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E W Newcomb

Publications and source records attributed to E W Newcomb.

At least 37 records · Page 2Linked to original sources
PubMed1995

N-methylnitrosourea-induced Ki-ras codon 12 mutations: early events in mouse thymic lymphomas.

N-Methylnitrosourea (NMU)-induced codon 12 Ki-ras mutations were analyzed in premalignant thymic lymphomas from C57BL/6J mice by using a selective polymerase chain reaction amplification strategy. The frequency of codon 12 Ki-ras mutations was 67% (16 of 24) in NMU-treated animals with premalignant stage I disease. Previously, animals with different stages of disease had been analyzed for cytogenetic changes and for mutations in the p53 tumor suppressor gene. The genetic changes observed were early-activating codon 12 G35-->A transition mutations of the Ki-ras gene, followed closely by trisomy 15 and infrequent mutation of the p53 gene late in tumor development. The consistent and early detection of Ki-ras mutations in NMU-treated animals but not in untreated controls suggests that the mutations result from direct carcinogen exposure. Alternate pathways of NMU-induced thymic lymphomagenesis were implicated. One pathway involved putative NMU-induced mutations in other, non-ras oncogenes that cooperate with trisomy 15 to produce similar T-cell tumors. The frequency of p53 gene mutations in human and murine T-cell tumors is similar but low.

Animals↗
PubMed1995

Alterations of multiple tumor suppressor genes (p53 (17p13), p16INK4 (9p21), and DBM (13q14)) in B-cell chronic lymphocytic leukemia.

B-cell chronic lymphocytic leukemia (B-CLL) samples were screened for alterations in multiple tumor suppressor genes (p53 (17p13), p16INK4 (9p21), and disrupted in B-cell malignancy (DBM) (13q14) by using polymerase chain reaction-based assays. Eleven percent (11 of 96) of the B-CLL cases analyzed in this study and a previous study had mutations in the p53 gene. In contrast, analysis of the p16 gene showed none of 80 B-CLL cases had mutations and five cases (6%) had homozygous deletions. Deletions of 13q14 (DBM) occurred in 18% (17 of 96) of patients surveyed. Thus, 28 of 96 cases showed an alteration in one or more of the three tumor suppressor loci examined. However, cases with p53 mutations rarely showed simultaneous loss of DBM. Our results suggest that inactivation of the tumor suppressor genes p53 and DBM may be mutually exclusive, thus providing alternate pathways for tumor development in B-CLL patients.

Base Sequence↗
PubMed1995

A unique spectrum of p53 mutations in B-cell chronic lymphocytic leukemia distinct from that of other lymphoid malignancies.

The spectrum and pattern of p53 mutations detected in 42 cases of B-cell chronic lymphocytic leukemia (B-CLL) were analyzed, and several interesting features were noted. Codon 209 in the p53 gene may be a new hot-spot for p53 mutation in B-CLL disease. Four of the 42 (10%) reported B-CLL p53 mutations occurred at codon 209 versus none in 214 cases of other lymphoid malignancies screened for p53 mutations (P = 0.0006). Transversion mutations predominated at codon 273 rather than the transition mutations that are known to occur at this CpG site. Four of six (67%) B-CLL cases had transversions at codon 273 compared with two of 17 (12%) of all other lymphoid tumors examined (P = 0.02). In addition, over 65% of the p53 mutations detected in B-CLL showed a strand bias for p53 mutations on the untranscribed DNA strand. This feature of DNA strand bias is notable in cancers of the lung, esophagus, and head and neck, which may result from high exposure to carcinogens. This spectrum of p53 mutations in B-CLL together with the high frequency of transversion mutations and DNA strand bias may implicate environmental carcinogens associated with p53 gene damage in some B-CLL patients.

Codon↗
PubMed1995

P53 gene mutations in lymphoid diseases and their possible relevance to drug resistance.

