Albany Research Institute

BACKGROUND: Moxifloxacin is a quinolone antimicrobial that has potent activity against Mycobacterium tuberculosis. To optimize moxifloxacin dose and dose regimen, pharmacodynamic antibiotic-exposure targets associated with maximal microbial kill and complete suppression of drug resistance in M. tuberculosis must be identified. METHODS: We used a novel in vitro pharmacodynamic infection model of tuberculosis in which we exposed M. tuberculosis to moxifloxacin with a pharmacokinetic half-life of decline similar to that encountered in humans. Data obtained from this model were mathematically modeled, and the drug-exposure breakpoint associated with the suppression of drug resistance was determined. Monte-Carlo simulations were performed to determine the probability that 10,000 clinical patients taking different doses of moxifloxacin would achieve or exceed the drug-exposure breakpoint needed to suppress resistance to moxifloxacin in M. tuberculosis. RESULTS: The ratio of the moxifloxacin-free (non-protein-bound) area under the concentration-time curve from 0 to 24 h to the minimum inhibitory concentration associated with complete suppression of the drug-resistant mutant population was 53. For patients taking moxifloxacin doses of 400, 600, or 800 mg/day, the calculated target-attainment rates to suppress drug resistance were 59%, 86%, and 93%, respectively. CONCLUSION: A moxifloxacin dose of 800 mg/day is likely to achieve excellent M. tuberculosis microbial kill and to suppress drug resistance. However, tolerability of this higher dose is still unknown.

Conventional chemotherapy not only kills tumor cells but also changes gene expression in treatment-damaged tissues, inducing production of multiple tumor-supporting secreted factors. This secretory phenotype was found here to be mediated in part by a damage-inducible cell-cycle inhibitor p21 (CDKN1A). We developed small-molecule compounds that inhibit damage-induced transcription downstream of p21. These compounds were identified as selective inhibitors of a transcription-regulating kinase CDK8 and its isoform CDK19. Remarkably, p21 was found to bind to CDK8 and stimulate its kinase activity. p21 and CDK8 also cooperate in the formation of internucleolar bodies, where both proteins accumulate. A CDK8 inhibitor suppresses damage-induced tumor-promoting paracrine activities of tumor cells and normal fibroblasts and reverses the increase in tumor engraftment and serum mitogenic activity in mice pretreated with a chemotherapeutic drug. The inhibitor also increases the efficacy of chemotherapy against xenografts formed by tumor cell/fibroblast mixtures. Microarray data analysis revealed striking correlations between CDK8 expression and poor survival in breast and ovarian cancers. CDK8 inhibition offers a promising approach to increasing the efficacy of cancer chemotherapy.

Expression of class III β-tubulin (βIII-tubulin) correlates with tumor progression and resistance to taxane-based therapies for several human malignancies, but its use as a biomarker of tumor behavior in prostate cancer (PCa) remains largely unexplored. Here, we describe βIII-tubulin immunohistochemical staining patterns of prostate tumors obtained from a broad spectrum of PCa patients, some of whom subsequently received docetaxel therapy for castration-resistant PCa (CRPC). Elevated βIII-tubulin expression was significantly associated with tumor aggressiveness in PCa patients with presumed localized disease, as it was found to be an independent marker of biochemical recurrence after treatment. Additionally, βIII-tubulin expression in tumor cells was an independent predictor of lower overall survival for patients receiving docetaxel-based chemotherapy for CRPC. Manipulation of βIII-tubulin expression in human PCa cell lines using a human βIII-tubulin expression vector or βIII-tubulin small interfering RNA altered cell survival in response to docetaxel treatment in a manner that supports a role for βIII-tubulin expression as a mediator of PCa cell resistance to docetaxel therapy. Our findings suggest a role for βIII-tubulin as candidate theranostic biomarker to predict the response to docetaxel-based chemotherapy as well as to target for treatment of docetaxel-resistant CRPC.

