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F H Sarkar

Publications and source records attributed to F H Sarkar.

At least 109 records · Page 6Linked to original sources
PubMed1992

Genetic markers as prognostic indicators in breast cancer.

BACKGROUND: Identifying markers that have the potential to predict tumor behavior is important in breast cancer because of the variability in clinical disease progression. Genetic alterations in tumors may appear as changes in total DNA content, individual chromosomes, single genes, or gene expression. Alteration in DNA content is an imprecise but accessible measurement of the genome. Diploid tumors have been associated with a better clinical outcome, and increased ploidy correlates with other indicators of poor prognosis. Concurrent analysis of DNA content with markers of genetic expression is feasible (e.g., myc oncogene) and may increase its prognostic power. Chromosomal studies could provide a more precise tool for localizing genetic damage, but there is little cytogenetic information about primary breast cancers, no convincing evidence has emerged to target locations in the karyotype that appear specifically altered, and many primary and cultured breast cancers contain cells that appear chromosomally normal. Attempts to define molecular markers have used probes of different chromosomal sites, some chosen because of logical associations with hormonal activity, known oncogenes, or tumor-suppressor genes, and some by chance. Currently, to the authors' knowledge, none has shown uniform changes by mutation, loss, or overexpression in all breast cancers, although a remarkable number of loci are altered to some extent. These lesions must be associated with particular disease subsets or, retrospectively, with differential survival if they are to have prognostic value. METHODS: The authors examined several loci (ERBB2, INT2, MUC1) for gene amplification or loss of heterozygosity by Southern blotting and for gene expression by immunohistochemistry in breast tumors from patient groups selected by survival. RESULTS AND CONCLUSIONS: A retrospective series showed gene amplification at the erbB2 locus in 22% of rapidly recurrent (RR) tumors and 13% of tumors from long-term tumor-free survivors (LTS), but the difference was not statistically significant (P = 0.18). The erbB2 product was displayed histochemically with equal frequency between those with RR tumors and LTS patients. Moreover, the correlation was poor between different analytic measures on the same tumors. This result was tested using a prospective study of erbB2 to correlate DNA analysis with western blot findings and frozen and fixed histochemical results. Another oncogene, int2, showed significant correlation between amplification and recurrence; 16% of RR tumors showing genetic amplification (P = 0.02). Loci on other chromosomes, 1 (muc1) and 17 (cmm86), also are being investigated in groups selected for differences in survival.

Breast Neoplasms↗
PubMed1992

Analysis of retinoblastoma (RB) gene deletion in human prostatic carcinomas.

The retinoblastoma tumor suppressor gene (RB gene) has been reported to be deleted and/or modified in a number of human cancers, indicating that dysfunction of this tumor suppressor gene is perhaps critical in the development of many human tumors. In addition to deletion of one copy and/or mutational inactivation of the RB gene, this gene has also been reported to be altered by a small deletion in the promoter sequence in one case of small cell mixed adenocarcinoma of the prostate. A deletion of 105 nucleotides of the RB gene in exon 21, leading to an aberrant short-sized mRNA transcript, has also been reported in one cell line (DU 145) derived from brain metastasis of prostatic adenocarcinoma. We have analyzed tissues from 10 prostate specimens (3 hyperplastic and 7 neoplastic) and one prostate cancer cell line (DU 145) for the presence of short-sized mRNA transcript (exon 21 alterations) by polymerase chain reaction (PCR) using total RNA extracted from frozen tumors and the cell line. None of the prostate tissue showed any evidence of aberrant short-sized mRNA, although it was confirmed in the DU 145 cell line. Simultaneously, we have used DNA-PCR to investigate RB promoter deletion in 23 adenocarcinomas and one small cell carcinoma of the prostate. We also failed to demonstrate any indication of RB promoter deletion at the DNA level in adenocarcinomas. The single case of small cell carcinoma failed to show evidence of any aberration in RB promoter. We therefore conclude that neither RB promoter alterations nor the exon 21 deletion are associated with typical prostate adenocarcinoma.

