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Biomedical subjects

Michael D Johnson

Publications and source records attributed to Michael D Johnson.

At least 19 recordsLinked to original sources
PubMed2007

Small-molecule agonists for the glucagon-like peptide 1 receptor.

The peptide hormone glucagon-like peptide (GLP)-1 has important actions resulting in glucose lowering along with weight loss in patients with type 2 diabetes. As a peptide hormone, GLP-1 has to be administered by injection. Only a few small-molecule agonists to peptide hormone receptors have been described and none in the B family of the G protein coupled receptors to which the GLP-1 receptor belongs. We have discovered a series of small molecules known as ago-allosteric modulators selective for the human GLP-1 receptor. These compounds act as both allosteric activators of the receptor and independent agonists. Potency of GLP-1 was not changed by the allosteric agonists, but affinity of GLP-1 for the receptor was increased. The most potent compound identified stimulates glucose-dependent insulin release from normal mouse islets but, importantly, not from GLP-1 receptor knockout mice. Also, the compound stimulates insulin release from perfused rat pancreas in a manner additive with GLP-1 itself. These compounds may lead to the identification or design of orally active GLP-1 agonists.

Animals↗
PubMed2007

Design and synthesis of redox stable analogues of sunflower trypsin inhibitors (SFTI-1) on solid support, potent inhibitors of matriptase.

[structure: see text] Matriptase is a member of the emerging class of type II transmembrane serine proteases. It was found that the sunflower trypsin inhibitor (SFTI-1), isolated from sunflower seeds, inhibits matriptase with a subnanomolar Ki of 0.92 nM. On the basis of this result, we designed and synthesized its proteolytically stable analogues, SFTI-2 and SFTI-3. SFTI-3 exhibited very good binding affinity to matriptase, and it was metabolically stable.

Cyclization↗
PubMed2006

MDA-MB-435 cells are derived from M14 melanoma cells--a loss for breast cancer, but a boon for melanoma research.

BACKGROUND: The tissue of origin of the cell line MDA-MB-435 has been a matter of debate since analysis of DNA microarray data led Ross et al. (2000, Nat Genet 24(3):227-235) to suggest they might be of melanocyte origin due to their similarity to melanoma cell lines. We have previously shown that MDA-MB-435 cells maintained in multiple laboratories are of common origin to those used by Ross et al. and concluded that MDA-MB-435 cells are not a representative model for breast cancer. We could not determine, however, whether the melanoma-like properties of the MDA-MB-435 cell line are the result of misclassification or due to transdifferention to a melanoma-like phenotype. METHODS: We used karyotype, comparative genomic hybridization (CGH), and microsatalite polymorphism analyses, combined with bioinformatics analysis of gene expression and single nucleotide polymorphism (SNP) data, to test the hypothesis that the MDA-MB-435 cell line is derived from the melanoma cell line M14. RESULTS: We show that the MDA-MB-435 and M14 cell lines are essentially identical with respect to cytogenetic characteristics as well as gene expression patterns and that the minor differences found can be explained by phenotypic and genotypic clonal drift. CONCLUSIONS: All currently available stocks of MDA-MB-435 cells are derived from the M14 melanoma cell line and can no longer be considered a model of breast cancer. These cells are still a valuable system for the study of cancer metastasis and the extensive literature using these cells since 1982 represent a valuable new resource for the melanoma research community.

Breast Neoplasms↗
PubMed2006

GREB1 is a novel androgen-regulated gene required for prostate cancer growth.

BACKGROUND: Gene regulated in breast cancer 1 (GREB1) is a novel estrogen-regulated gene shown to play a pivotal role in hormone-stimulated breast cancer growth. GREB1 is expressed in the prostate and its putative promoter contains potential androgen receptor (AR) response elements. METHODS: We investigated the effects of androgens on GREB1 expression and its role in androgen-dependent prostate cancer growth. RESULTS: Real-time PCR demonstrated high level GREB1 expression in benign prostatic hypertrophy (BPH), localized prostate cancer (L-PCa), and hormone refractory prostate cancer (HR-PCa). Androgen treatment of AR-positive prostate cancer cells induced dose-dependent GREB1 expression, which was blocked by anti-androgens. AR binding to the GREB1 promoter was confirmed by chromatin immunoprecipitation (ChIP) assays. Suppression of GREB1 by RNA interference blocked androgen-stimulated LNCaP cell proliferation. CONCLUSIONS: GREB1 is expressed in proliferating prostatic tissue and prostate cancer, is regulated by androgens, and suppression of GREB1 blocks androgen-induced growth suggesting GREB1 may be critically involved in prostate cancer proliferation.

