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Although chemotherapy using CHOP-based protocol induces remission in most cases of canine multicentric high-grade B-cell lymphoma (mhBCL), some cases develop early relapse during the first induction protocol. In this study, we examined the gene expression profiles of canine mhBCL before chemotherapy and investigated their associations with early relapse during the first whole CHOP-based protocol. Twenty-five cases of mhBCL treated with CHOP-based protocol as first induction chemotherapy were included in this study. Sixteen cases completed the first whole CHOPbased protocol without relapse (S-group), and nine developed relapse during the chemotherapy (R-group). RNA-seq was performed on samples from neoplastic lymph nodes. Differentially expressed genes (DEGs) were extracted by the comparison of gene expression profiles between S- and R-groups, and the differences in the expression levels of these genes were validated by RTqPCR. Extracted 179 DEGs included the genes related to chemokine CC motif ligand, T-cell receptor signaling pathway, and PD-L1 expression and PD-1 checkpoint pathway. We focused on chemokine CC motif ligand, and CCL4 was confirmed to be significantly downregulated in the R-group (P=0.039). We also focused on the genes related to T-cell signaling pathway, and CD3E (P=0.039), ITK (P=0.023), and LAT (P=0.023) genes were confirmed to be significantly upregulated in the R-group. The current results suggest that both changes in tumor cells and the interactions between tumor cells and immune cells are associated with the efficacy of the chemotherapy for first remission induction. [ABSTRACT FROM AUTHOR]

Celastrol, a bioactive molecule extracted from the Tripterygium wilfordii plant, has been shown to exhibit anti-inflammatory properties. However, its mechanism of action has not been fully elucidated. Here, we show that celastrol suppresses humoral immune responses and autoimmunity by disabling a protein complex consisting of copper metabolism MURR1 domain–containing (COMMD) 3 and COMMD8 (COMMD3/8 complex), a signaling adaptor for chemoattractant receptors. Having demonstrated the involvement of the COMMD3/8 complex in a mouse model of rheumatoid arthritis, we identified celastrol as a compound that covalently bound to and dissociated the COMMD3/8 complex. Celastrol inhibited B cell migration, reduced antibody responses, and blocked arthritis progression, recapitulating deficiency of the COMMD3/8 complex. These effects of celastrol were abolished in mice expressing a celastrol-resistant mutant of the COMMD3/8 complex. These findings establish that celastrol exerts immunosuppressive activity by targeting the COMMD3/8 complex. Our study suggests that the COMMD3/8 complex is a potentially druggable target in autoimmune diseases and points to celastrol as a lead pharmacologic candidate in this capacity. Targeting B cell migration in arthritis: COMMD3 and COMMD8, members of the copper metabolism MURR1 domain–containing (COMMD) protein family, form a heterodimer essential for chemoattractant-mediated B cell migration during induction of humoral immune responses. By screening a compound library for molecules capable of disrupting the COMMD3/8 complex, Shirai et al. identified the Chinese herbal medicine celastrol as a potent inhibitor of COMMD3 association with COMMD8. In a mouse model of rheumatoid arthritis, celastrol treatment initiated at the onset of arthritis prevented disease progression. Celastrol's ability to disrupt the COMMD3/8 complex and inhibit arthritis was lost when cysteine 170 of COMMD3 was replaced with alanine. These findings reveal a molecular mechanism for the anti-arthritic activity of celastrol and identify the COMMD3/8 complex as a potential drug target in B cell-dependent autoimmune diseases. —IRW [ABSTRACT FROM AUTHOR]