Humans die from age-related diseases, which are deadly manifestations of the aging process. In order to extend life span, an anti-aging drug must delay age-related diseases. All together age-related diseases are the best biomarker of aging. Once a drug is used for treatment of any one chronic disease, its effect against other diseases (atherosclerosis, cancer, prostate enlargement, osteoporosis, insulin resistance, Alzheimer's and Parkinson's diseases, age-related macular degeneration) may be evaluated in the same group of patients. If the group is large, then the anti-aging effect could be validated in a couple of years. Startlingly, retrospective analysis of clinical and preclinical data reveals four potential anti-aging modalities.

The echinocandins potentially have an important role in treatment of infections caused by Candida spp. and Aspergillus spp. in immunocompromised children. However, there are no population pharmacokinetic models of the echinocandins for pediatric patients. The safety and descriptive pharmacokinetics of micafungin in children were recently reported. However, a population pharmacokinetic model in children is needed in order to accurately determine the dosage of micafungin that produces an equivalent magnitude of drug exposure to that observed in adults. In order to explore the effect of weight on micafungin pharmacokinetics, a standard two-compartment pharmacokinetic model, a linear model, and an allometric power model were developed. For all three models, the fit to the data was excellent, with comparable measures of precision and bias. However, the superior log-likelihood value of the allometric power model suggested that it best reflected the data and was therefore chosen for a more detailed analysis of the magnitude and pattern of drug exposure which develop following the administration of micafungin. The allometric power model suggested that clearance in smaller children is higher than that predicted on the basis of weight alone. Consequently, a degree of dosage increase is required in smaller children to ensure comparable levels of drug exposure to those observed in larger children and adults. The allometric power model developed in this study enables identification of pediatric dosage regimens of micafungin which, based upon Monte Carlo simulations, result in equivalent drug exposures to those observed in adults, for which antifungal efficacy has been established.

BACKGROUND: Class III beta-tubulin (betaIII-tubulin) is expressed in tissues of neuronal lineage and also in several human malignancies, including non-small-cell lung carcinoma, breast and ovarian cancer. Overexpression of betaIII-tubulin in these tumours is associated with an unfavourable outcome and resistance to taxane-based therapies. At present, betaIII-tubulin expression remains largely uncharacterised in prostate cancer. METHODS: In this report, we evaluated the expression of betaIII-tubulin in 138 different human prostate tumour specimens by immunohistochemistry from patients with hormone-treated or hormone-untreated prostate cancer. betaIII-tubulin expression was also examined in various prostatic cancer cell lines including in androgen-sensitive human prostate cancer cells, LNCaP, grown in androgen-depleted medium in 2D cultures or as tumour xenografts when the host mouse was castrated. RESULTS: Whereas moderate-to-strong betaIII-tubulin expression was detected in only 3 out of 74 (4%) hormone-naive tumour specimens obtained from patients who never received hormone therapy, 6 out of 24 tumour specimens (25%) from patients treated for 3 months with neoadjuvant hormone therapy and 24 out of 40 (60%) castration-resistant tumour specimens from chronic hormone-treated patients were found to express significant levels of betaIII-tubulin. These findings were supported by in vitro and in vivo settings. CONCLUSION: Our data indicate that betaIII-tubulin expression is augmented in prostate cancer by androgen ablation and that the expression of this beta-tubulin isoform is associated with the progression of prostate cancer to the castration-resistant state, a stage largely responsible for mortality from prostate cancer.

The proliferative lifespan of normal somatic human cells in culture terminates in a permanent growth-arrested state known as replicative senescence. In this study, we show that RNA interference-mediated repression of the genes encoding the small ubiquitin-related modifier (SUMO)-specific proteases, Senp1, Senp2, and Senp7, induced low passage primary human fibroblasts to senesce rapidly. Following Senp1 repression, we observed a global increase in sumoylated proteins and in the number and size of nuclear SUMO-containing promyelocytic leukemia (PML) bodies. SUMO/PML bodies also increased during replicative senescence. p53 transcriptional activity was enhanced towards known p53 target genes following repression of Senp1, and inhibition of p53 function prevented senescence after Senp1 repression. These data indicate that Senp1 repression induces p53-mediated premature senescence and that SUMO proteases may thus be required for proliferation of normal human cells.

