Discovery and preclinical evaluations of TQB3616, a novel CDK4-biased inhibitor.

Among small-molecule CDK4/6 inhibitors (palbociclib, ribociclib, and abemaciclib) approved for metastatic breast cancers, abemaciclib has a more tolerable adverse effects in clinic. This is attributable to preferential inhibition of CDK4 over CDK6. In our search for a biased CDK4 inhibitor, we discovered a series of pyrimidine-indazole inhibitors. SAR studies led us to TQB3616 as a preferential CDK4 inhibitor. TQB3616 exhibited improvements in both enzymatic and cellular proliferation inhibitory potency when tested side-by-side with the FDA approved palbociclib and abemaciclib. TQB3616 also possessed favorable PK profile in multiple species. These differentiated properties, together with excellent GLP safety profile warranted TQB3616 moving to clinic. TQB3616 entered into clinical development in 2019 and currently in phase III clinical trials (NCT05375461, NCT05365178).

Discovery and preclinical evaluations of GST-HG131, a novel HBV antigen inhibitor for the treatment of chronic hepatitis B infection.

Chronic hepatitis B (CHB) remains a significant health challenge worldwide. The current treatments for CHB achieve less than 10% cure rates, majority of the patients are on therapy for life. Therefore, cure of CHB is a high unmet medical need. HBV surface antigen (HBsAg) loss and seroconversion are considered as the key for the cure. RG7834 is a novel, orally bioavailable small molecule reported to reduce HBV antigens. Based on RG7834 chemistry, we designed and discovered a series of dihydrobenzopyridooxazepine (DBP) series of HBV antigen inhibitors. Extensive SAR studies led us to GST-HG131 with excellent reduction of HBV antigens (both HBsAg and HBeAg) in vitro and in vivo. GST-HG131 improved safety in rat toxicology studies over RG7834. The promising inhibitory activity, together with animal safety enhancement, merited GST-HG131 progressed into clinical development in 2020 (NCT04499443).

Discovery and preclinical evaluations of JBD0131, a novel nitrodihydro-imidazooxazole anti-tuberculosis agent.

Multidrug-resistant pulmonary tuberculosis (MDR-TB) is a major health problem worldwide. The treatment for MDR-TB requires medications for a long duration (up to 20-24 months) with second-line drugs resulting in unfavorable outcomes. Nitroimidazoles are promising antimycobacterial agents known to inhibit both aerobic and anaerobic mycobacterial activity. Delamanid and pretomanid are two nitroimidazoles approved by the regulatory agencies for MDR-TB treatment. However, both agents possess unsatisfactory absorption and QTc prolongation. In our search for a safer nitroimidazole, we discovered JBD0131 (2). It exhibited excellent anti-mycobacterial activity against M. tuberculosis H37Rv in vitro and in vivo, improved PK and absorption, reduced QT prolongation potential of delamanid. JBD0131 is currently in clinical development in China for pulmonary tuberculosis (CTR20202308).

Discovery and preclinical profile of sudapyridine (WX-081), a novel anti-tuberculosis agent.

Multidrug resistant tuberculosis (MDR-TB) remains a major human health challenge. Bedaquiline was approved in 2012 by the US FDA, and listed by WHO as a treatment for multidrug-resistant tuberculosis (MDR-TB) in 2018. However, the side effects of bedaquiline including the risk of unexplained mortality, QTc prolongation and hepatotoxicity limit its wide clinical use. Based on bedaquiline, we describe herein discovery and development of a novel diarylpyridine series, which led to identification of WX-081 (sudapyridine, 21l). It displayed excellent anti-mycobacterial activity against M. tuberculosis H37Rv in vitro and in vivo and low cytotoxicity; additionally WX-081 had excellent pharmacokinetic parameters in animals, better lung exposure and lower QTc prolongation potential compared to bedaquiline. WX-081 is currently under clinical phase II development (NCT04608955).

Discovery and preclinical profile of LX-039, a novel indole-based oral selective estrogen receptor degrader (SERD).

We previously described the discovery of a novel indole series compounds as oral SERD for ER positive breast cancer treatment. Further SAR exploration focusing on substitutions on indole moiety of compound 12 led to the discovery of a clinical candidate LX-039. We report herein its profound anti-tumor activity, desirable ER antagonistic characteristics combined with favorable pharmacokinetic and preliminary safety properties. LX-039 is currently in clinical trial (NCT04097756).

