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Drugs targeting adenosine receptors (AR) can provide treatment for diseases. We report the identification of 7-(phenylamino)-pyrazolo[3,4

Undesirable activation of endosomal toll-like receptors TLR7 and TLR9 present in specific immune cells in response to host-derived ligands is implicated in several autoimmune diseases and other contexts of autoreactive inflammation, making them important therapeutic targets. We report a drug development strategy identifying a new chemotype for incorporating relevant structural subunits into the basic imidazopyridine core deemed necessary for potent TLR7 and TLR9 dual antagonism. We established minimal pharmacophoric features in the core followed by hit-to-lead optimization, guided by in vitro and in vivo biological assays and ADME. A ligand-receptor binding hypothesis was proposed, and selectivity studies against TLR8 were performed. Oral absorption and efficacy of lead candidate

Selective inhibition of the angiotensin-converting enzyme C-domain (cACE) and neprilysin (NEP), leaving the ACE N-domain (nACE) free to degrade bradykinin and other peptides, has the potential to provide the potent antihypertensive and cardioprotective benefits observed for nonselective dual ACE/NEP inhibitors, such as omapatrilat, without the increased risk of adverse effects. We have synthesized three 1-carboxy-3-phenylpropyl dipeptide inhibitors with nanomolar potency based on the previously reported C-domain selective ACE inhibitor lisinopril-tryptophan (LisW) to probe the structural requirements for potent dual cACE/NEP inhibition. Here we report the synthesis, enzyme kinetic data, and high-resolution crystal structures of these inhibitors bound to nACE and cACE, providing valuable insight into the factors driving potency and selectivity. Overall, these results highlight the importance of the interplay between the S1′ and S2′ subsites for ACE domain selectivity, providing guidance for future chemistry efforts toward the development of dual cACE/NEP inhibitors.

The development of novel and safe insecticides remains an important need for a growing world population to protect crops and animal and human health. New chemotypes modulating the insect nicotinic acetylcholine receptors have been recently brought to the agricultural market, yet with limited understanding of their molecular interactions at their target receptor. Herein, we disclose the first crystal structures of these insecticides, namely, sulfoxaflor, flupyradifurone, triflumezopyrim, flupyrimin, and the experimental compound, dicloromezotiaz, in a double-mutated acetylcholine-binding protein which mimics the insect-ion-channel orthosteric site. Enabled by these findings, we discovered novel pharmacophores with a related mode of action, and we describe herein their design, synthesis, and biological evaluation.

Nowadays, the simultaneous inhibition of two or more pathways plays an increasingly important role in cancer treatment due to the complex and diverse pathogenesis of cancer, and the combination of the cyclin-dependent kinase 6 (CDK6) inhibitor and PIM1 inhibitor was found to generate synergistic effects in acute myeloid leukemia (AML) treatment. Therefore, we discovered a novel lead

Visceral leishmaniasis (VL) is a parasitic disease endemic across multiple regions of the world and is fatal if untreated. Current therapies are unsuitable, and there is an urgent need for safe, short-course, and low-cost oral treatments to combat this neglected disease. The benzoxaborole chemotype has previously delivered clinical candidates for the treatment of other parasitic diseases. Here, we describe the development and optimization of this series, leading to the identification of compounds with potent in vitro and in vivo antileishmanial activity. The lead compound (DNDI-6148) combines impressive in vivo efficacy (>98% reduction in parasite burden) with pharmaceutical properties suitable for onward development and an acceptable safety profile. Detailed mode of action studies confirm that DNDI-6148 acts principally through the inhibition of Leishmania cleavage and polyadenylation specificity factor (CPSF3) endonuclease. As a result of these studies and its promising profile, DNDI-6148 has been declared a preclinical candidate for the treatment of VL.

The FGF19-FGFR4 signaling pathway has been extensively studied as a promising target for the treatment of hepatocellular carcinoma (HCC). Several FGFR4-selective inhibitors have been developed, but none of them receives approval. Additionally, acquired resistance caused by FGFR4 gatekeeper mutations is emerging as a serious limitation for these targeted therapies. Herein, we report a novel series of 5-formyl-pyrrolo[3,2

When the nucleoside analogue acyclovir was introduced in the early 1980s, it presented a game-changing treatment modality for herpes simplex virus infections. Since then, work has been ongoing to improve the weaknesses that have now been identified: a narrow time window for therapeutic success, resistance in immunocompromised patients, little influence on frequency of recurrences, relatively fast elimination, and poor bioavailability. The present Drug Annotation focuses on the helicase-primase inhibitor pritelivir currently in development for the treatment of acyclovir-resistant HSV infections and describes how a change of the molecular target (from viral DNA polymerase to the HSV helicase-primase complex) afforded improvement of the shortcomings of nucleoside analogs. Details are presented for the discovery process leading to the final drug candidate, the pivotal preclinical studies on mechanism of action and efficacy, and on how ongoing clinical research has been able to translate preclinical promises into clinical use.

