Marvin J. Meyers

Discovery of (3S,3aR)-2-(3-Chloro-4-cyanophenyl)-3-cyclopentyl-3,3a,4,5-tetrahydro-2H-benzo[g]indazole-7-carboxylic Acid (PF-3882845), an Orally Efficacious Mineralocorticoid Receptor (MR) Antagonist for Hypertension and Nephropathy

We have discovered a novel class of nonsteroidal pyrazoline antagonists of the mineralocorticoid receptor (MR) that show excellent potency and selectivity against other nuclear receptors. Early analogues were poorly soluble and had a propensity to inhibit the hERG channel. Remarkably, both of these challenges were overcome by incorporation of a single carboxylate moiety. Structural modification of carboxylate-containing lead R-4g with a wide range of substituents at each position of the pyrazoline ring resulted in R-12o, which shows excellent activity against MR and reasonable pharmacokinetic profile. Introduction of conformational restriction led to a novel series characterized by exquisite potency and favorable steroid receptor selectivity and pharmacokinetic profile. Oral dosing of 3S,3aR-27d (PF-3882845) in the Dahl salt sensitive preclinical model of salt-induced hypertension and nephropathy showed blood pressure attenuation significantly greater than that with eplerenone, reduction in urinary albumin, and renal protection. As a result of these findings, 3S,3aR-27d was advanced to clinical studies.

The Hepatitis B Virus Ribonuclease H Is Sensitive to Inhibitors of the Human Immunodeficiency Virus Ribonuclease H and Integrase Enzymes

Nucleos(t)ide analog therapy blocks DNA synthesis by the hepatitis B virus (HBV) reverse transcriptase and can control the infection, but treatment is life-long and has high costs and unpredictable long-term side effects. The profound suppression of HBV by the nucleos(t)ide analogs and their ability to cure some patients indicates that they can push HBV to the brink of extinction. Consequently, more patients could be cured by suppressing HBV replication further using a new drug in combination with the nucleos(t)ide analogs. The HBV ribonuclease H (RNAseH) is a logical drug target because it is the second of only two viral enzymes that are essential for viral replication, but it has not been exploited, primarily because it is very difficult to produce active enzyme. To address this difficulty, we expressed HBV genotype D and H RNAseHs in E. coli and enriched the enzymes by nickel-affinity chromatography. HBV RNAseH activity in the enriched lysates was characterized in preparation for drug screening. Twenty-one candidate HBV RNAseH inhibitors were identified using chemical structure-activity analyses based on inhibitors of the HIV RNAseH and integrase. Twelve anti-RNAseH and anti-integrase compounds inhibited the HBV RNAseH at 10 µM, the best compounds had low micromolar IC50 values against the RNAseH, and one compound inhibited HBV replication in tissue culture at 10 µM. Recombinant HBV genotype D RNAseH was more sensitive to inhibition than genotype H. This study demonstrates that recombinant HBV RNAseH suitable for low-throughput antiviral drug screening has been produced. The high percentage of compounds developed against the HIV RNAseH and integrase that were active against the HBV RNAseH indicates that the extensive drug design efforts against these HIV enzymes can guide anti-HBV RNAseH drug discovery. Finally, differential inhibition of HBV genotype D and H RNAseHs indicates that viral genetic variability will be a factor during drug development.

Recent advances in plasmepsin medicinal chemistry and implications for future antimalarial drug discovery efforts.

Plasmepsins are the aspartic proteases of Plasmodium that play key roles in the survival of the parasite in its host. The plasmepsins of the digestive vacuole play an important role in hemoglobin degradation, providing the parasite with a vital source of nutrients. Recently, plasmepsin V has been shown to be an essential protease, processing hundreds of parasite proteins for export into the host erythrocyte. The functions of the remaining plasmepsins have yet to be discovered. Over the past decade, much effort has been placed towards developing plasmepsin inhibitors as antimalarial agents, particularly targeting the digestive vacuole. This review will highlight some of the recent work in this field with a particular focus on target druggability and strategies for identifying plasmepsins inhibitors as effective antimalarial drugs. Given recent advances in understanding the fundamental roles of the various plasmepsins, it is likely that the most effective antimalarial plasmepsin targets will be the non-digestive vacuole plasmepsins.

Hydroxylated Tropolones Inhibit Hepatitis B Virus Replication by Blocking Viral Ribonuclease H Activity

ABSTRACT Hepatitis B virus (HBV) remains a major human pathogen despite the development of both antiviral drugs and a vaccine, in part because the current therapies do not suppress HBV replication far enough to eradicate the virus. Here, we screened 51 troponoid compounds for their ability to suppress HBV RNaseH activity and HBV replication based on the activities of α-hydroxytropolones against HIV RNaseH, with the goal of determining whether the tropolone pharmacophore may be a promising scaffold for anti-HBV drug development. Thirteen compounds inhibited HBV RNaseH, with the best 50% inhibitory concentration (IC50) being 2.3 μM. Similar inhibition patterns were observed against HBV genotype D and C RNaseHs, implying limited genotype specificity. Six of 10 compounds tested against HBV replication in culture suppressed replication via blocking of viral RNaseH activity, with the best 50% effective concentration (EC50) being 0.34 μM. Eighteen compounds inhibited recombinant human RNaseH1, and moderate cytotoxicity was observed for all compounds (50% cytotoxic concentration [CC50] = 25 to 79 μM). Therapeutic indexes ranged from 3.8 to 94. Efficient inhibition required an intact α-hydroxytropolone moiety plus one or more short appendages on the tropolone ring, but a wide variety of constituents were permissible. These data indicate that troponoids and specifically α-hydroxytropolones are promising lead candidates for development as anti-HBV drugs, providing that toxicity can be minimized. Potential anti-RNaseH drugs are envisioned to be employed in combination with the existing nucleos(t)ide analogs to suppress HBV replication far enough to block genomic maintenance, with the goal of eradicating infection.