Background Studies examining sex differences in CYP3A activity have produced conflicting results. Our objective is to investigate whether sex differences exist in CYP3A activity as assessed by intravenous (IV) or oral midazolam pharmacokinetic analysis in healthy volunteers. Methods Data from 13 previous studies were used. A single dose of IV midazolam (0.025 mg/kg) was administered to 66 white adults (37 women and 29 men; mean age, 36.3 ± 7.7 years). A single dose of oral midazolam, 0.075 mg/kg (5 studies), 0.15 mg/kg (1 study), or 5 mg (1 study), was administered to 72 adults (71 white and 1 Asian; 37 women and 35 men; mean age, 38.3 ± 8.9 years). Pharmacokinetic parameters were determined via population methods by use of a nonparametric adaptive grid program and a 2-compartment IV and 1-compartment oral absorption model. The maximum a posteriori probability Bayesian method was used to estimate each subject's pharmacokinetic parameters. Monte Carlo simulation was performed to determine the probability distribution of the area under the concentration-time curves (AUCs). Results Women exhibited 11% higher mean weight-corrected total body midazolam clearance and 28% higher oral clearance compared with men (P ≤ .01). The median AUC of IV midazolam was 0.05 mg · h/L (range, 0.02–0.14 mg · h/L) in women and 0.06 mg · h/L (range, 0.02–0.14 mg · h/L) in men. The median AUC of oral midazolam was 0.11 mg · h/L (range, 0.02–0.60 mg · h/L) in women and 0.12 mg · h/L (range, 0.04–0.45 mg · h/L) in men. Conclusions Although women showed significantly greater hepatic and intestinal CYP3A activity, only a minor sex difference in AUC was noted. Therefore the observed disparity may be of negligible clinical importance. Clinical Pharmacology & Therapeutics (2006) 80, 531–538; doi: 10.1016/j.clpt.2006.08.014

Anticancer drugs are effective against tumors that depend on the molecular target of the drug. Known targets of cytotoxic anticancer drugs are involved in cell proliferation; drugs acting on such targets are ineffective against nonproliferating tumor cells, survival of which leads to eventual therapy failure. Function-based genomic screening identified the coatomer protein complex ζ1 (COPZ1) gene as essential for different tumor cell types but not for normal cells. COPZ1 encodes a subunit of coatomer protein complex 1 (COPI) involved in intracellular traffic and autophagy. The knockdown of COPZ1, but not of COPZ2 encoding isoform coatomer protein complex ζ2, caused Golgi apparatus collapse, blocked autophagy, and induced apoptosis in both proliferating and nondividing tumor cells. In contrast, inhibition of normal cell growth required simultaneous knockdown of both COPZ1 and COPZ2. COPZ2 (but not COPZ1) was down-regulated in the majority of tumor cell lines and in clinical samples of different cancer types. Reexpression of COPZ2 protected tumor cells from killing by COPZ1 knockdown, indicating that tumor cell dependence on COPZ1 is the result of COPZ2 silencing. COPZ2 displays no tumor-suppressive activities, but it harbors microRNA 152, which is silenced in tumor cells concurrently with COPZ2 and acts as a tumor suppressor in vitro and in vivo. Silencing of microRNA 152 in different cancers and the ensuing down-regulation of its host gene COPZ2 offer a therapeutic opportunity for proliferation-independent selective killing of tumor cells by COPZ1-targeting agents.