Discovery and preclinical evaluations of WX-0593, a novel ALK inhibitor targeting crizotinib-resistant mutations.

ALK gene rearrangements are oncogenic drivers in approximately 5% of NSCLC. Crizotinib, a first generation ALK inhibitor, is widely prescribed for ALK-positive NSCLC in clinic. Resistance to crizotinib and other ALK inhibitors has been problematic. Addressing resistance, here we describe discovery and development of a novel, proprietary spirocyclic diamine-substituted aryl phosphine oxide series of inhibitors, which led to the identification of WX-0593 (16a) as a potent ALK inhibitor. WX-0593 inhibited the activity of both wild type and resistant mutants of ALK in vitro, showed strong antitumor activity in a crizotinib-resistant mouse PDX model. WX-0593 is currently under development in phase II/III clinical trials.

Design, syntheses and evaluations of novel indole derivatives as orally selective estrogen receptor degraders (SERD).

Most estrogen receptor positive (ER +) breast cancers depend on ER signaling pathway to develop. Clinical application of SERD fulvestrant effectively degraded ER, blocked its function and prolonged progression free survival of ER + breast cancer patients. However, current SERD suffers from limited bioavailability, therefore is given as intramuscular (IM) injection. In this paper, we report herein a novel indole series compounds with nanomolar range ER degradation potencies and oral systemic exposures. Selected compounds suppressed tumor growth in vivo in ER + MCF7 breast cancer CDX model via p.o. administration. All those data supported further optimizations of this analog to develop preclinical candidate as oral SERD for ER + breast cancer's treatment.

Therapeutic Effect and Mechanisms of the Novel Monosulfactam 0073.

This study aimed to evaluate the antimicrobial activity of the novel monosulfactam 0073 against multidrug-resistant Gram-negative bacteria in vitro and in vivo and to characterize the mechanisms underlying 0073 activity. The in vitro activities of 0073, aztreonam, and the combination with avibactam were assessed by MIC and time-kill assays. The safety of 0073 was evaluated using 3-(4,5-dimethylthizol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and acute toxicity assays. Murine thigh infection and pneumonia models were employed to define in vivo efficacy. A penicillin-binding protein (PBP) competition assay and confocal microscopy were conducted. The inhibitory action of 0073 against ß-lactamases was evaluated by the half-maximal inhibitory concentration (IC50), and resistance development was evaluated via serial passage. The monosulfactam 0073 showed promising antimicrobial activity against Enterobacteriaceae, Pseudomonas aeruginosa, and Acinetobacter baumannii isolates producing metallo-ß-lactamases (MBLs) and serine ß-lactamases. In preliminary experiments, compound 0073 exhibited safety both in vitro and in vivo In the murine thigh infection model and the pneumonia models in which infection was induced by P. aeruginosa and Klebsiella pneumoniae, 0073 significantly reduced the bacterial burden. Compound 0073 targeted several PBPs and exerted inhibitory effects against some serine ß-lactamases. Finally, 0073 showed a reduced propensity for resistance selection compared with that of aztreonam. The novel monosulfactam 0073 exhibited increased activity against ß-lactamase-producing Gram-negative organisms compared with the activity of aztreonam and showed good safety profiles both in vitro and in vivo The underlying mechanisms may be attributed to the affinity of 0073 for several PBPs and its inhibitory activity against some serine ß-lactamases. These data indicate that 0073 represents a potential treatment for infections caused by ß-lactamase-producing multidrug-resistant bacteria.

Synergistic Activity of Nitroimidazole-Oxazolidinone Conjugates against Anaerobic Bacteria.

The introductions of the bicyclic 4-nitroimidazole and the oxazolidinone classes of antimicrobial agents represented the most significant advancements in the infectious disease area during the past two decades. Pretomanid, a bicyclic 4-nitroimidazole, and linezolid, an oxazolidinone, are also part of a combination regimen approved recently by the US Food and Drug Administration for the treatment of pulmonary, extensively drug resistant (XDR), treatment-intolerant or nonresponsive multidrug-resistant (MDR) Mycobacterium tuberculosis (TB). To identify new antimicrobial agents with reduced propensity for the development of resistance, a series of dual-acting nitroimidazole-oxazolidinone conjugates were designed, synthesized and evaluated for their antimicrobial activity. Compounds in this conjugate series have shown synergistic activity against a panel of anaerobic bacteria, including those responsible for serious bacterial infections.

