The matrix metalloproteinase inhibitor marimastat inhibits seizures in a model of kainic acid-induced status epilepticus.

An intra-hippocampus injection of kainic acid serves as a model of status epilepticus and the subsequent development of temporal lobe epilepsy. Matrix metalloproteinase-9 (MMP-9) is an enzyme that controls remodeling of the extracellular milieu under physiological and pathological conditions. In response to brain insult, MMP-9 contributes to pathological synaptic plasticity that may play a role in the progression of an epileptic condition. Marimastat is a metalloproteinase inhibitor that was tested in clinical trials of cancer. The present study assessed whether marimastat can impair the development of epilepsy. The inhibitory efficacy of marimastat was initially tested in neuronal cultures in vitro. As a marker substrate, we used nectin-3. Next, we investigated the blood-brain barrier penetration of marimastat using mass spectrometry and evaluated the therapeutic potential of marimastat against seizure outcomes. We found that marimastat inhibited the cleavage of nectin-3 in hippocampal neuronal cell cultures. Marimastat penetrated the blood-brain barrier and exerted an inhibitory effect on metalloproteinase activity in the brain. Finally, marimastat decreased some seizure parameters, such as seizure score and number, but did not directly affect status epilepticus. The long-term effects of marimastat were evident up to 6 weeks after kainic acid administration, in which marimastat still inhibited seizure duration.

Discovery of Novel Inhibitors of LpxC Displaying Potent in Vitro Activity against Gram-Negative Bacteria.

UDP-3-O-((R)-3-hydroxymyristoyl)-N-glucosamine deacetylase (LpxC) is as an attractive target for the discovery and development of novel antibacterial drugs to address the critical medical need created by multidrug resistant Gram-negative bacteria. By using a scaffold hopping approach on a known family of methylsulfone hydroxamate LpxC inhibitors, several hit series eliciting potent antibacterial activities against Enterobacteriaceae and Pseudomonas aeruginosa were identified. Subsequent hit-to-lead optimization, using cocrystal structures of inhibitors bound to Pseudomonas aeruginosa LpxC as guides, resulted in the discovery of multiple chemical series based on (i) isoindolin-1-ones, (ii) 4,5-dihydro-6H-thieno[2,3-c]pyrrol-6-ones, and (iii) 1,2-dihydro-3H-pyrrolo[1,2-c]imidazole-3-ones. Synthetic methods, antibacterial activities and relative binding affinities, as well as physicochemical properties that allowed compound prioritization are presented. Finally, in vivo properties of lead molecules which belong to the most promising pyrrolo-imidazolone series, such as 18d, are discussed.

Optimization of LpxC Inhibitor Lead Compounds Focusing on Efficacy and Formulation for High Dose Intravenous Administration.

LpxC inhibitors were optimized starting from lead compounds with limited efficacy and solubility and with the goal to provide new options for the treatment of serious infections caused by Gram-negative pathogens in hospital settings. To enable the development of an aqueous formulation for intravenous administration of the drug at high dose, improvements in both solubility and antibacterial activity in vivo were prioritized early on. This lead optimization program resulted in the discovery of compounds such as 13 and 30, which exhibited high solubility and potent efficacy against Gram-negative pathogens in animal infection models.

Reducing Antibacterial Development Risk for GSK1322322 by Exploring Potential Human Dose Regimens in Nonclinical Efficacy Studies Using Immunocompetent Rats.