We review progress in the application of fragment-based drug discovery (FBDD) to epigenetic drug discovery (EPIDD) targeted at epigenetic writer and eraser enzymes as well as reader domains over the last 15 years. The greatest successes to date are in prospecting for bromodomain binding ligands. From a diverse array of fragment hits, multiple potent and selective compounds ensued, including the oncology clinical candidates mivebresib, ABBV-744, pelabresib, and PLX51107.

β-Site amyloid precursor protein-cleaving enzyme 1 (BACE1) is considered to be a promising target for treating Alzheimer's disease. However, all clinical BACE1 inhibitors have failed due to lack of efficacy, and some have even led to cognitive worsening. Recent evidence points to the importance of avoiding BACE2 inhibition along with careful dose titration. In this study, we focused on the fact that the 10s loop lining the S3 pocket in BACE1 can form both "open (up)" and "closed (down)" conformations, whereas in BACE2, it prefers to adopt a "closed" form; thus, more space is available in BACE1. By leveraging the difference, we designed fused pyridine analogues that could reach the 10s loop, leading to 6 with high selectivity and significant Aβ reduction. The cocrystal structures confirmed that 6 significantly increased B-factors of the 10s loop in BACE2 relative to those in BACE1. Thus, the destabilization of BACE2 seems to offer structural insights into the reduced BACE2 potency of 6, explaining the significant improvement in BACE1 selectivity.

Class F G protein-coupled receptors are characterized by a large extracellular domain (ECD) in addition to the common transmembrane domain (TMD) with seven α-helixes. For smoothened receptor (SMO), structural studies revealed dissected ECD and TMD, and their integrated assemblies. However, distinct assemblies were reported under different circumstances. Using an unbiased approach based on four series of cross-conjugated bitopic ligands, we explore the relationship between the active status and receptor assembly. Different activity dependency on the linker length for these bitopic ligands corroborates the various occurrences of SMO assembly. These results reveal a rigid "near" assembly for active SMO, which is in contrast to previous results. Conversely, inactive SMO adopts a free ECD, which would be remotely captured at "far" assembly by cholesterol. Altogether, we propose a mechanism of cholesterol flow-caused SMO activation involving an erection of ECD from far to near assembly.

We previously reported a potent tubulin inhibitor CH-2-77. In this study, we optimized the structure of CH-2-77 by blocking metabolically labile sites and synthesized a series of CH-2-77 analogues. Two compounds, 40a and 60c, preserved the potency while improving the metabolic stability over CH-2-77 by 3- to 4-fold (46.8 and 29.4 vs 10.8 min in human microsomes). We determined the high-resolution X-ray crystal structures of 40a (resolution 2.3 Å) and 60c (resolution 2.6 Å) in complex with tubulin and confirmed their direct binding at the colchicine-binding site. In vitro, 60c maintained its mode of action by inhibiting tubulin polymerization and was effective against P-glycoprotein-mediated multiple drug resistance and taxol resistance. In vivo, 60c exhibited a strong inhibitory effect on tumor growth and metastasis in a taxol-resistant A375/TxR xenograft model without obvious toxicity. Collectively, this work showed that 60c is a promising lead compound for further development as a potential anticancer agent.

Few targeted drugs were approved for treatment of colorectal cancer (CRC). Cyclin-dependent kinase 8 played a vital role in regulating transcription and was a key colorectal oncogene associated to colorectal cancer. Here, through de novo drug design and in depth structure-activity relationship analysis, title compound

Activated factor XI (FXIa) inhibitors are promising novel anticoagulants with low bleeding risk compared with current anticoagulants. The discovery of potent FXIa inhibitors with good oral bioavailability has been challenging. Herein, we describe our discovery effort, utilizing nonclassical interactions to improve potency, cellular permeability, and oral bioavailability by enhancing the binding while reducing polar atoms. Beginning with literature-inspired pyridine N-oxide-based FXIa inhibitor

