Judith Gilbert - Academia.edu (original) (raw)

Papers by Judith Gilbert

Research paper thumbnail of Gemcitabine Cytotoxicity: Interaction of Efflux and Deamination

Journal of drug metabolism & toxicology, Jan 2, 2011

Gemcitabine is a cytidine analogue used in the treatment of various solid tumors. Little is known... more Gemcitabine is a cytidine analogue used in the treatment of various solid tumors. Little is known about how gemcitabine and its metabolites are transported out of cells. We set out to study the efflux of gemcitabine and the possible consequences of that process in cancer cells. We observed the efflux of gemcitabine and its deaminated metabolite, 2',2'-difluorodeoxyuridine (dFdU) using high performance liquid chromatography and tandem mass spectrometry (LC-MS/MS) after gemcitabine treatment. Non-selective ABCC-transport inhibition with probenecid significantly increased intracellular dFdU concentrations, with a similar trend observed with verapamil, a non-selective ABCB1 and ABCG2 transport inhibitor. Neither probenecid nor verapamil altered intracellular gemcitabine levels after the inhibition of deamination with tetrahydrourudine, suggesting that efflux of dFdU, but not gemcitabine, was mediated by ABC transporters. MTS assays showed that probenecid increased sensitivity to...

Research paper thumbnail of Rapid Degradation of Neurotensin by Intact Murine Neuroblastoma Cells (Clone N1E-115)

Journal of Neurochemistry, 1987

Research paper thumbnail of Neurotensin and its analogs—Correlation of specific binding with stimulation of cyclic GMP formation in neuroblastoma clone N1E-115

Biochemical Pharmacology, 1986

The receptors which mediate neurotensin-stimulated intracellular cyclic GMP formation in murine n... more The receptors which mediate neurotensin-stimulated intracellular cyclic GMP formation in murine neuroblastoma clone NlE-115 [J. A. Gilbert and E. Richelson, Eur. J. Pharmnc. 99,245 (1984)] were further characterized. The binding of [3H]neurotensin to intact NlE-115 cells at 0" displayed specificity, saturability, reversibility, and tissue linearity. A single class of neurotensin receptors was demonstrated with an apparent KD of 9-11 nM and a B,, of 180-250 fmoles/106 cells, determined by the type of serum employed in the cellular culture medium. A number of neurotensin analogs and fragments were compared for their ability to inhibit [3H]neurotensin binding and stimulate intracellular cyclic GMP formation with intact NlE-115 cells. A direct correlation was found to exist between the KD and EC5~ for each peptide. The carboxyl-terminal portion of neurotensin proved to be responsible for the binding and biochemical activities of this peptide with clone NlE-115. Neurotensin@-13) was, in fact, fifty times more potent than native neurotensin in stimulating intracellular cyclic GMP formation and had an 18-fold higher affinity for the neurotensin receptor on this neuronal cell type.

Research paper thumbnail of Molecular Markers for Novel Therapeutic Strategies in Pancreatic Endocrine Tumors

Pancreas, 2013

Objectives: Pancreatic endocrine tumors (PETs) share numerous features with gastrointestinal neur... more Objectives: Pancreatic endocrine tumors (PETs) share numerous features with gastrointestinal neuroendocrine (carcinoid) tumors. Targets of novel therapeutic strategies previously assessed in carcinoid tumors were analyzed in PETs (44 cases). Methods: Activating mutations in EGFR, KIT, and PDGFRA and nonresponse mutations in KRAS were evaluated. Copy number of EGFR and HER-2/neu was quantified by fluorescence in situ hybridization. Expression of EGFR, PDGFRA, VEGFR1, TGFBR1, Hsp90, SSTR2A, SSTR5, IGF1R, mTOR, and MGMTwas measured immunohistochemically. Results: Elevated EGFR copy number was found in 38% of cases but no KRAS nonresponse mutations. VEGFR1, TGFBR1, PDGFRA, SSTR5, SSTR2A, and IGF1R exhibited the highest levels of expression in the largest percentages of PETs. Anticancer drugs BMS-754807 (selective for IGF1R/IR), 17-(allylamino)-17-demethoxygeldanamycin (17-AAG, targeting Hsp90), and axitinib (directed toward VEGFR1-3/PDGFRA-B/KIT) induced growth inhibition of human QGP-1 PET cells with IC 50 values (nM) of 273, 723, and 743, respectively. At growth-inhibiting concentrations, BMS-754807 inhibited IGF1R phosphorylation; 17-AAG induced loss of EGFR, IGF1R, and VEGFR2; and axitinib increased p21 Waf1/Cip1 (CDKN1A) expression without inhibiting VEGFR2 phosphorylation. Conclusions: Results encourage further research into multidrug strategies incorporating inhibitors targeting IGF1R or Hsp90 and into studies of axitinib combined with conventional chemotherapeutics toxic to tumor cells in persistent growth arrest.