Simple Summary: Cardiogenic pulmonary edema (CPE) is the leading cause of cats visiting emergency hospitals with respiratory distress. The association of physical examination and venous blood gas parameters with the survival of cats with CPE was retrospectively investigated. Thirty-six cats were ultimately included, and eight of them died within 12 h after their presentation to our hospital. The Mann–Whitney U test with Bonferroni correction was used to compare the parameters. Cats that died within 12 h had significantly lower rectal temperatures and higher PvCO2 than those that did not die within 12 h. Moreover, vasoconstrictor use was related to death within 12 h of presentation and higher PvCO2. A large number of prospective studies should be performed to validate these results. Cats urgently visit emergency hospitals due to respiratory distress, and the chief cause is cardiogenic pulmonary edema (CPE). Although cats with CPE were frequently encountered in clinics, the prognostic factors were poorly reported. The objective of this retrospective study was to investigate the association of physical examination and venous blood gas parameters with the survival of cats with CPE in an emergency hospital. Thirty-six cats with CPE were ultimately included in the present study, and eight of them died within 12 h after their presentation to our hospital. Statistical analyses of clinical parameters between cats that died within 12 h and those that survived for 12 h were conducted using Mann–Whitney U test with Bonferroni correction. Cats that died within 12 h had significantly lower rectal temperatures and higher PvCO2 than those that did not die within 12 h. Moreover, hypotension and vasoconstrictor use were related to death within 12 h of presentation and higher PvCO2. These findings indicated the prognostic utility of body temperature and PvCO2, and the association between hypercapnia and the severity of CPE or hypotension. A large number of prospective studies should be performed to validate these results. [ABSTRACT FROM AUTHOR]

Canine histiocytic proliferative disorders include aggressive and fatal diseases, such as histiocytic sarcoma (HS) and histiocytosis (SyH). The molecular mechanisms underlying cell proliferation need to be elucidated for the development of effective treatments. In the present study, mRNA expression levels were comprehensively analysed in cell lines derived from localized HS, disseminated HS, SyH and Langerhans cell histiocytosis (LCH) in dogs. Based on the results obtained, the growth inhibitory effects of palbociclib, a CDK4/6 inhibitor, were verified with the cell lines in vitro and in xenograft mouse model. Hierarchical clustering and principal component analysis plots of mRNA expression profiles divided the cell lines into three groups: a localized HS group, disseminated HS/SyH group, and LCH. The results of an ingenuity pathway analysis suggested that the MAPK signalling pathway was activated in the localized HS and LCH cell lines, and the PI3K signalling pathway in the disseminated and localized HS cell lines. In all cell lines, the expression of the tumour suppressor genes TP53, CDKN2A and CDKN1A was down‐regulated, whereas that of Rb was preserved. In vitro assessments revealed the growth inhibitory effects of palbociclib in all cell lines examined. In a xenograft mouse model using a cell line from disseminated HS, palbociclib exerted significant growth inhibitory effects. These results suggest the potential of palbociclib as a therapeutic drug candidate for the treatment of malignant histiocytic proliferative disorders of the dog. [ABSTRACT FROM AUTHOR]

Ectopic gene expression is an indispensable tool in biology and medicine, but is often limited by the low efficiency of DNA transfection. We previously reported that depletion of the autophagy receptor p62/SQSTM1 enhances DNA transfection efficiency by preventing the degradation of transfected DNA. Therefore, p62 is a potential target for drugs to increase transfection efficiency. To identify such drugs, a nonbiased high‐throughput screening was applied to over 4,000 compounds from the Osaka University compound library, and their p62 dependency was evaluated. The top‐scoring drugs were mostly microtubule inhibitors, such as colchicine and vinblastine, and all of them showed positive effects only in the presence of p62. To understand the p62‐dependent mechanisms, the time required for p62‐dependent ubiquitination, which is required for autophagosome formation, was examined using polystyrene beads that were introduced into cells as materials that mimicked transfected DNA. Microtubule inhibitors caused a delay in ubiquitination. Furthermore, the level of phosphorylated p62 at S405 was markedly decreased in the drug‐treated cells. These results suggest that microtubule inhibitors inhibit p62‐dependent autophagosome formation. Our findings demonstrate for the first time that microtubule inhibitors suppress p62 activation as a mechanism for increasing DNA transfection efficiency and provide solutions to increase efficiency. [ABSTRACT FROM AUTHOR]

