Search PubMed⌕ Search

Biomedical subjects

S R Wolman

Publications and source records attributed to S R Wolman.

At least 55 records · Page 3Linked to original sources
PubMed1994

Fluorescence in situ hybridization: a new tool for the pathologist.

DNA in situ hybridization methods permit investigations of genetic alterations within the context of cell morphology and tissue architecture and are of particular importance in tumor pathology. Detection of specific chromosome aberrations may be a critical adjunct to diagnosis or prognosis when morphology is ambiguous. This approach will lead to better understanding of heterogeneity in tumors and to better definition of premalignant lesions.

Humans↗
PubMed1993

Xenograft model of progressive human proliferative breast disease.

BACKGROUND: Progression of proliferative breast disease has been associated with increased risk for development of invasive carcinoma. Cell lines have been developed to facilitate the study of this process. Human cell line MCF10A originated from spontaneous immortalization of breast epithelial cells obtained from a patient with fibrocystic disease, and cell lines MCF10AneoN and MCF10AneoT were created by stable transfection of these cells with the neomycin-resistance gene and either the HRAS gene or the mutated T-24 HRAS gene, respectively. PURPOSE: Our goal was to develop an experimental model of progressive human proliferative breast disease. METHODS: MCF10A, MCF10AneoN, and MCF10AneoT cells were injected subcutaneously into the dorsal flank of male nude/beige (C57/BALB/c nu/nu bg/bg) mice (12 mice for each cell type). These mice were examined periodically for formation and persistence or growth of palpable nodules. One mouse per group was killed 1 week after cell injection; thereafter, mice were observed as long as possible. Cells were recovered from palpable lesions by enzymatic dissociation of the excised lesions. Cells re-established in tissue culture from a week-14 tumor (MCF10AneoT.TG1) were injected into 12 male nude/beige mice. Southern blot hybridization analysis of the HRAS gene locus and cytogenetic analyses were performed. RESULTS: Transplanted MCF10A and MCF10AneoN cells formed transient, small palpable nodules that regressed and disappeared during the 4th and 5th weeks. In 10 of the 12 mice, T-24 HRAS gene-transfected MCF10A cells (MCF10AneoT) formed small, flat nodules that persisted for at least 1 year. Three of these xenografts became carcinomas. One (removed 7 weeks after transplantation) was an undifferentiated carcinoma composed of polygonal cells with large, vesicular nuclei and numerous mitoses. The second (removed after 14 weeks) was an invasive squamous cell carcinoma. The third (removed after 56 weeks) was a moderately differentiated adenocarcinoma. Initially, xenografts of MCF10AneoT.TG1 cells showed intraductal proliferative changes; after 23 weeks, the lesions showed histologic features resembling those seen in atypical hyperplasia of the human breast, and later lesions showed characteristics of carcinoma in situ. The MCF10 lineage of cells of three MCF10AneoT.TG1 xenografts was confirmed by DNA fingerprinting and karyotype analysis. CONCLUSIONS: MCF10AneoT and MCF10AneoT.TG1 comprise a transplantable xenograft model that produces a broad spectrum of human proliferative breast disease. IMPLICATIONS: The reproducible establishment of representative stages in early breast cancer progression from the MCF10 model offers a new opportunity to analyze critical events of carcinogenesis and progression in breast cancer.

Adult↗
PubMed1993

Xenografts of primary human prostatic carcinoma.

BACKGROUND: Prostatic carcinoma is both the most common invasive cancer and the second most common cause of cancer deaths in men in the United States. Before 1991, attempts to propagate prostatic carcinoma from primary tumors for periods longer than 3 months were unsuccessful in vivo and in vitro with rare exceptions. In 1991, we reported establishment of slowly growing tumors for six of 10 human primary prostatic carcinomas approximately 2-6 months after transplantation. However, none of the tumors were larger than 5 mm or serially transplantable. PURPOSE: Our purpose in this study was to determine whether human primary prostatic carcinoma could be grown as serially transplantable xenografts. METHODS: Cells from primary prostatic carcinomas obtained from transurethral prostatic resections or total prostatectomies in 20 patients were injected subcutaneously into male nude mice on the day of surgery. Sustained-release testosterone pellets were placed subcutaneously in the mice 2-24 days before transplantation of tumors and at intervals of 10-12 weeks. Serial transplantations in subsequent generations of mice were carried out by similar methods. Chromosome analysis was performed on six tumors. RESULTS: Six of 20 primary prostatic carcinomas have grown sufficiently to permit serial transplantation into second mice; four have been documented histopathologically in the second mouse and serially transplanted into three or more successive mice. When a single primary tumor was injected into several mice by the same procedure, tumors failed to grow in some recipients but became serially transplantable in others. Growth of these tumors is slow and irregular, with frequent regressions. Short-term cultures of 10 tumors, eight of which were injected into mice in parallel, were initiated on the day of surgery; CWR31, which was successfully transplanted serially, exhibited only aberrant metaphases and showed clonal, chromosomal changes in culture. Including CWR31, three of the six tumors for which chromosomal analysis was successful contained clonal aberrations. Preliminary studies of SCID (severe combined immunodeficient) mice suggest that they are not superior to nude mice for establishment of serially transplantable prostatic carcinoma xenografts. CONCLUSIONS: A proportion of human primary prostatic carcinomas can be grown as xenografts. Four new serially transplantable xenografts (CWR21, CWR31, CWR91, and CWR22) are currently propagated in our laboratory, a resource that was not previously available. IMPLICATIONS: Our experience suggests that the most important factor in serial transplantation is the collaboration of urologists and pathologists in expediting placement of the tumor in cold saline, examination of the frozen section, and transplantation.