Adenocarcinoma↗
PubMed1992

Purified prostaglandin synthase activates aromatic amines to derivatives that are mutagenic to Salmonella typhimurium.

Prostaglandin H synthase (PHS) is widely distributed in mammalian tissues and has the ability to oxidize a variety of mutagens and carcinogens. It may therefore play a key role in the metabolic activation of xenobiotics. The present study documents that highly purified PHS can be used in conjunction with 5-phenyl-4-pentenyl-1-hydroperoxide (PPHP), a relatively stable and non-mutagenic hydroperoxide substrate, for the metabolic activation of aromatic amines to mutagenic derivatives that can be detected in short-term Salmonella typhimurium mutagenesis assays. The PHS-based activation system alone was not mutagenic for these tester strains, nor were the test compounds significantly toxic for the bacteria over the concentration range tested. When used in conjunction with Salmonella strains TA98 and TA100 in a modified Ames assay, this system should prove useful for screening of a wide range of compounds for metabolic activation by this mammalian peroxidase. The potential broad utility of this purified PHS-dependent metabolic activation system was investigated by evaluating the activation of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), which are representative of a group of mutagenic and carcinogenic heterocyclic arylamines to which humans are exposed via their diet. Both IQ and MeIQ were activated by PHS to potent mutagens and confirm the utility of the PPHP/PHS system for the activation of premutagens. Whereas the extent of activation of aromatic amines by S9-based systems is significantly greater than for the PHS activation system described herein, PHS may play a significant role in target tissues in which it is present at significantly greater levels than P450 isoenzymes. Moreover, it is likely that the substrate specificity of PHS differs sufficiently from that of P450 isoenzymes so that PHS may activate some compounds that are not efficiently activated by mixed-function oxidase based systems.

Alkenes↗
PubMed1992

Detection of human papillomavirus in squamous neoplasm of the penis.

Infection of the external human urogenital system with human papillomavirus has been implicated with the development of genital cancer. A modified polymerase chain reaction technique has been used to evaluate type specific deoxyribonucleic acid (DNA) sequences of unique E6 to E7 transforming regions of human papillomavirus genomes (types 6b/11, 16 and 18) in a morphological spectrum of in situ (carcinoma in situ) and invasive neoplasm of the penis. We studied 15 examples of carcinoma in situ [7 bowenoid and 8 nonbowenoid (squamoid or simplex)], 11 of invasive squamous carcinoma, 1 of verrucous carcinoma, 2 of verrucous hyperplasia, 1 of urethral adenocarcinoma and 1 solitary papilloma. Viral DNA was not detected in any of the nonbowenoid specimens of carcinoma in situ, the verrucous carcinoma, the adenocarcinoma or the papilloma of the penis. Human papillomavirus types 6b/11 and 18 specific sequences also were not detectable in any of the specimens examined. However, all 7 of the bowenoid forms of carcinoma in situ were positive for human papillomavirus type 16 DNA. The presence of human papillomavirus type 16 was also detected in 9 of 11 invasive squamous carcinomas and in both verrucous hyperplasias. Our results confirm that the bowenoid forms of intraepithelial neoplasms and most invasive squamous carcinomas contain the E6 to E7 portion of type 16 human papillomavirus genome.

Blotting, Southern↗
PubMed1992

Detection of HPV 16/18 DNA in cervical adenocarcinoma using polymerase chain reaction (PCR) methodology.