Trelagliptin (SYR-472), a novel dipeptidyl peptidase-4 inhibitor, shows sustained efficacy by once-weekly dosing in type 2 diabetes patients. In this study, we characterized in vitro properties of trelagliptin, which exhibited approximately 4- and 12-fold more potent inhibition against human dipeptidyl peptidase-4 than alogliptin and sitagliptin, respectively, and >10,000-fold selectivity over related proteases including dipeptidyl peptidase-8 and dipeptidyl peptidase-9. Kinetic analysis revealed reversible, competitive and slow-binding inhibition of dipeptidyl peptidase-4 by trelagliptin (t1/2 for dissociation ≈ 30 minutes). X-ray diffraction data indicated a non-covalent interaction between dipeptidyl peptidase and trelagliptin. Taken together, potent dipeptidyl peptidase inhibition may partially contribute to sustained efficacy of trelagliptin. [ABSTRACT FROM AUTHOR]

Approximately 60–70% of EWSR1-negative small blue round cell sarcomas harbour a rearrangement of CIC, most commonly CIC-DUX4. CIC-DUX4 sarcoma (CDS) is an aggressive and often fatal high-grade sarcoma appearing predominantly in children and young adults. Although cell lines and their xenograft models are essential tools for basic research and development of antitumour drugs, few cell lines currently exist for CDS. We successfully established a novel human CDS cell line designated Kitra-SRS and developed orthotopic tumour xenografts in nude mice. The CIC-DUX4 fusion gene in Kitra-SRS cells was generated by t(12;19) complex chromosomal rearrangements with an insertion of a chromosome segment including a DUX4 pseudogene component. Kitra-SRS xenografts were histologically similar to the original tumour and exhibited metastatic potential to the lungs. Kitra-SRS cells displayed autocrine activation of the insulin-like growth factor 1 (IGF-1)/IGF-1 receptor (IGF-1R) pathway. Accordingly, treatment with the IGF-1R inhibitor, linsitinib, attenuated Kitra-SRS cell growth and IGF-1-induced activation of IGF-1R/AKT signalling both in vitro and in vivo. Furthermore, upon screening 1134 FDA-approved drugs, the responses of Kitra-SRS cells to anticancer drugs appeared to reflect those of the primary tumour. Our model will be a useful modality for investigating the molecular pathology and therapy of CDS. [ABSTRACT FROM AUTHOR]

Vasoactive intestinal peptide receptor 2 (VIPR2, also known as VPAC2) is a class B G-protein coupled receptor (GPCR) and plays important roles in the physiology of central nervous system (CNS) by interaction with natural ligands; vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP). Because it has been reported that high-expression and/or overactivation of VIPR2 link to schizophrenic symptoms, VIPR2 antagonists could be good drug candidates for schizophrenia therapeutics. In this study, we discovered several artificial peptides that antagonize both human and rodent VIPR2 with selectivities against receptor subtypes VIPR1 (also known as VPAC1) and pituitary adenylate cyclase-activating polypeptide type-1 receptor (PAC1). Of them, the representative 16-mer cyclic peptide VIpep-3 (Ac-CPPYLPRRLCTLLLRS-OH) exhibited strong binding affinity with K D value of 41 nM to extracellular domain of human VIPR2 in SPR analysis and showed potent antagonist activity with IC 50 values of 47 nM (human), 180 nM (mouse), and 44 nM (rat) against VIP–VIPR2 signal in cell-based Ca influx assay. This is not only the first report on artificial VIPR2-selective antagonist peptides but also good example of the effective approach to discover novel antagonist against class B GPCR. Our peptides will contribute to study and development of the novel CNS drugs targeting to VIPR2. [ABSTRACT FROM AUTHOR]

We report a design strategy to obtain potent DPP-4 inhibitors by incorporating salt bridge formation with Lys554 in the S1′ pocket. By applying the strategy to the previously identified templates, quinoline 4 and pyridines 16a , 16b , and 17 have been identified as subnanomolar or nanomolar inhibitors of human DPP-4. Docking studies suggested that a hydrophobic interaction with Tyr547 as well as the salt bridge interaction is important for the extremely high potency. The design strategy would be useful to explore a novel design for DPP-4 inhibitors having a distinct structure with a unique binding mode. [ABSTRACT FROM AUTHOR]