Androgens↗
PubMed2006

In vivo magnetic resonance volumetric and spectroscopic analysis of mouse prostate cancer models.

BACKGROUND: Mouse prostate cancer modeling presents unique obstacles to the study of spontaneous tumor initiation and progression due to the anatomical location of the tissue. RESULTS: High resolution (130 microm(x) x 130 microm(y) x 300 microm(z)), three-dimensional MRI allowed for the visualization, segmentation, and volumetric measurement of the prostate from normal and genetically engineered animals, in vivo. Additionally, MRS performed on the prostate epithelia of probasin-ErbB-2Delta x Pten(+/-) mice identified changes in the relative concentrations of the metabolites choline and citrate, which was not observed in TRAMP mice. METHODS: T1-weighted MRI was performed on normal, TRAMP, probasin-ErbB-2/Her2/Neu (probasin-ErbB-2Delta), and probasin-ErbB-2Delta in the context of decreased Pten activity (probasin-ErbB-2Delta x Pten(+/-)) mice. Volume-localized single-voxel proton magnetic resonance spectroscopy (SVS (1)H MRS) was also performed. CONCLUSIONS: The data presented supports the use of combined MRI and MRS for the measurement of biochemical and morphometric alterations in mouse models of prostate cancer.

Animals↗
PubMed2006

Genes regulated by estrogen in breast tumor cells in vitro are similarly regulated in vivo in tumor xenografts and human breast tumors.

BACKGROUND: Estrogen plays a central role in breast cancer pathogenesis. Although many studies have characterized the estrogen regulation of genes using in vitro cell culture models by global mRNA expression profiling, it is not clear whether these genes are similarly regulated in vivo or how they might be coordinately expressed in primary human tumors. RESULTS: We generated DNA microarray-based gene expression profiles from three estrogen receptor alpha (ERalpha)-positive breast cancer cell lines stimulated by 17beta-estradiol (E2) in vitro over a time course, as well as from MCF-7 cells grown as xenografts in ovariectomized athymic nude mice with E2 supplementation and after its withdrawal. When the patterns of genes regulated by E2 in vitro were compared to those obtained from xenografts, we found a remarkable overlap (over 40%) of genes regulated by E2 in both contexts. These patterns were compared to those obtained from published clinical data sets. We show that, as a group, E2-regulated genes from our preclinical models were co-expressed with ERalpha in a panel of ERalpha+ breast tumor mRNA profiles, when corrections were made for patient age, as well as with progesterone receptor. Furthermore, the E2-regulated genes were significantly enriched for transcriptional targets of the myc oncogene and were found to be coordinately expressed with Myc in human tumors. CONCLUSION: Our results provide significant validation of a widely used in vitro model of estrogen signaling as being pathologically relevant to breast cancers in vivo.

Age Factors↗
PubMed2006

Reduction of vanadium(V) by L-ascorbic acid at low and neutral pH: kinetic, mechanistic, and spectroscopic characterization.

L-Ascorbic acid interacts with vanadium(V) over the pH range of 0.4-7.0 to form three different coordination complexes. Both inner- and outer-sphere electron-transfer pathways are proposed to form vanadium(IV) complexes with L-ascorbate or dehydroascorbate, respectively. Effects of the pH on the coordination of L-ascorbic acid to the vanadium(V) center were observed and are presumably related to the speciation of the vanadium(V) ion. Three vanadium(IV) complexes were observed using ambient-temperature electron paramagnetic resonance spectroscopy. Two of these complexes are proposed to be vanadium(IV) L-ascorbate complexes, and one is consistent with a vanadium(IV) dehydroascorbic acid complex proposed earlier. These reduction reactions will occur under physiological conditions and could be important to the reduction of vanadium(V)-containing coordination complexes used as insulin-enhancing agents for treatment of diabetes.

Ascorbic Acid↗
PubMed2006

Matriptase activation and shedding with HAI-1 is induced by steroid sex hormones in human prostate cancer cells, but not in breast cancer cells.