Research paper thumbnail of Afatinib monotherapy in EGFR-mutant lung adenocarcinoma

The Lancet Oncology, 2013

Research paper thumbnail of The genomic landscape of small intestine neuroendocrine tumors

Journal of Clinical Investigation, 2013

Small intestine neuroendocrine tumors (SI-NETs) are the most common malignancy of the small bowel... more Small intestine neuroendocrine tumors (SI-NETs) are the most common malignancy of the small bowel. Several clinical trials target PI3K/Akt/mTOR signaling; however, it is unknown whether these or other genes are genetically altered in these tumors. To address the underlying genetics, we analyzed 48 SI-NETs by massively parallel exome sequencing. We detected an average of 0.1 somatic single nucleotide variants (SNVs) per 10 6 nucleotides (range, 0-0.59), mostly transitions (C>T and A>G), which suggests that SI-NETs are stable cancers. 197 protein-altering somatic SNVs affected a preponderance of cancer genes, including FGFR2, MEN1, HOOK3, EZH2, MLF1, CARD11, VHL, NONO, and SMAD1. Integrative analysis of SNVs and somatic copy number variations identified recurrently altered mechanisms of carcinogenesis: chromatin remodeling, DNA damage, apoptosis, RAS signaling, and axon guidance. Candidate therapeutically relevant alterations were found in 35 patients, including SRC, SMAD family genes, AURKA, EGFR, HSP90, and PDGFR. Mutually exclusive amplification of AKT1 or AKT2 was the most common event in the 16 patients with alterations of PI3K/Akt/ mTOR signaling. We conclude that sequencing-based analysis may provide provisional grouping of SI-NETs by therapeutic targets or deregulated pathways. Conflict of interest: The authors have declared that no conflict of interest exists.

Research paper thumbnail of Molecular markers for novel therapies in neuroendocrine (carcinoid) tumors

Endocrine-related Cancer, 2010

Neuroendocrine (carcinoid) tumors (NETs) are endocrine neoplasms occurring most frequently in gas... more Neuroendocrine (carcinoid) tumors (NETs) are endocrine neoplasms occurring most frequently in gastrointestinal and bronchopulmonary (BP) systems. The majority of patients present with advanced disease for which few treatment options exist. We assessed 104 NETs (74 cases) for biomarkers targeted by anticancer drugs under development for other forms of cancer. Activating mutations were assessed in epidermal growth factor receptor (EGFR), stem cell factor receptor (KIT), and platelet-derived growth factor receptor alpha (PDGFRA), as well as non-response mutations in KRAS. Copy number of EGFR and HER-2/neu was quantified with fluorescence in situ hybridization. Immunohistochemical analyses were performed for EGFR, KIT, PDGFRA, somatostatin receptor subtypes 2A and 5 (SSTR5), vascular endothelial growth factor receptor 1, mammalian target of rapamycin (mTOR), insulin-like growth factor 1 receptor (IGF1R), heat shock protein 90 (Hsp90), and transforming growth factor-b receptor 1 (TGFBR1). NETs lacked HER2-overexpression predictive of anti-HER2 response and KIT and PDGFRA activating mutations indicative of imatinib sensitivity. High EGFR aneusomy (20% of all cases) and elevated EGFR copy number (39%) were found, but few KRAS mutations associated with non-response to anti-EGFR therapy (3%). Hsp90, TGFBR1, IGF1R, and SSTR5 exhibited highest levels of immunohistochemical staining in the largest percents of tumors. In subsequent in vitro studies, anticancer drug 17-(allylamino)-17-demethoxygeldanamycin (17-AAG) (targeting Hsp90) inhibited proliferation of BP NET lines NCI-H727, NCI-H720, and NCI-H835 with IC 50 values of 70.4, 310, and 788 nM respectively; BMS-754807 (targeting IGF1R/IR) inhibited growth with IC 50 values of 428 nM, 2.8 mM, and 1 mM. At growth-inhibiting concentrations, 17-AAG (24 h) induced loss of EGFR and IGF1R in the IGF1R-expressing NCI-H727 line, and BMS-754807 (24 h) inhibited constitutive IGF1R autophosphorylation. Our results support further research into Hsp90, IGF1R, and EGFR as targets for developing new anticancer therapeutics for some NETs.

