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Biomedical subjects

R G Martin

Publications and source records attributed to R G Martin.

At least 91 records · Page 5Linked to original sources
PubMed1984

Mammalian DNA enriched for replication origins is enriched for snap-back sequences.

Using the instability of replication loops as a method for the isolation of double-stranded nascent DNA, extruded DNA enriched for replication origins was obtained and denatured. Snap-back DNA, single-stranded DNA with inverted repeats (palindromic sequences), reassociates rapidly into stem-loop structures with zero-order kinetics when conditions are changed from denaturing to renaturing, and can be assayed by chromatography on hydroxyapatite. Origin-enriched nascent DNA strands from mouse, rat and monkey cells growing either synchronously or asynchronously were purified and assayed for the presence of snap-back sequences. The results show that origin-enriched DNA is also enriched for snap-back sequences, implying that some origins for mammalian DNA replication contain or lie near palindromic sequences.

Animals↗
PubMed1984

Epidermoid cancer of the anorectum.

The optimal treatment for squamous and cloacogenic tumors of the anorectum is controversial. Radical surgery, limited surgery, and radiotherapy (XRT) are all potentially curative. This study was undertaken to determine which patients are candidates for each type of treatment and which would benefit from combined treatment. The records of 192 patients treated at this institution between 1954 and 1979 with the diagnosis of squamous or cloacogenic carcinoma of the anorectum were retrospectively reviewed. A subgroup of 132 patients undergoing abdominal perineal resection (APR) was analyzed to determine prognostic factors for these tumors. No survival difference was observed between the two histologic types (P = 0.51). Prognostic variables significant at P = 0.05 or better were sex, size, nodal status, and level of invasion. A new staging system is proposed that utilizes tumor size, invasion, grade, and nodal status. Actuarial 10-year survival was 100%, 76%, 29%, and 0% for Stages A, B, C and D, respectively (P values 0.22, 0.0007, and 0.01, respectively). Twelve patients undergoing APR received postoperative XRT; when compared by stage with APR alone no survival difference can be shown, although there is a trend towards fewer local recurrences. Of 26 patients (14 Stage B, 12 Stage C) receiving preoperative XRT (average 4000 R) before APR, 10 had inoperable tumors prior to XRT. All became operable. Eight patients had negative surgical margins and survival was equivalent stage for stage to the operable group (Stage B 78%, 5-year survival; Stage C 43%, 5-year survival). Eleven patients had no demonstrable primary tumor after XRT, although three had nodal metastasis. Five-year survival was 83% for this group. Thirty-one local recurrences were retreated for cure by surgery, XRT, or combination. Actuarial 5-year survival after retreatment was 38%. Thirty metachronous inguinal metastases were seen, 20 were retreated for cure, 18 by inguinal lymphadenectomy. Actuarial 5-year survival was 42%. Using a new staging system based on analysis of prognostic parameters for this disease, the outcome of combined surgery and XRT is compared. The efficacy of preoperative XRT for inoperable tumors is demonstrated. An appreciable salvage rate for local or inguinal recurrence was observed.

Carcinoma, Squamous Cell↗
PubMed1984

Adjuvant chemotherapy with fluorouracil, doxorubicin, and cyclophosphamide, with or without Bacillus Calmette-Guerin and with or without irradiation in operable breast cancer. A prospective randomized trial.

Between May 1977 and April 1980, 238 patients with operable breast cancer were treated with adjuvant fluorouracil, doxorubicin, and cyclophosphamide (FAC) chemotherapy. All patients were randomized to receive FAC alone or FAC with nonspecific immunotherapy with Bacillus Calmette-Guerin (BCG) vaccine. A randomization for routine postoperative irradiation was included in the study in May 1978. At the median follow-up of 33 months, 53 patients had developed recurrent disease. Up to the present time, there have been no significant differences in the disease-free survival of patients treated with FAC alone from those treated with FAC + BCG (P = 0.21). The disease-free survival for patients treated with and without routine postoperative irradiation was similar (P = 0.99). Disease-free survival of premenopausal and postmenopausal women was similar. The overall estimate of disease-free survival was 72% at 3 years.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1984

The management of locally advanced breast cancer: a combined modality approach.