Amino-acid mutations of Gly 12 ( e.g. G12D, G12V, G12C) of V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (K-Ras), the most promising drug target in cancer therapy, are major growth drivers in various cancers. Although over 30 years have passed since the discovery of these mutations in most cancer patients, effective mutated K-Ras inhibitors have not been marketed. Here, we report novel and selective inhibitory peptides to K-Ras(G12D). We screened random peptide libraries displayed on T7 phage against purified recombinant K-Ras(G12D), with thorough subtraction of phages bound to wild-type K-Ras, and obtained KRpep-2 (Ac-RRCPLYISYDPVCRR-NH 2 ) as a consensus sequence. KRpep-2 showed more than 10-fold binding- and inhibition-selectivity to K-Ras(G12D), both in SPR analysis and GDP/GTP exchange enzyme assay. K D and IC 50 values were 51 and 8.9 nM, respectively. After subsequent sequence optimization, we successfully generated KRpep-2d (Ac-RRRRCPLYISYDPVCRRRR-NH 2 ) that inhibited enzyme activity of K-Ras(G12D) with IC 50 = 1.6 nM and significantly suppressed ERK-phosphorylation, downstream of K-Ras(G12D), along with A427 cancer cell proliferation at 30 μM peptide concentration. To our knowledge, this is the first report of a K-Ras(G12D)-selective inhibitor, contributing to the development and study of K-Ras(G12D)-targeting drugs. [ABSTRACT FROM AUTHOR]

The phospholipid hydroperoxidase glutathione peroxidase (GPX4) is an enzyme that reduces lipid hydroperoxides in lipid membranes. Recently, GPX4 has been investigated as a target molecule that induces iron-dependent cell death (ferroptosis) selectively in cancer cells that express mutant Ras. GPX4 inhibitors have the potential to become novel anti-cancer drugs. However, there are no druggable pockets for conventional small molecules on the molecular surface of GPX4. To generate GPX4 inhibitors, we examined the use of peptides as an alternative to small molecules. By screening peptide libraries displayed on T7 phages, and analyzing the X-ray crystal structures of the peptides, we successfully identified one peptide that binds to near Sec73 of catalytic site and two peptides that bind to another site on GPX4. To our knowledge, this is the first study reporting GPX4 inhibitory peptides and their structural information. [ABSTRACT FROM AUTHOR]

Abstract: Spiro-pyrazolidinedione derivatives without quaternary chiral center were discovered by structure-based drug design and characterized as potent acetyl-CoA carboxylase (ACC) inhibitors. The high metabolic stability of the spiro-pyrazolo[1,2-a]pyridazine scaffold and enhancement of the activity by incorporation of a 7-methoxy group on the benzothiophene core successfully led to the identification of compound 4c as an orally bioavailable and highly potent ACC inhibitor. Oral administration of 4c significantly decreased the values of the respiratory quotient in rats, indicating the stimulation of fatty acid oxidation. [Copyright &y& Elsevier]

Abstract: Spiro-lactone (S)-1 is a potent acetyl-CoA carboxylase (ACC) inhibitor and was found to be metabolically liable in human hepatic microsomes. To remove one of the risk factors in human study by improving the metabolic stability, we focused on modifying the spiro-lactone ring and the benzothiophene portion of the molecule. Spiro-imide derivative 8c containing a 6-methylthieno[2,3-b]pyridine core exhibited potent ACC inhibitory activity and favorable pharmacokinetic profiles in rats. [Copyright &y& Elsevier]

Abstract: The co-crystal structure of the human acetyl-coenzyme A 2 (ACC2) carboxyl transferase domain and the reported compound CP-640186 (1b) suggested that two carbonyl groups are essential for potent ACC2 inhibition. By focusing on enhancing the interactions between the two carbonyl groups and the amino acid residues Gly2162 and Glu2230, we used ligand- and structure-based drug design to discover spirolactones bearing a 2-ureidobenzothiophene moiety [Copyright &y& Elsevier]