Preclinical Safety Evaluation and Human Dosimetry of [18F]MK-6240, a Novel PET Tracer for Imaging Neurofibrillary Tangles.

PURPOSE: [18F]MK-6240 is a selective, high-affinity positron emission tomography tracer for imaging neurofibrillary tangles, a key pathological signature that correlates with cognitive decline in Alzheimer disease. This report provides safety information from preclinical toxicology studies and first-in-human whole-body biodistribution and dosimetry studies of [18F]MK-6240 for its potential application in human brain imaging studies. PROCEDURES: MK-6240 was administered intravenously (IV) in a 7-day rat toxicity study at × 50, × 100, and × 1000 dose margins relative to projected highest clinical dose of 0.333 µg/kg. The IV formulation of MK-6240 for clinical use and the formulation used in the 7-day rat toxicity study was tested for hemolysis potential in human and Wistar rat whole blood. Sequential whole-body positron emission tomography scans were performed in three healthy young subjects after IV bolus injection of 180 ± 0.3 MBq [18F]MK-6240 to characterize organ biodistribution and estimate whole-body radiation exposure (effective dose). RESULTS: MK-6240 administered IV in a 7-day rat toxicity study did not show any test article-related changes. The no-observed-adverse-effect level in rats was ≥ 333 µg/kg/day which provides a margin 1000-fold over an anticipated maximum clinical dose of 0.333 µg/kg. Additionally, the MK-6240 formulation was not hemolytic in human or Wistar rat blood. [18F]MK-6240 activity was widely distributed to the brain and the rest of the body, with organ absorbed doses largest for the gall bladder (202 µGy/MBq). The average (±SD) effective dose was 29.4 ± 0.6 µSv/MBq, which is in the typical range for F-18 radiolabeled ligands. CONCLUSIONS: Microdoses of [18F]MK-6240 are safe for clinical positron emission tomography imaging studies. Single IV administration of 185 MBq (5 mCi) [18F]MK-6240 is anticipated to result in a total human effective dose of 5.4 mSv and thus allows multiple positron emission tomography scans of the same subject per year.

The novel gamma secretase inhibitor N-[cis-4-[(4-chlorophenyl)sulfonyl]-4-(2,5-difluorophenyl)cyclohexyl]-1,1,1-trifluoromethanesulfonamide (MRK-560) reduces amyloid plaque deposition without evidence of notch-related pathology in the Tg2576 mouse.

There is a substantial body of evidence indicating that beta-amyloid peptides (Abeta) are critical factors in the onset and development of Alzheimer's disease (AD). One strategy for combating AD is to reduce or eliminate the production of Abeta through inhibition of the gamma-secretase enzyme, which cleaves Abeta from the amyloid precursor protein (APP). We demonstrate here that chronic treatment for 3 months with 3 mg/kg of the potent, orally bioavailable and brain-penetrant gamma-secretase inhibitor N-[cis-4-[(4-chlorophenyl)-sulfonyl]-4-(2,5-difluorophenyl)cyclohexyl]-1,1,1-trifluoromethanesulfonamide (MRK-560) attenuates the appearance of amyloid plaques in the Tg2576 mouse. These reductions in plaques were also accompanied by a decrease in the level of reactive gliosis. The morphometric and histological measures agreed with biochemical analysis of Abeta(40) and Abeta(42) in the cortex. Interestingly, the volume of the plaques across treatment groups did not change, indicating that reducing Abeta levels does not significantly alter deposit growth once initiated. Furthermore, we demonstrate that these beneficial effects can be achieved without causing histopathological changes in the ileum, spleen, or thymus as a consequence of blockade of the processing of alternative substrates, such as the Notch family of receptors. This indicates that in vivo a therapeutic window between these substrates seems possible--a key concern in the development of this approach to AD. An understanding of the mechanisms whereby MRK-560 shows differentiation between the APP and Notch proteolytic pathway of gamma-secretase should provide the basis for the next generation of gamma-secretase inhibitors.

The effects of ad libitum feeding and marked dietary restriction on spontaneous skeletal muscle pathology in Sprague-Dawley rats.

The effects of ad libitum (AL) feeding and marked dietary restriction (DR) on spontaneous age-related skeletal muscle changes in male Sprague-Dawley (SD) rats were evaluated at 1 and 2 years. SD rats were fed Certified UAR A04C Rodent Chow ad libitum (AL), or DR at 50% of AL for (106 weeks). Body weights and organ weights were measured at the 1-year interim and 2-year final necropsies. In addition to the routine histopathologic examination, determination of 5 stereologic parameters was done in the vastus lateralis muscle after histochemistry of ATPase activity at 1 and 2 years. Body and skeletal muscle weights were proportional to the food intake. In AL-fed rats, muscle weights decreased between 1 and 2 years, in correlation with decreased type 2 myofiber numbers. In this group, fibrovascular index markedly increased with aging and muscle degeneration occurred at 2 years. In DR rats, there were no significant changes in muscle weights between 1 and 2 years. No histopathological changes were observed and the fibrovascular index was unchanged. These results demonstrated a protective effect of DR on the age-related skeletal muscle pathology in SD rats.

