A 90-day tenofovir reservoir intravaginal ring for mucosal HIV prophylaxis.

A vaginal gel containing the antiretroviral tenofovir (TFV) recently demonstrated 39% protection against HIV infection in women. We designed and evaluated a novel reservoir TFV intravaginal ring (IVR) to potentially improve product effectiveness by providing a more controlled and sustained vaginal dose to maintain cervicovaginal concentrations. Polyurethane tubing of various hydrophilicities was filled with a high-density TFV/glycerol/water semisolid paste and then end-sealed to create IVRs. In vitro, TFV release increased with polyurethane hydrophilicity, with 35 weight percent water-swelling polyurethane IVRs achieving an approximately 10-mg/day release for 90 days with mechanical stiffness similar to that of the commercially available NuvaRing. This design was evaluated in two 90-day in vivo sheep studies for TFV pharmacokinetics and safety. Overall, TFV vaginal tissue, vaginal fluid, and plasma levels were relatively time independent over the 90-day duration at approximately 10(4) ng/g, 10(6) ng/g, and 10(1) ng/ml, respectively, near or exceeding the highest observed concentrations in a TFV 1% gel control group. TFV vaginal fluid concentrations were approximately 1,000-fold greater than levels shown to provide significant protection in women using the TFV 1% gel. There were no toxicological findings following placebo and TFV IVR treatment for 28 or 90 days, although slight to moderate increases in inflammatory infiltrates in the vaginal epithelia were observed in these animals compared to naïve animals. In summary, the controlled release of TFV from this reservoir IVR provided elevated sheep vaginal concentrations for 90 days to merit its further evaluation as an HIV prophylactic.

An assessment of the antinociceptive efficacy of intrathecal and epidural contulakin-G in rats and dogs.

Contulakin-G is a novel conopeptide with an incompletely defined mechanism of action. To assess nociceptive activity we delivered Contulakin-G as a bolus intrathecally (0.03, 0.1, 0.3, 3 nmol) or epidurally (10, 30, 89 nmol) in rats. Intrathecal Contulakin G significantly decreased Phase II and, to a lesser degree, Phase I paw flinching produced by intradermal formalin. Intrathecal and epidural doses of ED50s were 0.07 nmol and 45 nmol, respectively, giving an epidural/intrathecal ED50 ratio = 647). In dogs, intrathecal Contulakin-G (50-500 nmoL) produced a dose-dependent increase in the thermally evoked skin twitch latency by 30 min after administration, as did morphine (150 and 450 nmol). Epidural morphine (750 and 7500 nmol), but not epidural 1000 nmol Contulakin-G, also significantly decreased skin twitch in dogs. No changes in motor function were seen in any rats or dogs receiving these doses of Contulakin-G. In dogs, no physiologically significant dose-dependent changes in motor function, heart rate, arterial blood pressure, or body temperature were found. Contulakin-G is a potent antinociceptive drug when delivered intrathecally with no observable negative side effects in rats or dogs and may provide an alternative to opioid spinal analgesics.

Powerful antinociceptive effects of the cone snail venom-derived subtype-selective NMDA receptor antagonists conantokins G and T.

Subunit non-selective N-methyl-D-aspartate (NMDA) receptor antagonists reduce injury-induced pain behavior, but generally produce unacceptable side effects. In this study, we examined the antinociceptive and motor effects of cone snail venom-derived peptides, conantokins G and T (conG and conT), which are selective inhibitors of the NR2B or NR2A and NR2B subtypes of the NMDA receptor, respectively. We tested the effects of conG and conT in models of tissue (formalin test), nerve injury (partial sciatic nerve ligation) and inflammation-induced (intraplantar Complete Freund's Adjuvant; CFA) pain in mice. In the formalin test, intrathecal (i.t.) conG or conT suppressed the ongoing pain behavior (ED(50) and 95% confidence intervals (CI), 11 (7-19) and 19 (11-33), respectively) at doses that were 17-27 times lower than those required to impair motor function (accelerating rotarod treadmill test: ED(50) and 95% CI, 300 (120-730) and 320 (190-540) pmol, respectively). By comparison, SNX-111, an N-type voltage-sensitive calcium channel antagonist that is also derived from cone snail venom, produced significant motor impairment at a dose (3.0 pmol, i.t.) that was only partially efficacious in the formalin test. Furthermore, conG reversed the allodynia produced by nerve injury, with greater potency on thermal (ED50 and 95% CI, 24 (10-55) pmol) than on mechanical allodynia (59 (33-105) pmol). Finally, a single dose of conG (100 pmol, i.t.) also reduced CFA-evoked thermal and mechanical allodynia. Taken together, these results demonstrate that conantokins exhibit potent antinociceptive effects in several models of injury-induced pain. The study supports the notion that drugs directed against subtypes of the NMDA receptor, by virtue of their reduced side-effect profile, hold promise as novel therapeutic agents for the control of pain.

Intrathecal CGX-1007 is neuroprotective in a rat model of focal cerebral ischemia.

The NMDA antagonist CGX-1007 (Conantokin-G) has previously been shown to possess potent neuroprotective properties when administered intracranially following experimental ischemic brain injury. Using the same model of middle cerebral artery occlusion (MCAo) in rats we now report the neuroprotective effects of CGX-1007 when delivered intrathecally (i.t.). When given 4 h post-occlusion, a reduction in brain infarction was measured along with significant neurological recovery. Furthermore, we describe an i.t. neuroprotective therapeutic window lasting > or = 8 h from the start of the injury. Critically, this is the first comprehensive report of a neuroprotective agent that can be administered i.t. to ameliorate experimental brain injury and potentially provide an excellent therapeutic window as a neuroprotection treatment.

