Jay George - Academia.edu (original) (raw)

Papers by Jay George

Journal of Biological Chemistry, 1985

Arginyl residues in BamHI endonuclease were examined because of their alleged role in proteins th... more Arginyl residues in BamHI endonuclease were examined because of their alleged role in proteins that contain nucleotide- or phosphate-binding sites. Butanedione, an arginine-specific reagent, inhibited the endonuclease in the presence of sodium borate. The inhibition was decreased by preliminary incubation of the enzyme with DNA or competitive inhibitors which were the 5'-phosphoryl deoxydinucleotide subsets of the BamHI recognition sequence. The dinucleotide pdGpdG protected the enzyme most efficiently against the butanedione modification. Dinucleotides that were unrelated to the recognition sequence failed to protect the enzyme from inactivation. These studies indicate that arginine residues may reside in the enzyme's active site and might function in the sequence-specific recognition of the BamHI palindrome.

Journal of Biological Chemistry, 1980

The specificity of cleavage of Bam HI is altered in the presence of hydrophobic reagents, such as... more The specificity of cleavage of Bam HI is altered in the presence of hydrophobic reagents, such as glycerol and M2SO. The enzyme with altered specificity, designated Bam HI.1, generated digestion patterns of various DNAs, which were distinct from those generated by Bam HI. Cleavage sites recognized in phiX174 RF DNA in the presence of these hydrophobic reagents are not related to the Bam HI palindrome. Bam HI.1 appears to be an endogenous form of Bam HI that can be expressed by altering the hydrophobicity of the reaction.

Photochemistry and Photobiology, 1984

The effect of vacuum UV-laser irradiation on Simian virus 40 (SV40) DNA I was studied in vitro. F... more The effect of vacuum UV-laser irradiation on Simian virus 40 (SV40) DNA I was studied in vitro. Following exposure of SV40 DNA I to an argon fluoride (ArF) laser (A = 193 nm). single-strand breaks occur in either strand of the double-stranded superhelical molecule (DNA I) and convert it to the DNA I1 conformation. Upon increased irradiation times, additional breaks occur in already scissioned strands resulting in the production of smaller fragments as analyzed by alkaline sucrose centrifugation. These results suggest that short periods of irradiation lead to conformational alterations of SV40 DNA I.

In standard culture, the cells or tissue are place inside BME, plated on a dish where the BME gel... more In standard culture, the cells or tissue are place inside BME, plated on a dish where the BME gels, and then media is added. An organoid forms within days. For the air liquid interface culture, cells or tissue fragments are plated on top of gelled collagen I in an insert which is placed into a chamber with media such that the organoids are at the air liquid interface. Trevigen’s Cultrex® Basement Membrane Extract (BME) for in vitro Organoid/Spheroid Studies

3-Nitropyrrole (M) was introduced as a non-discrimi-nating ‘universal ’ base in nucleic acid dupl... more 3-Nitropyrrole (M) was introduced as a non-discrimi-nating ‘universal ’ base in nucleic acid duplexes by virtue of small size and a presumed tendency to stack but not hydrogen bond with canonical bases. However, the absence of thermally-induced hyperchromic changes by single-stranded deoxyoligomers in which M alternates with A or C residues shows that M does not stack strongly with A or C nearest neighbors. Yet, the insertion of a centrally located M opposite any canonical base in a duplex is sometimes even less destabilizing than that of some mismatches, and the variation in duplex stability is small. In triplexes, on the other hand, an M residue centrally located in the third strand reduces triplex stability drastically even when the X·Y target base pair is A·T or G·C in a homopurine· homopyrimidine segment. But, when the target duplex opposition is M-T and the third strand residue is T, the presence of M in the test triplet has little effect on triplex stability. Therefore, a lac...