Mutations in the p53 tumor suppressor gene occur with a frequency of 12.5% in lymphoid malignancies. The viral-associated diseases, Adult T-cell Leukemia (ATL) and Burkitt's lymphoma, showed higher p53 mutation frequencies of 24% and 41%, respectively. Mutations occurred in the highly conserved regions of the p53 gene. Two new hot spots for mutation were noted in exon 7 at codons 239 and 245. The spectrum of p53 mutations differs among different cancers. Transition mutations occurring in colon and brain tumors also predominated in the majority of the lymphoid malignancies. However, B-cell chronic lymphocytic leukemia (B-CLL) and non-Hodgkin's lymphoma (NHL) had an unusually high frequency of G to T transversions. Among carcinomas of the lung, liver, breast and esophagus there is also a high frequency of G to T transversions. The differences in mutation spectra between different lymphoid diseases may be due to differences in mutagenic factors or differences in the biological properties of the p53 protein in different lymphoid compartments. Mutation of the p53 gene is associated with advanced stage of lymphoid disease and poor prognosis. For B-CLL disease, p53 mutations are associated with drug resistance. Overexpression of the bcl-2 protein is also associated with a block in apoptosis. Resistance to apoptosis could be a general mechanism for drug resistance in B-CLL and other lymphoid diseases.

Drug Resistance↗
PubMed1994

Chemosensitivity of lymphocytes from patients with B-cell chronic lymphocytic leukemia to chlorambucil, fludarabine, and camptothecin analogs.

Chemosensitivity of B lymphocytes, obtained from 65 patients with B-cell chronic lymphocytic leukemia (B-CLL), Rai stages 0 through IV, was determined using the MTT assay. The results were expressed by the drug concentration required for 50% inhibition of cell viability (IC50). The cytotoxicity of chlorambucil (CLB) was compared with that of fludarabine and the DNA topoisomerase I inhibitors, camptothecin, 9-aminocamptothecin, 10,11-methylenedioxy-20(S)-camptothecin (10,11-MDC) and 9-amino-10,11-methylenedioxy-20(S)-campthothecin (9-A-10,11-MDC), and topotecan. Considerable heterogeneity in sensitivity to CLB was observed, with a median IC50 of 40.5 mumol/L in untreated patients. B-CLL cells from patients treated with CLB had a significantly higher median IC50 of 86.0 mumol/L (P < .01). Untreated as well as CLB-treated patients were divided into two subsets. For the purpose of this study, B-CLL lymphocytes with an IC50 CLB of less than 61.0 mumol/L were designated as "sensitive" and those with an IC50 CLB of > or = 61.0 mumol/L were designated as "resistant." After baseline assays, 15 untreated patients received CLB; after treatment, the IC50 increased in B-CLL lymphocytes from 13 of 15 patients. The response to CLB treatment, determined by its effect on the absolute lymphocyte count and by the Eastern Cooperative Oncology Group clinical criteria, was significantly better in patients whose lymphocytes had an IC50 CLB of less than 61.0 mumol/L before therapy (P < .01). B-CLL lymphocytes also had a variable degree of sensitivity in vitro to each of the other drugs. There was significant cross-resistance between CLB and fludarabine (P < 0.01). Whereas only 29% of CLB-resistant B-lymphocyte specimens obtained from individual patients were sensitive to fludarabine in vitro, 52% and 67% of CLB-resistant lymphocyte samples were sensitive to 10,11-MDC and 9-A-10,11-MDC, respectively. We have previously reported that p53 gene mutations were associated with aggressive B-CLL and a poor prognosis. B lymphocytes from seven patients with these mutations were resistant to CLB, and five of six were resistant to fludarabine. Lymphocytes from four of seven were resistant to 10,11-MDC, and three of four were resistant to 9-A-10,11-MDC. This study implies that the MTT assay may be useful in identifying subsets of CLL patients resistant to conventional chemotherapy. However, definitive conclusions can not be drawn in view of the small number of patients studied prospectively. In addition, these results suggest the potential of camptothecin-based therapy for patients unresponsive to standard treatment.