OBJECTIVE: To report a case series of high-dose continuous infusion beta-lactam antibiotics for the treatment of resistant Pseudomonas aeruginosa infections. CASE SUMMARY: Continuous infusion ceftazidime or aztreonam was administered to achieve target drug concentrations at or above the minimum inhibitory concentration, when possible, in 3 patients with P. aeruginosa infections. The maximal calculated target drug concentration was 100 mg/L. In the first patient, with primary immunodeficiency, neutropenia, and aggressive cutaneous T-cell lymphoma/leukemia, continuous infusion ceftazidime (6.5-9.6 g/day) was used to successfully treat multidrug-resistant P. aeruginosa bacteremia. In the second patient, with leukocyte adhesion deficiency type 1, continuous infusion aztreonam (8.4 g/day) was used to successfully treat multidrug-resistant P. aeruginosa wound infections. In the third patient, with severe aplastic anemia, continuous infusion ceftazidime (7-16.8 g/day) was used to treat P. aeruginosa pneumonia and bacteremia. In each patient, bacteremia cleared, infected wounds healed, and pneumonia improved in response to continuous infusion ceftazidime or aztreonam. DISCUSSION: Treatment strategies for multidrug-resistant P. aeruginosa infections are limited. A novel treatment strategy, when no other options are available, is the continuous infusion of existing beta-lactam antibiotics to maximize their pharmacodynamic activity. High-dose continuous infusion ceftazidime or aztreonam was used for the successful treatment of resistant systemic P. aeruginosa infections in 3 chronically immunocompromised patients. CONCLUSIONS: Continuous infusion beta-lactam antibiotics are a potentially useful treatment strategy for resistant P. aeruginosa infections in immunocompromised patients.

The polyphenol piceatannol has shown inhibition against tyrosine and serine/threonine kinases. Whether piceatannol also exerts activity on the mammalian target of rapamycin (mTOR), a kinase involved in growth control of eukaryotic cells, is not known. In this study, we tested the effects of piceatannol on proliferation of androgen-dependent (AD) LNCaP and androgen-independent (AI) DU145 and PC-3 prostate cancer (CaP) cells. Suppression of AD and AI CaP cell growth by piceatannol was accompanied by cell cycle blockade in G(1)/S and S phases for LNCaP and PC-3 and induction of apoptosis in DU145 cells. Induction of apoptosis by piceatannol in DU145 cells was evident by reduced expression of poly(ADP-ribose) polymerase (PARP), cleavage of caspase 3 and apoptosis inducing factor AIF, and an increase in cytochrome c. The apoptotic changes occurred in concordance with DNA damage, supported by increased phosphorylated histone H2AX. Immunoblot analyses showed that exposure of different-stage CaP cells to piceatannol also resulted in cell-type-specific downregulation of mTOR and its upstream and downstream effector proteins, AKT and eIF-4E-BP1. We propose that the observed AKT and mTOR changes are new targets of piceatannol possibly contributing to its inhibitory activities on proliferation of CaP cells.

The potent carcinogen aflatoxin B(1) is a weak mutagen but a strong recombinagen in Saccharomyces cerevisiae. Aflatoxin B(1) exposure greatly increases frequencies of both heteroallelic recombination and chromosomal translocations. We analyzed the gene expression pattern of diploid cells exposed to aflatoxin B(1) using high-density oligonucleotide arrays comprising specific probes for all 6218 open reading frames. Among 183 responsive genes, 46 are involved in either DNA repair or in control of cell growth and division. Inducible growth control genes include those in the TOR signaling pathway and SPO12, whereas PKC1 is downregulated. Eleven of the 15 inducible DNA repair genes, including RAD51, participate in recombination. Survival and translocation frequencies are reduced in the rad51 diploid after aflatoxin B(1) exposure. In mec1 checkpoint mutants, aflatoxin B(1) exposure does not induce RAD51 expression or increase translocation frequencies; however, when RAD51 is constitutively overexpressed in the mec1 mutant, aflatoxin B(1) exposure increased translocation frequencies. Thus the transcriptional profile after aflatoxin B(1) exposure may elucidate the genotoxic properties of aflatoxin B(1).