Matriptase and its cognate inhibitor, hepatocyte growth factor activator inhibitor-1 (HAI-1), have been implicated in carcinoma onset and malignant progression. However, the pathological mechanisms of matriptase activation are not defined. Steroid sex hormones play crucial roles in prostate and breast cancer. Therefore, we investigated the questions of whether and how steroid sex hormones regulate matriptase activation in these cancer cells. Treatment of cells with 17beta-estradiol had no effect on activation of matriptase in hormone-starved breast cancer cells, in part due to their high constitutive level of activated matriptase. In striking contrast, very low levels of activated matriptase were detected in hormone-starved lymph node prostatic adenocarcinoma (LNCaP) cells. Robust activation of matriptase was observed as early as 6 h after exposure of these cells to 5alpha-dihydrotestosterone (DHT). Activation of matriptase was closely followed by shedding of the activated matriptase with >90% of total activated matriptase present in the culture media 24 h after DHT treatment. Activated matriptase was shed in a complex with HAI-1 and may result from simultaneously proteolytic cleavages of both membrane-bound proteins. Latent matriptase and free HAI-1 were also shed into culture media. As a result of shedding, the cellular levels of matriptase and HAI-1 were significantly reduced 24 h after exposure to DHT. DHT-induced matriptase activation and shedding were significantly inhibited by the androgen antagonist bicalutamide, by the RNA transcription inhibitor actinomycin D, and by the protein synthesis inhibitor cycloheximide. These results suggest that in LNCaP cells, androgen induces matriptase activation via the androgen receptor, and requires transcription and protein synthesis.

Breast Neoplasms↗
PubMed2006

Contrast-enhanced in vivo imaging of breast and prostate cancer cells by MRI.

The development of effective cancer therapies has been hampered, in part, by the inability to noninvasively follow tumor progression from the initial cancerous lesion through to metastasis. We have previously shown that superparamagnetic iron oxide particles can be used as magnetic resonance imaging contrast agents to label embryonic, mesenchymal and hematopoietic stem cells in vivo. Improving the capacity to non-invasively image cancer progression is an appealing method that could be useful for assessing the efficacy of anticancer therapies. We have established that human prostate (LNCaP, DU145, PC3), rodent prostate (TRAMPC1, YPEN-1), human breast (MDA-MB-231) and mouse mammary (Myc/VEGF) cancer cell lines were readily labeled by fluorescent superparamagnetic sub-micron particles of iron oxide (MPIOs). The MPIOs were essentially inert with respect to cell proliferation and tumor formation. Fluorescence stereomicroscopy and three dimensional magnetic resonance imaging (MRI) determined that subcutaneous, intramuscular or orthotopically implanted labeled cancer cells could be imaged, in vivo, despite in some cases being undetectable by manual palpation. The MPIO-labeled cancer cells could also be imaged, in vivo, at least 6 weeks after implantation. The fluorescent MPIOs further allowed for the ex vivo identification of tumors cells from histological sections. This study demonstrates the feasibility of using fluorescent MPIOs in prostate and breast cancer cell lines as both a negative contrast agent for in vivo MRI as well as a fluorescent tumor marker for optical imaging in vivo and ex vivo.

Animals↗
PubMed2006

A collection of breast cancer cell lines for the study of functionally distinct cancer subtypes.

Recent studies suggest that thousands of genes may contribute to breast cancer pathophysiologies when deregulated by genomic or epigenomic events. Here, we describe a model "system" to appraise the functional contributions of these genes to breast cancer subsets. In general, the recurrent genomic and transcriptional characteristics of 51 breast cancer cell lines mirror those of 145 primary breast tumors, although some significant differences are documented. The cell lines that comprise the system also exhibit the substantial genomic, transcriptional, and biological heterogeneity found in primary tumors. We show, using Trastuzumab (Herceptin) monotherapy as an example, that the system can be used to identify molecular features that predict or indicate response to targeted therapies or other physiological perturbations.

Breast Neoplasms↗
PubMed2006

Multiple professional identities: examining differences in identification across work-related targets.

Although there is a growing literature on organizational identification, relatively little research has investigated other possible targets of identification. In a sample of veterinarians working in a wide range of organizations, the authors compared their identification with the veterinary profession, their organization, and their workgroup. The authors found different patterns of identification across these targets depending on whether the individual (a) worked in a veterinary medicine or nonveterinary medicine organization and (b) was an owner/partner or an associate. Owners of veterinary medicine organizations identified more with the organization than with either the profession or their workgroup. Associates in veterinary medicine organizations identified more with the organization and the workgroup than with the profession. Veterinarians in nonveterinary medicine organizations identified more with the profession and their workgroup than with the organization. Identification with each of the targets provided independent predictive validity of job satisfaction.