Twenty-two tissue samples of primary adenocarcinoma (adenoCA) of the uterine cervix were evaluated for the presence of HPV 16/18 DNA using the polymerase chain reaction (PCR). PCR was used to specifically amplify the E6-E7 gene region of HPV 16/18 DNA. The amplification products were analyzed using gel electrophoresis and Southern dot blotting with 32p labeled type-specific oligonucleotide probes. HPV 18 DNA was identified in 13/22 (59%) and HPV 16 DNA was identified in 5/22 (23%) of the tumors. There were no tumors with mixed infections. In three patients, two different specimens were evaluated, and there was concordance of HPV typing. The presence of squamous carcinoma in situ, koilocytosis and younger patient age were associated with an increased incidence of HPV 16/18 DNA detection. HPV 16/18 DNA was not detected in six metastatic adenoCA to cervix (four endometrial, two ovarian). We conclude that HPV 16/18 DNA is present in a significant proportion of primary adenoCA of the cervix, and we have identified some clinicopathologic associations. The detection of HPV DNA may be useful in distinguishing primary from metastatic adenoCA of the cervix.

Adenocarcinoma↗
PubMed1992

Sinonasal Schneiderian papillomas: human papillomavirus typing by polymerase chain reaction.

A series of 35 formalin-fixed, paraffin-embedded Schneiderian papillomas (24 inverted, nine cylindrical cell type, and two fungiform) of the nasal cavity were evaluated for the presence of human papillomavirus (HPV) types 6b/11, 16, and 18 DNA sequences using both a highly sensitive and specific modification of the polymerase chain reaction technique and conventional in situ hybridization. The HPV gene sequences (E6-E7 portions) were not detected in any of the 24 inverted or nine cylindrical cell papillomas. One of the fungiform papillomas was positive for HPV 6b/11. We conclude: (a) the origin of most Schneiderian sinonasal papillomas is not associated with HPV infection of these common types, and (b) fungiform papilloma is a distinctive clinicopathologic subtype of Schneiderian papilloma that may be HPV-related.

Base Sequence↗
PubMed1992

Comparative method for detection of RNA-PCR-amplified signals.

During the development of a micro-method for the quantitative analysis of gene expression, we observed that the sensitivity for detection of PCR-amplified product was higher when silver staining was used, compared with either ethidium bromide staining (as reported previously) or Southern hybridization with 32P-labeled oligonucleotide probe. This observation may have an important impact on the analysis of RNA-PCR-amplified signal for the quantitation of mRNA expression, simply by densitometric scanning of silver-stained polyacrylamide gel. We believe that this is the first report to show such a comparison, demonstrating the greater sensitivity of silver staining compared with either ethidium bromide staining or Southern hybridization utilizing an oligonucleotide probe prepared by 3'-end labeling with 32P-dATP.

Base Sequence↗
PubMed1991

HPV detection by polymerase chain reaction (PCR) in verrucous lesions of the upper aerodigestive tract.

Nineteen formalin-fixed, paraffin-embedded verrucous lesions of the upper aerodigestive tract (UADT) were evaluated for the presence of human papillomavirus (HPV) 6b/11, 16, and 18 DNA sequences using the polymerase chain reaction (PCR), in-situ hybridization, and dot blot analysis. HPV DNA was confirmed in two dysplastic papillomas only; both cases contained HPV 6b/11. E6-E7 portions of HPV DNA was not reproducibly detected in any of the 11 verrucous carcinomas, 4 verrucous hyperplasias, or 2 mature papillomas. In-situ hybridization and dot blot analysis confirmed HPV 6b/11 in the two dysplastic papillomas and failed to identify HPV in the other verrucous lesions.

Adult↗
PubMed1991

Correlation of DNA ploidy with c-erbB-2 expression in preinvasive and invasive breast tumors.

Detection of c-erbB-2 oncogene product expression by monoclonal antibody staining (avidin-biotin technique) in formalin-fixed, paraffin-embedded atypical hyperplasias (AH, n = 20), intraductal carcinomas (IDCA, n = 27) and invasive carcinomas (INVCA, n = 48) was compared to ploidy determinations obtained by flow cytometry (INVCA) or image analysis (AH, IDCA). Cytoplasmic membrane staining was present in 11/48 (23%) INVCA and 8/27 (30%) IDCA but none of the AH. Tumors with abnormal DNA content expressed c-erbB-2 more frequently: INVCA, 2/19 (11%) diploid range versus 9/29 (31%) aneuploid; IDCA, 1/7 (14%) diploid range versus 7/20 (35%) aneuploid. Poorly differentiated (nuclear grade) IDCA or INVCA were also more frequently stained (14/35, 40%) than were well or moderately differentiated cases (5/40, 12.5%). Oncogene product expression and DNA content derangements may be related biologic parameters in breast neoplasia, and both are highly associated with cytologic nuclear abnormalities.