Discovery and evaluation of novel nitrodihydroimidazooxazoles as promising anti-tuberculosis agents.

New analogues of antitubercular drug Delamanid were prepared, seeking drug candidates with enhanced aqueous solubility and high efficacy. The strategy involved replacement of phenoxy linker proximal to the 2-nitroimidazooxazole of Delamanid by piperidine fused 5 or 6-membered ring heterocycles (ring A). The new compounds were all more hydrophilic than Delamanid, and several class of analogues showed remarkable activities against M. bovis. And among these series, the tetrahydro-naphthyridine-linked nitroimidazoles displayed excellent antimycobacterial activity against both replicating (MABA) and nonreplicating (LORA) M. tb H37Rv and low cytotoxicity. Compared to Delamanid, these new compounds (6, 7, 45) demonstrated dramatically improved physicochemical properties and are suitable for further in vitro and in vivo evaluation.

Cell Active Hydroxylactam Inhibitors of Human Lactate Dehydrogenase with Oral Bioavailability in Mice.

A series of trisubstituted hydroxylactams was identified as potent enzymatic and cellular inhibitors of human lactate dehydrogenase A. Utilizing structure-based design and physical property optimization, multiple inhibitors were discovered with <10 µM lactate IC50 in a MiaPaca2 cell line. Optimization of the series led to 29, a potent cell active molecule (MiaPaca2 IC50 = 0.67 µM) that also possessed good exposure when dosed orally to mice.

Discovery of Novel Oral Protein Synthesis Inhibitors of Mycobacterium tuberculosis That Target Leucyl-tRNA Synthetase.

The recent development and spread of extensively drug-resistant and totally drug-resistant resistant (TDR) strains of Mycobacterium tuberculosis highlight the need for new antitubercular drugs. Protein synthesis inhibitors have played an important role in the treatment of tuberculosis (TB) starting with the inclusion of streptomycin in the first combination therapies. Although parenteral aminoglycosides are a key component of therapy for multidrug-resistant TB, the oxazolidinone linezolid is the only orally available protein synthesis inhibitor that is effective against TB. Here, we show that small-molecule inhibitors of aminoacyl-tRNA synthetases (AARSs), which are known to be excellent antibacterial protein synthesis targets, are orally bioavailable and effective against M. tuberculosis in TB mouse infection models. We applied the oxaborole tRNA-trapping (OBORT) mechanism, which was first developed to target fungal cytoplasmic leucyl-tRNA synthetase (LeuRS), to M. tuberculosis LeuRS. X-ray crystallography was used to guide the design of LeuRS inhibitors that have good biochemical potency and excellent whole-cell activity against M. tuberculosis Importantly, their good oral bioavailability translates into in vivo efficacy in both the acute and chronic mouse models of TB with potency comparable to that of the frontline drug isoniazid.

Structure-Based Design of GNE-495, a Potent and Selective MAP4K4 Inhibitor with Efficacy in Retinal Angiogenesis.

Diverse biological roles for mitogen-activated protein kinase kinase kinase kinase 4 (MAP4K4) have necessitated the identification of potent inhibitors in order to study its function in various disease contexts. In particular, compounds that can be used to carry out such studies in vivo would be critical for elucidating the potential for therapeutic intervention. A structure-based design effort coupled with property-guided optimization directed at minimizing the ability of the inhibitors to cross into the CNS led to an advanced compound 13 (GNE-495) that showed excellent potency and good PK and was used to demonstrate in vivo efficacy in a retinal angiogenesis model recapitulating effects that were observed in the inducible Map4k4 knockout mice.

Synthesis and biological evaluation of novel pyrrolidine acid analogs as potent dual PPARα/γ agonists.

