In several pedigrees of early onset familial Alzheimer's disease (FAD), point mutations in the beta-amyloid precursor protein (APP) gene are genetically linked to the disease. This finding implicates APP in the pathogenesis of Alzheimer's disease in these individuals. To understand the in vivo function of APP and its processing, we have generated an APP-null mutation in mice. Homozygous APP-deficient mice were viable and fertile. However, the mutant animals weighed 15%-20% less than age-matched wild-type controls. Neurological evaluation showed that the APP-deficient mice exhibited a decreased locomotor activity and forelimb grip strength, indicating a compromised neuronal or muscular function. In addition, four out of six homozygous mice showed reactive gliosis at 14 weeks of age, suggesting an impaired neuronal function as a result of the APP-null mutation.

Summary. Intratracheal instillation (IT) of bleomycin is a widely used experimental model for lung fibrosis. In this study we describe the time‐course of bleomycin‐induced lung fibrosis in mice using computer‐assisted morphometry. C57Bl/6J mice were treated with a single IT dose of bleomycin or control saline. Animals were killed 3, 6, 14 and 21 days post‐IT. Lung injury was evaluated by analysis of bronchoalveolar lavage (BAL) fluid, hydroxyproline concentration in the lung, routine light microscopic examination resulting in a semiquantitative morphological index (SMI) of lung injury, and quantitative morphological measurements (fibrosis fraction and alveolar wall area fraction) aided by optimas image analysis software. Changes in BAL fluid attributed to bleomycin treatment include increased total cell count (days 14 and 21), and increased percentage of neutrophils (days 3 and 6) followed by a sustained increase in lymphocytes (days 6, 14 and 21). Hydroxyproline levels increased in bleomycin‐treated mice on days 14 and 21. Median SMI grades were significantly elevated on days 3, 14 and 21. Computer‐assisted morphometry demonstrated a 3‐fold increase in fibrosis fraction and a 1.3‐fold increase in wall area fraction in bleomycin‐treated mice on day 14, with no further increase on day 21. These data also demonstrate that the most suitable time point for assessing lung fibrosis in this model is 14 days after IT instillation of bleomycin, based on the observation that at 14 days the animals developed extensive fibrosis, but had less variability in the fibrotic response and lower mortality than later at 21 days. Computer‐assisted morphometry provides objective and quantitative measurements that are a useful tool for the evaluation of bleomycin‐induced lung injury.

For Ras oncoproteins to transform mammalian cells, they must be post-translationally modified with a farnesyl group in a reaction catalysed by the enzyme farnesyl-protein transferase (FPTase). Inhibitors of FPTase have therefore been proposed as anti-cancer agents. We show that L-744,832, which mimics the CaaX motif to which the farnesyl group is added, is a potent and selective inhibitor of FPTase. In MMTV-v-Ha-ras mice bearing palpable tumours, daily administration of L-744,832 caused tumour regression. Following cessation of treatment, tumours reappeared, the majority of which regressed upon retreatment. No systemic toxicity was found upon necropsy of L-744,832-treated mice. This first demonstration of anti-FPTase-mediated tumour regression suggests that FPTase inhibitors may be safe and effective anti-tumour agents in some cancers.

The posttranslational addition of a farnesyl moiety to the Ras oncoprotein is essential for its transforming activity. Cell-active inhibitors of the enzyme that catalyzes this reaction, protein farnesyltransferase, have been shown to selectively block ras-dependent transformation of cells in culture. Here we describe the protein farnesyltransferase inhibitor