Longevity in humans is determined by multiple environmental and genetic factors. We have investigated possible associations between longevity and Single Nucleotide Polymorphisms (SNPs) in the p21 (CDKN1A) gene, a stress-inducible senescence-associated cell cycle inhibitor, expression of which upregulates genes implicated in several age-related diseases. By sequencing the promoter and exons of p21 in genomic DNA of ten individuals over 90 years old, we have identified 30 SNPs, many of which had not been previously characterized. A cluster of minor alleles within the -4547/-3489 bp region did not alter the basal activity or p53 responsiveness of the p21 promoter. We then compared the frequency of 41 p21 SNPs between 184 centenarians and 184 younger subjects in the Italian population. Rare alleles of two exon-derived SNPs, rs1801270 and rs1059234, were significantly under-represented among the centenarians; no significant differences were found for 39 non-exonic SNPs. SNP rs1801270 causes Ser to Arg substitution at amino acid 31 and SNP rs1059234 leads to a nucleotide change in the 3'-untranslated region. Previous studies showed that the rare alleles of these two SNPs may play a role in cancer. These p21 alleles may be potentially detrimental to longevity and therefore are rare in centenarians.

Stromal-epithelial interactions may control the growth and initiation of cancers. Here, we not only test the hypothesis that bone marrow-derived cells may effect development of cancers arising from other tissue cells by forming tumor stroma but also that sarcomas may arise by transformation of stem cells from the bone marrow and epithelial cancers may arise by transdifferentiation of bone marrow stem cells to epithelial cancers. Lethally irradiated female FVB/N mice were restored with bone marrow (BM) transplants from a male transgenic mouse carrying the polyoma middle T-oncoprotein under the control of the mouse mammary tumor virus promoter (MMTV-PyMT) and followed for development of lesions. All of 8 lethally irradiated female FVB/N recipient mice, restored with BM transplants from a male MMTV-PyMT transgenic mouse, developed Y-chromosome negative (Y-) cancers of various organs surrounded by Y+ stroma. One of the female FVB/N recipient mice also developed fibrosarcoma and 1, a diploid breast adenocarcinoma containing Y chromosomes. In contrast, only 1 of 12 control female mice restored with normal male BM developed a tumor (lymphoma) during the same time period. These results indicate not only that the transgenic BM-derived stromal cells may indirectly contribute to development of tumors in recipient mice but also that sarcomas may arise by transformation of BM stem cells and that breast cancers arise by transdifferentiation of BM stem cells, presumably by mesenchymal-epithelial transition.

As a general strategy for function-based gene identification, an shRNA library containing approximately 150 shRNAs per gene was enzymatically generated from normalized (reduced-redundance) human cDNA. The library was constructed in an inducible lentiviral vector, enabling propagation of growth-inhibiting shRNAs and controlled activity measurements. RNAi activities were measured for 101 shRNA clones representing 100 human genes and for 201 shRNAs derived from a firefly luciferase gene. Structure-activity analysis of these two datasets yielded a set of structural criteria for shRNA efficacy, increasing the frequencies of active shRNAs up to 5-fold relative to random sampling. The same library was used to select shRNAs that inhibit breast carcinoma cell growth by targeting potential oncogenes. Genes targeted by the selected shRNAs were enriched for 10 pathways, 9 of which have been previously associated with various cancers, cell cycle progression, or apoptosis. One hundred nineteen genes, enriched through this selection and represented by two to six shRNAs each, were identified as potential cancer drug targets. Short interfering RNAs against 19 of 22 tested genes in this group inhibited cell growth, validating the efficiency of this strategy for high-throughput target gene identification.

Abstract An iron aluminide (Fe3Al) intermetallic coating was deposited onto F22 (2.25Cr-1Mo) steel substrate using a JP-5000 high velocity oxy-fuel (HVOF) thermal spray system. The as-sprayed coating was characterized by electron microscopy, X-ray diffraction, oxidation, and adhesion. Fe3Al coated steel specimens were exposed to a mixed oxidizing/sulfidizing environment of N2-10%CO-5%CO2-2%H2O-0.12%H2S (by volume) at 500, 600, 700, and 800°C for approximately seven days. All specimens gained mass after exposure, inversely proportional to temperature increases. Representative cross-sectioned specimens from each temperature underwent scanning electron microscopy (SEM) and X-ray mapping examination. Results are presented in terms of corrosion weight gain and product formation. The research evaluated the effectiveness of an HVOF-sprayed Fe3Al coating in protecting a steel substrate exposed to a fossil energy environment.