Salt-inducible kinases (SIKs) are key metabolic regulators. The imbalance in SIK function is associated with the development of diverse cancers, including breast, gastric, and ovarian cancers. Chemical tools to clarify the roles of SIK in different diseases are, however, sparse and are generally characterized by poor kinome-wide selectivity. Here, we have adapted the pyrido[2,3-d]pyrimidin-7-one-based p21-activated kinase (PAK) inhibitor G-5555 for the targeting of SIK, by exploiting differences in the back-pocket region of these kinases. Optimization was supported by high-resolution crystal structures of G-5555 bound to the known off-targets, MST3 and MST4, leading to a chemical probe, MRIA9, with dual SIK/PAK activity and excellent selectivity over other kinases. Furthermore, we show that MRIA9 sensitizes ovarian cancer cells to treatment with the mitotic agent paclitaxel, confirming earlier data from genetic knockdown studies and suggesting a combination therapy with SIK inhibitors and paclitaxel for the treatment of paclitaxel-resistant ovarian cancer.

The enterovirus genus of the picornavirus family contains many important human pathogens. EV-D68 primarily infects children, and the disease manifestations range from respiratory illnesses to neurological complications such as acute flaccid myelitis (AFM). EV-A71 is a major pathogen for the hand, foot, and mouth disease (HFMD) in children and can also lead to AFM and death in severe cases. CVB3 infection can cause cardiac arrhythmias, acute heart failure, as well as type 1 diabetes. There is currently no FDA-approved antiviral for any of these enteroviruses. In this study, we report our discovery and development of pyrazolopyridine-containing small molecules with potent and broad-spectrum antiviral activity against multiple strains of EV-D68, EV-A71, and CVB3. Serial viral passage experiments, coupled with reverse genetics and thermal shift binding assays, suggested that these molecules target the viral protein 2C. Overall, the pyrazolopyridine inhibitors represent a promising class of candidates for the urgently needed non-polio enterovirus antivirals.

Activation of the toll-like receptors 7 and 8 has emerged as a promising strategy for cancer immunotherapy. Herein, we report the design and synthesis of a series of pyrido[3,2-d]pyrimidine-based toll-like receptor 7/8 dual agonists that exhibited potent and near-equivalent agonistic activities toward TLR7 and TLR8. In vitro, compounds 24e and 25a significantly induced the secretion of IFN-α, IFN-γ, TNF-α, IL-1β, IL-12p40, and IP-10 in human peripheral blood mononuclear cell assays. In vivo, compounds 24e, 24m, and 25a significantly suppressed tumor growth in CT26 tumor-bearing mice by remodeling the tumor microenvironment. Additionally, compounds 24e, 24m, and 25a markedly improved the antitumor activity of PD-1/PD-L1 blockade. In particular, compound 24e combined with the anti-PD-L1 antibody led to complete tumor regression. These results demonstrated that TLR7/8 agonists (24e, 24m, and 25a) held great potential as single agents or in combination with PD-1/PD-L1 blockade for cancer immunotherapy.

Drug reabsorption following biliary excretion is well-known as enterohepatic recirculation (EHR). Renal tubular reabsorption (RTR) following renal excretion is also common but not easily assessed. Intestinal excretion (IE) and enteroenteric recirculation (EER) have not been recognized as common disposition mechanisms for metabolically stable and permeable drugs. IE and intestinal reabsorption (IR:EHR/EER), as well as RTR, are governed by dug concentration gradients, passive diffusion, active transport, and metabolism, and together they markedly impact disposition and pharmacokinetics (PK) of small molecule drugs. Disruption of IE, IR, or RTR through applications of active charcoal (AC), transporter knockout (KO), and transporter inhibitors can lead to changes in PK parameters. The impacts of intestinal and renal reabsorption on PK are under-appreciated. Although IE and EER/RTR can be an intrinsic drug property, there is no apparent strategy to optimize compounds based on this property. This review seeks to improve understanding and applications of IE, IR, and RTR mechanisms.