Adenocarcinoma↗
PubMed1990

Mutagenic response of mouse lymphoma cells after activation of benzidine and 2-aminofluorene with purified prostaglandin H synthase.

The mutagenic response of L5178Y mouse lymphoma cells to the model aromatic amine carcinogens, benzidine (BNZ) and 2-aminofluorene (2-AF) in the presence of the mammalian peroxidase prostaglandin H synthase (PHS) was examined. Standard incubation conditions for mouse lymphoma cells and the PHS system were developed. The cells were exposed to BNZ and 2-AF with purified PHS in the presence or absence of a peroxide, 5-phenyl-4-pentenyl hydroperoxide (PPHP) which is required for PHS-dependent amine oxidation. Incubations were carried out in a medium consisting of Hanks' balanced salt solution with calcium and magnesium and 0.1% pluronic F-68. BNZ by itself or in the presence of PPHP induced a weak mutagenic response in mouse lymphoma cells, but the addition of PHS or PHS and its co-factor PPHP increased the mutagenic response approximately 5-fold over that observed in the absence of PHS. A maximal mutagenic response for BNZ was observed after incubation with the complete activating system, PHS and PPHP. These data are in agreement with the fact that BNZ is an excellent substrate for PHS. When 2-AF was incubated with mouse lymphoma cells, only a minimal mutagenic response was observed. Incubation of 2-AF with either PPHP or PPHP and PHS (complete peroxidase system produced a significant enhancement in mutagenic response. Thus, the mutagenic response of the mouse lymphoma cells to 2-AF was dependent on the peroxide, PPHP but not the enzyme PHS. These data suggest that 2-AF, which is a poor PHS substrate, is oxidized by a different catalyst than PHS. This work demonstrates that BNZ and 2-AF are converted by peroxide-dependent mechanisms to mutagens that can be detected in mammalian cells.

Alkenes↗
PubMed1990

Detection of human papilloma virus DNA sequences by polymerase chain reaction.

We have developed a polymerase chain reaction-based procedure for reproducible detection of the E6-E7 gene in human papilloma virus DNA sequences using formalin-fixed, paraffin-embedded tissue sections. This procedure is a simple one-step procedure which does not require any elaborate hybridization following polymerase chain reaction amplification. The protocol combines modified tissue treatment and proper primer selection for efficient amplification of target DNA in a highly specific manner allowing identification in ethidium bromide-stained gels. The procedure described here is useful for a variety of tissue preparations, particularly formalin-fixed, paraffin-embedded archival tissues.

Base Sequence↗
PubMed1989

Purification and immunochemical characterization of the cytoplasmic androgen-binding protein of rat liver.

The cytoplasmic androgen-binding (CAB) protein of the male rat liver has been implicated to play a role in the androgen-dependent regulation of alpha 2u-globulin synthesis. The liver of the adult male rat contains about 50 fmol of specific high-affinity androgen-binding activity per milligram of total cytosolic protein. Photoaffinity labeling with [3H]R-1881 followed by SDS-polyacrylamide gel electrophoresis and autoradiography shows that the CAB is a 31-kilodalton protein. By means of DEAE-cellulose chromatography and preparative SDS-polyacrylamide gel electrophoresis, we have purified the CAB protein to electrophoretic homogeneity and have raised polyclonal rabbit antiserum that is monospecific to this protein. In the sucrose density gradient, the antiserum reacted with the androgen-binding component of the male liver cytosol prelabeled with tritiated dihydrotestosterone. Western blot analysis of the liver cytosol showed that the antiserum recognizes only the 31-kDa androgen-binding component. Such immunoblotting also showed that unlike the young adult, the androgen-insensitive states during prepuberty and senescence are associated with a marked reduction in the hepatic concentration of the immunoreactive CAB protein. No immuno-chemical cross-reactivity between CAB and another androgen-binding component of Mr 29K (which is associated with androgen insensitivity during prepuberty and senescence) was observed. The latter finding favors the possibility that 31- and 29-kDa androgen-binding components may have distinct sequence structure.