Research paper thumbnail of Gemcitabine Cytotoxicity: Interaction of Efflux and Deamination

Journal of drug metabolism & toxicology, Jan 2, 2011

Gemcitabine is a cytidine analogue used in the treatment of various solid tumors. Little is known... more Gemcitabine is a cytidine analogue used in the treatment of various solid tumors. Little is known about how gemcitabine and its metabolites are transported out of cells. We set out to study the efflux of gemcitabine and the possible consequences of that process in cancer cells. We observed the efflux of gemcitabine and its deaminated metabolite, 2',2'-difluorodeoxyuridine (dFdU) using high performance liquid chromatography and tandem mass spectrometry (LC-MS/MS) after gemcitabine treatment. Non-selective ABCC-transport inhibition with probenecid significantly increased intracellular dFdU concentrations, with a similar trend observed with verapamil, a non-selective ABCB1 and ABCG2 transport inhibitor. Neither probenecid nor verapamil altered intracellular gemcitabine levels after the inhibition of deamination with tetrahydrourudine, suggesting that efflux of dFdU, but not gemcitabine, was mediated by ABC transporters. MTS assays showed that probenecid increased sensitivity to...

Research paper thumbnail of Rapid Degradation of Neurotensin by Intact Murine Neuroblastoma Cells (Clone N1E-115)

Journal of Neurochemistry, 1987

Research paper thumbnail of Neurotensin and its analogs—Correlation of specific binding with stimulation of cyclic GMP formation in neuroblastoma clone N1E-115

Biochemical Pharmacology, 1986

The receptors which mediate neurotensin-stimulated intracellular cyclic GMP formation in murine n... more The receptors which mediate neurotensin-stimulated intracellular cyclic GMP formation in murine neuroblastoma clone NlE-115 [J. A. Gilbert and E. Richelson, Eur. J. Pharmnc. 99,245 (1984)] were further characterized. The binding of [3H]neurotensin to intact NlE-115 cells at 0" displayed specificity, saturability, reversibility, and tissue linearity. A single class of neurotensin receptors was demonstrated with an apparent KD of 9-11 nM and a B,, of 180-250 fmoles/106 cells, determined by the type of serum employed in the cellular culture medium. A number of neurotensin analogs and fragments were compared for their ability to inhibit [3H]neurotensin binding and stimulate intracellular cyclic GMP formation with intact NlE-115 cells. A direct correlation was found to exist between the KD and EC5~ for each peptide. The carboxyl-terminal portion of neurotensin proved to be responsible for the binding and biochemical activities of this peptide with clone NlE-115. Neurotensin@-13) was, in fact, fifty times more potent than native neurotensin in stimulating intracellular cyclic GMP formation and had an 18-fold higher affinity for the neurotensin receptor on this neuronal cell type.

Research paper thumbnail of Molecular Markers for Novel Therapeutic Strategies in Pancreatic Endocrine Tumors

Pancreas, 2013

Objectives: Pancreatic endocrine tumors (PETs) share numerous features with gastrointestinal neur... more Objectives: Pancreatic endocrine tumors (PETs) share numerous features with gastrointestinal neuroendocrine (carcinoid) tumors. Targets of novel therapeutic strategies previously assessed in carcinoid tumors were analyzed in PETs (44 cases). Methods: Activating mutations in EGFR, KIT, and PDGFRA and nonresponse mutations in KRAS were evaluated. Copy number of EGFR and HER-2/neu was quantified by fluorescence in situ hybridization. Expression of EGFR, PDGFRA, VEGFR1, TGFBR1, Hsp90, SSTR2A, SSTR5, IGF1R, mTOR, and MGMTwas measured immunohistochemically. Results: Elevated EGFR copy number was found in 38% of cases but no KRAS nonresponse mutations. VEGFR1, TGFBR1, PDGFRA, SSTR5, SSTR2A, and IGF1R exhibited the highest levels of expression in the largest percentages of PETs. Anticancer drugs BMS-754807 (selective for IGF1R/IR), 17-(allylamino)-17-demethoxygeldanamycin (17-AAG, targeting Hsp90), and axitinib (directed toward VEGFR1-3/PDGFRA-B/KIT) induced growth inhibition of human QGP-1 PET cells with IC 50 values (nM) of 273, 723, and 743, respectively. At growth-inhibiting concentrations, BMS-754807 inhibited IGF1R phosphorylation; 17-AAG induced loss of EGFR, IGF1R, and VEGFR2; and axitinib increased p21 Waf1/Cip1 (CDKN1A) expression without inhibiting VEGFR2 phosphorylation. Conclusions: Results encourage further research into multidrug strategies incorporating inhibitors targeting IGF1R or Hsp90 and into studies of axitinib combined with conventional chemotherapeutics toxic to tumor cells in persistent growth arrest.