The ultrastructural localisation of the N-methyl-D-aspartate NR2B receptor subunit in rat lumbar spinal cord.

Glutamate together with its N-methyl-d-aspartate (NMDA) receptors has an important role in the transmission of stimuli in the spinal cord. Whilst the expression of the various NMDA receptor subunits within the spinal cord has been investigated the subcellular location of the NMDA NR2B subunit has yet to be definitively established. Both mRNA and light microscopical studies have failed to unequivocally demonstrate the proposed pre-synaptic location of this subunit. This has been proposed from pharmacological data and is thought to underlie the apparent analgesic properties of selective NR2B antagonists. Using pre-embedding immunohistochemistry combined with electron microscopy our findings provide the first definitive morphological evidence for both a pre- and post-synaptic localisation of NR2B/containing NMDA receptors, and suggest expression by astrocytes, in the rat lumbar spinal cord.

Mechanisms of action of the antidepressants fluoxetine and the substance P antagonist L-000760735 are associated with altered neurofilaments and synaptic remodeling.

Antidepressants are widely prescribed in the treatment of depression, although the mechanism of how they exert their therapeutic effects is poorly understood. To shed further light on their mode of action, we have attempted to identify a common proteomic signature in guinea pig brains after chronic treatment with two different antidepressants. Both fluoxetine and the substance P receptor (NK(1)R) antagonist (SPA) L-000760735 altered cortical expression of multiple heat shock protein 60 forms along with neurofilaments and related proteins that are critical determinants of synaptic structure and function. Analysis of NK(1)R-/- mice showed similar alterations of neurofilaments confirming the specificity of the effects observed with chronic NK(1)R antagonist treatment. To determine if these changes were associated with structural modification of synapses, we carried out electron microscopic analysis of cerebral cortices from fluoxetine-treated guinea pigs. This showed an increase in the percentage of synapses with split postsynaptic densities (PSDs), a phenomenon that is characteristic of activity-dependent synaptic rearrangement. These findings suggest that cortical alterations of the neurofilament pathway and increased synaptic remodeling are associated with the mechanism of these two antidepressant drug treatments and may contribute to their psychotherapeutic actions.

The effects of ad libitum overfeeding and moderate and marked dietary restriction on age-related spontaneous pituitary gland pathology in Sprague-Dawley rats.

This study compared the effects of ad libitum (AL) overfeeding and moderate or marked dietary restriction (DR) on the pathogenesis of aged-related pituitary gland changes in Sprague-Dawley (SD) rats. SD rats were fed Purina Certified Rodent Diet AL (group 1), DR at 72-79% of AL (group 2), DR at 68-72% of AL (group 3) or DR at 47-48% of AL (group 4) for 106 weeks. Interim necropsies were performed at 13, 26, and 53 weeks, after a 7-day 5-bromo-2-deoxyuridine (BrdU)-filled minipump implantation. Body weights, organ weights and insulin-like growth factor 1 (IGF-1) serum levels were measured at interim and final necropsies. Serum levels of prolactin (PRL), progesterone, estradiol, luteinizing hormone (LH) and follicle stimulating hormone (FSH) were measured at 53 and/or 106 weeks. In addition to the routine histopathologic examination, determination of 7 stereologic parameters after pituitary immunohistochemistry of PRL, growth hormone (GH) and BrdU was done in both sexes at 13, 26, and 53 weeks. Body and pituitary weights were proportional to the food intake. In AL-fed rats, hyperplastic and neoplastic changes developed early and progressed with age, affecting almost all animals by 106 weeks. These changes were associated with high PRL serum levels. Pituitary adenomas were the most common cause of death in both sexes. In DR rats, a delayed onset and a decreased incidence of pituitary tumors were observed in association with decreased serum IGF-1, PRL, estradiol, and LH levels. The results of the stereological analysis demonstrated that, compared to AL-fed rats, pituitary glands from DR rats contained lower PRL and GH secreting cell volumes, and a lower epithelial cell BrdU labeling index, which correlated with a lower incidence of pituitary tumors at study termination. Moderate and marked degrees of DR delayed the onset of pituitary tumors in a temporal- and dose-related manner. In contrast to marked DR, which dramatically reduced the incidence of hyperplastic and neoplastic pituitary gland changes, moderate DR delayed the onset but did not prevent the development of pituitary tumors.