The design, synthesis and structure-activity relationships of a novel series of 3,4-disubstituted pyrrolidine acid analogs as PPAR ligands is outlined. In both the 1,3- and 1,4-oxybenzyl pyrrolidine acid series, the preferred stereochemistry was shown to be the cis-3R,4S isomer, as exemplified by the potent dual PPARα/γ agonists 3k and 4i. The N-4-trifluoromethyl-pyrimidinyl pyrrolidine acid analog 4i was efficacious in lowering fasting glucose and triglyceride levels in diabetic db/db mice.

Optimization of 5-(2,6-dichlorophenyl)-3-hydroxy-2-mercaptocyclohex-2-enones as potent inhibitors of human lactate dehydrogenase.

Optimization of 5-(2,6-dichlorophenyl)-3-hydroxy-2-mercaptocyclohex-2-enone using structure-based design strategies resulted in inhibitors with considerable improvement in biochemical potency against human lactate dehydrogenase A (LDHA). These potent inhibitors were typically selective for LDHA over LDHB isoform (4­10 fold) and other structurally related malate dehydrogenases, MDH1 and MDH2 (>500 fold). An X-ray crystal structure of enzymatically most potent molecule bound to LDHA revealed two additional interactions associated with enhanced biochemical potency.

Identification of 3,6-disubstituted dihydropyrones as inhibitors of human lactate dehydrogenase.

A series of 3,6-disubstituted dihydropyrones were identified as inhibitors of human lactate dehydrogenase (LDH)-A. Structure activity relationships were explored and a series of 6,6-spiro analogs led to improvements in LDHA potency (IC50 <350 nM). An X-ray crystal structure of an improved compound bound to human LDHA was obtained and it illustrated additional opportunities to enhance the potency of these compounds, resulting in the identification of 51 (IC50=30 nM).

Identification of substituted 3-hydroxy-2-mercaptocyclohex-2-enones as potent inhibitors of human lactate dehydrogenase.

A novel class of 3-hydroxy-2-mercaptocyclohex-2-enone-containing inhibitors of human lactate dehydrogenase (LDH) was identified through a high-throughput screening approach. Biochemical and surface plasmon resonance experiments performed with a screening hit (LDHA IC50=1.7 µM) indicated that the compound specifically associated with human LDHA in a manner that required simultaneous binding of the NADH co-factor. Structural variation of this screening hit resulted in significant improvements in LDHA biochemical inhibition activity (best IC50=0.18 µM). Two crystal structures of optimized compounds bound to human LDHA were obtained and explained many of the observed structure-activity relationships. In addition, an optimized inhibitor exhibited good pharmacokinetic properties after oral administration to rats (F=45%).

Identification of 2-amino-5-aryl-pyrazines as inhibitors of human lactate dehydrogenase.

A 2-amino-5-aryl-pyrazine was identified as an inhibitor of human lactate dehydrogenase A (LDHA) via a biochemical screening campaign. Biochemical and biophysical experiments demonstrated that the compound specifically interacted with human LDHA. Structural variation of the screening hit resulted in improvements in LDHA biochemical inhibition and pharmacokinetic properties. A crystal structure of an improved compound bound to human LDHA was also obtained and it explained many of the observed structure-activity relationships.

Identification of substituted 2-thio-6-oxo-1,6-dihydropyrimidines as inhibitors of human lactate dehydrogenase.

A novel 2-thio-6-oxo-1,6-dihydropyrimidine-containing inhibitor of human lactate dehydrogenase (LDH) was identified by high-throughput screening (IC50=8.1 µM). Biochemical, surface plasmon resonance, and saturation transfer difference NMR experiments indicated that the compound specifically associated with human LDHA in a manner that required simultaneous binding of the NADH co-factor. Structural variation of the screening hit resulted in significant improvements in LDHA biochemical inhibition activity (best IC50=0.48 µM). A crystal structure of an optimized compound bound to human LDHA was obtained and explained many of the observed structure-activity relationships.

Synthesis and antibacterial evaluation of a novel tricyclic oxaborole-fused fluoroquinolone.

We have designed and synthesized a novel class of compounds based on fluoroquinolone antibacterial prototype. The design concept involved the replacement of the 3-carboxylic acid in ciprofloxacin with an oxaborole-fused ring as an acid-mimicking group. The synthetic method employed in this work provides a good example of incorporating boron atom in complex molecules with multiple functional groups. The antibacterial activity of the newly synthesized compounds has been evaluated.