The prognosis for patients who have locally advanced breast cancer and are treated with conventional locoregional therapy is poor. Between 1974 and 1981, 93 evaluable patients with stages III (nine patients) and IV (84 patients) disease who had received no prior therapy were placed on a combined regimen of 5-fluorouracil, 500 mg/m2 on days 1 and 8, doxorubicin (adriamycin), 50 mg/m2, and cyclophosphamide, 500 mg/m2 on day 1 (FAC) every 3 weeks for three cycles or until maximum tumor reduction occurred. This treatment was followed by mastectomy, local irradiation or both. FAC chemotherapy was then resumed until a total of 450 mg/m2 of adriamycin had been given. At this time adriamycin was discontinued and replaced with methotrexate, 30 mg/m2 on days 1 and 8 (CMF), in 3-week cycles until the patient had received therapy for a total of 24 months. The complete and partial response rate to initial FAC chemotherapy was 86%, allowing 89 of 93 patients who had tumors considered to be technically unresectable to become free of clinically detectable disease with subsequent local therapy. After a median follow-up of 53 months disease recurred in 47 patients, six of whom had attained a second disease-free status following local therapy. The estimated median length of survival for the whole group was 66 months, and the median disease-free interval (DFI) was 30 month. Age, race, menopausal status and the presence of supraclavicular lymphadenopathy had no effect on prognosis. In patients with stage III disease DFI and survival duration were increased, as was true also for patients with residual tumors of less than or equal to 1 cm in their mastectomy specimens. Patients in whom the tumors were estrogen receptor (ER)-positive also had longer DFI than those with ER-negative tumors. In spite of the encouraging results obtained with the combined modality approach to treating locally advanced breast cancer, future improvement should be pursued through additional hormonal therapy, optimal local radiation therapy timing and early resumption of intensive chemotherapy following locoregional treatment.

Adult↗
PubMed1983

Binding of simian virus 40 large T antigen from virus-infected monkey cells to wild-type and mutant viral replication origins.

The binding of purified simian virus 40 (SV40) large T antigen (T) from monkey cells infected with wild-type SV40 virus to viral replication origin-containing DNA fragments was studied by DNase footprinting and restriction endonuclease protection methods. A strong affinity binding site (site 1) of 30 base-pairs and a second, adjacent 40 base-pair lower affinity binding site (site 2), which includes the origin of replication, were detected in these assays. These sites appear identical to those previously noted in similar assays performed with the Ad2 + D2 (D2) T protein. Heating T prior to incubation with DNA significantly increased the binding to these two sites, and the order of binding did not change. Moreover, protection of sequences was observed on both strands in these two sites suggesting that both strands can participate in binding of T to these two sites. Studies with DNAs from two internal site 2 deletion mutants as well as with a DNA fragment lacking the distal 13 base-pairs of site 2 revealed that sequences in the "early" portion of site 2 are sufficient for T binding to the intact site. Furthermore, use of a new assay that measures protection of DNA sequences from specific restriction enzyme cleavage revealed that site 2 can be subdivided into two subsites, 2A and 2B, where 2A corresponds to the above-noted early segment of this locus. In titration experiments, the affinity of 2A for T was greater than that of 2B. Hence, binding to a major portion of the replication initiation sequence (i.e. site 2) is the product of at least two interactions. Finally, analyses performed with DNA from a site 1 deletion mutant, cs1085, revealed that prior binding of T to this locus did not facilitate its binding to site 2. The opposite effect was observed when D2T was employed in these assays. Thus, although similar in many respects, these proteins display a detectable difference in their DNA binding mechanisms.

Animals↗
PubMed1983

Mapping of the 3'-end positions of simian virus 40 nascent strands.

Using the instability of replication loops as the basis for the isolation of replication origins, we have undertaken an analysis of the 3' ends of the extruded nascent strands of replicating simian virus 40 (SV40) DNA. DNA fragments containing the SV40 origin of replication were obtained by digesting highly purified replicative intermediates of SV40 with BamHI and then heating at 55 degrees C for 16h. The origin-containing fragments extruded under these conditions were purified and cloned into pBR322. We used restriction mapping to analyze 640 clones of the 674 that contained SV40 sequences. A large majority of the clones were found to contain rearrangements in the sequences of either pBR322 or SV40 and were disregarded. Those clones that contained legitimate SV40 and pBR322 sequences were presumed to have been derived from the extruded SV40 nascent strands and were further analyzed. A combination of restriction enzymes was used that allowed us to define the 3' ends with an accuracy of +/- 20 base-pairs. The results of restriction analysis were confirmed by nucleotide sequence analysis of selected clones. The results show that the replication forks move with a high degree of symmetry, with respect to the initiation site of DNA replication, and are consistent with the existence of pause sites for the extension of replication forks. From the clones analyzed, it appears that the center of the replication bubble is to the early side of the BglI site.

