ABSTRACT
Dysfunction of the circadian clock has been implicated in the pathogenesis of cardiovascular disease. The CLOCK protein is a core molecular component of the circadian oscillator, so that mice with a mutated Clock gene (Clk/Clk) exhibit abnormal rhythms in numerous physiological processes. However, here we report that chronic kidney disease (CKD)-induced cardiac inflammation and fibrosis are attenuated in Clk/Clk mice even though they have high blood pressure and increased serum angiotensin II levels. A search for the underlying cause of the attenuation of heart disorder in Clk/Clk mice with 5/6 nephrectomy (5/6Nx) led to identification of the monocytic expression of G protein-coupled receptor 68 (GPR68) as a risk factor of CKD-induced inflammation and fibrosis of heart. 5/6Nx induces the expression of GPR68 in circulating monocytes via altered CLOCK activation by increasing serum levels of retinol and its binding protein (RBP4). The high-GPR68-expressing monocytes have increased potential for producing inflammatory cytokines, and their cardiac infiltration under CKD conditions exacerbates inflammation and fibrosis of heart. Serum retinol and RBP4 levels in CKD patients are also sufficient to induce the expression of GPR68 in human monocytes. Our present study reveals an uncovered role of monocytic clock genes in CKD-induced heart failure.
Subject(s)
CLOCK Proteins/genetics , Circadian Clocks/genetics , Circadian Rhythm/physiology , Heart Diseases/pathology , Monocytes/metabolism , Renal Insufficiency, Chronic/pathology , ARNTL Transcription Factors/metabolism , Animals , CLOCK Proteins/metabolism , Cells, Cultured , Circadian Rhythm/genetics , Cytokines/biosynthesis , Fibrosis/pathology , Hypertension/genetics , Hypertension/pathology , Inflammation/genetics , Inflammation/pathology , Male , Mice , Mice, Inbred C57BL , Mice, Inbred ICR , Mice, Transgenic , Receptors, G-Protein-Coupled/metabolismABSTRACT
Irreversible inhibition of disease-associated proteins with small molecules is a powerful approach for achieving increased and sustained pharmacological potency. Here, we introduce α-chlorofluoroacetamide (CFA) as a novel warhead of targeted covalent inhibitor (TCI). Despite weak intrinsic reactivity, CFA-appended quinazoline showed high reactivity toward Cys797 of epidermal growth factor receptor (EGFR). In cells, CFA-quinazoline showed higher target specificity for EGFR than the corresponding Michael acceptors in a wide concentration range (0.1-10 µM). The cysteine adduct of the CFA derivative was susceptible to hydrolysis and reversibly yielded intact thiol but was stable in solvent-sequestered ATP-binding pocket of EGFR. This environment-dependent hydrolysis can potentially reduce off-target protein modification by CFA-based drugs. Oral administration of CFA quinazoline NS-062 significantly suppressed tumor growth in a mouse xenograft model. Further, CFA-appended pyrazolopyrimidine irreversibly inhibited Bruton's tyrosine kinase with higher target specificity. These results demonstrate the utility of CFA as a new class warheads for TCI.
Subject(s)
Acetamides/chemical synthesis , Cysteine/metabolism , Quinazolines/chemical synthesis , Acetamides/chemistry , Acetamides/pharmacology , Animals , Antineoplastic Agents , Cell Line , ErbB Receptors , Humans , Mice , Mice, Nude , Neoplasms , Phosphotransferases/physiology , Protein Kinase Inhibitors/metabolism , Protein Kinase Inhibitors/pharmacology , Pyrimidines/antagonists & inhibitors , Quinazolines/chemistry , Structure-Activity Relationship , Xenograft Model Antitumor AssaysABSTRACT
Clock genes encoding transcription factors that regulate circadian rhythms may inform chronomodulated chemotherapy, where time-dependent dose alterations might affect drug efficacy and reduce side effects. For example, inhibiting the essential cystine transporter xCT with sulfasalazine induces growth arrest in cancer cells. Although the anticancer effects of sulfasalazine have been studied extensively, its effects on transcriptional control of xCT expression have not been studied. Here, we show that sulfasalazine administration during the period of increased xCT expression improves its anticancer effects and that the Clock gene itself induces xCT expression and regulates its circadian rhythm. Our findings highlight the clinical potential of chronomodulated chemotherapy and the importance of xCT-mediated transcriptional regulation in the utility of such strategies. Cancer Res; 77(23); 6603-13. ©2017 AACR.
Subject(s)
ARNTL Transcription Factors/genetics , Amino Acid Transport System y+/antagonists & inhibitors , Antineoplastic Agents/pharmacology , Circadian Clocks/physiology , Drug Chronotherapy , Sulfasalazine/pharmacology , Animals , Cell Line, Tumor , Cell Proliferation/drug effects , Gene Expression Regulation , Male , Mice , Mice, Inbred BALB C , Mice, Inbred ICR , Organoplatinum Compounds/pharmacology , Oxaliplatin , RNA Interference , RNA, Small Interfering/genetics , Transcription, Genetic/geneticsABSTRACT
Chronic kidney disease (CKD) is a global health problem, and novel therapies to treat CKD are urgently needed. Here, we show that inhibition of G0/G1 switch 2 (G0s2) ameliorates renal inflammation in a mouse model of CKD. Renal expression of chemokine (C-C motif) ligand 2 (Ccl2) was increased in response to p65 activation in the kidneys of wild-type 5/6 nephrectomy (5/6Nx) mice. Moreover, 5/6Nx Clk/Clk mice, which carry homozygous mutations in the gene encoding circadian locomotor output cycles kaput (CLOCK), did not exhibit aggravation of apoptosis or induction of F4/80-positive cells. The renal expression of G0s2 in wild-type 5/6Nx mice was important for the transactivation of Ccl2 by p65. These pathologies were ameliorated by G0s2 knockdown. Furthermore, a novel small-molecule inhibitor of G0s2 expression was identified by high-throughput chemical screening, and the inhibitor suppressed renal inflammation in 5/6Nx mice. These findings indicated that G0s2 inhibitors may have applications in the treatment of CKD.