Nuclear export factor chromosome region maintenance 1 (CRM1) is an attractive anticancer and antiviral drug target that spurred several research efforts to develop its inhibitor. Noncovalent CRM1 inhibitors are desirable, but none is reported to date. Here, we present the crystal structure of yeast CRM1 in complex with S109, a substructure of CBS9106 (under clinical test). Superimposition with the LFS-829 (another covalent CRM1 inhibitor) complex inspired the design of a noncovalent CRM1 inhibitor. Among nine synthesized compounds, noncovalent CRM1 inhibitor 1 (NCI-1) showed a high affinity to human and yeast CRM1 in the absence or presence of GST-bound Ras-related nuclear protein (RanGTP). Unlike covalent inhibitors, the crystal structure showed that NCI-1 is bound in the "open" nuclear export signal (NES) groove of CRM1, simultaneously occupying two hydrophobic pockets. NCI-1 additionally inhibited the nuclear export and proliferation of cells harboring the human CRM1-C528S mutant. Our work opens up the avenue of noncovalent CRM1 inhibitor development toward a more potent, less toxic, and broad-spectrum anticancer/antiviral therapy.

Terpyridine platinum (TP)-based chemotherapeutic agents target three-dimensional structures on DNA known as G-quadruplexes. We report the rational design and synthesis of a TP conjugate combined with copper-64 (64Cu), the decay characteristics of which include emission of β- and Auger electrons for radiotherapy and β+ particles for positron emission tomography (PET) imaging. The present experimental studies show that the novel [64Cu]Cu-1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA)-TP is stable, permitting selective killing of cancer cells. The antitumor activity of [64Cu]Cu-NOTA-TP at high apparent molar activity is in the low nanomolar range and 27,800-fold greater than that of natCu-NOTA-TP at 24 h post treatment. These results suggest that this combination of a cytotoxic TP agent with 64Cu has considerable potential for cancer treatment and PET imaging.

Despite the therapeutic relevance of δ-containing γ-aminobutyric acid type A receptors (GABAARs) and the need for δ-selective compounds, the structural determinants for the mode and molecular site of action of δ-selective positive allosteric modulator imidazo[1,2-a]pyridine DS2 remain elusive. To guide the quest for insight, we synthesized a series of DS2 analogues guided by a structural receptor model. Using a fluorescence-based fluorometric imaging plate reader membrane potential assay, we found that the δ-selectivity and the pharmacological profile are severely affected by substituents in the 5-position of the imidazopyridine core scaffold. Interestingly, the 5-methyl, 5-bromo, and 5-chloro DS2 analogues, 30, 35, and 36, were shown to be superior to DS2 at α4β1δ as mid-high nanomolar potency δ-selective allosteric modulators, displaying 6-16 times higher potency than DS2. Of these, 30 also displayed at least 60-fold selectivity for α4β1δ over α4β1γ2 receptor subtypes representing a potential tool for the selective characterization of δ-containing GABAARs in general.

The connection with acute myelogenous leukemia (AML) of dihydroorotate dehydrogenase (hDHODH), a key enzyme in pyrimidine biosynthesis, has attracted significant interest from pharma as a possible AML therapeutic target. We recently discovered compound 1, a potent hDHODH inhibitor (IC50 = 1.2 nM), able to induce myeloid differentiation in AML cell lines (THP1) in the low nM range (EC50 = 32.8 nM) superior to brequinar's phase I/II clinical trial (EC50 = 265 nM). Herein, we investigate the 1 drug-like properties observing good metabolic stability and no toxic profile when administered at doses of 10 and 25 mg/kg every 3 days for 5 weeks (Balb/c mice). Moreover, in order to identify a backup compound, we investigate the SAR of this class of compounds. Inside the series, 17 is characterized by higher potency in inducing myeloid differentiation (EC50 = 17.3 nM), strong proapoptotic properties (EC50 = 20.2 nM), and low cytotoxicity toward non-AML cells (EC30(Jurkat) > 100 μM).

Neomorphic mutations in isocitrate dehydrogenase 1 (IDH1) are oncogenic for a number of malignancies, primarily low-grade gliomas and acute myeloid leukemia (AML). We report a medicinal chemistry campaign around a 7,7-dimethyl-7,8-dihydro-2H-1λ(2)-quinoline-2,5(6H)-dione screening hit against the R132H and R132C mutant forms of isocitrate dehydrogenase (IDH1). Systematic SAR efforts produced a series of potent pyrid-2-one mIDH1 inhibitors, including the atropisomer (+)-119 (NCATS-SM5637, NSC 791985). In an engineered mIDH1-U87-xenograft mouse model; after a single oral dose of 30 mg/kg, 16 h post dose, between 16-48 h, (+)-119 showed higher tumoral concentrations that corresponded to lower 2-HG concentrations, when compared with the approved drug AG-120 (ivosidenib).