Aging↗
PubMed1989

Advances in the diagnosis of urothelial neoplasia.

Urothelial neoplasia is a unique cancer in that is consists of a spectrum of tumors with different biologic behaviors. The most common urothelial neoplasm is the low grade superficial papillary carcinoma or papilloma which may recur numerous times but does not result in significant morbidity or mortality. A variant of the superficial papillary carcinoma, which represents approximately 10% of the tumors, is the noninvasive papillary neoplasm which progresses to a less differentiated invasive transitional cell carcinoma (TCC). Considerable effort has been directed at identifying which of the superficial well differentiated papillary tumors will persist, recur, and progress to invasive cancer. Current approaches to identifying such tumors include cytogenetics, molecular biology, and flow cytometric DNA analysis. In the final group of bladder carcinomas, the high grade invasive neoplasms, evidence suggests that these life-threatening tumors arise de novo without identifiable precursors. Unfortunately, 75% to 90% of invasive TCCs are classified in this group, with the remaining minority progressing from preexisting recurrent superficial papillary carcinomas. Obviously the biologic behavior of these aggressive poorly differentiated tumors is life-threatening, and application of traditional diagnostic procedures and new technologies need to be directed at early diagnosis.

Biomarkers, Tumor↗
PubMed1987

Cytoplasmic androgen binding protein of rat liver: molecular characterization after photoaffinity labeling and functional correlation with the age-dependent synthesis of alpha 2u-globulin.

The liver of the mature male rat contains a moderate affinity (Kd = 10(-8)M), low-capacity, cytoplasmic androgen binding protein (CAB) whose appearance during puberty and disappearance during senescence correlate with the androgen-dependent synthesis of alpha 2u-globulin. Molecular properties of CAB were examined by photoaffinity labeling with tritiated methyltrienolone (R-1881), a synthetic androgen, and by its localization within the hepatocytes which are competent to produce alpha 2u-globulin. Photoaffinity labeling of the liver cytosol derived from postpubertal male rats, followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fluorography, showed a predominant androgen binding band corresponding to Mr 31,000. This 31-kilodalton (kDa) binding component was conspicuously absent in the liver of androgen-insensitive prepubertal and senescent male rats and in adult male rats treated with estradiol-17 beta. In addition, unlike the cytoplasmic extract, the nuclear lysate of the male rat hepatocytes did not contain the 31-kDa androgen binder. Disappearance of the 31-kDa androgen binding band from the cytosolic fraction of androgen-insensitive animals was associated with a concomitant appearance of a minor androgen binding component of apparent Mr 29,000. The livers of postpubertal male rats normally contain two subpopulations of hepatocytes, only one of which is highly active (competent) in alpha 2u-globulin synthesis. Separation of these two subpopulations through a fluorescence-activated cell sorter followed by whole cell labeling showed more than a 2-fold higher uptake of R-1881 by the competent cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Affinity Labels↗
PubMed1987

Androgenic regulation of hepatic gene expression.