Hepatitis B virus replication is blocked by a 2-hydroxyisoquinoline-1,3(2H,4H)-dione (HID) inhibitor of the viral ribonuclease H activity.

Nucleos(t)ide analog drugs profoundly suppress Hepatitis B virus (HBV) replication but rarely cure the infection, so therapy is usually life-long. The nucleos(t)ide analogs inhibit the viral DNA polymerase and often push HBV to the brink of extinction, so it may be possible to eradicate HBV by suppressing HBV replication further. The HBV ribonuclease H (RNaseH) is a logical new drug target because it is the second of only two viral enzymes essential for viral replication. We recently developed a low throughput screening pipeline for inhibitors of the HBV RNaseH and viral replication. Here, we screened a series of twenty-three nitrogen-based polyoxygenated heterocycles including sixteen 2-hydroxyisoquinoline-1,3(2H,4H)-dione derivatives for anti-HBV RNaseH activity. Nine compounds inhibited the HBV RNaseH, but activity was marginal for eight of them. Compound #1 [2-hydroxyisoquinoline-1,3(2H,4H)-dione, HID] was the best hit with an IC50 of 28.1μM and an EC50 of 4.2μM. It preferentially suppressed accumulation of the viral plus-polarity DNA strand in replication inhibition assays, indicating that replication was blocked due to suppression of HBV RNaseH activity. It had a CC50 of 75μM, yielding a therapeutic index of ∼18. The EC50 value was 7-fold lower than the IC50, possibly due to cellular retention or metabolism of the compound, or higher affinity for the full-length enzyme than the recombinant form used for screening. These data indicate that the 2-hydroxyisoquinoline-1,3(2H,4H)-diones will have different structure-activity relationships for the HBV and HIV RNaseHs. Therefore, HID compounds may provide a foundation for development of more effective RNaseH inhibitors of HBV replication.

Quinine bis-conjugates with quinolone antibiotics and peptides: synthesis and antimalarial bioassay

Benzotriazole-mediated syntheses led to novel bis-conjugates of quinine with quinolone antibiotics and amino acid linkers which were successfully prepared by two alternative routes with excellent yields and retention of chirality. These bis conjugates retain in vitro antimalarial activity with IC(50) values ranging from 12 to 207 nM, similar to quinine itself.

Evaluation of Aminohydantoins as a Novel Class of Antimalarial Agents

Given the threat of drug resistance, there is an acute need for new classes of antimalarial agents that act via a unique mechanism of action relative to currently used drugs. We have identified a set of druglike compounds within the Tres Cantos Anti-Malarial Set (TCAMS) which likely act via inhibition of a Plasmodium aspartic protease. Structure-activity relationship analysis and optimization of these aminohydantoins demonstrate that these compounds are potent nanomolar inhibitors of the Plasmodium aspartic proteases PM-II and PM-IV and likely one or more other Plasmodium aspartic proteases. Incorporation of a bulky group, such as a cyclohexyl group, on the aminohydantion N-3 position gives enhanced antimalarial potency while reducing inhibition of human aspartic proteases such as BACE. We have identified compound 8p (CWHM-117) as a promising lead for optimization as an antimalarial drug with a low molecular weight, modest lipophilicity, oral bioavailability, and in vivo antimalarial activity in mice.

Identification of (R)-6-(1-(4-cyano-3-methylphenyl)-5-cyclopentyl-4,5-dihydro-1H-pyrazol-3-yl)-2-methoxynicotinic acid, a highly potent and selective nonsteroidal mineralocorticoid receptor antagonist.

A novel series of nonsteroidal mineralocorticoid receptor (MR) antagonists identified as part of our strategy to follow up on the clinical candidate PF-03882845 (2) is reported. Optimization departed from the previously described pyrazoline 3a and focused on improving the selectivity for MR versus the progesterone receptor (PR) as an approach to avoid potential sex-hormone-related adverse effects and improving biopharmaceutical properties. From this effort, (R)-14c was identified as a potent nonsteroidal MR antagonist (IC50 = 4.5 nM) with higher than 500-fold selectivity versus PR and other related nuclear hormone receptors, with improved solubility as compared to 2 and pharmacokinetic properties suitable for oral administration. (R)-14c was evaluated in vivo using the increase of urinary Na(+)/K(+) ratio in rat as a mechanism biomarker of MR antagonism. Treatment with (R)-14c by oral administration resulted in significant increases in urinary Na(+)/K(+) ratio and demonstrated this novel compound acts as an MR antagonist.

Synthesis and antimalarial bioassay of quinine - peptide conjugates.

Amino acid and peptide conjugates of quinine were synthesized using microwave irradiation in 52–95% yields using benzotriazole methodology. The majority of these conjugates retain in vitro antimalarial activity with IC50 values below 100 nm, similar to quinine.

Synthesis, antimalarial properties and 2D-QSAR studies of novel triazole-quinine conjugates

Click chemistry technique led to novel 1,2,3-triazole-quinine conjugates 8a-g, 10a-o, 11a-h and 13 utilizing benzotriazole-mediated synthetic approach with excellent yields. Some of the synthesized analogs (11a, 11d-h) exhibited antimalarial properties against Plasmodium falciparum strain 3D7 with potency higher than that of quinine (standard reference used) through in vitro standard procedure bio-assay. Statistically significant BMLR-QSAR model describes the bio-properties, validates the observed biological observations and identifies the most important parameters governing bio-activity.