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STAT5 is a member of the signal transducer and activator of transcription family of proteins and is widely recognized as an oncogenic master regulator of hematological malignancies. To date, therapeutic approaches to attenuate aberrant activity have focused on upstream kinase targets such as JAK2, Bcr-Abl, Flt3, and Flt3-ITD. To date, there is no STAT5-selective inhibitor in clinical development. More recently, PROTAC approaches have been deployed successfully against the STAT3 isoform and demonstrated efficacy and safety in preclinical models. We herein report the identification of a potent and selective, non-PROTAC, small molecule degrader of STAT5 protein, JPX-0750. Upon binding, JPX-0750 covalently and selectively binds to a cysteine residue on STAT5 via an electrophilic warhead, which induces profound destabilization of STAT5. The unfolding/destabilization effect can be observed via Western blot, isothermal denaturation of purified protein and HDX exchange. In a range of AML and TPLL cell lines exposure to JPX-750 at nM concentrations leads to a rapid and dose-dependant degradation of both phospho-STAT5 and total STAT5 with resultant IC50's between 80-200 nM. Washout experiments determined that exposure to JPX-0750 at 1 µM for 2-4 h is sufficient to degrade STAT5, inhibit downstream targets, and induce apoptosis. Moreover, in the same washout experiments, 50% STAT5 recovery post-washout required >72 h suggesting a long pharmacodynamic effect. Against STAT3, degrading effects are observed later, 24 h post washout (6 h treatment), indicating that anti-STAT5 activity is more rapid in AML cells and in line with the inherent stability of each protein. No effect was observed on either STAT1 or pSTAT1 upon exposure to JPX-750. Importantly, compared to standard AML cell lines JPX-750 demonstrated similar low nM potency in 25/30 primary AML blasts and TPLL patient samples, including those with poor prognostic markers. JPX-750 exhibits a large therapeutic window for AML versus pooled human fibroblasts and hematopoietic stem cells (ca 100 fold). In a preclinical MV4;11 luciferase model, structurally related JPX-700, at 5 mg/kg (IP, qd) significantly reduced leukemic burden, suppressed tumour dissemination to both the lung and liver, and had no effect on body weight, organ histology or blood parameters. In summary, JPX-750 represents a new class of potent and selective small molecule degraders of STAT5 protein. Citation Format: Ji Sung Park, Gary Tin, Elvin D. de Araujo, Anna Orlova, Helena Sorger, Florian Grebien, Elizabeth Heyes, Mulu Geletu, Ruth Villalonga, Angel Sampedro, Abootaleb Sedighi, Marco Herling, Satu Mustjoki, Mohammad S. Eram, Siawash Ahmar, Richard Moriggl, Jeff A. O'Meara, John Proudfoot, Kay Noel, Dziyana Kraskouskaya, Roman Fleck, Patrick T. Gunning. A potent and selective small molecule degrader of STAT5 for the treatment of hematological malignancies [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr LB-108.

Discovery and optimization of potency and selectivity of a non-Zn-chelating MMP-13 inhibitor with the aid of protein co-crystal structural information is reported. This inhibitor was observed to have a binding mode distinct from previously published MMP-13 inhibitors. Potency and selectivity were improved by extending the hit structure out from the active site into the S1' pocket.

We report on the nuclear receptor binding affinities, cellular activities of transrepression and transactivation, and anti-inflammatory properties of a quinol-4-one and other A-ring mimetic containing nonsteroidal class of glucocorticoid agonists.

A systematic structure-permeability relationship study on a set of small drug-like molecules with log D values in the range -2.5 to 3 and carrying a diverse array of functionality reveals that the compounds with log D>0 and

Synthesis and structure-activity relationship (SAR) of a series of alkyl and cycloalkyl containing non-steroidal dissociated glucocorticoid receptor (GR) agonists is reported. This series of compounds was identified as part of an effort to replace the CF3 group in a scaffold represented by 1a. The study culminated in the identification of compound 14, a t-butyl containing derivative, which has shown potent activity for GR, selectivity against the progesterone receptor (PR) and the mineralocorticoid receptor (MR), in vitro anti-inflammatory activity in an IL-6 transrepression assay, and dissociation in a MMTV transactivation counter-screen. In a collagen-induced arthritis mouse model, 14 displayed prednisolone-like efficacy, and lower impact on body fat and free fatty acids than prednisolone at an equivalent anti-inflammatory dose.