Androgen-dependent synthesis of alpha 2u globulin in the rat liver has been used in our laboratory as a model for studying the effect of sex hormones on hepatic gene expression. alpha 2u Globulin is a group of low molecular weight (Mr approximately 18,000) male specific urinary proteins synthesized and secreted by hepatocytes. In the male rat hepatic synthesis of alpha 2u globulin begins at puberty (approximately 40 days), reaches a peak level (approximately 20 mg/day) at about 75 days and declines during old age. Androgens can induce alpha 2u globulin in ovariectomized female rats in vivo and in the liver perfusion system in vitro. However, both prepubertal and senescent (greater than 800 days) male rats not only do not produce alpha 2u globulin but are also refractory to androgen administration. alpha 2u Globulin is coded by a multigene family comprising about 20-30 gene copies per haploid genome. All of these gene copies seem to express translationally active mRNAs giving rise to individual isoforms of alpha 2u globulin. Appearance and disappearance of the cytoplasmic androgen-binding protein (CAB) correlates with the androgen responsiveness of hepatocytes. Photoaffinity labeling of the hepatic cytosol shows that the biologically active binding protein, found in the cytosol of the mature male rat liver, has a molecular weight of 31 kDa. A molecular transition of the 31-kDa CAB to a biologically inactive 29-kDa form may be the basis of hepatic androgen insensitivity during prepuberty and senescence.

Alpha-Globulins↗
PubMed1987

Sex-independent synthesis of alpha 2u-globulin and its messenger ribonucleic acid in the rat preputial gland: biochemical and immunocytochemical analyses.

alpha 2u-Globulin is the principal urinary protein of the mature male rat. The major urinary source of this protein is the liver where it is synthesized and secreted by hepatocytes under hormonal regulation. High levels of alpha 2u-globulin and its messenger RNA (mRNA) are also present in the preputial gland of both male and female rats, and neither castration nor ovariectomy significantly alters the preputial concentration of this protein and its mRNA. Per unit mass of RNA and protein, the preputial gland as compared to liver contains about 3-fold higher level of alpha 2u-globulin mRNA and about 300-fold higher level of alpha 2u-globulin. Despite a 3-fold (300%) difference in the content of alpha 2u-globulin mRNA, nuclear run-off experiments show only a 30% higher rate of alpha 2u-globulin gene transcription in the preputial gland than in the liver. Immunocytochemical analyses reveal that the liver possesses two alpha 2u-globulin cell populations, one showing higher immunoreactivity than the other. In contrast, the preputial gland contains only one type of alpha 2u-globulin containing acinar cells, and a large amount of alpha 2u-globulin accumulates in the ductal lumen. From these results we conclude that the 300% higher level of alpha 2u-globulin mRNA in the preputial gland is not due to a corresponding difference in the rate of transcription of alpha 2u-globulin gene. Such a difference may represent tissue-specific regulation at a posttranscriptional level of mRNA metabolism. Furthermore, the huge difference in the alpha 2u-globulin content of the preputial gland and the liver is primarily due to the cellular and ductal accumulation of this protein in the preputial gland vs. its rapid secretion by the liver.

Alpha-Globulins↗
PubMed1986

Cellular interactions in the hormonal induction of alpha 2u-globulin in rat liver.

The role of hepatocellular interaction in the androgen-dependent synthesis of rat alpha 2u-globulin was examined by immunochemical analysis of liver sections. Both after androgen administration to the ovariectomized female and during puberty in the male, only a subpopulation of hepatocytes became competent to synthesize alpha 2u-globulin. These competent hepatocytes first appeared as discontinuous patches along the wall of the central vein. After the formation of a confluent layer around the central vein, cellular competency was seen to propagate toward the periportal direction through the cords of hepatic cells. Although the periportal progression of cellular competency for the synthesis of alpha 2u-globulin appeared to be an all-or-none phenomenon, it did not require cell division. From these results we conclude that certain components of the central vein are necessary for the hormonal induction of alpha 2u-globulin in the rat liver. We also propose that a primary endocrine influence on the hepatic vein results in the production of a secondary paracrine mediator which can trigger the synthesis of alpha 2u-globulin in adjacent hepatocytes. Perivenous to periportal flow of this putative secondary mediator can explain cell recruitment for the synthesis of alpha 2u-globulin along the hepatic cords.

Alpha-Globulins↗