Directly testing proposed clinical dosing regimens in nonclinical studies can reduce the risk during the development of novel antibacterial agents. Optimal dosing regimens can be identified in animal models by testing recreated human pharmacokinetic profiles. An example of this approach using continuous intravenous infusions of GSK1322322 in immunocompetent rats to evaluate recreated human exposures from phase I trials in pneumonia models with Streptococcus pneumoniae and Haemophilus influenzae and an abscess model with Staphylococcus aureus is presented. GSK1322322 was administered via continuous intravenous infusion to recreate 1,000- or 1,500-mg oral doses every 12 h in humans. Significant reductions (P ≤ 0.05 for all comparisons) in bacterial numbers compared with those for the baseline controls were observed for S. pneumoniae and H. influenzae (mean log10 reductions, 1.6 to ≥2.7 and 1.8 to 3.3 CFU/lungs, respectively) with the recreated 1,000-mg oral dose. This profile was also efficacious against S. aureus (mean log10 reduction, 1.9 to 2.4 CFU/abscess). There was a nonsignificant trend for improved efficacy against S. aureus with the 1,500-mg oral dose (mean log10 reduction, 2.4 to 3.1 CFU/abscess). These results demonstrate that the human oral 1,000- or 1,500-mg exposure profiles of GSK1322322 recreated in rats were effective against representative community-associated pathogens and supported selection of the 1,500-mg oral dose given every 12 h for a phase II clinical skin infection study. Furthermore, this work exemplifies how the testing of recreated human pharmacokinetic profiles can be incorporated into the development process and serve as an aid for selecting optimal dosing regimens prior to conducting large-scale clinical studies.

Panobinostat for the treatment of acute myelogenous leukemia.

INTRODUCTION: Therapeutic strategies in patients with acute myeloid leukemia (AML) have not changed significantly over the last decades. Appropriate strategies are ultimately driven by the assessment of patients' fitness to define suitability for intensive induction chemotherapy, which produces high initial remission rates but, increased likelihood of relapse. Old/unfit AML patients still represent an urgent and unmet therapeutic need. Epigenetic deregulation represents a strategic characteristic of AML pathophysiology whereby aberrant gene transcription provides an advantage to leukemic cell survival. Efforts to re-establish impaired epigenetic regulation include hypomethylating agents and histone deacetylase inhibitors (HDACi). AREAS COVERED: The review discusses the underlying mechanisms leading to disruption of lysine acetyltransferases (KAT or HAT)/deacetylase (KDAC or HDAC) balance and the rationale for using the HDACi panobinostat (LBH-589) in AML. EXPERT OPINION: Although panobinostat has produced significant results in myeloma, its efficacy remains limited in AML. Panobinostat exerts pleiotropic activity and lack of specificity, which likely contributes to its inadequate safety in elderly AML patients. Phase I-II trials, utilizing panobinostat associated with well-known chemotherapeutic agents are ongoing and combinations with other druggable targets may likely be evaluated in future trials. The clinical use of this HDACi in AML the near future does not appearing promising.

Pyridone methylsulfone hydroxamate LpxC inhibitors for the treatment of serious gram-negative infections.

The synthesis and biological activity of a new series of LpxC inhibitors represented by pyridone methylsulfone hydroxamate 2a is presented. Members of this series have improved solubility and free fraction when compared to compounds in the previously described biphenyl methylsulfone hydroxamate series, and they maintain superior Gram-negative antibacterial activity to comparator agents.

A randomized phase II study of two doses of vorinostat in combination with 5-FU/LV in patients with refractory colorectal cancer.

BACKGROUND: Vorinostat is synergistic with 5-FU in vitro and in vivo models. A combination of these two agents was associated with clinical activity in 5-FU refractory colorectal cancer patients in a phase I clinical trial, therefore warranting the conduct of this prospective phase II study. PATIENTS AND METHODS: Patients with refractory metastatic colorectal cancer were randomized in a two-stage design to receive vorinostat at 800 or 1,400 mg/day once a day × 3, every 2 weeks. 5-FU, preceded by leucovorin, was administered as a bolus followed by a 46-h infusion on days 2 and 3 of vorinostat. A pre-specified 2-month progression-free survival (PFS) rate of 27/43 patients per arm was needed to deem an arm interesting for further investigation. RESULTS: The high-dose vorinostat arm did not reach the needed efficacy endpoint at completion of the first stage, with only 8 out of 15 patients being alive and progression free at 2 months. The low-dose vorinostat arm proceeded to accrue 43 patients with a 2-month PFS rate of 53% (23 out 43), including one partial response. The median PFS and overall survival on the low-dose arm were 2.4 and 6.5 months, respectively. Both treatment arms were well tolerated. No differences were noted in the pharmacokinetics of vorinostat at the 800- or 1,400-mg dose-levels, suggesting bioavailability saturation. CONCLUSIONS: While the addition of vorinostat to 5-FU resulted in 1 partial response and in some disease stabilizations, the limited activity does not warrant the unselected use of this combination in chemotherapy-refractory colorectal cancer.