Base Sequence↗
PubMed1983

Effect of a stem-loop structure within the SV40 replication origin upon SV40 T antigen binding to origin region sequences.

Heteroduplexes were formed between SV40 replication origin-containing DNA fragments derived from wild-type genomic DNA and a viral deletion mutant (dl295) lacking 18 base pairs from a 27 bp inverted repeat that is imbedded within the minimal replication origin sequence. Among these structures were molecules bearing a 7 bp stem and a 3 nucleotide loop on the wild-type strand between nucleotides 5230 and 6. By nondenaturing gel electrophoresis, heteroduplex molecules bearing such a structure on the E-strand could be separated from those with the complementary structure on the L-strand, and either heteroduplex could be so separated from either the wild-type or dl295 homoduplex fragment. Each of these forms was singly 5' or 3' end-labeled and then evaluated as a target for specific binding to purified SV40 large T antigen isolated from wild-type virus-infected cells. The results of DNAase footprint protection analyses showed that the existence of both the E- and L-strand stem-loop structures inhibited T binding to site 2 which composes much of the minimal origin sequence. By contrast, T bound readily to this site in both homoduplexes. Furthermore, T protected both E- and L-strand sequences of its strongest binding site (site 1)--which abuts the early side of site 2--when the site 2 stem-loop was on the L-strand, and protected L-strand site 1 sequences normally when the stem-loop was on the E-strand. However, a marked alteration of binding to site 1 E-strand sequences was noted when the stem-loop was on the E-strand. Thus this alteration in replication origin secondary structure resulted in discrete local and vicinal effects on T binding. Furthermore, the results suggest that within site 1--a sequence employed as an early transcriptional regulatory locus--T antigen can bind specifically and tightly to one strand without exhibiting similar behavior on the other strand.

Antigens, Viral, Tumor↗
PubMed1983

DNA stem-loop structures bind poorly to histone octamer cores.

Heteroduplex DNA molecules were generated in which one of the two strands contained a 7-base-pair (double-stranded) stem and 3-base-pair (single-stranded) loop. The heteroduplexes and their corresponding homoduplex parental molecules, each of approximately 260 base pairs, were used for nucleosomes reconstitution. Protection from restriction endonuclease digestion was used to probe the structure of the resulting dinucleosomes. Although 50% or more of the potential cleavage sites in the homoduplex DNAs and in the linear portion of the heteroduplex DNAs were inaccessible to nuclease digestion, no protection of the stem-loop structures was observed. The results imply that a stem-loop structure preferentially occupies the spacer region between nucleosomes, but if it is part of a core particle the stem-loop is always pointed "outward" in such a way as to be accessible to nuclease digestion.

Animals↗
PubMed1983

Flat revertants of temperature-insensitive transformants induced by simian virus 40 tsA mutants lose their ability to express T-antigen.

Temperature-insensitive transformants that contained simian virus 40 sequences at only one or a few sites in the rat chromosome and that were induced by a temperature-sensitive A gene mutant of simian virus 40 were used to select flat revertants (revertants that had lost the transformed phenotype). The isolation was performed at the nonpermissive temperature so as not to select against temperature-sensitive transformants. Nonetheless, all of the revertants examined had lost their ability to express the T-antigen at both temperatures, and all contained rearrangements of the integrated simian virus 40 sequences. These results are most compatible with the hypothesis that the T-antigen of simian virus 40 is required for the maintenance of the transformed state even in temperature-insensitive cell lines.

Animals↗
PubMed1983

Relationship of simian virus 40 tumor antigens to virus-induced mutagenesis.

We analyzed the mutation frequency to 8-azaguanine (8AZ) resistance in rat FR3T3 cells acutely infected with simian virus 40 wild type and tsA and early deletion mutants and in a series of temperature-sensitive (N) and temperature-insensitive (A) transformants derived from Chinese hamster lung (CHL) cells. Upon acute infection, the frequency of mutation to 8AZ resistance was raised at most by two- to eightfold over the spontaneous frequency, and it was independent of the presence of a functional 90,000-molecular-weight T antigen or 20,000-molecular-weight t antigen or both. Similarly, in the stable transformants of CHL cells, no correlation was found between functional T antigens and mutation to 8AZ resistance. It therefore seems unlikely that simian virus 40-induced transformation results from any mutagenic activity of this virus.

