Saccharomyces cerevisiae GSC1 (also called FKS1) and GSC2 (also called FKS2) have been identified as the genes for putative catalytic subunits of ␤-1,3-glucan synthase. We have cloned three Candida albicans genes, GSC1, GSL1, and GSL2, that have significant sequence homologies with S. cerevisiae GSC1/FKS1, GSC2/FKS2, and the recently identified FKSA of Aspergillus nidulans at both nucleotide and amino acid levels. Like S. cerevisiae Gsc/Fks proteins, none of the predicted products of C. albicans GSC1, GSL1, or GSL2 displayed obvious signal sequences at their N-terminal ends, but each product possessed 10 to 16 potential transmembrane helices with a relatively long cytoplasmic domain in the middle of the protein. Northern blotting demonstrated that C. albicans GSC1 and GSL1 but not GSL2 mRNAs were expressed in the growing yeastphase cells. Three copies of GSC1 were found in the diploid genome of C. albicans CAI4. Although we could not establish the null mutation of C. albicans GSC1, disruption of two of the three GSC1 alleles decreased both GSC1 mRNA and cell wall ␤-glucan levels by about 50%. The purified C. albicans ␤-1,3-glucan synthase was a 210-kDa protein as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and all sequences determined with peptides obtained by lysyl endopeptidase digestion of the 210-kDa protein were found in the deduced amino acid sequence of C. albicans Gsc1p. Furthermore, the monoclonal antibody raised against the purified ␤-1,3-glucan synthase specifically reacted with the 210-kDa protein and could immunoprecipitate ␤-1,3-glucan synthase activity. These results demonstrate that C. albicans GSC1 is the gene for a subunit of ␤-1,3-glucan synthase.

Osimertinib is the only EGFR-tyrosine kinase inhibitor (TKI) capable of overcoming EGFR-T790M-mutated NSCLC, but osimertinib-resistant EGFR triple mutations (Del19/T790M/C797S or L858R/T790M/C797S) have been reported. Although allosteric EGFR TKIs (eg. EAI-045) which potentially overcome L858R/T790M/C797S have been identified, there are no effective inhibitors against Del19/T790M/C797S. In this study, we identified CH7233163 as having the potential to overcome EGFR-Del19/T790M/C797S. CH7233163 showed potent antitumor activities against tumor with EGFR-Del19/T790M/C797S in vitro and in vivo. In addition to EGFR-Del19/T790M/C797S, the characterization assays showed that CH7233163 more selectively inhibits various types of EGFR mutants (eg. L858R/T790M/C797S, L858R/T790M, Del19/T790M, Del19 and L858R) over wild-type (WT). Furthermore, crystal structure analysis suggested that CH7233163 is a non-covalent ATP competitive inhibitor for EGFR-Del19/T790M/C797S that utilizes multiple interactions with the EGFR's αC-helix-in conformation to achieve potent inhibitory activity and mutant selectivity. Therefore, we conclude that CH7233163 is a potentially effective therapy for osimertinib resistant patients, especially in cases of EGFR-Del19/T790M/C797S.

Purpose: The phosphatidylinositol 3-kinase (PI3K) pathway plays a central role in cell proliferation and survival in human cancer. PIK3CA mutations, which are found in many cancer patients, activate the PI3K pathway, resulting in cancer development and progression. We previously identified CH5132799 as a novel PI3K inhibitor. Thus, this study aimed to clarify the biochemical and antitumor activity of CH5132799 and elucidate the correlation between CH5132799 response and genetic alterations in the PI3K pathway.Experimental Design: Kinase inhibitory activity was profiled in cell-free assays. A large panel of human breast, ovarian, prostate, and endometrial cancer cell lines, as well as xenograft models, were used to evaluate the antitumor activity of CH5132799, followed by analysis for genetic alterations. Effects on Akt phosphorylation induced by mTORC1 inhibition were tested with CH5132799 and compared with mTORC1 and PI3K/mTOR inhibitors.Results: CH5132799 selectively inhibited class I PI3Ks and PI3Ka mutants in in vitro kinase assays. Tumors harboring PIK3CA mutations were significantly sensitive to CH5132799 in vitro and were remarkably regressed by CH5132799 in in vivo mouse xenograft models. In combination with trastuzumab, tumors disappeared in the trastuzumab-insensitive breast cancer model with the PIK3CA mutation. Moreover, CH5132799 did not reverse a negative feedback loop of PI3K/Akt/mTOR signaling and induced regression against tumors regrown after long-term mTORC1 inhibitor treatment.Conclusions: CH5132799 is a selective class I PI3K inhibitor with potent antitumor activity against tumors harboring the PIK3CA mutations. Prediction of CH5132799 response on the basis of PIK3CA mutations could enable patient stratification in clinical settings. Clin Cancer Res; 17(10); 3272-81. Ó2011 AACR.