Cooperative Behavior↗
PubMed2005

What does an orphan G-protein-coupled receptor have to do with estrogen?

Estrogen affects multiple aspects of human physiology, including the normal growth and development of female reproductive tissues, bone integrity, cardiovascular and central nervous system functions, and plays a central role in normal mammary development and breast pathogenesis. It modulates diverse cell signaling pathways, some of which appear to be independent of the known estrogen receptors (ERs). Although many of estrogen's actions can be explained by the nuclear ERs (ER-alpha and ER-beta) functioning as ligand-activated RNA transcription factors, there are numerous rapid biochemical and physiological responses that cannot be explained by the classical genomic effects of estrogen signaling. It has long been postulated that the rapid effects of estrogen are due to a membrane-bound ER, and two recent reports suggest that it is in fact a G-protein-coupled receptor named 'GPR30'.

Breast Neoplasms↗
PubMed2005

HAI-1 regulates activation and expression of matriptase, a membrane-bound serine protease.

Hepatocyte growth factor activator inhibitor-1 (HAI-1) was initially identified as cognate inhibitor of matriptase, a membrane-bound serine protease. Paradoxically, HAI-1 is also required for matriptase activation, a process that requires sphingosine 1-phosphate (S1P)-mediated translocation of the protease to cell-cell junctions in human mammary epithelial cells. In the present study, we further explored how HAI-1 regulates this protease. First, we observed that after S1P treatment HAI-1 was cotranslocated with matriptase to cell-cell junctions and that the cellular ratio of HAI-1 to matriptase was maintained during this process. However, when this ratio was changed by cell treatment with HAI-1 small interfering RNA or anti-HAI-1 MAb M19, spontaneous activation of matriptase occurred in the absence of S1P-induced translocation; S1P-induced matriptase activation was also enhanced. These results support a role for HAI-1 in protection of cell from uncontrolled matriptase activation. We next expressed matriptase, either alone or with HAI-1 in breast cancer cells that do not endogenously express either protein. A defect in matriptase trafficking to the cell surface occurred if wild-type matriptase was expressed in the absence of HAI-1; this defect appeared to result from matriptase toxicity to cells. Coexpression with matriptase of wild-type HAI-1, but not HAI-1 mutants altered in its Kunitz domain 1, corrected the trafficking defect. In contrast, catalytically defective matriptase mutants were normal in their trafficking in the absence of HAI-1. These results are also consistent with a role for HAI-1 to prevent inappropriate matriptase proteolytic activity during its protein synthesis and trafficking. Taken together, these results support multiple roles for HAI-1 to regulate matriptase, including its proper expression, intracellular trafficking, activation, and inhibition.

Blotting, Western↗
PubMed2005

GREB 1 is a critical regulator of hormone dependent breast cancer growth.

BACKGROUND: Estrogen plays a central role in breast cancer pathogenesis and many potent risk factors for the development of the disease can be explained in terms of increased lifetime exposure to estrogen. Although estrogen regulated genes have been identified, those critically involved in growth regulation remain elusive.METHODS. To identify candidate genes involved in estrogen stimulated breast cancer growth, DNA microarray based gene expression profiles were generated from three estrogen receptor alpha (ER alpha) positive breast cancer cell lines grown under multiple stimulatory and inhibitory conditions. RESULTS: Only three genes were significantly induced by 17beta-estradiol (E2) relative to control in all three cell lines: GREB 1, stromal cell-derived factor 1 (SDF-1) and trefoil factor 1 (pS2). Quantitative real-time PCR assays confirmed that in all three cell lines, GREB 1 was induced by E2, but not by the antiestrogens tamoxifen (TAM) or ICI 182,780. GREB 1 expression level was strongly correlated with ER alpha positivity in 39 breast cancer cell lines of known ER alpha expression status. GREB 1 induction by E2 was rapid (7.3 fold by 2 h for MCF-7) and mirrored the fraction of cells entering S-phase when released from an estrogen deprivation induced cell arrest. Suppression of GREB 1 using siRNA blocked estrogen induced growth in MCF-7 cells and caused a paradoxical E2 induced growth inhibition. CONCLUSION: These data suggest that GREB 1 is critically involved in the estrogen induced growth of breast cancer cells and has the potential of being a clinical marker for response to endocrine therapy as well as a potential therapeutic target.

Animals↗
PubMed2005

Reduction of lunar landing fuel requirements by utilizing lunar ballistic capture.