Antineoplastic Agents↗
PubMed1994

Pathways leading to glioblastoma multiforme: a molecular analysis of genetic alterations in 65 astrocytic tumors.

To characterize some of the genetic events underlying the development of glioblastoma multiforme, the authors analyzed 65 astrocytic tumors (seven pilocytic astrocytomas, eight astrocytomas, 16 anaplastic astrocytomas, and 34 glioblastomas multiforme) for loss of heterozygosity for chromosome 17p, loss of heterozygosity for chromosomes 10p and 10q, amplification of the epidermal growth factor receptor (EGFR) gene, and amplification of the oncogenes N-myc, c-myc, and N-ras using Southern blot analysis. Alterations of the p53 gene (positive immunostaining for p53 protein in tumors with or without p53 gene mutations) in these 65 tumors were analyzed previously. None of the 65 tumors showed amplification or rearrangement of N-myc, c-myc, or N-ras oncogenes. The molecular analysis presented here demonstrates distinct variants of astrocytic tumors, with at least three genetic pathways leading to glioblastoma multiforme. One pathway was characterized by 43 astrocytomas with alterations in p53. Glioblastomas with p53 alterations may represent tumors that progress from lower-grade astrocytomas. This variant was more likely to show loss of chromosome 17p than tumors without p53 alterations (p < 0.04). Seventy-five percent of tumors with loss of one 17p allele demonstrated mutations in the p53 gene. Loss of chromosome 10 was associated with progression from anaplastic astrocytoma (13%) to glioblastoma (38%) (p < 0.04). Amplification of the EGFR gene was a rare (7%) but late event in tumor progression (p < 0.03). A second pathway was characterized by six astrocytomas without p53 alterations and may represent clinically de novo high-grade tumors. These tumors were more likely to show amplification of the EGFR gene (83%) than tumors with p53 alterations. Sixty percent of tumors with EGFR amplification also showed loss of chromosome 10; loss of chromosome 17p was infrequent in this variant. One or more alternative pathways were characterized by 16 astrocytomas without p53 alterations and with none of the genetic changes analyzed in this study. Glioblastomas are a heterogeneous group of tumors that may arise via multiple genetic pathways.

Adolescent↗
PubMed1994

Laparoscopic surgery in a mobile army surgical hospital deployed to the former Yugoslavia.

The 212th Mobile Army Surgical Hospital was deployed from a United States military base in Germany to the former Yugoslavia to provide medical support for more than 25,000 United Nations soldiers. This medical unit was the first mobile hospital ever to deploy with a capability for laparoscopic surgery. During a 2-month trial period, seven laparoscopic procedures were performed for various abdominal emergencies. There were no complications, and all patients returned to full duty within 1 week. No equipment problems or other logistical obstacles were identified. This preliminary report suggests that laparoscopic surgery is quite feasible in a forward-deployed field hospital, allowing combat soldiers to return to full duty in a much shorter time than after conventional surgery.

Abdomen↗
PubMed1994

High frequency of p53 protein accumulation without p53 gene mutation in human juvenile pilocytic, low grade and anaplastic astrocytomas.

We analysed 31 non-glioblastoma astrocytomas for alterations in p53 protein expression and for mutations in the p53 gene. Immunohistochemistry detected p53 protein accumulation in 71% (five of seven) of juvenile pilocytic astrocytomas (WHO grade I), 63% (five of eight) of astrocytomas (WHO grade II), and 63% (10 of 16) of anaplastic astrocytomas (WHO grade III). The single strand conformation polymorphism (SSCP) assay of exons 2-11 of the p53 gene and direct DNA sequencing identified p53 mutations in 14% (one of seven) of grade I, 25% (two of eight) of grade II, and 19% (three of 16) of grade III astrocytomas. This is the first report of a p53 mutation in grade I juvenile pilocytic astrocytomas. Immunohistochemistry and SSCP analyses gave concordant results in 55% (17 of the 31) of the tumors. A total of 14 tumors, 60-80% within each grade, showed p53 protein accumulation in the absence of detectable mutations of the p53 gene. No mdm-2 gene amplification was found in these tumors. The similar frequency of p53 alterations in tumors of grades I-III suggests that the p53 gene plays a significant role early in the formation of astrocytomas rather than late in tumor progression to higher grade. The data suggest that mechanisms other than p53 gene inactivation by mutation or mdm-2 complex formation result in the accumulation of P53 protein in > 70% of non-glioblastoma astrocytomas.

