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S Tommasi

Publications and source records attributed to S Tommasi.

At least 37 records · Page 2Linked to original sources

Oxidative phosphorylation and F(O)F(1) ATP synthase activity of human hepatocellular carcinoma.

A study of mitochondrial oxidative phosphorylation in biopsies from human hepatocellular carcinoma is presented. Tumour mitochondria as compared to control liver mitochondria, besides a reduced activity of complex IV (cytochrome c oxidase) of the respiratory chain, show a decreased phosphorylative capacity. This appears to be mainly related to a defective F o F(1) ATP synthase complex. Use of an antibody against the F(1) portion of the complex demonstrate a definite decrease for the beta subunit of F(1) in tumour mitochondria.

Adenosine Triphosphatases↗

In vivo structure of the human cdc2 promoter: release of a p130-E2F-4 complex from sequences immediately upstream of the transcription initiation site coincides with induction of cdc2 expression.

In quiescent cells, cdc2 mRNA is almost undetectable. Stimulation of cells to reenter the cell cycle results in induction of cdc2 expression, beginning at the G1-to-S transition and reaching maximum levels during late S and G2 phases. To investigate cdc2 transcriptional regulation throughout cell cycle progression, we monitored protein-DNA interactions by in vivo footprinting along 800 bp of the human cdc2 promoter in quiescent fibroblasts and at different time points following serum stimulation. We found 11 in vivo protein-binding sites, but no protein binding was observed at a high-affinity E2F site that had previously been implicated in cdc2 regulation. Nine of the identified in vivo binding sites (among them were two inverted CCAAT boxes, two Sp1 sites, and one ets-2 site) bind transcription factors constitutively throughout the cell cycle. However, at two elements located at positions -60 and -20 relative to the transcription start site, the binding pattern changes significantly as the cells are entering S phase. A G0- and G1-specific protein complex disappears at the -20 element at the beginning of S phase. This sequence deviates at one base position from known E2F consensus binding sites. We found that the major E2F activity in human fibroblasts contains E2F-4 and p130. The -20 element of the cdc2 gene specifically interacts with a subset of E2F-4-p130 complexes present in G0 cells but does not interact with S-phase-specific E2F complexes. Transient-transfection experiments with wild-type and mutant cdc2 promoter constructs indicate that the -20 element is involved in suppressing cdc2 activity in quiescent cells. We suggest that the presence of the p130-E2F-4 complex in G0/G1 blocks access of components of the basal transcription machinery or prevents transaction by the constitutively bound upstream activator proteins.

Base Sequence↗

Detection of novel transcripts in the human mitochondrial DNA region coding for ATPase8-ATPase6 subunits.

We have analyzed the tRNA(Lys), ATPase8, ATPase6, COIII region of mitochondrial DNA in several human tissues. Beside the mature tRNA(Lys), ATPase8 and ATPase6 common mRNA, and COIII mRNA, we have characterized two new transcripts, called RNA 20 and RNA 21. The RNA 20 is a precursor species which contains the tRNA(Lys) plus the ATPase8 and ATPase6 common mRNA; the RNA 21 is an RNA species shorter than the ATPase8 and ATPase6 common mRNA. The relative concentration of the mature with respect to that of the new species proved different in the various tissues. These findings provide new insights into the mitochondrial transcription mechanism opening the question of a possibly regulatory role of the processing on the expression of the mitochondrial genome.

Adenosine Triphosphatases↗

Methylation analysis by genomic sequencing of 5' region of mouse Pgk-1 gene and a cautionary note concerning the method.

We have used genomic sequencing aided by ligation-mediated PCR (LMPCR) to assay for 5-methylcytosine in the CpG-rich promoter region of the mouse X-linked phosphoglycerate kinase gene (Pgk-1). Earlier studies showed that there was very heavy methylation of CpG dinucleotides in the CpG-rich promoter of the human PGK1 gene on the inactive X chromosome (the Xi), but that these same sites were completely unmethylated on the active X chromosome (the Xa). For mouse Pgk-1, previous restriction enzyme analysis had shown apparently complete methylation of only one cytosine in the promoter region on the Xi, at HpaII site H7, which is located in the untranslated region, 28 nucleotides upstream of the translation start site. We analyzed this potentially critical region by combining the use of HpaII with LMPCR, and find that the CpG dinucleotides near H7 are either unmethylated or only partially methylated on the Xi. LMPCR analysis of male and female DNA over a 490-bp sequence including the promoter and enhancer extend the finding of relative hypomethylation on the mouse Xi to include all CpG dinucleotides in this region. These results are relevant to the role of DNA methylation in stabilizing the inactive state of chromatin. In addition, we find that caution must be exercised in using LMPCR for methylation analysis of some sequences. A DNA concentration-dependent band-suppression artifact can incorrectly suggest methylation of both CpG and nonCpG dinucleotides.