Abstract Open air burn pits were used extensively during military operations in Iraq and Afghanistan, potentially exposing millions of US Veterans to toxic airborne hazards. Many of the airborne toxins released have been shown to induce lung inflammation and lung injury and are mutagenic. This is the first large-scale study of associations between self-reported burn pit exposures and the development of cancer. Using data from the Airborne Hazards and Open Burn Pit Registry, we found that Veterans reporting burn pit exposures are associated with a higher odds of developing cancer. However, investigations into the development of specific type of cancer and into a burn pit exposure dose-response effect were inconclusive.

A device and process strategy for achieving reliable indium gallium zinc oxide and indium oxide transistors compatible with a 400oC BEOL thermal budget and without performance degradation is demonstrated by fully exploiting intrinsic oxide material properties. An indium oxide transistor with a novel amorphous In2O3 mixed with SiO2 capping layer exhibits a positive threshold voltage, high extrinsic saturation mobility 33.1 cm2/V.s ,and only a 5mV Vt shift after positive-bias stress at 3 MV/cm for 1000s at room temperature, superior to conventional SiO2 encapsulation.

Objective: Brain-computer interface (BCI)-based color vision (CV) assessment is a new passive, objective, precise, and automatic method for identifying people with congenital redgreen CV deficiencies. At present, BCI-based CV assessment has limited practical utility because it requires lengthy data collection sessions. Here, we demonstrate a 97.5% reduction in the time required to perform BCI-based CV assessment, enabling identification of people with congenital red-green CV deficiencies in less than one minute with >99% accuracy. Methods: Previously collected BCI-based CV assessment data from 31 participants (20 controls, 6 protans, 5 deutans) were analyzed. Each participant completed four experimental runs sampling a 10×10 two-dimensional color grid of red-green intensity combinations around a yellow reference. Each grid location corresponded to a 3.75-s epoch of electroencephalography (EEG) recorded during alternation of the yellow source and the red-green mixture, resulting in 100 epochs per run (1500 s total). Baseline EEG was recorded before and after each run (60 s total), and 120-s breaks were provided between runs. A total 1920 s per participant. Results: Classification accuracies exceeding 99% were achieved using 2 runs of data from 5 grid locations. These locations correspond to regions previously identified in our prior work as showing the strongest inter-group differences. Conclusion: Our feature selection method reduced the time required for passive BCI-based CV assessment by 97.7% (i.e., from 1920 s to 45 s). Significance: The optimized BCI-based assessment accurately identifies people with red-green CV deficiencies in under one minute, greatly enhancing its clinical utility.

We demonstrate brain-computer interface (BCI)- based color vision (CV) assessment for the identification of congenital red-green CV deficiencies. Experiments were based on the identification of metamers—light sources with different spectral distributions perceived to be the same color. Metamers elicit steady-state visual evoked potentials (SSVEPs) of minimal size and are different for people with versus without CV deficiencies. Methods: Thirty-one participants (20 control (CTR), 11 color vision deficient (CVdef)) completed behaviorand BCI-based CV assessments. Experiments used a visual stimulus that alternated between a monochromatic light source (yellow; fixed luminance) and a dichromatic light source (red and green; varying luminances) at a fixed frequency. During behavior-based CV assessment, participants identified metamers by manually adjusting the dichromatic source’s settings until its color matched that of the monochromatic source. During BCI-based CV assessment, participants attended a sequence of stimuli (each with a different dichromatic source setting) while electroencephalography was recorded; metamers were defined as the dichromatic source settings that minimized SSVEP size. Results: The behavior- and BCIidentified metamers were identical within each group, but different across groups (i.e., CTR and CVdef and for people with protan- versus deutan-type CV deficiencies). Automatic identification of CVdef individuals (and type of deficiency) was demonstrated using a classification analysis (93% accuracy). Conclusion: Experiments validated BCI-based CV assessment for the identification of congenital red-green CV deficiencies. Significance: BCI-based CV assessment does not require behavioral responses and can be automated, making it suitable for people with cognitive/motor deficits. With further development, BCI-based CV assessment may enable automatic identification of many types of congenital and acquired CV deficiencies.