Adolescent↗
PubMed1994

Analysis of alterations of oncogenes and tumor suppressor genes in chronic lymphocytic leukemia.

B cell chronic lymphocytic leukemia (B-CLL) represents the most frequent adult leukemia in the Western world. The molecular pathogenesis of B-CLL is largely unknown. Although initial reports on small panels of cases had suggested a role for Bcl-1 and Bcl-2 oncogene activation in B-CLL, later investigations failed to confirm these data. Among tumor suppressor genes, p53 mutations have been reported in a fraction of cases. In this study, we have attempted a conclusive definition of the involvement of dominantly acting oncogenes (Bcl-1 and Bcl-2) and tumor suppressor loci (p53, 6q-) in 100 cases of B-CLL selected for their CD5 positivity and Rai's stage (0 to IV). Rearrangements of Bcl-1 and Bcl-2 and deletions of 6q and 17p were analyzed by Southern blot using multiple probes. Mutational analysis (single strand conformation polymorphism and polymerase chain reaction direct sequencing) was used to assay p53 inactivation. No alterations of Bcl-1 or Bcl-2 were detected in the 100 cases tested. Mutations of p53 were found in 10/100 cases without any significant association with clinical stage. Deletions of 6q were present in 4/100 cases. Overall, our data indicate that: 1) contrary to previous reports, Bcl-1 and Bcl-2 rearrangements are not involved in CD5+ B-CLL pathogenesis and 2) p53 mutations are present in 10% of cases at all stages of the disease.

Blotting, Southern↗
PubMed1993

p53 gene mutation in B-cell chronic lymphocytic leukemia is associated with drug resistance and is independent of MDR1/MDR3 gene expression.

We studied 53 patients with B-cell chronic lymphocytic leukemia (B-CLL) and found mutations of the p53 gene in 15%. Patients with p53 gene mutations were found to have an aggressive form of B-CLL disease characterized by advanced Rai stage, rapid lymphocyte doubling time (LDT), and resistance to chemotherapy. While 27 of 29 treated patients (93%) without p53 mutations achieved a partial remission, only one of seven treated patients (14%) with p53 mutations achieved a partial remission (P = .00009). Adjusting for prognostic factors (age, sex, race, and Rai stage), patients with p53 gene mutations had a 13-fold greater risk of death than patients without p53 mutations (P = .013). In addition to examining the clinical relevance of p53 gene mutations in B-CLL, we investigated the possible role of p53 gene regulation in the expression of the multidrug resistance genes MDR1 and MDR3. We quantitated MDR1 and MDR3 mRNA expression by reverse transcription-polymerase chain reaction (RT-PCR). Expression of both the MDR1 and MDR3 genes was independent of p53 gene mutation or prior drug treatment, and did not predict for clinical response. Our findings indicate that p53 gene mutations in B-CLL are associated with a poor clinical outcome and may be a prognostic indicator for drug resistance.

Adult↗
PubMed1993

A correlative study of p53 protein alteration and p53 gene mutation in glioblastoma multiforme.