51 p.-6 fig.-1 tab.-4 schem.-1 graph. abst., Bacterial resistance to antibiotics makes previously manageable infections again disabling and lethal, highlighting the need for new antibacterial strategies. In this regard, inhibition of the bacterial division process by targeting key protein FtsZ has been recognized as an attractive approach for discovering new antibiotics. Binding of small molecules to the cleft between the N-terminal guanosine triphosphate (GTP)-binding and the C-terminal subdomains allosterically impairs the FtsZ function, eventually inhibiting bacterial division. Nonetheless, the lack of appropriate chemical tools to develop a binding screen against this site has hampered the discovery of FtsZ antibacterial inhibitors. Herein, we describe the first competitive binding assay to identify FtsZ allosteric ligands interacting with the interdomain cleft, based on the use of specific high-affinity fluorescent probes. This novel assay, together with phenotypic profiling and X-ray crystallographic insights, enables the identification and characterization of FtsZ inhibitors of bacterial division aiming at the discovery of more effective antibacterials., Thiswork was supported by grants BFU 2014-51823-R (J.M.A.),SAF2016-78792-R, PID2019-106279RB-I00 (M.L.L.-R.), BFU2017-87387-P (C.F.-T.), and predoctoral FPU fellowshipsfrom MECD (A.E.-P. and M.A.).

There is an urgent need for new treatments for visceral leishmaniasis (VL), a parasitic infection which impacts heavily large areas of East Africa, Asia, and South America. We previously reported on the discovery of GSK3494245/DDD01305143 (1) as a preclinical candidate for VL and, herein, we report on the medicinal chemistry program that led to its identification. A hit from a phenotypic screen was optimized to give a compound with in vivo efficacy, which was hampered by poor solubility and genotoxicity. The work on the original scaffold failed to lead to developable compounds, so an extensive scaffold-hopping exercise involving medicinal chemistry design, in silico profiling, and subsequent synthesis was utilized, leading to the preclinical candidate. The compound was shown to act via proteasome inhibition, and we report on the modeling of different scaffolds into a cryo-EM structure and the impact this has on our understanding of the series' structure-activity relationships.

Accumulation of amyloid β peptides (Aβ) is thought to be one of the causal factors of Alzheimer's disease (AD). The aspartyl protease β-site amyloid precursor protein cleaving enzyme 1 (BACE1) is the rate-limiting protease for Aβ production, and therefore, BACE1 inhibition is a promising therapeutic approach for the treatment of AD. Starting with a dihydro-1,3-thiazine-based lead, Compound J, we discovered atabecestat 1 (JNJ-54861911) as a centrally efficacious BACE1 inhibitor that was advanced into the EARLY Phase 2b/3 clinical trial for the treatment of preclinical AD patients. Compound 1 demonstrated robust and dose-dependent Aβ reduction and showed sufficient safety margins in preclinical models. The potential of reactive metabolite formation was evaluated in a covalent binding study to assess its irreversible binding to human hepatocytes. Unfortunately, the EARLY trial was discontinued due to significant elevation of liver enzymes, and subsequent analysis of the clinical outcomes showed dose-related cognitive worsening.

Persistent androgen receptor (AR) activation drives therapeutic resistance to second-generation AR pathway inhibitors and contributes to the progression of advanced prostate cancer. One resistance mechanism is point mutations in the ligand binding domain of AR that can transform antagonists into agonists. The AR F877L mutation, identified in patients treated with enzalutamide or apalutamide, confers resistance to both enzalutamide and apalutamide. Compound 4 (JNJ-pan-AR) was identified as a pan-AR antagonist with potent activity against wild-type and clinically relevant AR mutations including F877L. Metabolite identification studies revealed a latent bioactivation pathway associated with 4. Subsequent lead optimization of 4 led to amelioration of this pathway and nomination of 5 (JNJ-63576253) as a clinical stage, next-generation AR antagonist for the treatment of castration-resistant prostate cancer (CRPC).

Cystic fibrosis (CF) is a life-threatening recessive genetic disease caused by mutations in the gene encoding for the cystic fibrosis transmembrane conductance regulator (CFTR). With the discovery of Ivacaftor and Lumacaftor, it has been shown that administration of one or more small molecules can partially restore the CFTR function. Correctors are small molecules that enhance the amount of CFTR on the cell surface, while potentiators improve the gating function of the CFTR channel. Herein, we describe the discovery and optimization of a novel potentiator series. Scaffold hopping, focusing on retaining the different intramolecular contacts, was crucial in the whole discovery process to identify a novel series devoid of genotoxic liabilities. From this series, the clinical candidate GLPG2451 was selected based on its pharmacokinetic properties, allowing QD dosing and based on its low CYP induction potential.

