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Biomedical subjects

C Talmadge

Publications and source records attributed to C Talmadge.

18 recordsLinked to original sources
PubMed2001

Ex vivo purging by adenoviral p53 gene therapy does not affect NOD-SCID repopulating activity of human CD34+ cells.

Co-incubation of a replication-deficient, recombinant adenovirus carrying the wild-type p53 gene (rAd-p53) and hematopoietic stem cell (HSC) products from patients with breast cancer can significantly reduce tumor cell contamination. Whereas this approach provides a powerful tumor cell purging strategy, potential detrimental effects on the HSC population have not been investigated. The ability of human HSC to reconstitute hematopoiesis in severe combined immunodeficient (SCID) mice and to undergo secondary transplantation provides the only nonclinical measure of self-renewing, stem cell function. The objective of this study was to investigate whether co-incubation with rAd-p53 compromised the SCID repopulating activity (SRA) of HSC. Granulocyte colony-stimulating factor-mobilized human CD34+ cells were co-cultured with rAd-p53 at our targeted clinical dose, and the ability of these cells to establish multilineage hematopoiesis in sublethally irradiated, nonobese diabetic (NOD)-SCID mice was investigated. The persistence of human cells in the mice was investigated by flow cytometry, granulocyte-macrophage colony-forming unit assay, and polymerase chain reaction of human Alu sequences. Further, limiting dilution analysis provided a quantitative comparison between the SRA of CD34+ cells co-incubated with rAd-p53 and control CD34+ cells (no rAd-p53 co-incubation). We conclude that co-incubation with rAd-p53 has little effect on the SRA of HSC.

Adenoviridae↗
PubMed2000

The gene for a novel transmembrane protein containing epidermal growth factor and follistatin domains is frequently hypermethylated in human tumor cells.

A DNA fragment frequently hypermethylated in tumor cells was isolated using a novel screening strategy termed methylation-sensitive arbitrarily primed PCR. The isolated sequence corresponded to a CpG island at the 5' end of a previously unknown gene, TPEF (transmembrane protein containing epidermal growth factor and follistatin domains). Expression of TPEF was observed using Northern master blot analysis of a variety of normal tissues including colon, bladder, and prostate tissue. TPEF maps to human chromosome 2q33, where frequent loss of heterozygosity is seen in various human tumors, and TPEF was not expressed in most human colon and various other tumor cell lines examined by reverse transcription-PCR. Nine of 11 tumor cell lines were highly methylated in the 5' region and the first exon of the gene that demonstrated features characteristic of a CpG island. However the other two cell lines, which expressed TPEF, were hypomethylated in the 5' end of the gene. The region was also hypermethylated in 11 of 16 primary bladder tumors and in 3 of 4 primary colon tumors when compared with adjacent normal tissue. Our results suggest that potential tumor suppressor genes can be isolated from human tumors by virtue of their altered methylation patterns.

Amino Acid Sequence↗
PubMed1999

The human DNA methyltransferases (DNMTs) 1, 3a and 3b: coordinate mRNA expression in normal tissues and overexpression in tumors.

DNA methylation in mammals is required for embryonic development, X chromosome inactivation and imprinting. Previous studies have shown that methylation patterns become abnormal in malignant cells and may contribute to tumorigenesis by improper de novo methylation and silencing of the promoters for growth-regulatory genes. RNA and protein levels of the DNA methyltransferase DNMT1 have been shown to be elevated in tumors, however murine stem cells lacking Dnmt1 are still able to de novo methylate viral DNA. The recent cloning of a new family of DNA methyltransferases (Dnmt3a and Dnmt3b) in mouse which methylate hemimethylated and unmethylated templates with equal efficiencies make them candidates for the long sought de novo methyltransferases. We have investigated the expression of human DNMT1, 3a and 3b and found widespread, coordinate expression of all three transcripts in most normal tissues. Chromosomal mapping placed DNMT3a on chromosome 2p23 and DNMT3b on chromosome 20q11.2. Significant overexpression of DNMT3b was seen in tumors while DNMT1 and DNMT3a were only modestly over-expressed and with lower frequency. Lastly, several novel alternatively spliced forms of DNMT3b, which may have altered enzymatic activity, were found to be expressed in a tissue-specific manner.