Purpose. To investigate the mechanisms involved in transport of sulfasalazine in Caco-2 cells. Methods. Permeability coefficients of sulfasalazine and its analogs across Caco-2 cell monolayers were measured as a function of direction of transport, energy and concentration dependence, and in the presence of inhibitors of various cellular efflux pumps and transporters. Results. Permeability coefficients of sulfasalazine across Caco-2 cell monolayers were approximately 342-, 261-, and 176-fold higher from basolateral to apical direction (BL→AP) than from apical to basolateral direction (AP→BL) at 100, 200, and 500 μM, respectively. Carrier permeability coefficient, non-saturable membrane permeability coefficient, and Michaelis constant were estimated to be 1.4×10−5 cm/s, 1.9×10−8 cm/s, and 369 μM, respectively. The efflux of sulfasalazine was completely blocked at 4°C and in the presence of an uncoupler of oxidative phosphorylation. Using cellular efflux inhibitors, the permeability of sulfasalazine was shown to depend on multidrug resistance-associated protein and anion sensitive transport mechanisms. Structure-permeability studies showed that the affinity of sulfasalazine for the cellular efflux pumps and transporters in Caco-2 cells depended strongly on the carboxylic acid functional group. Conclusions. The permeability of sulfasalazine across Caco-2 cell monolayer is very low due to its strong interaction with multiple cellular efflux pumps and transporters. This may partially explain its low absorption in vivo.

Recent progress in the area of non-nucleoside inhibitors of human immunodeficiency virus Type 1 (HIV-1) reverse transcriptase (RT) is covered. Inhibitors of this enzyme, which plays an essential role in the virus life cycle, constitute a key component of current anti-HIV-1 therapies. Recently, nevirapine (Viramune™) and delavirdine (Rescriptor™), non-nucleoside inhibitors with a mechanism of action different to that used by nucleoside analogues such as azidothymidine (AZT), have been approved for use in the treatment of acquired immunodeficiency syndrome (AIDS). Efforts to improve these and other non-nucleoside inhibitors and to develop more effective structures are surveyed.

Molecular modeling analysis of the recently published X-ray crystal structure of nevirapine bound to wild type human immunodeficiency virus type 1 reverse transcriptase (WT-RT) indicated the presence of a lipophilic cavity proximal to the 4-position of the inhibitor. A series of 4-substituted derivatives of nevirapine were thus synthesized to assess structure-activity relationships (SARs) and to see if increased binding to this region might translate into greater activity against mutant RTs. The results show that compounds with an appropriately spaced aryl ring appended to the 4-position of the dipyridodiazepinone ring system show good activity against WT-RT. Furthermore certain derivatives appear to inhibit the Y181C mutant RT. Attempts to combine these results with the recent discovery that 2-substituents enhance activity against the Y181C mutant led to a few compounds with moderate activity against both enzymes. The SAR of these two positions, however, could not be combined in a simple fashion.

The synthesis of 11-ethyl-5-methyldipyrido[3,2-b:3′,4′-e][1,4]diazepin-6-one 6, the first representative of a C-ring dipyridodiazepinone isomer of the HIV-1 reverse transcriptase inhibitor nevirapine 1 is described. The key step involves the regiospecific lithiation of 3-(tbutoxycarbonylamino)pyridine 7 followed by trapping with N-(2-chloropyridyl)-N-methylcarbamoyl chloride 10.

The synthesis of 11-ethyl-5-methyldipyrido[3,2-b:3′,4′-e][1,4]diazepin-6-one 6, the first representative of a C-ring dipyridodiazepinone isomer of the HIV-1 reverse transcriptase inhibitor nevirapine 1 is described. The key step involves the regiospecific lithiation of 3-(tbutoxycarbonylamino)pyridine 7 followed by trapping with N-(2-chloropyridyl)-N-methylcarbamoyl chloride 10.