5-Methylcytosine↗

Different assays for HER-2/neu evaluation in breast cancer.

To evaluate different methodologic approaches for HER-2/neu analysis, we performed Southern, Northern, Western blot and histochemical assay on 112 samples from 86 primary tumors and 26 synchronous axillary metastatic lymph nodes of patients affected by operable breast cancer. Simultaneous statistical analysis of data obtained with the four methods (31 samples) showed that Western blot detected a higher percentage of alterations than the other assays (Cochran and Victor tests, 0.01 less than p less than 0.05). The same result was emphasized by pair analysis (McNemar, p less than 0.05), which evaluated the assay data two by two. Immunohistochemical evaluations were more in accord with immunoprecipitation data when performed on frozen or Bouin-fixed, paraffin-embedded tissues than on formalin-fixed, paraffin-embedded tissues.

Blotting, Northern↗

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History, Modern 1601-↗

Tumor-proliferative activity, progesterone receptor status, estrogen receptor level, and clinical outcome of estrogen receptor-positive advanced breast cancer.

The relations among pretreatment tumor-proliferative activity, progesterone receptor (PgR) status, estrogen receptor (ER) level, and clinical outcome were analyzed in a series of 45 ER-positive advanced breast cancer postmenopausal women treated with tamoxifen (20 mg/day) until disease progression. Tritiated thymidine ([3H]dThd) Labeling Index (LI) by autoradiographic assay was utilized for proliferative activity analysis, whereas the dextran-coated charcoal method was used for ER and PgR evaluation (cutoff value, 10 fmol/mg of protein cytosol). The median [3H]dThd LI value was 1.8%; 73% of cases were PgR positive, and 53% were highly ER positive (greater than 100 fmol/mg of protein cytosol). Clinical responses were more frequently observed in slowly than in quickly proliferating tumors (86% versus 60%; P less than 0.05) but were similar for PgR-positive and -negative cases, as well as for those with high and low ER positivity. Only [3H]dThd LI was found to individualize patients with different survival rates (at 40 mo of follow-up, 78% versus 40% in slowly and quickly proliferating tumors, respectively). The [3H]dThd LI, monitored in ten patients by a second tumor biopsy after 14 days of tamoxifen therapy, was found to have a significantly lower median value (P = 0.03). These data indicate that pretreatment [3H]dThd LI provides information, which is not available in a study of PgR and ER status, on the clinical outcome of ER-positive advanced breast cancer patients treated by hormone therapy.

Breast Neoplasms↗

Sequential alternate administration of tamoxifen and medroxyprogesterone acetate in advanced breast cancer: clinical-biological randomized study.

From January 1985 to September 1988, 60 women with advanced breast cancer were randomized in two arms to receive: A) tamoxifen (TAM) (20 mg/die) until progression or B) TAM (20 mg/die for 14 days) then medroxyprogesterone acetate (MPA) (1500 mg/die p.o. for 14 days) followed by 7 days of wash-out before repeating the TAM/MPA treatment. All patients were post-menopausal, previously untreated with hormone therapy, and with tumor receptor status determined immediately before randomization; all had objectively evaluable lesions. In order to verify hormone receptor variations due to the antiestrogen, when possible a second biopsy was performed after the initial 14 day cycle of TAM. Thirty-one and 29 patients were included respectively in arms A and B. Objective regression (CR + PR) was observed in 58% of group A and 75% of group B, with CR in 11% and 23%, respectively. Differences were not statistically significant. Median time to progression was 12 months for group A and 9 for group B. Overall survival has not yet been reached in group A while it was 34 months for patients of group B. Metrorrhagia was observed in two cases of group A and in 6 of group B, and thrombophlebitis in 1 and 3 cases, respectively. The second biopsy confirmed a clear increase of PgR content in 8/11 cases (75%). Plasma level variations of TAM, N-desmethyl TAM and MPA were checked at various intervals on 3 patients of group B, and confirmed that our schedule is able to produce a drug wash-out period for tumor cells. In conclusion, our study demonstrated that while the manipulation of hormone receptors seems possible, results indicating better overall survival and time to progression were not obtained with alternate sequential TAM-MPA therapy.

Antineoplastic Combined Chemotherapy Protocols↗

[The relationship between the receptor status and the mammographic picture in operable breast cancer].