Alternative Splicing↗
PubMed1994

Somatic cell hybrid panel and NotI linking clones for physical mapping of human chromosome 3.

To identify by positional cloning the putative tumor-suppressor genes on the short arm of human chromosome 3 that are involved in the initiation or progression of several human malignancies, we have developed human mouse somatic cell hybrids and NotI linking libraries. The somatic cell hybrids contain either the intact human chromosome 3 or its derivatives as the only human genetic material in rodent background. The somatic cell hybrid panel defines five chromosomal regions on the short arm and two chromosomal regions on the long arm of chromosome 3. Two hundred sixty-one NotI linking probes from three independently constructed linking libraries were assigned to the seven chromosomal regions. The somatic cell hybrid panel and the regionally localized NotI linking probes should facilitate the construction of genetic linkage and physical maps to identify various tumor-suppressor and disease-related genes not only on the chromosome 3p, but on the entire chromosome.

Animals↗
PubMed1987

Tumor cells transfected with the neomycin resistance gene (neo) contain unique genetic markers useful for identification of tumor recurrence and metastasis.

Studies in tumor biology often require techniques to accurately identify tumors or tumor cell lines. Transfection of plasmid DNA into mammalian cells has been shown to introduce unique genetic markers into recipient cells. These unique genetic markers are detected by Southern transfer. To determine the usefulness of such markers for in vivo studies, a bacterial gene, neo, was transfected into mouse and guinea pig tumor cells. Inbred mice and guinea pigs were injected with syngeneic tumor lines marked with neo. DNA extracted from primary and recurrent tumors and metastases was tested for the presence of the characteristic markers. Integration of neo in cell lines was shown to be stable during tumor growth, recurrence and metastasis, and therefore to be useful for identification of tumors growing in vivo.

Animals↗
PubMed1986

In vivo immunologic selection of proviral gene deletion variants from a nonproducer clone.

The mechanism by which tumors recur at sites of injection of retrovirus-infected fibrosarcoma cell lines was investigated. Previously, it was established that tumor recurrences reflect outgrowth of rare cells that lack viral antigens and are susceptible to superinfection with the homologous retrovirus. In the present study clones isolated from a retrovirus-infected cell line were evaluated as precursors for tumor recurrence. Under conditions of in vivo immunologic selection, a clone that contained a single abbreviated copy of the provirus formed variants that lacked the proviral gene. Tumor variants lacking the proviral gene grew progressively in both nonimmune and virus-immune male Sewall Wright strain 2 guinea pigs. Tumor recurrence could be prevented by superinfection of the virus-infected fibrosarcoma cell line or by superinfection of the precursor for tumor recurrence. Cell lines infected with retroviruses varied in frequency of tumor recurrence formation. This model may be useful in analyzing gene deletion as a mechanism of tumor escape from host immunologic attack.

Animals↗
PubMedDate not supplied

Loss of alleles of loci on the short arm of chromosome 3 in renal cell carcinoma.

Loss of genes at specific chromosomal loci is a characteristic of retinoblastoma, Wilms' tumour, transitional cell carcinoma of the bladder, embryonal tumours and small cell carcinoma of the lung. The significance of nonrandom gene loss in these neoplasms is that gene loss on one chromosome may uncover null mutations at corresponding loci of the homologous chromosome. Loss of specific gene products from somatic cells may be critical in the origin or evolution of certain human tumours. Clues to identification of new loci of gene loss in common adult solid tumours may be found in literature that describes chromosomal abnormalities in rare heritable cancers. Karyotypes of tumours in two families with hereditary renal carcinoma showed translocations involving the short arm of chromosome 3 (refs 10 and 11). We have examined tumours from 18 patients with non-hereditary renal cell carcinomas and found loss of alleles at loci on the short arm of chromosome 3 in all eleven of the patients who could be evaluated.

Adult↗