Animals↗
PubMed1981

Conservative surgery and postoperative radiotherapy in 300 adults with soft-tissue sarcomas.

From January 1963 through December 1977, 300 adults with soft-tissue sarcomas were treated by a conservative surgical excision and postoperative radiotherapy. The absolute two- and five-year disease-free survival rates are 74% (222/300) and 61.3% (103/168), respectively. The five-year survival rate varies with: (1) anatomic site, e.g., 69.4% (75/108) for extremity lesions vs. 33% (5/15) for abdominal lesions; (2) histopathologic diagnosis, e.g., 86.4% (19/22) for fibrosarcoma vs. 50% (15/30) for neurofibrosarcoma; and (3) stage of the lesion. The overall local recurrence rate was 22.3% (67/300) and 27% (81/300) of the patients developed distant metastases. The incidence of lymph node metastases as an initial site of spread was only 2.7% (8/300); therefore, elective treatment of the regional lymphatics is not indicated. The 6.5% incidence of significant complications in extremities is low and might be further diminished by careful treatment planning. The combination of conservative surgery and postoperative radiation therapy maintains a functional limb in 84.5% (169/200) of patients with extremity lesions. This rate is comparable to the five-year survival rate attained with radical surgery.

Abdomen↗
PubMed1981

The remarkable instability of replication loops provides a general method for the isolation of origins of DNA replication.

Closed superhelical replicative intermediates of SV40 DNA were stable during incubation at elevated temperatures for prolonged times. However, when the replicative intermediates were nicked or made linear by digestion either outside or inside of the replication loop, the nascent DNA strands were spontaneously extruded at a measurable rate even at 37 degrees C. The extruded DNA was double-stranded. The rate-limiting step in the extrusion was not the rate of branch migration. The instability of replication loops suggested a method for the isolation of replication origins. The method has been tested by a reconstruction experiment in which a small amount of linear SV40 replicative intermediates was mixed with a vast excess of cellular DNA. The mixture was heated overnight at 55 degrees C, and the spontaneously extruded nascent strands were isolated and inserted into pBR322. Plasmids containing SV40 DNA of less than genomic size, symmetrically distributed about the SV40 replication origin, were isolated. This result suggests that the method is generally applicable to the isolation of replication origins.

DNA Replication↗
PubMed1980

Laparotomy findings in lymphangiogram-staged I and non-Hodgkin's lymphomas.

Ninety patients with Stage I or II, including extranodal (E), presentations of non-Hodkin's lymphoma were investigated by laparotomy, either for the definitive staging of patients with peripheral presentations or for diagnosis of abdominal disease. Sixty-eight patients had staging laparotomies after extensive work-up, which included lymphangiography and bone marrow biopsy. The remaining 22 patients had laparotomies for diagnosis of abdominal symptoms or for management of unrelated surgical problems, with lyphoma being found incidentally. The incidence of positive abdominal findings in the 68 patients who had staging laparotomies depended on histologic subtype. In nodular lymphoma patients, abdominal disease was found in 17 of 28 (61%) and in diffuse lymphoma patients, in 9 of 40 (22.5%). For patients with nodular lymphomas upstaged by laparotomy, the positive findings tended to cluster around an axis consisting of the celiac, porta-hepatic, para-aortic, and root of the mesentery lymph nodes. In addition, 64% of these patients had splenic involvement. For the diffuse lymphoma upstaged by laparotomy, the distribution of positive findings was similar to that of the nodular lymphoma group; however, only 1 patient had splenic involvement. By contrast, in approximately half of the diffuse lymphoma patients who had laparotomies for diagnosis, the gastrointestinal tract was found to be involved in addition to the previously cited lymph nodes.

Abdominal Neoplasms↗
PubMed1980

Evaluation of the significance of prognostic factors in stage III Hodgkin's disease treated with MOPP and radiotherapy.