A CDK9 inhibitor having short target engagement would enable a reduction of Mcl-1 activity, resulting in apoptosis in cancer cells dependent on Mcl-1 for survival. We report the optimization of a series of amidopyridines (from compound 2), focusing on properties suitable for achieving short target engagement after intravenous administration. By increasing potency and human metabolic clearance, we identified compound 24, a potent and selective CDK9 inhibitor with suitable predicted human pharmacokinetic properties to deliver transient inhibition of CDK9. Furthermore, the solubility of 24 was considered adequate to allow i.v. formulation at the anticipated effective dose. Short-term treatment with compound 24 led to a rapid dose- and time-dependent decrease of pSer2-RNAP2 and Mcl-1, resulting in cell apoptosis in multiple hematological cancer cell lines. Intermittent dosing of compound 24 demonstrated efficacy in xenograft models derived from multiple hematological tumors. Compound 24 is currently in clinical trials for the treatment of hematological malignancies.

Cholesterol 24-hydroxylase (CH24H or CYP46A1) is a brain-specific cytochrome P450 enzyme that metabolizes cholesterol into 24

A series of novel heteroaromatic biphenyl-methyl-pyrimidine analogues were designed via hybridization of privileged structures of two HIV-1 inhibitors. Among them, compound 7a containing 4-pyridinyl-phenyl and methyl-pyrimidine fragments revealed excellent wild-type HIV-1 inhibitory activity with low cytotoxicity. 7a had favorable solubility and liver microsome stability; moreover, no apparent CYP enzymatic inhibitory activity or acute toxicity was observed. However, its inhibitory activity toward mutant strains and the pharmacokinetic (PK) profiles were still unsatisfactory. Further optimizations resulted in a highly potent compound 9d without methyl on the pyrimidine but a heteroaromatic dimethyl-biphenyl on the left rings of difluoro-pyridinyl-diarylpyrimidines (DAPYs). A broad-spectrum activity (EC50 = 2.0-57 nM) of 9d against resistant strains was revealed. This compound also exhibited good solubility and safety profiles and a good PK profile with an oral bioavailability of 59% in rats. Collectively, these novel heteroaromatic dimethyl-biphenyl-DAPYs represent promising drug candidates for HIV clinical therapy. ispartof: JOURNAL OF MEDICINAL CHEMISTRY vol:65 issue:3 pages:2122-2138 ispartof: location:United States status: published

FGF19 signaling through the FGFR4/β-klotho receptor complex has been shown to be a key driver of growth and survival in a subset of hepatocellular carcinomas, making selective FGFR4 inhibition an attractive treatment opportunity. A kinome-wide sequence alignment highlighted a poorly conserved cysteine residue within the FGFR4 ATP-binding site at position 552, two positions beyond the gate-keeper residue. Several strategies for targeting this cysteine to identify FGFR4 selective inhibitor starting points are summarized which made use of both rational and unbiased screening approaches. The optimization of a 2-formylquinoline amide hit series is described in which the aldehyde makes a hemithioacetal reversible-covalent interaction with cysteine 552. Key challenges addressed during the optimization are improving the FGFR4 potency, metabolic stability, and solubility leading ultimately to the highly selective first-in-class clinical candidate roblitinib.

Despite the broad implications of the cannabinoid type 2 receptor (CB2) in neuroinflammatory processes, a suitable CB2-targeted probe is currently lacking in clinical routine. In this work, we synthesized 15 fluorinated pyridine derivatives and tested their binding affinities toward CB2 and CB1. With a sub-nanomolar affinity (Ki for CB2) of 0.8 nM and a remarkable selectivity factor of >12,000 over CB1, RoSMA-18-d6 exhibited outstanding in vitro performance characteristics and was radiofluorinated with an average radiochemical yield of 10.6 ± 3.8% (n = 16) and molar activities ranging from 52 to 65 GBq/μmol (radiochemical purity > 99%). [18F]RoSMA-18-d6 showed exceptional CB2 attributes as demonstrated by in vitro autoradiography, ex vivo biodistribution, and positron emission tomography (PET). Further, [18F]RoSMA-18-d6 was used to detect CB2 upregulation on postmortem human ALS spinal cord tissues. Overall, these results suggest that [18F]RoSMA-18-d6 is a promising CB2 PET radioligand for clinical translation.