The p53 tumor suppressor gene is frequently mutated in glioblastomas. Mutations within the p53 gene often result in aberrant expression of the p53 protein leading to protein accumulation within the nucleus of the cells which can be detected by immunochemistry. Many studies have correlated alterations of p53 protein expression with p53 gene mutations. Positive staining of tumor cells for p53 protein has been widely assumed, perhaps incorrectly, to signify the presence of p53 gene mutations. This study compared the immunostaining patterns for p53 protein in 37 glioblastomas with the molecular genetic data obtained by the single strand conformation polymorphism assay. p53 gene mutations were detected in 46% (17 of 37) of glioblastomas, while 65% (24 of 37) of glioblastomas were positive for protein accumulation by immunohistochemistry. Although 30 of 37 glioblastomas analyzed showed concordance for p53 protein expression and p53 gene mutations, a subset of seven glioblastomas showed discordant accumulation of the p53 protein in the absence of any detectable p53 gene mutations. The mdm-2 gene was assessed in 17 glioblastomas for gene rearrangements or amplification, but none were found. This result suggests that a mechanism other than p53 gene mutation can result in altered p53 protein expression.

Antibodies, Monoclonal↗
PubMed1992

p53 mutations in C57BL/6J murine thymic lymphomas induced by gamma-irradiation and N-methylnitrosourea.

Genomic DNA from thymus tissue obtained from 47 C57BL/6J animals treated with the DNA alkylating agent N-methylnitrosourea or gamma-irradiation were screened for the presence of p53 mutations by using the single strand conformation polymorphism assay. Mutations were detected in 13% (4 of 30) of primary thymic lymphomas but none of 17 early stage lymphomas. The frequency of p53 mutations was the same in tumors induced by N-methylnitrosourea (2 of 15) or by gamma-irradiation (2 of 15). Mutations occurred in the highly conserved regions of the p53 gene in exons 5, 7, and 8. G:C to A:T transitions were commonly observed. One of 4 of the tumors analyzed contained two p53 mutations in exons 7 and 8. A previous study of the same tumors showed that ras mutations occurred with high frequency (greater than 50%) (E. W. Newcomb et al., Cancer Res., 48:5514-5521, 1988). Our data suggest that p53 mutations do not play a major role in carcinogen-induced thymic lymphomas studied here.

Animals↗
PubMed1992

p53 mutations in human malignant gliomas: comparison of loss of heterozygosity with mutation frequency.

Mutations in the p53 gene were analyzed in 40 gliomas using the single strand conformation polymorphism assay together with restriction fragment length polymorphism analysis to assess loss of heterozygosity for 17p alleles in the same tumors. Mutations occurred in 40% of the gliomas and were found in exons 4-8 of the p53 gene. G:C to T:A transversions, which occur in high frequency in some lung (greater than 50%), liver (greater than 80%), breast (30%), and esophageal cancers (25%), were noted in greater than 25% of the gliomas studied here. These transversions were clustered in exon 5 from codons 156 to 168, a region of the p53 gene not previously associated with a high frequency of mutation, and may represent a new hot spot for mutations in certain cancers. The majority of gliomas (27 of 38) analyzed here retained both 17p alleles. The frequency of p53 mutations was 37% in this group of tumors and increased to 64% in tumors with one 17p allele. Allelic loss for chromosome 17p occurred in 4 of 11 gliomas independently of mutations in the p53 gene. Absence of p53 mutations in 36% of the tumors with one 17p allele suggests that a tumor suppressor gene other than p53 may be located on chromosome 17p and involved in progression to malignancy of some gliomas.

Adolescent↗
PubMed1991

p53 mutations in nonastrocytic human brain tumors.

Genomic DNA from 51 primary human brain tumors was screened for the presence of mutations in the tumor suppressor gene, p53, using the polymerase chain reaction and single strand conformation polymorphism analysis, followed by direct DNA sequencing. Mutations leading to an amino acid change were found in 2 of 17 (12%) oligodendrogliomas and 2 of 19 (11%) medulloblastomas but none of 15 ependymomas. Sites of mutations were in exon 5 (codon 141), exon 6 (codon 193 and 213), and exon 7 (codon 246). In addition, there were silent mutations in exon 6 (codon 213) in one oligodendroglioma and in one ependymoma. This study points to the possible role of the p53 tumor suppressor gene in some central nervous system neoplasms of divergent histogenesis.