Ballistic lunar capture trajectories have been successfully utilized for lunar orbital missions since 1991. Recent interest in lunar landing trajectories has occurred due to a directive from President Bush to return humans to the Moon by 2015. NASA requirements for humans to return to the lunar surface include separation of crew and cargo missions, all lunar surface access, and anytime-abort to return to Earth. Such requirements are very demanding from a propellant standpoint. The subject of this paper is the application of lunar ballistic capture for the reduction of lunar landing propellant requirements. Preliminary studies of the application of weak stability boundary (WSB) trajectories and ballistic capture have shown that considerable savings in low Earth orbit (LEO) mission mass may be realized, on the order of 36% less than conventional Hohmann transfer orbit missions. Other advantages, such as reduction in launch window constraints and reduction of lunar orbit maintenance propellant requirements, have also surfaced from this study.

Journal Article↗
PubMed2004

Mechanistic variations in the oxidation of Piloty's acid by metal complexes.

The reactions of N-hydroxybenzenesulfonamide (Piloty's acid, PA) with a variety of metal oxidants are reported. Either nitric oxide or nitrite is the final reaction product, along with benzenesulfinate and the reduced metal compound. The nitrogen product depends on the oxidation potential of the metal oxidant and its ability to further oxidize NO to nitrite. The observation and preliminary interpretation of unusual kinetic behavior of Piloty's acid as a reductant is also described. Analogues of PA were also prepared and found to show similar reactivity.

Journal Article↗
PubMed2004

A phase I study of oral BMS-275291, a novel nonhydroxamate sheddase-sparing matrix metalloproteinase inhibitor, in patients with advanced or metastatic cancer.

PURPOSE: BMS-275291 is a novel broad-spectrum inhibitor of matrix metalloproteinase (MMPs) rationally designed to spare a class of closely related metalloproteinases known as sheddases. Inadvertent sheddase inhibition is hypothesized to play a role in the dose-limiting joint toxicities occurring with hydroxamate-based MMP inhibitors. This trial was conducted to establish the recommended phase II dose; determine safety, toxicity, and pharmacokinetics of BMS-275291; and to assess potential markers of sheddase activity [tumor necrosis factor-alpha (TNFalpha) release and TNFalpha-RII shedding]. EXPERIMENTAL DESIGN: This was an open label, single arm, phase I study conducted at two centers. Patients with advanced or metastatic cancer were treated with once-daily oral BMS-275291 at doses escalating from 600 to 2400 mg/day. Six to eight patients/dose level were to be studied with the recommended phase II dose level expanded to a total of 15 patients. Pharmacokinetic sampling was performed on days 1, 15, and 29 at 0, 0.5, 1, 2, 4, 6, 8, and 24 h after dosing. Radiological tumor assessment was performed every 8 weeks. RESULTS: Forty-four evaluable patients were enrolled in this study with the most frequent tumor types being colorectal cancer and non-small cell lung cancer. Dose limiting toxicities were observed at 600 mg/day (one of eight patients with grade 3 transaminitis) and at 1200 mg/day (1 of 15 patients with grade 3 rash and grade 4 shortness of breath), both in the context of predisposing conditions. No dose-limiting toxicities occurred at 900, 1800, or 2400 mg/day. The most frequent adverse events considered possibly, probably, or definitely drug-related were joint toxicity (myalgia/arthralgia), rash, fatigue, headache, nausea, and taste change, all of which were mild, grade 1, grade 2, and not dose-limiting. No objective tumor responses were observed. Twelve of forty-four patients received treatment for 4+ months, six for 8+ months, three for >1 year. Desired trough levels of parent BMS-275291 were maintained with once daily dosing. The mean plasma concentration of parent BMS-275291 at trough exceeded the calculated in vitro IC(80) value for MMP-2 and IC(90) value for MMP-9 at the recommended phase II dose of 1200 mg/day. No major changes in serum concentrations of sheddase enzymatic products, TNFalpha or TNFalpha-RII, were observed. CONCLUSIONS: BMS-275291 is a nonhydroxamate MMP inhibitor with a novel mercaptoacyl zinc-binding group. In this study, plasma concentrations of BMS-275291 continuously exceeded in vitro MMP IC(50) values without dose-limiting joint toxicity. In this refractory patient population, a suggestion of disease stabilization was observed in 12 patients. On the basis of preclinical, clinical, and pharmacokinetic data, the recommended phase II dose for future study is 1200 mg/day.

Adult↗