The farnesyltransferase inhibitor L-744,832 selectively blocks the transformed phenotype of cultured cells expressing a mutated H-ras gene and induces dramatic regression of mammary and salivary carcinomas in mouse mammary tumor virus (MMTV)-v-Ha-ras transgenic mice. To better understand how the farnesyltransferase inhibitors might be used in the treatment of human tumors, we have further explored the mechanisms by which L-744,832 induces tumor regression in a variety of transgenic mouse tumor models. We assessed whether L-744,832 induces apoptosis or alterations in cell cycle distribution and found that the tumor regression in MMTV-v-Ha-ras mice could be attributed entirely to elevation of apoptosis levels. In contrast, treatment with doxorubicin, which induces apoptosis in many tumor types, had a minimal effect on apoptosis in these tumors and resulted in a less dramatic tumor response. To determine whether functional p53 is required for L-744,832-induced apoptosis and the resultant tumor regression, MMTV-v-Ha-ras mice were interbred with p53 ؊/؊ mice. Tumors in ras/p53 ؊/؊ mice treated with L-744,832 regressed as efficiently as MMTV-v-Haras tumors, although this response was found to be mediated by both the induction of apoptosis and an increase in G 1 with a corresponding decrease in the S-phase fraction. MMTV-v-Ha-ras mice were also interbred with MMTV-c-myc mice to determine whether ras/myc tumors, which possess high levels of spontaneous apoptosis, have the potential to regress through a further increase in apoptosis levels. The ras/myc tumors were found to respond nearly as efficiently to L-744,832 treatment as the MMTV-v-Ha-ras tumors, although no induction of apoptosis was observed. Rather, the tumor regression in the ras/myc mice was found to be mediated by a large reduction in the S-phase fraction. In contrast, treatment of transgenic mice harboring an activated MMTVc-neu gene did not result in tumor regression. These results demonstrate that a farnesyltransferase inhibitor can induce regression of v-Ha-ras-bearing tumors by multiple mechanisms, including the activation of a suppressed apoptotic pathway, which is largely p53 independent, or by cell cycle alterations, depending upon the presence of various other oncogenic genetic alterations.

The efficacy of telavancin, a bactericidal lipoglycopeptide, was compared to that of vancomycin and linezolid against methicillin-resistant Staphylococcus aureus (MRSA) in a murine pneumonia model. Telavancin produced greater reductions in lung bacterial titer and mortality than did vancomycin and linezolid at human doses equivalent to those described by the area under the concentration-time curve. These results suggest the potential utility of telavancin for treatment of MRSA pneumonia.Nosocomial pneumonia due to methicillin-resistant Staphylococcus aureus (MRSA) has been reported with increasing frequency over the past 2 decades (13). Vancomycin and linezolid, two drugs that are slowly bactericidal and bacteriostatic, respectively, are frequently used in the treatment of nosocomial pneumonia caused by Staphylococcus aureus, including MRSA. However, overall clinical cure rates attained with these two drugs are less than optimal (8).Telavancin is a novel lipoglycopeptide that operates through multiple mechanisms to produce potent and rapid bactericidal activity against clinically relevant gram-positive bacteria, including MRSA (4, 5, 7). We have previously shown that telavancin exhibits potent antibacterial activity against a range of gram-positive bacteria, including MRSA, in two animal models of soft-tissue infection, with the area under the concentrationtime curve (AUC)/MIC ratio being the pharmacodynamically linked parameter (3). In the studies described here, we assessed the efficacy of telavancin in an immunocompromised murine model of MRSA-induced pneumonia.Telavancin for injection (250 mg/vial) was reconstituted in 5% dextrose in water. Vancomycin (Sigma-Aldrich, St. Louis, MO) and linezolid (Pharmacia, Kalamazoo, MI) were dissolved in 5% dextrose in water and hydroxypropyl-␤-cyclodextrin, respectively. MRSA ATCC 33591 was obtained from the American Type Culture Collection (Manassas, VA). MICs were determined by the broth microdilution method according to protocol M7-A5 of the National Committee for Clinical Laboratory Standards (6). The MICs of telavancin, vancomycin, and linezolid against MRSA 33591 were 0.5, 1, and 1 g/ml, respectively. The doses used in pharmacodynamic studies were chosen to approximate human doses equivalent to those described by AUCs as opposed to other pharmacokinetic parameters since the AUC/MIC ratio is believed to be the key pharmacodynamically linked parameter for all three drugs against S. aureus (1, 3).All studies were approved by the Institutional Animal Care and Use Committee at Theravance, Inc. Female BALB/c mice (Harlan, Indianapolis, IN), weighing between 16 and 26 g, were rendered neutropenic by treatment with 250 mg/kg of cyclophosphamide intraperitoneally at 4 and 2 days prior to infection. Neutropenic animals were lightly anesthetized with isoflurane gas and then held in an upright position to be inoculated. Each animal was inoculated by placing 50 l of inoculum containing approximately 10 7 CFU of MRSA onto the tip of the nares. The animals were allowed to inhale t...

To understand the in vivo function of the amyloid precursor protein (APP) we generated an APP null mutation in mice by homologous recombination in embryonic stem (ES) cells. We show here that homozygous APP deficient mice were produced at expected frequencies. Neither APP mRNA nor protein could be detected in these animals. Yet the homozygous APP mutant mice are fertile and do not show overt abnormalities at up to 12 weeks of age. Neuroanatomical studies of the brain did not reveal significant differences in the knockout mice as compared to the wild-type controls. These results argue against an essential function of APP in mouse embryonic and early neuronal development.

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