2-[2-[4-[2-[2-[ 1,3-Dihydro- 1,1-bis (4-hydroxyphenyl)-3-oxo-5-isobenzofuranthioureidyl]ethylaminocarbonyl]ethyl]phenyl] ethylamino]-5'-N-ethylcarboxamidoadenosine (FITC-APEC): A Fluorescent Ligand For A2a-Adenosine Receptors.

The fluorescein conjugate, FITC-APEC (2-[2-[4-[2-[2-[1,3-dihydro-l,l-bis(4-hydroxyphenyl)-3-oxo-5-isobenzofuranthioureidyl]ethylaminocarbonyl]ethyl]phenyl]ethylamino]-5'-N-ethylcarboxamidoadenosine), is a novel ligand derived from a series of functionalized congeners that act as selective A2a-adenosine receptor agonists. The binding of FITC-APEC to bovine striatal A2a,-adenosine receptors measured by fluorescence techniques was saturable and of a high affinity, with a Bmax, of 2.3 ± 0.3 pmol/mg protein and KD of 57 ± 2 nM. The KD value estimated by fluorescence was consistent with the Ki (11 ± 0.3 nM) obtained by competition studies with [3H]CGS 21680. Additionally, the Bmax, value found by FITC-APEC measurement was in agreement with Bmax, values obtained using radioligand binding. FITC-APEC exhibited rapid and reversible binding to bovine striatum. The potencies of chemically diverse A2a-adenosine receptor ligands estimated by inhibition of FITC-APEC binding were in good agreement with their potencies determined using radioligand binding techniques (r = 0.97, P = 0.0003). FITC-APEC binding was not altered by purine derivatives that do not recognize A2a-adenosine receptors. These findings demonstrate that the novel fluorescent ligand FITC-APEC can be used in the quantitative characterization of ligand binding to A2a-adenosine receptors.

A hot-melt extruded intravaginal ring for the sustained delivery of the antiretroviral microbicide UC781.

Microbicide intravaginal rings (IVRs) are a promising woman-controlled strategy for preventing sexual transmission of human immunodeficiency virus (HIV). An IVR was prepared and developed from polyether urethane (PU) elastomers for the sustained delivery of UC781, a highly potent nonnucleoside reverse transcriptase inhibitor of HIV-1. PU IVRs containing UC781 were fabricated using a hot-melt extrusion process. In vitro release studies of UC781 demonstrated that UC781 release profiles are loading dependent and resemble matrix-type, diffusion-limited kinetics. The in vitro release methods employed over predicted the in vivo release rates of UC781 in rabbits. Accelerated stability studies showed good chemical stability of UC781 in prototype formulations, but surface crystallization of UC781 was observed following long-term storage at higher UC781 loadings, unless formulated with a polyvinylpyrrolidone/glycerol surface coating. Mechanical stability testing of prototype rings showed moderate stiffening upon storage. The PU and UC781 had minimal to no impact on viability, tissue integrity, barrier function, or cytokine expression in the tissue irritation model, and UC781 was shown to be delivered to and permeate through this tissue construct in vitro. Overall, UC781 was formulated in a stable PU IVR and provided controlled release of UC781 both in vitro and in vivo.

Correlation of Pap smear, cervical biopsy, and clinical follow-up with an HPV typing microarray system.

A human papillomavirus (HPV) microarray system allows the determination of HPV type in clinical samples. The purpose of this study was to determine the presence of HPV in liquid-based Pap smears with the MyGene MyHPV Chip Kit HPV genotyping microarray test (MyGene Assay), and to correlate this with the cytology and biopsy diagnoses, clinical follow-up, HPV Hybrid Capture data, and HPV sequence analyses. Four hundred and two Pap smears (93 ThinPrep, 309 SurePath) were available for study. Correlation of HPV DNA detection by the MyGene Assay with the Pap smear diagnosis showed a detection rate of 19/97 (19%) for normal Pap smears, 181/242 (74%) for atypical squamous cells of undetermined significance (ASCUS), and 61/63 (97%) for squamous intraepithelial lesions (SILs). Biopsy data on 248 women were available. HPV was noted by the MyGene Assay in the Pap smear in 98/100 (98%) of the cases, for which the corresponding biopsy had been diagnosed as SIL, compared with 103/148 (69%) of the cases for which the biopsy had been negative for SIL. Clinical follow-ups were available for 200 women with ASCUS Pap smears. A significant increase was observed in the rate of biopsy-proven SILs in women with ASCUS Pap smears that were HPV-positive (63/66=95%) as compared with those that were HPV-negative (96/134=71%, P<0.05). The MyGene Assay and Hybrid Capture system gave equivalent results for all the categories studied, except for the presence of multiple infections, as determined by viral sequence analysis. Specifically, the Hybrid Capture system overestimated the presence of dual infection (low-risk and high-risk positive) by 48% and missed many cases of multiple infections, especially when 2 or more high-risk types were present. It is concluded that the MyGene Assay allows for the routine typing of HPV in liquid-based Pap smears, and that the presence of HPV DNA in ASCUS Pap smears is strongly correlated with a biopsy-proven SIL.