This platform, which mimics more the natural state of cells existing in vivo and provides biologi... more This platform, which mimics more the natural state of cells existing in vivo and provides biological sur-faces rather than plastics for adher-ence and growth, has become an invaluable tool in the study of nor-mal cells, as well as in the study of cancer genesis and progression. Over the last decade a newtechnology has emergedthat allows cells to grow in three-dimensional structures. This platform, which mimics more the natural state of cells existing in vivo and provides biological surfaces rather than plastics for adherence and growth, has become an invalu-

The development of molecularly targeted anti-cancer drugs is a result of decades of basic researc... more The development of molecularly targeted anti-cancer drugs is a result of decades of basic research [1-5] and the efforts of the human and cancer genome projects. Such therapeutics offer improved efficacy and fewer cytotoxic side effects compared to previous classes of therapies. The regulatory approval of molecularly targeted drugs such as imatinib (Gleevec), trastuzumab (Herceptin), gefitinib (Iressa), erlotinib (Tarceva), bevacizumab (Avastin), cetuximab (Erbitux), sorafenib (Nexavar), and sunitinib (Sutent ) has provided clinical validation for this approach. More targeted compounds such as the PARP inhibitors, based upon their synthetic lethal relationship with BRCA1/2, are currently in clinical trials demonstrating efficacy with little or no toxic side effects at therapeutic dosages [6]. The management of cancer patients is transforming from empirically based approaches to individualized treatments based on tumor specific molecular targets [4, 7, 8].

Cancer Research, 2010

We have developed a panel of human cell lines, each deficient in one of the eleven DNA glycosylas... more We have developed a panel of human cell lines, each deficient in one of the eleven DNA glycosylase enzymes. To expand background diversity, the same shRNA lentiviruses were also used to develop parallel cell line panels in different tumor backgrounds, including glioma and breast cancer cell lines. Depletion of target mRNA ranged from 67-99% with similar target mRNA depletion across tumor cell backgrounds. Gene expression knockdown was sufficient to functionally diminish enzyme activity, protein levels and mRNA profiling for each exemplified the impact of DNA repair defects on the human transcriptome. As an example of the far reaching potential for a panel of DNA repair deficient cell lines, we show that DNA glycosylase deficiency modulated both the transcriptome and epigenome, implicating some DNA glycosylases in methylation maintenance and genome expression diversity. In addition we demonstrate the impact on DNA repair in glycosylase depleted backgrounds. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3939.

Currently, the high cost and limited capacity for bringing new drugs to market is attributed to i... more Currently, the high cost and limited capacity for bringing new drugs to market is attributed to ineffective methods utilized early in the drug development process. The most commonly used practice involves culturing a single cell type in an artificial environment and evaluating the impact of drug treatment on cell viability. This approach vastly oversimplifies the complexity of the tumor tissue. Within these monocultures, the cells adhere to hard plastic through non-specific charged interactions, and as a result, the cells grow in monolayers and exhibit a cobblestone-like morphology. Both the conditions and the resulting cultures are unlike a tumor, so why would they function or respond like one? To craft a predictive model, it must possess all of the key attributes of the system that it represents. At the time of diagnosis, tumors have sufficient size where multiple cell layers restrict diffusion within the tumor leading to the formation of physiological gradients for nutrients, was...

Molecular and Cellular Biology, Genetics

Radiotherapy and Oncology

Cancer Research, Dec 1, 1995

Few molecular genetic alterations have been identified in endometrial cancers that are associated... more Few molecular genetic alterations have been identified in endometrial cancers that are associated with poor clinical outcome. Overexpression of HER-2/neu, transforming growth factor a, and p53 proteins have all been associated with poor prognosis in women with endometrial cancer. In this study, the level of HER-2lneu gene amplification and expression was characterized in 92 endometrial cancers. Fluorescence in situ hybridiza tion (FISH) was used to characterize HER-2/neu gene copy number, and immunohistochemistry was used to characterize expression. Forty-seven of the 90 (52%) endometrial cancers were characterized as showing moderate or high immunostaining. HER-2lneu gene amplification was detected in 17 ofSl (21%) cases. Immunohistochemical staining and FISH results were both available for 80 cases. Fourteen of these cases showed both moderate or high immunostaining and gene amplification. Clinical follow-up information was available for 76 women in this study. Women whose endometrial cancer exhibited HER-2/neu gene amplification by FISH had a shorter overall survival than women whose endometrial cancer lacked amplification (P = 0.018). Likewise, tumors with moderate or high HER-2lneu immunostaining were associated with a lower cumu lative overall survival than tumors with low immunostaining by log rank analysis (P < 0.0001). Multivariate analysis of survival rates revealed RER-2lneu overexpression to be an independent predictor of overall survival (P = 0.0163). Among those patients with HER-Zfneuoverexpres sion, adjuvant chemotherapy or radiation therapy was associated with an improved overall survival (P 0.039). However, among those women whose tumor lacked overexpression, overall survival was not improved by adjuvant treatment.