One hundred consecutive Stage IIIA and IIIB Hodgkin's disease patients were treated between July 1969 and September 1976, on a combined chemotherapy-radiotherapy program consisting of two courses of MOPP followed by radiotherapy to the mantle, abdomen, and pelvis. Eighty-eight patients completed radiotherapy to all disease sites. At the time of this analysis, the mean follow-up period for the 100 patients was 45 months; 70 patients had been followed three to seven years. Projected five-year survival and disease-free survival figures for the 100 patients were 80% and 68% respectively; corresponding results for the 88 patients were 90% and 78%. To determine whether any of the following prognostic factors influenced results for either the 100 or the 88 patients, survival curves were calculated for age, sex, constitutional symptoms, histopathology, method of staging, and status of the mediastinum. Other than age, none of these factors influenced survival significantly. However, disease-free survival figures were influenced adversely by mediastinal disease as well as age.

Adult↗
PubMed1980

Laparotomy staged I and II non-Hodgkin's lymphomas: preliminary results of radiotherapy and adjunctive chemotherapy.

Fifty-three non-Hodgkin's lymphoma patients staged I, IE, II, and IIE after laparotomy, received involved field radiotherapy with or without adjunctive chemotherapy. These cases have been analyzed for survival, disease-free survival, sites of relapse, and subsequent management. For nodular lymphoma patients, the five-year survival figure was 87%. While this represents only modest improvement over survival figures for patients staged I and II by lymphangiography, results for patients with upper torso disease were significantly better. Although there was a definite incidence of relapse, early failures common in the lymphangiogram series, were not seen in this study. Of the total group of 14 patients, 4 relapsed after a minimum complete remission period of 20 months. The five-year survival figure for all presentations of stages I and II diffuse histiocytic lymphoma was 85%. This represents a substantial improvement over the corresponding figure of 36% for patients staged clinically after lymphangiography. The improvement in results is credited to more precise staging of patients with peripheral presentations, the use of adjunctive multiple-agent chemotherapy for abdominal and mediastinal presentations, and more effective treatment for relapse. To date, too few patients with peripheral presentations have been treated to determine the value of adjunctive chemotherapy for nodular or diffuse histiocytic patients.

Aged↗
PubMed1980

Regulation of the Balb/c-3T3 cell cycle-effects of growth factors.

The platelet-derived growth factor (PDGF), which is found in serum but not in plasma, has been purified to homogeneity; it stimulates replication at a concentration of 10(-10) M. Brief treatment with PDGF causes density-inhibited Balb/c-3T3 cells to become competent to synthesize DNA; pituitary fibroblast growth factor (FGF) or precipitates of calcium phosphate also induce competence. Continuous treatment with plasma allows competent, but not incompetent, cells to synthesize DNA. A critical component of plasma is somatomedin, a group of hormones with insulin-like activity; multiplication-stimulating activity (MSA) or insulin replace plasma somatomedin in promoting DNA synthesis. We have studied the molecular correlates of competence and the role of SV40 gene A products in regulating DNA synthesis. Treatment of quiescent cells with pure PDGF or FGF causes the preferential synthesis of five cytoplasmic proteins (approximate molecular weight 29,000, 35,000, 45,000, 60,000, and 72,000 detected by SDS-PAGE under reducing conditions). Two of these competence-associated proteins (29,000 and 35,000 daltons) are insulin, or epidermal growth factor (EGF). PDGF, FGF, or calcium phosphate induce an ultrastructure change within the centriole of 3T3 cells; this ultrastructural modification of the centriole is detectable by immunofluorescence within 2 h or PDGF treatment. Plasma, EGF, or MSA do not modify the centriole. SV40 induces replicative DNA synthesis in growth-arrested 3T3 cells but does not cause this alteration in centriole structure. Gene A variants of SV40, including a mutant with temperature-sensitive (ts) T-antigen (ts A209), a deletion in t-antigen (dl 884), and several ts A209 strains containing t-antigen deletions were used to induce DNA synthesis in Balb/c-3T3 cells. Like wild type SV40, all strains induced DNA synthesis equally well under permissive or nonpermissive conditions. Addition of PDGF or plasma had little effect on SV40-induced DNA synthesis. Thus, the viral function that induces replicative DNA synthesis in Balb/c-3T3 cells. Like wild type SV40, all strains induced DNA synthesis equally well under permissive or nonpermissive conditions. Addition of PDGF or plasma had little effect on SV40-induced DNA synthesis. Thus, the viral function that induces replicative DNA synthesis in Balb/c-3T3 cells is not t and is not temperature sensitive. This SV40 gene function overrides the cellular requirement for hormonal growth factors. It does not induce transient centriole deciliation, a hormonally regulated event.

Animals↗