No abstract

Targeting genetic alterations of oncogenes by moleculartargeted agents (MTA) is an effective approach for treating cancer. However, there are still no clinical MTA options for many cancers, including esophageal cancer. We used a short hairpin RNA library to screen for a new oncogene in the esophageal cancer cell line KYSE70 and identified YES proto-oncogene 1 (YES1) as having a significant impact on tumor growth. An analysis of clinical samples showed that YES1 gene amplification existed not only in esophageal cancer but also in lung, head and neck, bladder, and other cancers, indicating that YES1 would be an attractive target for a cancer drug. Because there is no effective YES1 inhibitor so far, we generated a YES1 kinase inhibitor, CH6953755. YES1 kinase inhibition by CH6953755 led to antitumor activity against YES1-amplified cancers in vitro and in vivo. Yes-associated protein 1 (YAP1) played a role downstream of YES1 and contributed to the growth of YES1amplified cancers. YES1 regulated YAP1 transcription activity by controlling its nuclear translocation and serine phosphorylation. These findings indicate that the regulation of YAP1 by YES1 plays an important role in YES1-amplified cancers and that CH6953755 has therapeutic potential in such cancers. Significance: These findings identify the SRC family kinase YES1 as a targetable oncogene in esophageal cancer and describe a new inhibitor for YES1 that has potential for clinical utility. See related commentary by Rai, p. 5702

Members of the tropomyosin receptor kinase (TRK) family are expressed in their constitutively activated forms as a result of a gene fusion that occurs across a wide variety of cancer types. We have identified CH7057288 as a potent and selective TRK inhibitor that belongs to a novel chemical class. CH7057288 showed selective inhibitory activity against TRKA, TRKB, and TRKC in cell-free kinase assays and suppressed proliferation of TRK fusion-positive cell lines, but not that of TRK-negative cell lines. Strong in vivo tumor growth inhibition was observed in subcutaneously implanted xenograft tumor models of TRK fusion-positive cells. Furthermore, in an intracranial implantation model mimicking brain metastasis, CH7057288 significantly induced tumor regression and improved event-free survival. Recently, resistant mutations in the kinase domain of TRK have been reported in patients who show disease progression after treatment with the TRK inhibitors now under clinical development. Our compound maintained similar levels of in vitro and in vivo activity against one of these resistant mutants as it did to wild-type TRK. An X-ray crystal structure of the TRKA and CH7057288 complex supported the activity against the mutant. In addition, gene expression analysis revealed that CH7057288 suppressed MAPK and E2F pathways as downstream signaling of TRK fusion. Therefore, CH7057288 could be a promising therapeutic agent for TRK fusion-positive cancer.IC 50 values of in vitro kinase assay were shown as mean AE standard deviation (n ¼ 3). Analysis and interpretation of data (e.g., statistical analysis, biostatistics, computational analysis):

Two metabolites (M4 and M1b) in plasma and four metabolites (M4, M6, M1a and M1b) in faeces were detected through the human ADME study following a single oral administration of [14C]alectinib, a small-molecule anaplastic lymphoma kinase inhibitor, to healthy subjects. In the present study, M1a and M1b, which chemical structures had not been identified prior to the human ADME study, were identified as isomers of a carboxylate metabolite oxidatively cleaved at the morpholine ring. In faeces, M4 and M1b were the main metabolites, which shows that the biotransformation to M4 and M1b represents two main metabolic pathways for alectinib. In plasma, M4 was a major metabolite and M1b was a minor metabolite. The contribution to in vivo pharmacological activity of these circulating metabolites was assessed from their in vitro pharmacological activity and plasma protein binding. M4 had a similar cancer cell growth inhibitory activity and plasma protein binding to that of alectinib, suggesting its contribution to the antitumor activity of alectinib, whereas the pharmacological activity of M1b was insignificant.

No abstract