cancer xenograft mouse models.

The MAP kinase pathway is one of the most important pathways involved in cell proliferation and differentiation, and its components are promising targets for antitumor drugs. Design and synthesis of a novel MEK inhibitor, based on the 3D-structural information of the target enzyme, and then multidimensional optimization including metabolic stability, physicochemical properties and safety profiles were effectively performed and led to the identification of a clinical candidate for an orally available potent MEK inhibitor, CH4987655, possessing a unique 3-oxo-oxazinane ring structure at the 5-position of the benzamide core structure. CH4987655 exhibits slow dissociation from the MEK enzyme, remarkable in vivo antitumor efficacy both in mono- and combination therapy, desirable metabolic stability, and insignificant MEK inhibition in mouse brain, implying few CNS-related side effects in human. An excellent PK profile and clear target inhibition in PBMC were demonstrated in a healthy volunteer clinical study.

A new antifungal substance, azoxybacilin (an unusual amino acid with an azoxy moiety) and its derivatives have been synthesized from Boc-L-Asp-OtBu utilizing the Moss procedure for the preparation of the azoxy moiety. The ester derivative, Ro 09-1824, showed more potent antifungal activity and a broader antifungal spectrum than azoxybacilin did.

Capecitabine (N4-pentyloxycarbonyl-5'-deoxy-5-fluorocytidine) is a novel oral fluoropyrimidine carbamate, which was designed to be sequentially converted to 5-fluorouracil (5-FU) by three enzymes located in the liver and in tumors. N4-alkoxycarbonyl-5'-deoxy-5-fluorocytidine derivatives including capecitabine pass intact through the intestinal tract and are sequentially converted to 5-FU by a cascade of the three enzymes. The first step is the conversion to 5'-deoxy-5-fluorocytidine (5'-DFCR) by carboxylesterase located in the liver, then to 5'-deoxy-5-fluorouridine (5'-DFUR) by cytidine deaminase highly expressed in the liver and various solid tumors, and finally to 5-FU by thymidine phosphorylase (dThdPase) preferentially located in tumor tissues. Among large numbers of the derivatives, capecitabine was selected based on its susceptibility to hepatic carboxylesterase, oral bioavailability in monkeys and efficacy in a human cancer xenograft. Capecitabine given orally yielded substantially higher concentrations of 5-FU within tumors than in plasma or normal tissue (muscle). The tumor 5-FU levels were also much higher than those achieved by intraperitoneal administration of 5-FU at equi-toxic doses. This tumor selective delivery of 5-FU ensured greater efficacy and a more favourable safety profile than with other fluoropyrimidines. In 24 human cancer xenograft models studied, capecitabine was more effective at a wider dose range and had a broader spectrum of antitumor activity than 5-FU, UFT or its intermediate metabolite 5'-DFUR. The susceptibility of the xenografts to capecitabine correlated with tumor dThdPase levels. Moreover, the conversion of 5'-DFUR to 5-FU by dThdPase in tumor was insufficient in a xenograft model refractory to capecitabine. In addition, the efficacy of capecitabine was enhanced by dThdPase up-regulators, such as by taxanes and cyclophosphamide and by X-ray irradiation. The efficacy of capecitabine may, therefore, be optimized by selecting the most appropriate patient population based on dThdPase status and/or by combining it with dThdPase up-regulators. Capecitabine has additional characteristics not found with 5-FU, such as potent antimetastatic and anticachectic actions in mouse tumor models. With these profiles, capecitabine may have substantial potential in cancer treatment.

Pharmacokinetic (PK) parameters of N-myristoyltransferase (Nmt) inhibitors were measured, and a multivariate quantitative structure−pharmacokinetic relationship (QSPKR) model for predicting rat elimination half-life (t1/2) values was constructed. One hundred seven benzofuran derivatives have been selected as the data set for QSPKR analysis. The correlation between the t1/2 values and 30 physicochemical descriptors was examined by a stepwise multiple linear regression method. The statistical analysis gives a significant QSPKR model (r = 0.843) with the following three variables: partial negative surface area (PNSA), atomic-based octanol/water partition coefficient (AlogP), and the number of rotational bonds (Rotlbonds). The QSPKR model obtained is predictive and simple, and would give a direction for designing new Nmt inhibitors having good PK profiles.

Identification of a novel topoisomerase I inhibitor which shows superior efficacy and less individual variation than irinotecan hydrochloride (CPT-11). A novel camptothecin analog that is effective against breast cancer resistance protein (BCRP)-positive cells was screened, and a water soluble prodrug was generated. Antitumor activity of the prodrug was examined in BCRP-positive and -negative xenografts both as a single agent and in combination with other anti-cancer drugs. A novel camptothecin analog, CH0793076, was discovered. Because CH0793076 was found to be highly lipophilic, a water soluble prodrug (TP300) was generated. TP300 is stable in an acidic solution but is rapidly converted to CH0793076 under physiological pH conditions such as in sera. This efficient prodrug activation would minimize interpatient differences in pharmacokinetic and toxicity profiles. Unlike CPT-11, TP300 does not exhibit cholinergic interaction or cause acute diarrhea at effective doses. In mouse xenograft models, TP300 showed antitumor activity against both BCRP-positive and -negative xenografts, whereas CPT-11 was less active against BCRP-positive xenografts. In addition, the effective dose range (MTD/ED50) for TP300 was wider than for CPT-11 and TP300 showed additive or synergistic antitumor effects in combination with other anti-cancer drugs such as capecitabine, oxaliplatin, cisplatin, bevacizumab and cetuximab. It is therefore expected that TP300 will provide an additional treatment option for patients who will undergo chemotherapy with camptothecins.

Vinaxanthone is a novel phospholipase C inhibitor produced by Penicillium vinaceum Gilman and About NR6815. Its structre (MW 576, C28H16O14) has been elucidated as a polycyclic xanthone with poly acidic functional groups based on various NMR studies including HMBC, COLOC, 2D-INADEQUATE and selective 1D-INADEQUATE.

Novel hexacyclic camptothecin analogs containing cyclic amidine, urea, or thiourea moiety were designed and synthesized based on the proposed 3D-structure of the topoisomerase I (Topo I)/DNA/camptothecin ternary complex. The analogs were prepared from 9-nitrocamptothecin via 7,9-diaminocamptothecin derivatives as a key intermediate. Among them, 7c exhibited in vivo antitumor activities superior to CPT-11 in human cancer xenograft models in mice at their maximum tolerated doses though its in vitro antiproliferative activity was comparable to SN-38 against corresponding cell lines.

This section describes recent examples of prodrugs of functional groups not commonly associated with prodrug strategies, including nitrogen-containing heteroaromatic rings, thiols, nucleic acids, and enediynes. The prodrugs for several addional functional groups that are not covered in this book were reviewed in the book “Design of Prodrugs,” edited by Bundgaard (1985); examples include derivatives of CH acidic compounds, imine bisulphite addition products, and glycosidic products.

Capecitabine (2), is an off-white crystalline solid with and aqueous solubility of 26 mg/mL. It is sold as Xeloda® and is a prodrug said to achieve selective delivery of 5-fluorouracil (5-FU, 1) to tumors by sequential activation of the prodrug with endogenous enzymes preferentially located in the human liver and tumor tissues see Fig. 1). Capecitabine was first approved in the US in 1998 for the treatment of metastatic breast cancer that is refractory to standard chemotherapy with paclitaxel and anthracycline-containing regimen; it has now been used for the treatment of metastatic breast and colorectal cancers in more than 70 countries. It was designed to achieve greater selectivity than its active form, 5-FU (1)