Clinical development of panobinostat in classical Hodgkin's lymphoma.

Deacetylase (DAC) inhibitors are promising new anticancer drugs that have complex mechanisms of action, including induction of cell-cycle arrest and apoptosis, inhibition of angiogenesis and induction of a favorable anti-tumor immune response. Panobinostat, a potent inhibitor of DAC 1-11 enzymatic activity, has demonstrated a significant in vitro antiproliferative activity against classical Hodgkin's lymphoma (cHL) cell lines in addition to a promising clinical activity in early Phase I studies in patients with relapsed cHL. In a recently completed large Phase II study in patients with relapsed cHL, panobinostat reduced tumor measurements in 74% of patients, including 23% partial and 4% complete remissions. In this article, we review the status of panobinostat drug development and compare its activity to those of other DAC inhibitors in patients with relapsed cHL. Future investigations should focus on designing rational combination regimens and identifying predictive markers that will assist in selecting patients who are likely to benefit from this novel therapy.

Nonclinical safety assessment of the histone deacetylase inhibitor vorinostat.

Vorinostat (SAHA, Zolinza), a histone deacetylase inhibitor, is assessed in nonclinical studies to support its approval for cutaneous T-cell lymphoma. Vorinostat is weakly mutagenic in the Ames assay; is clastogenic in rodent (ie, CHO) cells but not in normal human lymphocytes; and is weakly positive in an in vivo mouse micronucleus assay. No effects are observed on potassium ion currents in the hERG assay up to 300 microM (safety margin approximately 300-fold the approximately 1 microM serum concentration associated with the 400 mg/d maximum recommended human dose. No rat respiratory or central nervous system effects are found at 150 mg/kg (>2-fold maximum recommended human dose). No cardiovascular effects, including effects on QTc interval, are observed after a single oral dose (150 mg/kg) in dogs. Vorinostat is orally dosed daily in rats (controls, 20, 50, or 150 mg/kg/d) and dogs (controls, 60, 80, or 100/125/160 mg/kg/d) for 26 weeks with a 4-week recovery. Rat vorinostat-related adverse findings are decreased food consumption, weight loss, and hematologic changes; a no observed adverse effects level is not established. In dogs, adverse effects are primarily gastrointestinal; the no observed adverse effects level is 60 mg/kg/d (approximately 6-fold maximum recommended human dose). Toxicities are reversible and can be monitored in the clinic.

In vitro phase I cytochrome P450 metabolism, permeability and pharmacokinetics of SB639, a novel histone deacetylase inhibitor in preclinical species.

In vitro liver microsomal stability, permeability, pharmacokinetics (PK) and oral bioavailability of SB639, a novel HDACi (Histone Deacetylase inhibitor), were determined. The in vitro metabolism was examined in mouse, rat, dog and human liver microsomes. The permeability and efflux potential of SB639 were determined using Caco-2 cell monolayers. To determine pharmacokinetics and oral bioavailability, blood samples were drawn at pre-determined intervals up to 24 h post-dose after single intravenous (i.v.) or oral (p.o.) administration of SB639 to mouse or rat. The concentrations of SB639 in plasma samples were determined by a validated LC-MS/MS method. In vitro liver microsomal stability data revealed that SB639 was stable in human and dog liver microsomes, unstable in mouse and rat liver microsomes. The Caco-2 data has shown that SB639 is highly permeable with an apparent permeability of 3.01.10(-6) cm/s at 10 microM. After oral administration, maximum concentrations of SB639 were achieved within 0.5 h of post dose. Following i.v. administration, the concentration of SB639 declined in a bi-exponential fashion with terminal elimination half-life of 1.67 h for mice and 1.12 h for rats. The systemic clearance and volume of distribution of SB639 in mice were 15.8 l/h/kg and 38 l/kg, respectively, while the respective values in rats were 3.84 l/h/kg and 3.67 l/kg. Elimination half-life in rats ranged between 1.12-2.26 h. Absolute oral bioavailability of SB639 in mouse and rat was 13% and 10%, respectively. In conclusion, the superior potency, physicochemical and PK properties of SB639 compared to the recently FDA approved drug Zolinza (Suberoylanilide hydroxamic acid or Vorinostat) in the preclinical setting makes it a potential clinical candidate.

