LATS2 degradation promoted fibrosis damage and rescued by vitamin K3 in lupus nephritis.

BACKGROUND: Lupus nephritis (LN) is the most common complication of systemic lupus erythematosus (SLE). The limited treatment options for LN increase the economic burdens on patients. Because fibrotic progression leads to irreversible renal damage in LN patients and further progresses to chronic kidney disease (CKD) and the end stage of renal disease (ESRD), developing new targets to prevent LN fibrotic progression could lead to a feasible treatment strategy for LN patients. METHODS: In this study, we examined YAP activation and LATS2 downregulation in LN kidney biopsy samples (LN: n = 8, normal: n = 2) and lupus-prone MRL/lpr mice (n = 8 for each disease stage). The function of LATS2 was further investigated by in situ injection of Ad-LATS2 into mice with LN (n = 6 mice per group). We examined the role of SIAH2-LATS2 regulation by IP-MS and co-IP, and the protective effect of the SIAH2 inhibitor was investigated in mice with LN. RESULTS: Restoring LATS2 by an adenovirus in vivo alleviated renal fibrotic damage in mice with LN. Moreover, we found that LATS2 was degraded by a K48 ubiquitination-proteasome pathway mediated by SIAH2 and promoted YAP activation to worsen fibrosis progression in LN. The H150 region of the substrate binding domain (SBD) is an important site for SIAH2-LATS2 binding. The SIAH2-specific inhibitor vitamin K3 protected against LN-associated fibrotic damage in vivo. CONCLUSION: In summary, we identified the SIAH2-LATS2 axis as an attractive intervention target in LN to alter the resistance to fibrosis.

An allosteric modulator of the adenosine A1 receptor potentiates the antilipolytic effect in rat adipose tissue.

The adenosine A1 receptor plays important roles in tuning free fatty acid (FFA) levels and represents an attractive target for metabolic disorders. Though remarkable progress has been achieved in the exploitation of effective (orthosteric) A1 receptor agonists in modulating aberrant FFA levels, the effect of A1 receptor allosteric modulation on lipid homeostasis is less investigated. Herein we sought to explore the effect of an allosteric modulator on the action of an A1 receptor orthosteric agonist in regulating the lipolytic process in vitro and in vivo. We examined the binding kinetics of a selective A1 receptor agonist 2-chloro-N6-cyclopentyladenosine (CCPA) in the absence or presence of an allosteric modulator (2-amino-4,5-dimethyl-3-thienyl)-[3-(trifluoromethyl)-phenyl]methanone (PD81,723) on rat adipocyte membranes. We also examined the allosteric effects of PD81,723 on mediating the CCPA-induced inhibition of cAMP accumulation, HSL (hormone-sensitive lipase) phosphorylation and FFA production in in vitro and in vivo models. Our results demonstrated that PD81,723 slowed down the dissociation of CCPA from the A1 receptor, which, consequently, potentiated the antilipolytic action of CCPA through downregulating the cAMP/HSL pathway. Our study exemplified the application of A1 receptor allosteric modulators as an alternative for metabolic disease treatments.

Structure-Affinity and Structure-Kinetic Relationship Studies of Benzodiazepine Derivatives for the Development of Efficacious Vasopressin V2 Receptor Antagonists.

Vasopressin V2 receptors (V2R) are a promising drug target for autosomal dominant polycystic kidney disease (ADPKD). As previous research demonstrated that the residence time of V2R antagonists is critical to their efficacy in both ex vivo and in vivo models of ADPKD, we performed extensive structure-kinetic relationship (SKR) analyses on a series of benzodiazepine derivatives. We found that subtle structural modifications of the benzodiazepine derivatives dramatically changed their binding kinetics but not their affinity. Compound 18 exhibited a residence time of 77 min, which was 7.7-fold longer than that of the reference compound tolvaptan (TVP). Accordingly, compound 18 exhibited higher efficacy compared to TVP in an in vivo model of ADPKD. Overall, our study exemplifies a kinetics-directed medicinal chemistry effort for the development of efficacious V2R antagonists. We envision that this strategy may also have general applicability in other therapeutic areas.

Optical-Controlled Kinetic Switch: Fine-Tuning of the Residence Time of an Antagonist Binding to the Vasopressin V2 Receptor in In Vitro, Ex Vivo, and In Vivo Models of ADPKD.