To verify the relationship of mammographic (Mx) patterns of breast cancer to hormone receptor content (ER, PgR) we studied 129 women (59% in postmenopause; average age 55) in the last 3 years. All women had operable breast cancer and were submitted to mammography before surgery. The tumors were classified, according to Mx characteristics, in 5 classes (Broberg, 1983). ER and PgR contents (cut-off for positivity: 10 fmol/mg prot cyt) were analyzed in all patients by means of DCC method. The percentage of ER+ cases was significantly higher in class I than in all other Mx classes (85% vs 57%; p = 0.02), whereas it was lowest in class IV (51% vs 70%; p = 0.03). The percentage of PgR+ cases was significantly different only in class I with respect to class IV (70% vs 41%; p = 0.002). As for ER and PgR mean tumor content, no statistically significant difference was observed between the 5 classes. When the 2 receptors were simultaneously considered the percentage of ER+ PgR+ cases was higher in class I than in all the extant classes (p = 0.01), and the percentage of ER- PgR- was higher in class IV than in the extant ones (p = 0.02). In selected subgroups of patients, Mx classification of breast cancer can help the physician predict the hormone receptor tumor status with sufficient reliability.

Adult↗

Cloning and cDNA sequence of the rat X-chromosome linked phosphoglycerate kinase.

This paper reports the isolation and the sequence determination of rat phosphoglycerate kinase (PGK) cDNA clones. This cDNA, derived from an X-linked PGK gene transcript, contains a reading frame of 1254 nt and 5' and 3' non coding regions of 40 and 380 nt respectively. Analysis of the nucleotide sequence at the three codon position shows a biased codon usage with a prevalence of the triplet G non G N. Comparison of the inferred rat amino acid sequence with that of other organisms makes possible the calculation of the unit evolutionary period (UEP) for this enzyme, placing it at around 40 million years (My). Thus PGK is one of the oldest housekeeping enzymes.

Amino Acid Sequence↗

Cell kinetics and hormonal receptor status in inflammatory breast carcinoma. Comparison with locally advanced disease.

The biological and prognostic role of hormone receptor status and proliferative activity have been studied in two series of patients affected by inflammatory breast carcinoma (IBC, 28 patients) and locally advanced breast cancer (LABC, 50 patients). Estrogen receptor (ER) and progesterone receptor (PgR) were measured by dextran-coated charcoal (DCC) method whereas proliferative activity was measured by 3H-thymidine autoradiographic labeling index (LI). The percentages of ER+ and PgR+ cases resulted lower in IBC than in LABC (ER+, 44% versus 64%; PgR+, 30% versus 51%, respectively), pertaining to both premenopausal and postmenopausal women. Inflammatory breast carcinoma showed a higher median LI value than LABC (3.5% versus 1.6%; P = 0.006). Regarding clinical aspects, time to progression (TTP) in IBC patients was not affected by hormone receptor status (19 evaluable patients) or by LI (17 evaluable patients); PgR+ status and low LI resulted important for individualizing women with a longer median overall survival (OS). Inflammatory breast carcinoma has been verified to be a heterogeneous biological entity for which hormone receptors and cell kinetics could be useful in identifying patients with different prognoses and therefore candidates for a personalized therapy.

Adult↗

Relevance of cell kinetics to hormonal response of receptor-positive advanced breast cancer.

The relationship between cell kinetics and hormonal status and the relevance of the cell kinetic variable on success of hormone-therapy in estrogen receptor positive (ER+) breast tumors were analyzed in patients with advanced disease. Cell kinetics were evaluated as in vitro 3H-thymidine labeling index (LI), and estrogen receptor (ER) and progesterone receptor (PgR) with the dextran-coated charcoal technique. The analyses performed on primary tumor or soft tissue metastases from 52 patients showed a general association between the presence of hormone receptors and low proliferative activity, or the absence of receptors and high proliferative activity (ER and L.I.: p greater than 0.05; PgR and L.I.: p = 0.05). However, hormonal status and cell kinetic status were unrelated in about 40% of the cases. Clinical response to additive hormonotherapy was analyzed in relation to pretreatment LI in 29 patients with ER+ tumors. Time to reach maximum response was significantly longer in slow than in fast proliferating tumors, but complete remission was reached in 88% of slow proliferating tumors compared to only 46% of fast proliferating tumors. These preliminary results show that ER+ fast proliferating tumors largely escape hormonal control, and if confirmed on larger series, could identify cell kinetics as an important tool to select patients who will benefit from hormonal treatment.