Cancer Research, 2013

Current in vitro models for evaluating cancer cell dissemination discount the importance of initi... more Current in vitro models for evaluating cancer cell dissemination discount the importance of initial tumor physiology as an effector in cancer cell invasion. Solid tumors exist in vivo as cell aggregates and are most accurately modeled by multicellular tumor spheroids (MCTS)[1]. As the tumor grows, proliferating cells create additional layers that isolate the cells contained within, limiting access for nutrients, growth factors, and oxygen for these cells; at the same time, products of catabolism become concentrated within the center and slowly diffuse out. The diffusion of nutrients inward and waste products outward creates physiological gradients containing heterogeneous cell populations (Figure 1) with differing gene expression and cell cycle regulation[2, 3]. As a result of the low nutrient levels and build up of waste products, the environment within the core becomes cytotoxic, and the cells in this region become quiescent and eventually die via apoptosis or necrosis. Alternatively, the cells in the surface layers, having access to all of the essentials elements and no waste build up, continue to proliferate. Between these two layers are other populations of cells that become quiescent, having resources that are adequate for viability but insufficient for replication. As a result of these chemical gradients and the concentric arrangement of heterogeneous cell populations, certain cell populations may react differently to pharmacological agents; furthermore, drug penetration may also present an obstacle due to the multitude of cell layers[4]. In vitro cell models that do not exhibit these properties will not predict a physiological mechanism or response. As a consequence, there is a growing demand for in vitro assay formats that incorporate the complexity of the tumor tissue architecture to provide more physiologically predictive models. To build a more physiological cancer assay platform, well-established methods for multicellular tumor spheroid formation were incorporated into 3-D culture models. Spheroid cultures have been utilized for cancer cell culture for over 40 years[5, 6]; however, they were eventually incorporated into a limited number of cancer models due to the inability of many of cancer cell lines to spontaneously assemble into spheroids. To promote cell-cell interactions, spheroid formation is usually conducted in an environment that increases the proximity of cells to one another and prevents cell-surface interactions. Common methods for promoting spheroid formation include hanging drops[7] or round bottom wells treated with cell adhesion resistant surfaces such as agarose[8] or poly-HEMA[9]. In addition, maintaining cells in suspension through continuous aggregation is also effective, although there appears to be less uniformity[10]. More recently, it has been shown that the addition of ECM proteins promotes spheroid formation through the assembly of cell-cell bonds, inducing cell aggregation and spheroid formation for many of these cancer cell models that were previously deemed incompatible[11], such as MDA-MB-231 (Figure 2) thus emphasizing the importance of the extracellular environment in tumor growth. The composition of the ECM is an also important determinant for cancer dissemination. Using a traditional, hydrogelbased breast cancer model, we have previously demonstrated that the composition of the ECM is an important determinant for the expression of cadherin proteins, the formation of cell-cell adhesion, and subsequent morphology[12]. Cells that were cultured on basement membrane proteins exhibited a tumor phenotype, increasing collagen I concentration decreased cell-cell adhesion and induced a migratory phenotype. Previously, MCTS have been embedded in ECM and utilized to evaluated cell invasion[11, 13, 14], and this model was used to further assess cell morphology, spheroid architecture, and relative invasion as a function of ECM composition for breast, prostate, fibrosarcoma, and glioblastoma cell lines.