SAR studies to improve the selectivity and metabolic stability of a class of recently discovered MMP-13 inhibitors are reported. Improved selectivity was achieved by modifying interactions with the S1' pocket. Metabolic stability was improved through reduction of inhibitor lipophilicity. This translated into lower in vivo clearance for the preferred compound.

We have recently reported the discovery of a novel class of glucocorticoid receptor (GR) antagonists, exemplified by 3, containing a 1,2-dihydroquinoline molecular scaffold. Further SAR studies of these antagonists uncovered chemical modifications conveying agonist functional activity to this series. These agonists exhibit good GR binding affinity and are selective against other nuclear hormone receptors.

A new series of ligands for the glucocorticoid receptor (GR) is described. SAR development was guided by docking 3 into the GR active site and optimizing an unsubstituted phenyl ring for key interactions found in the steroid A-ring binding pocket. To identify compounds with an improved side effect profile over marketed steroids the functional activity of compounds was evaluated in cell based assays for transactivation (aromatase) and transrepression (IL-6). Through this effort, 36 has been identified as a partial agonist with a dissociated profile in these cell based assays.

The article contains sections titled: 1. Introduction - HIV and AIDS 1.1. HIV Cytopathogenesis and AIDS 1.2. Virus Life Cycle 1.3. Opportunities for Drug Intervention 2. Reverse Transcriptase Inhibitors 2.1. Introduction 2.2. Nucleoside Analogs 2.3. Nonnucleoside Analogs 2.4. Compounds in Development 3. Protease Inhibitors 3.1. Introduction 3.2. Hydroxyethylamine Isosteres 3.3. Hydroxyethylene Isosteres 3.4. Compounds in Development 4. Emerging Concepts in AIDS Therapy 5. Acknowledgement

The syntheses of 11-ethyl and 11-cyclopropyldipyrido[2,3- b :3′,2′- f ]azepines, analogs of the HIV-1 reverse transcriptase inhibitor nevirapine 1 , are described. These compounds exhibit potency equivalent to nevirapine in the inhibition of wild-type and some mutant RT enzymes.

A novel dipyridodiazepinone, 6,11-dihydro-11-cyclopropyl-4-methyldipyrido(2,3-b:2{prime},3{prime}-e)-(1,4)diazepin-6-one (BI-RG-587), is a selective noncompetitive inhibitor of HIV-1 reverse transcriptase (RT-1). An azido photoaffinity analogue of BI-RG-587 was synthesized and found to irreversibly inhibit the enzyme upon UV irradiation. BI-RG-587 and close structural analogues competitively protected RT-1 from inactivation by the photoaffinity label. A thiobenzimidazolone (TIBO) derivative, a nonnucleoside inhibitor of RT-1, also protected the enzyme from photoinactivation, which suggests a common binding site for these compounds. Substrates dGTP, template-primer, and tRNA afforded no protection from enzyme inactivation. A tritiated photoaffinity probe was found to stoichiometrically and selectively label p66 such that 1 mol of probe inactivates 1 mol of RT-1.

Petrosterol, an unusual cyclopropane-containing marine sponge sterol, is shown, unexpectedly, to be derived by SAM biomethylation of 24-methylenecholesterol via a ccunplex rearrangement process.

The isolation, structure determination and synthesis of 23(R),24(R)-methylene-cholesterol—a possible biosynthetic precursor of brassicasterol—is described.

The title compounds were synthesized from a readily available steroidal acetylene by a protocol employing selenosulfonation, introduction of the appropriate side chain at C-24 via an alkyl selenocuprate, and reductive desulfonylation.

The partial synthesis of the marine cyclopropyl-sterol dihydrocalysterol (1), which has allowed the confirmation of the originally postulated absolute stereochemistry around the cyclopropane ring, is described. The firm knowledge of this stereochemistry and that of the other marine sterols petrosterol (7), dehydroaplysterol (21), ficisterol (22), nicasterol (8), and hebesterol (23) has allowed us to construct a unified biosynthetic scheme for these sterols of diverse side-chain structure. Our central postulate, that the intermediate in sterol side biomethylation may be described in terms of a protonated cyclopropane species [i.e. protonated dihydrocalysterol (1)], enables us to interrelate the absolute stereochemistries of the above sterols and predict the biosynthetic origin of the side-chain carbon atoms.