Acquired resistance to fulvestrant and palbociclib is a new challenge to treatment of estrogen receptor positive (ER+) breast cancer. ER is expressed in most resistance settings; thus, bromodomain and extra-terminal protein inhibitors (BETi) that target BET-amplified ER-mediated transcription have therapeutic potential. Novel pyrrolopyridone BETi leveraged novel interactions with L92/L94 confirmed by a cocrystal structure of 27 with BRD4. Optimization of BETi using growth inhibition in fulvestrant-resistant (MCF-7:CFR) cells was confirmed in endocrine-resistant, palbociclib-resistant, and ESR1 mutant cell lines. 27 was more potent in MCF-7:CFR cells than six BET inhibitors in clinical trials. Transcriptomic analysis differentiated 27 from the benchmark BETi, JQ-1, showing downregulation of oncogenes and upregulation of tumor suppressors and apoptosis. The therapeutic approach was validated by oral administration of 27 in orthotopic xenografts of endocrine-resistant breast cancer in monotherapy and in combination with fulvestrant. Importantly, at an equivalent dose in rats, thrombocytopenia was mitigated.

Diffuse intrinsic pontine glioma is an aggressive pediatric cancer for which no effective chemotherapeutic drugs exist. Analysis of the genomic landscape of this disease has led to the identification of the serine/threonine kinase ALK2 as a potential target for therapeutic intervention. In this work, we adopted an open science approach to develop a series of potent type I inhibitors of ALK2 which are orally bio-available and brain-penetrant. Initial efforts resulted in the discovery of M4K2009, an analogue of the previously reported ALK2 inhibitor LDN-214117. Although highly selective for ALK2 over the TGF-βR1 receptor ALK5, M4K2009 is also moderately active against the hERG potassium channel. Varying the substituents of the trimethoxyphenyl moiety gave rise to an equipotent benzamide analogue M4K2149 with reduced off-target affinity for the ion channel. Additional modifications yielded 2-fluoro-6-methoxybenzamide derivatives (26a-c), which possess high inhibitory activity against ALK2, excellent selectivity, and superior pharmacokinetic profiles.

Speckle-type POZ protein (SPOP) is overexpressed in the nucleus and misallocated in the cytoplasm in almost all the clear-cell renal cell carcinomas (ccRCCs), which leads to kidney tumorigenesis. Previously, we elucidated that the oncogenic SPOP-signaling pathway in ccRCC could be suppressed by 6b that inhibits SPOP-mediated protein interactions. Herein, we have established a structure-activity relationship for 6b analogues as SPOP inhibitors. Compound 6lc suppresses the viability and inhibits the colony formation of ccRCC cell lines driven by cytoplasmic SPOP, superior to 6b. Compound 6lc binds to the SPOP protein in vitro and disrupts SPOP binding to phosphatase-and-tensin homologue (PTEN) in HEK293T cells, which causes the observable phenomena: a decline in the ubiquitination of PTEN, elevated levels of both PTEN and dual-specificity phosphatase 7, and decreased levels of phosphorylated AKT and ERK when ccRCC cell lines are exposed to 6lc in a dose-response manner. Taken together, compound 6lc is a potent candidate against kidney tumorigenesis.

Development of new photosensitizers (PSs) with high photodynamic efficacy and minimal side effects is of great interest in photodynamic therapy (PDT). In this work, we reported several pyridine-embedded phenothiazinium (pyridophenothiazinium) dyes, which could be conveniently synthesized in a few short steps and acted as highly efficient and potent PSs to selectively localize to lysosomes and photosensitively kill cancer cells. Among them, compound 5, which possessed the ability of promoting intracellular reactive oxygen species (ROS) upon light irradiation by almost 40-fold higher than that of methylene blue (MB, a general phenothiazinium-based PS), exhibited a remarkable phototherapeutic index (PI = 53.8) against HT29 cancer cells, leading to eradication of large solid tumors (∼300 mm3) in a xenograft mouse model without apparent side effects. These results suggest that the pyridophenothiazinium dyes developed herein, especially compound 5, may serve as promising lysosome-targeted PSs for efficient photodynamic antitumor therapy.