Research paper thumbnail of Afatinib monotherapy in EGFR-mutant lung adenocarcinoma

The Lancet Oncology, 2013

Research paper thumbnail of The genomic landscape of small intestine neuroendocrine tumors

Journal of Clinical Investigation, 2013

Small intestine neuroendocrine tumors (SI-NETs) are the most common malignancy of the small bowel... more Small intestine neuroendocrine tumors (SI-NETs) are the most common malignancy of the small bowel. Several clinical trials target PI3K/Akt/mTOR signaling; however, it is unknown whether these or other genes are genetically altered in these tumors. To address the underlying genetics, we analyzed 48 SI-NETs by massively parallel exome sequencing. We detected an average of 0.1 somatic single nucleotide variants (SNVs) per 10 6 nucleotides (range, 0-0.59), mostly transitions (C>T and A>G), which suggests that SI-NETs are stable cancers. 197 protein-altering somatic SNVs affected a preponderance of cancer genes, including FGFR2, MEN1, HOOK3, EZH2, MLF1, CARD11, VHL, NONO, and SMAD1. Integrative analysis of SNVs and somatic copy number variations identified recurrently altered mechanisms of carcinogenesis: chromatin remodeling, DNA damage, apoptosis, RAS signaling, and axon guidance. Candidate therapeutically relevant alterations were found in 35 patients, including SRC, SMAD family genes, AURKA, EGFR, HSP90, and PDGFR. Mutually exclusive amplification of AKT1 or AKT2 was the most common event in the 16 patients with alterations of PI3K/Akt/ mTOR signaling. We conclude that sequencing-based analysis may provide provisional grouping of SI-NETs by therapeutic targets or deregulated pathways. Conflict of interest: The authors have declared that no conflict of interest exists.

Research paper thumbnail of Molecular markers for novel therapies in neuroendocrine (carcinoid) tumors

Endocrine-related Cancer, 2010

Neuroendocrine (carcinoid) tumors (NETs) are endocrine neoplasms occurring most frequently in gas... more Neuroendocrine (carcinoid) tumors (NETs) are endocrine neoplasms occurring most frequently in gastrointestinal and bronchopulmonary (BP) systems. The majority of patients present with advanced disease for which few treatment options exist. We assessed 104 NETs (74 cases) for biomarkers targeted by anticancer drugs under development for other forms of cancer. Activating mutations were assessed in epidermal growth factor receptor (EGFR), stem cell factor receptor (KIT), and platelet-derived growth factor receptor alpha (PDGFRA), as well as non-response mutations in KRAS. Copy number of EGFR and HER-2/neu was quantified with fluorescence in situ hybridization. Immunohistochemical analyses were performed for EGFR, KIT, PDGFRA, somatostatin receptor subtypes 2A and 5 (SSTR5), vascular endothelial growth factor receptor 1, mammalian target of rapamycin (mTOR), insulin-like growth factor 1 receptor (IGF1R), heat shock protein 90 (Hsp90), and transforming growth factor-b receptor 1 (TGFBR1). NETs lacked HER2-overexpression predictive of anti-HER2 response and KIT and PDGFRA activating mutations indicative of imatinib sensitivity. High EGFR aneusomy (20% of all cases) and elevated EGFR copy number (39%) were found, but few KRAS mutations associated with non-response to anti-EGFR therapy (3%). Hsp90, TGFBR1, IGF1R, and SSTR5 exhibited highest levels of immunohistochemical staining in the largest percents of tumors. In subsequent in vitro studies, anticancer drug 17-(allylamino)-17-demethoxygeldanamycin (17-AAG) (targeting Hsp90) inhibited proliferation of BP NET lines NCI-H727, NCI-H720, and NCI-H835 with IC 50 values of 70.4, 310, and 788 nM respectively; BMS-754807 (targeting IGF1R/IR) inhibited growth with IC 50 values of 428 nM, 2.8 mM, and 1 mM. At growth-inhibiting concentrations, 17-AAG (24 h) induced loss of EGFR and IGF1R in the IGF1R-expressing NCI-H727 line, and BMS-754807 (24 h) inhibited constitutive IGF1R autophosphorylation. Our results support further research into Hsp90, IGF1R, and EGFR as targets for developing new anticancer therapeutics for some NETs.