Adult↗

Correlation between ERICA and DCC assay in hormone receptor assessment of human breast cancer.

To study the basic relationships between estrogen receptor immunocytochemical assay (ERICA) and dextran-coated charcoal (DCC) techniques, 116 women affected by breast cancer were admitted to the study from June 1985. In 56% of cases the tumor sample came from patients with operable disease and in 44% from patients with advanced inoperable disease. We found an overall agreement of 80% between the two methods characterized by a high sensitivity (95%) and a low specificity of ERICA versus DCC. ER/ICA negative results were found in 20 out of 78 ER/DCC positive cases; on the contrary, discordant results were found in only 3 out of 38 cases described by DCC as ER negative. 29 patients with advanced disease and ER/DCC positive tumors were considered for clinical analysis and reliability of each hormone receptor assay. A clinical response was reached in 69% of ER/DCC positive patients and on 64% of ER/ICA positive ones (16/25). However, it must be noted that 4/29 cases described as negative by ERICA were clinically responsive to hormone therapy.

Breast Neoplasms↗

Transition and transversion rate in the evolution of animal mitochondrial DNA.

We present a further application of the stochastic model previously described (Lanave et al., 1984, 1985) for measuring the nucleotide substitution rate in the mammalian evolution of the mitochondrial DNA (mtDNA). The applicability of this method depends on the validity of "stationarity conditions" (equal nucleotide frequencies at first, second and third silent codon positions in homologous protein coding genes). In the comparison of homologous sequences satisfying the stationarity condition at the silent sites, only the four codon families (quartets) for which both transitions and transversions are silent at the third position are considered here. This has allowed us to estimate the transition and transversion rates for any pair of species. We have analyzed the third silent codon position of the triplet rat-mouse-cow, of a series of slightly divergent primates and of two Drosophila species. In terms of two external dating input we have then determined the phylogenetic trees for rat, mouse, and cow as well as for a number of primates including man. The phylogenetic tree that we have derived for the triplet rat, mouse and cow agrees with that we had previously determined by analyzing the first, second and third silent codon positions (in both duets and quartets) of mt genes (Lanave et al., 1985). For primates our method leads to the following branching order from the oldest to the most recent: Gibbon, Orangutan, Gorilla, Chimpanzee and Man. In absolute time, fixing the distance Chimpanzee-Man as 5 million years (Myr) we estimate the dating of the divergence nodes as: Gorilla 7 Myr; Orangutan 16 Myr; Gibbon 20 Myr. In all cases analyzed, the transition rate has been found to be substantially higher than the transversion rate. Moreover we have found that the transition/transversion ratio is different in the various lineages. We suggest that this fact is probably related to the nucleotide frequencies at the third silent codon position.

Animals↗

Chronobiological study on serum innumoreactive gastrin RIA levels in patients with liver cirrhosis.

After days of standardized conditions of life and standardized diets, serum gastrin levels were measured in 6 patients with liver cirrhosis on 3 consecutive days and in 14 normal controls on 1 day. Blood samples were drawn every 4 h. In patients as well as in controls, a significant serum gastrin circadian rhythm was noted. The daily mean serum gastrin levels of patients were not significantly different from those of controls. These results suggest a secondary role of the liver in gastrin metabolism.

Aged↗

Mammographic aspect, cell kinetics and hormone receptor status of operable breast cancer.

To investigate the relationships between the mammographic aspects (according to Broberg), proliferative activity and hormone receptor status, we studied 165 patients subjected to radical mastectomy for operable breast cancer. The highest percentage of estrogen receptor-positive cases was observed in mammograph class I (p < 0.02) while classes III and V were highest in progesterone receptor-positive cases. Proliferative activity (111 cases) was significantly lower in class I and IV with respect to all other classes (p < 0.03 and p < 0.01, respectively). This study suggests that certain mammographic signs can be related to the biological characteristics of breast cancer.

Breast Neoplasms↗

HER-2/neu gene in primary and local metastatic axillary lymph nodes in human breast tumors.

In order to verify whether the HER-2/neu gene is involved in the initial phases of neoplastic disease or in its progression, we evaluated the amplification and overexpression of this gene in the primary tumor and in synchronous metastatic axillary lymph nodes of 26 women with operable breast cancer. HER-2/neu was amplified in 35% and overexpressed in 33% of the primary sites; similar percentages were found in lymph nodes. The clear correlation between the two disease sites regarding gene, mRNA and protein levels, supports the hypothesis that this gene is involved in the initial and invasive phases of neoplasia. Its actual role with respect to other biological tumor characteristics during the metastatic process should be investigated further.

Adult↗