The pharmacological activity of a small-molecule ligand is linked to its receptor residence time. Therefore, precise control of the duration for which a ligand binds to its receptor is highly desirable. Herein, we designed photoswitchable ligands targeting the vasopressin V2 receptor (V2R), a validated target for autosomal dominant polycystic kidney disease (ADPKD). We adapted the photoswitching trait of azobenzene to the parent V2R antagonist lixivaptan (LP) to generate azobenzene lixivaptan derivatives (aLPs). Among them, aLPs-5g was a potential optical-controlled kinetic switch. Upon irradiation, cis-aLPs-5g displayed a 4.3-fold prolonged V2R residence time compared to its thermally stable trans configuration. The optical-controlled kinetic variations led to distinct inhibitory effects on cellular functional readout. Furthermore, conversion of the cis/trans isomer of aLPs-5g resulted in different efficacies of inhibiting renal cystogenesis ex vivo and in vivo. Overall, aLPs-5g represents a photoswitch for precise control of ligand-receptor residence time and, consequently, the pharmacological activity.

Discovery of Novel N-(5-(Pyridin-3-yl)-1H-indazol-3-yl)benzamide Derivatives as Potent Cyclin-Dependent Kinase 7 Inhibitors for the Treatment of Autosomal Dominant Polycystic Kidney Disease.

Recent evidence suggests that CDK7 is a novel potential drug target for autosomal dominant polycystic kidney disease (ADPKD) treatment. Herein, on the basis of structural analysis, a hit compound 3 with a novel scaffold was designed and subsequent medicinal chemistry efforts by a rational design strategy were conducted to improve CDK7 inhibitors' potency and selectivity. The representative compound B2 potently inhibited CDK7 with an IC50 value of 4 nM and showed high selectivity over CDKs. Compound B2 showed high potency to inhibit cyst growth and exhibited lower cytotoxicity than THZ1 in an in vitro Madin-Darby canine kidney cyst model. In addition, compound B2 was also highly efficacious in suppressing renal cyst development in an ex vivo embryonic kidney cyst model and in vivo ADPKD mouse model. These results indicate that compound B2 represents a promising lead compound that deserves further investigation to discover novel therapeutic agents for ADPKD.

PF-06409577 inhibits renal cyst progression by concurrently inhibiting the mTOR pathway and CFTR channel activity.

Renal cyst development and expansion in autosomal dominant polycystic kidney disease (ADPKD) involves over-proliferation of cyst-lining epithelial cells and excessive cystic fluid secretion. While metformin effectively inhibits renal cyst growth in mouse models of ADPKD it exhibits low potency, and thus an adenosine monophosphate-activated protein kinase (AMPK) activator with higher potency is required. Herein, we adopted a drug repurposing strategy to explore the potential of PF-06409577, an AMPK activator for diabetic nephropathy, in cellular, ex vivo and in vivo models of ADPKD. Our results demonstrated that PF-06409577 effectively down-regulated mammalian target of rapamycin pathway-mediated proliferation of cyst-lining epithelial cells and reduced cystic fibrosis transmembrane conductance regulator-regulated cystic fluid secretion. Overall, our data suggest that PF-06409577 holds therapeutic potential for ADPKD treatment.

Benzodiazepine Derivatives as Potent Vasopressin V2 Receptor Antagonists for the Treatment of Autosomal Dominant Kidney Disease.

Cyst formation and enlargement in autosomal dominant kidney disease (ADPKD) is mainly driven by aberrantly increased cytosolic cAMP in renal tubule epithelial cells. Because the vasopressin V2 receptor (V2R) regulates intracellular cAMP levels in kidneys, a series of benzodiazepine derivatives were developed targeting the V2R. Among these derivatives, compound 25 exhibited potent binding affinity to the V2R (Ki = 9.0 ± 1.5 nM) and efficacious cAMP inhibition (IC50 = 9.2 ± 3.0 nM). This led to the suppression of cyst formation and growth in both an MDCK cell model and an embryonic kidney cyst model. Further advancing compound 25 in a murine model of ADPKD demonstrated a significantly improved in vivo efficacy compared with the reference compound tolvaptan. Overall, compound 25 holds therapeutic potential for the treatment of ADPKD.

Long Residence Time at the Vasopressin V2 Receptor Translates into Superior Inhibitory Effects in Ex Vivo and In Vivo Models of Autosomal Dominant Polycystic Kidney Disease.

Prevailing strategies directing early-phase drug discovery heavily rely on equilibrium-based metrics such as affinity, which overlooks the kinetic process of a drug molecule interacting with its target. Herein, we developed a number of vasopressin V2 receptor (V2R) antagonists with divergent binding affinities and kinetics for autosomal dominant polycystic kidney disease (ADPKD). Surprisingly, the residence time of the V2R antagonists, but not their affinity, was correlated with the efficacy in both ex vivo and in vivo models of ADPKD. We envision that the kinetics-directed drug candidate selection and development may have general applicability for ADPKD and other therapeutic areas as well.