Adult↗
PubMed1991

Differential expression of the normal and mutated K-ras alleles in chemically induced thymic lymphomas.

The presence of point mutations in the K-ras gene was examined in murine thymic lymphomas induced by a single dose of N-methylnitrosourea by the RNase A mismatch cleavage method and by allelic-specific oligonucleotide hybridization of in vitro amplified DNA by polymerase chain reaction. The results show that the frequency of mutations is lower than that of tumors induced by multiple N-methylnitrosourea treatments. Four mutations identified were the aspartic acid at codon 12, a G:C to A:T transition in its second position. A G:C to T:A transversion in codon 146 was also found in one thymic lymphoma, changing the amino acid alanine to serine. The use of the RNase A assay allowed an estimation of the relative expression levels of both normal and mutant K-ras alleles. The results show that in approximately one half of the tumors the mutant allele is predominantly expressed, suggesting that the normal allele has been lost or that the mutant allele has been amplified relative to the normal. Altogether, these findings are consistent with ras mutations occurring in some instances during tumor development and with a ras effect being not strictly dominant but favoring selection for increasing levels of expression from the oncogenic allele.

Animals↗
PubMed1991

p53 mutations in human lymphoid malignancies: association with Burkitt lymphoma and chronic lymphocytic leukemia.

We have investigated the frequency of p53 mutations in B- and T-cell human lymphoid malignancies, including acute lymphoblastic leukemia, the major subtypes of non-Hodgkin lymphoma, and chronic lymphocytic leukemia. p53 exons 5-9 were studied by using genomic DNA from 197 primary tumors and 27 cell lines by single-strand conformation polymorphism analysis and by direct sequencing of PCR-amplified fragments. Mutations were found associated with (i) Burkitt lymphoma (9/27 biopsies; 17/27 cell lines) and its leukemic counterpart L3-type B-cell acute lymphoblastic leukemia (5/9), both of which also carry activated c-myc oncogenes, and (ii) B-cell chronic lymphocytic leukemia (6/40) and, in particular, its stage of progression known as Richter's transformation (3/7). Mutations were not found at any significant frequency in other types of non-Hodgkin lymphoma or acute lymphoblastic leukemia. In many cases, only the mutated allele was detectable, implying loss of the normal allele. These results suggest that (i) significant differences in the frequency of p53 mutations are present among subtypes of neoplasms derived from the same tissue; (ii) p53 may play a role in tumor progression in B-cell chronic lymphocytic leukemia; (iii) the presence of both p53 loss/inactivation and c-myc oncogene activation may be important in the pathogenesis of Burkitt lymphoma and its leukemic form L3-type B-cell acute lymphoblastic leukemia.

Base Sequence↗
PubMed1991

ras activation in human tumors and in animal model systems.

Environmental agents such as radiation and chemicals are known to cause genetic damage. Alterations in a limited set of cellular genes called proto-oncogenes lead to unregulated proliferation and differentiation. We have studied the role of the ras gene family in carcinogenesis using two different animal models. In one case, thymic lymphomas were induced in mice by either gamma or neutron radiation, and in the other, keratoacanthomas were induced in rabbit skin with dimethylbezanthracene. Human keratoacanthomas similar to the ones induced in rabbits were also analyzed. We found that different types of radiation such as gamma rays and neutrons, induced different point mutations in ras genes. A novel K-ras mutation in codon 146 has been found in thymic lymphomas induced by neutrons. Keratoacanthomas induced in rabbit skin by dimethylbenzanthracene show a high frequency of H-ras-activated genes carrying a mutation in codon 61. The same is observed in human keratoacanthomas, although mutations are in both the 12th and the 61st codons of the H-ras gene. H-ras activation is less frequent in human squamous cell carcinomas than in keratoacanthomas, suggesting that ras genes could play a role in vivo in differentiation as well as in proliferation.

3T3 Cells↗