Infrared spectroscopy of serum as a potential diagnostic screening approach for naturally occurring canine osteoarthritis associated with cranial cruciate ligament rupture.

OBJECTIVE: To evaluate infrared (IR) spectroscopy of serum as a screening tool to differentiate dogs affected by naturally occurring osteoarthritis (OA) associated with cranial cruciate ligament rupture (CrCLR) and controls. METHOD: 104 adult dogs with CrCLR (affected group) and 50 adult control dogs were recruited for this prospective observational study. Serum samples were collected preoperatively from CrCLR dogs and from a subset of these dogs at 4-, and 12-week post-surgical intervention to stabilize the affected stifles. Serum was collected once from control dogs. Dry films were made from serum samples, and IR absorbance spectra acquired. Data preprocessing, principal component analysis and multivariate analysis of covariance were performed to separate samples from the two groups, and to evaluate temporal differences. Weighted logistic regression with L1 regularization method was used to develop a predictive model. Model performance based on an independent test set was evaluated. RESULTS: Spectral data analysis revealed significant separation between the sera of CrCLR and control dogs (P < 0.0001), but not amongst different time points in the OA group. The sensitivity, specificity, AUC and accuracy of the test set were 84.62%, 96.15%, 93.20% and 92.31% respectively. CONCLUSIONS: These findings confirm the potential of IR-spectroscopy of serum with chemometrics methods to differentiate controls from dogs with OA associated with CrCLR. This is the first step in development of an economic, and comparatively simple IR-based screening serum test for OA. Utility of this tool as a clinical screening and diagnostic test requires further investigation and validation.

Tetrahydrobenzothiophene inhibitors of hepatitis C virus NS5B polymerase.

A novel series of selective HCV NS5B RNA dependent RNA polymerase inhibitors has been disclosed. These compounds contain an appropriately substituted tetrahydrobenzothiophene scaffold. This communication will detail the SAR and activities of this series.

Nasal dermoid sinus in an American cocker spaniel.

An American cocker spaniel was presented for a subcutaneous mass and draining tract located between its eyes. Contrast radiography and surgical excision showed communication of the tract with the left frontal sinus and rostral cerebral dura, respectively. A dermoid sinus was diagnosed by a combination of gross and histologic findings.

Antiviral properties of 3-quinolinecarboxamides: a series of novel non-nucleoside antiherpetic agents.

Novel antiherpetic 3-quinolinecarboxamides were discovered as part of a drug discovery program at Sterling Winthrop Inc. A major goal of this research was to identify novel non-nucleoside agents possessing activity against acyclovir resistant herpes simplex virus. From screening compound libraries in an HSV-2 plaque reduction assay, 1-ethyl-1,4-dihydro-4-oxo-7-(4-pyridinyl)-3-quinolinecarboxamide (1) emerged as an attractive lead structure. By modifying the quinoline ring at the 1-, 2-, 3-, 4-, and 7-positions, analogues were identified that have up to 5-fold increased in vitro potency relative to acyclovir. In a single dose mouse model of infection the 1-(4-FC6H4) analogue 17, one of the most potent derivatives in vitro, displayed comparable oral antiherpetic efficacy to acyclovir at 1/16 the dose; in a multiple dose regimen, however, it was 2-fold less potent. Mechanism of action studies indicate that these new compounds interact with a different, as yet undefined, molecular target than acyclovir.

Keto/enol epoxy steroids as HIV-1 Tat inhibitors: structure-activity relationships and pharmacophore localization.

Inhibition of the HIV-1 nuclear regulatory protein tat could potentially yield particularly useful drugs because it functions as an activator of transcription. It has no known cellular counterpart, and deletions in the tat gene destroy the ability of HIV-1 to replicate. We recently reported that a structurally unique class of tat inhibitors, 3-keto/enol 4,5-alpha-epoxy steroids bearing electron-withdrawing substituents at position 2, specifically inhibit tat-induced gene expression in virus free transfected SW480 cells. In this paper, we report on additional SAR (structure-activity relationships) for the steroid series and the localization of the pharmacophore to the A-ring functionality. There is a weak enantioselective preference for the natural steroid stereochemistry and hints of additional SAR in the electron-withdrawing group. Compound 34a is of particular interest in that it inhibits HIV replication in H9 cells at a concentration equivalent to its inhibitory level in the primary tat assay.

Structures of four methyltetrazole-containing antiviral compounds in human rhinovirus serotype 14.

Four novel antiviral WIN compounds, that contain a methyl tetrazole ring as well as isoxazole, pyridazine or acetylfuran rings, have had their structures determined in human rhinovirus serotype 14 at 2.9 A resolution. These compounds bind in the VP1 hydrophobic pocket, but are shifted significantly towards the pocket pore when compared to previously examined WIN compounds. A putative water network at the pocket pore is positioned to hydrogen bond with these four WIN compounds, and this network can account for potency differences seen in structurally similar WIN compounds.

An evaluation of the antirhinoviral activity of acetylfuran replacements for 3-methylisoxazoles. Are 2-acetylfurans bioisosteres for 3-methylisoxazoles?