Disposition of vorinostat, a novel histone deacetylase inhibitor and anticancer agent, in preclinical species.

The disposition of vorinostat, an anticancer agent, was investigated in rats and dogs. Vorinostat possessed high serum clearance, a short elimination half-life and low oral bioavailability in both species. The renal route played an important role in the elimination of drug-related material and vorinostat was eliminated primarily by metabolic biotransformation.

[Development of new hypoxic cell radiosensitizer in Japan].

Basic studies in hypoxic cell radiosensitizers developed in Japan were reviewed. Because of the unsuccessful clinical trials of misonidazole (MISO), many efforts have been made to find a new hypoxic cell sensitizer which is more effective and/or less toxic than MISO. Already over a thousand drugs were tested, but the majority of them did not work in vivo or were very toxic to the animals. Finally four drugs, KU-2285, KIH-801 (802), RK-28 and RP-170, have been proven to be effective both in vitro and in vivo. All of them are derivatives of 2-nitroimidazole. The first two have different side chains of fluorinated amide and acetohydroxamic acid, respectively. The second two both have sugar moieties. Investigations are now under way to find possible clinical applicability.

Selective inhibition of arachidonate 5-lipoxygenase by novel acetohydroxamic acids: effects on bronchial anaphylaxis in anaesthetized guinea-pigs.

1. The effect of a novel series of orally-active acetohydroxamic acid inhibitors of arachidonate 5-lipoxygenase on 'leukotriene-dependent' anaphylactic bronchoconstriction has been investigated in anaesthetized, pump-ventilated guinea-pigs actively sensitized to ovalbumin (OA). In a complementary series of experiments, the pharmacokinetics of these compounds in the plasma compartment following oral administration to guinea-pigs has also been investigated. 2. In animals pretreated with mepyramine (2 mg kg-1, i.v.) and indomethacin (10 mg kg-1, i.v.) and challenged with antigen aerosol (OA 10 mg ml-1; 5 s) compounds BW A4C, BW A137C and BW A797C (10-200 mg kg-1, p.o., 1 h pre-challenge) markedly reduced that component of anaphylactic bronchoconstriction shown to be 'leukotriene-dependent'. 3. The maximum degree of inhibition (up to 75%) of 'leukotriene-dependent' anaphylactic bronchoconstriction by these three compounds was equivalent to that seen with the leukotriene antagonist FPL 55712 (10 mg kg-1, i.v.). 4. The peak levels of unchanged acetohydroxamic acids in the plasma compartment occurred 0.5 h after their oral administration and were as follows: BW A4C: 11.3 +/- 3.9; BW A137C: 7.6 +/- 2.4; BW A797C: 3.9 +/- 1.3 micrograms ml-1 plasma. 5. The inhibition by BW A4C and BW A137C (50 mg kg-1, p.o.) of 'leukotriene-dependent' anaphylactic bronchospasm persisted for up to 3 and 4 h respectively but did not extend to 6 h. The decline in inhibitory activity paralleled the fall in the concentration of unchanged drug in the plasma compartment over this time period. 6. The results of the present study are consistent with BW A4C, BW A137C and BW A797C attenuating 'leukotriene-dependent' bronchial anaphylaxis in anaesthetized guinea-pigs by selective inhibition of arachidonate 5-lipoxygenase.