As a probe of the 3-methylisoxazole portion of our broad-spectrum antipicornaviral series, a panel of 2-acetylfuran analogues was prepared as replacements for the 3-methylisoxazole ring. Comparison of the two series showed remarkable similarity in potency, spectrum of activity, logP, and electrostatic parameters. X-ray studies of 21b bound to human rhinovirus-14 showed that the 2-acetyl group adopted a syn conformation and the carbonyl oxygen acts as a hydrogen bond acceptor with ASN219 in much the same way as the nitrogen of the isoxazole. The importance of the syn conformation and the hydrogen-bonding capability was confirmed by the reduced antiviral activity of the 2-methylfuran and 2-formylfuran analogues. From the results of this study, it is apparent that the syn-2-acetylfuran ring is acting as a bioisostere for the 3-methylisoxazole.

Oxadiazoles as ester bioisosteric replacements in compounds related to disoxaril. Antirhinovirus activity.

A series of 1,2,4-oxadiazoles has been prepared as ester bioisosteres and tested against 15 human rhinovirus serotypes, and the MIC80, the concentration which inhibits 80% or 12 of the serotypes tested, was determined. Homologation of the alkyl group attached to the oxadiazole ring resulted in a reduction in activity with increased chain length. Introduction of hydrophilic groups in this position rendered the compounds inactive. Increasing the length of the side chain attached to the isoxazole ring resulted in an increase in activity. Replacement of the methyl with alkoxyalkyl substituents retained activity; however, introduction of a hydroxyl group on to the side chain reduced activity. Compound 8a, where both the isoxazole and oxadiazole rings were substituted with methyl groups, was one of the most active compounds in the series. A comparison was made between 8a and the two isomeric oxadiazoles 41 and 46, and an attempt was made to explain the difference in activity by examining electrostatic potential maps and by an energy profiling study. No conclusive results were obtained from these studies.

Antipicornavirus activity of tetrazole analogues related to disoxaril.

A series of tetrazole analogues of Win 54954, a broad-spectrum antipicornavirus compound, has been synthesized to address the acid lability of the oxazoline ring of this series of compounds. The results of X-ray crystallography studies of several members of the oxazoline series bound to human rhinovirus type 1A and 14 have been used to design compounds in the tetrazole series with a broad spectrum of activity. Compound 16b, which has a three-carbon linkage between the isoxazole and phenyl rings and a propyl chain extending from the isoxazole ring, exhibiting an MIC80 for 15 rhinovirus serotypes of 0.20 microM as compared to 0.40 microM for Win 54954. X-ray studies of 16b bound to human rhinovirus-14 show that the propyl side chain extends into a pore in the binding site with the possibility of hydrophobic interactions with a pocket formed by Leu106 and a portion of Ser107.

3-Quinolinecarboxamides. A series of novel orally-active antiherpetic agents.

A series of novel 3-quinolinecarboxamides that are structurally similar to the quinolone class of antibacterial agents possess excellent antiherpetic properties. By modifying the quinoline ring at the 1-, 2-, 3-, and 7-positions, analogues were identified that have up to 5-fold increased HSV-2 plaque-reduction potency relative to acyclovir. In a single-dose mouse model of infection, one of the most potent derivatives in vitro, 1-(4-fluorophenyl)-1,4-dihydro-4-oxo-7-(4-pyridinyl)-3-quinolinecarbo xamide (97), displayed comparable oral antiherpetic efficacy to acyclovir at 1/16 the dose; in a multiple-dose regimen, however, 97 was 2-fold less potent. In mice dosed orally with 97, sustained plasma drug levels were evident that may account for the high efficacy observed. The molecular mechanism of action of these agents is not known; however, based on in vitro studies with acyclovir resistant mutants, it is likely that the mechanism differs from that of acyclovir. In vitro plaque-reduction potency was not generally predictive of oral efficacy in mice. An X-ray crystal structure of 97 corroborated the assignment of structure and provided useful insights as to the effect of conformation on plaque-reduction potency.

Antirhinoviral activity of heterocyclic analogs of Win 54954.

A variety of heterocyclic analogs of Win 54954 have been synthesized and tested in vitro against human rhinovirus type 14 (HRV-14) in a plaque reduction assay. The more active compounds were tested against 14 additional serotypes, and the concentration which inhibited 80% of the serotypes tested (MIC80) was measured. One compound, 37, exhibited activity comparable to Win 59454. Physicochemical as well as electrostatic parameters were calculated and the results subjected to a QSAR analysis in an effort to explain differences in activity observed between these compounds; however, no meaningful correlation with biological activity was found with any of these parameters.

A model for compounds active against human rhinovirus-14 based on X-ray crystallography data.

A number of (oxazolinylphenyl)isoxazoles have been synthesized and tested against human rhinovirus-14 (HRV-14). Several of the more active compounds have been examined by X-ray crystallography and their orientation in the compound binding site on the capsid protein of HRV-14 has been determined. Based on the minimum inhibitory concentration against HRV-14 and the X-ray conformation of the compounds, a model has been developed which distinguishes between the space-filling properties of the active and inactive compounds in this series. The model was generated by overlaying composite structures and comparing the van der Waals generated volume maps. The results of this study indicate that inactive compounds display areas of excessive bulk particularly around the phenyl ring, while the active compounds occupy space below the pore area of the compound binding site.