Animals↗
PubMed1993

Complementarity of interphase and metaphase chromosome analysis in human renal tumors.

Fluorescence in situ hybridization (FISH) was used as a complement to earlier cytogenetic studies of human renal tumors. Chromosome-specific para-centromeric probes were applied to cells disaggregated from tissue blocks of tumors, fresh samples from which had yielded cytogenetic results after short-term culture. Cells were dissociated from thick sections of paraffin-embedded, formalin-fixed tissues. Biotin-labeled probes specific for chromosomes 1, 3, 7, 11, 12, 15, 17, and the Y chromosome were applied in individual cases and were detected by fluorescence. Probes for chromosomes 1, 7, and 17 yielded clean signals with disomic control frequencies near 90%, and 97% of controls showed a single bright spot for the Y chromosome. The 9 cases selected all provided ample numbers of dissociated cells which were hybridized successfully, although some chromosomal signals were poor in individual cases. Abnormal copy numbers of chromosomes 1 and/or 17, not identified in culture, were observed in 7 cases. Trisomy 7 observed in culture was substantiated by FISH on the original tissues, as was loss of the Y, in each of 4 cases, respectively. Our results include limited validation of culture cytogenetics, evidence of selection in culture of tumor subpopulations, and demonstration that interphase cytogenetics by FISH is applicable to archival tissue blocks after prolonged periods of storage. Conditions of culture before harvest and inherent heterogeneity within tumors permit selection for nonrepresentative subgroups within tumors and emphasize the need for multiple approaches to evaluation.

Adenocarcinoma↗
PubMed1993

Extensive genetic alterations in prostate cancer revealed by dual PCR and FISH analysis.

The genetic alterations that underlie prostate tumorigenesis are assumed to comprise gain or loss of specific chromosomal regions, whole chromosomes, or sequence-specific mutations. Existing data have not demonstrated clear specificity of whole chromosome or regional chromosomal gain or loss that characterizes entire individual malignant lesions, or all malignant lesions, within a cancerous prostate. We have analyzed tissues from 13 patients for target sequences by using PCR and FISH techniques on paired malignant or prostatic intraepithelial neoplastic (PIN) and benign samples (usually from different areas of the same paraffin section). We exercised stringent histologic control over these samples by examining small (< 5 mm2), discrete regions of sectioned benign, malignant, and PIN tissue. The same histologic region was examined on serial sections by FISH and PCR analysis. The tissues were examined for numerical aberrations involving chromosomes 4 (as a control), 7, 8, 10, and the Y by FISH analysis, and for gain or loss of chromosome 7 and chromosomal arms 8p, 10q, and Yp by PCR analysis. The concurrent application of PCR and FISH to microdissected prostatic tissues yielded evidence of higher frequencies of genetic aberration in prostate cancers than those found with either method alone or by other approaches. These results indicate the power of simultaneous genetic assays that are closely linked to specific tumor histology.

Base Sequence↗
PubMed1993

Extrachromosomal gene amplification in acute myeloid leukemia; characterization by metaphase analysis, comparative genomic hybridization, and semi-quantitative PCR.

A case of acute myeloid leukemia (M-3) with complex karyotypic aberrations and double minute (dmin) chromosomes is presented. The patient had no history of prior exposure to mutagenic or carcinogenic agents or of other malignancies. She died from CNS involvement six weeks after the initial diagnosis. We used comparative genomic hybridization to identify the amplified sequences presumed to represent the dmin of the leukemic cells; the tumor/normal ratios indicated increased signal intensity at 8q24. This localization prompted investigation by semi-quantitative PCR that revealed amplification of the MYC oncogene. The extent of chromosome aberrations and the oncogene amplification, both linked with poor prognosis, may relate to the rapid course of this patient's disease.

Base Sequence↗
PubMed1993

Chromosomal aberrations in two sporadic gastrinomas.

Results of cell culture and cytogenetic analysis (standard and fluorescent in situ hybridization, FISH) of two sporadic gastrinomas are reported. Maintenance of hormonal activity was assessed by detection of gastrin levels during the first 3 months in culture. Case 1 showed clonal aberrations consisting of two marker chromosomes: marker 1 is a large metacentric chromosome and marker 2 is a small acrocentric chromosome. Case 2 showed a constitutional polymorphism with chromosome 15p+ and a clone in the tumor cell culture with trisomy for chromosome 3. To our knowledge, this is the first cytogenetic report of sporadic gastrinomas (Zollinger-Ellison syndrome).

Aged↗
PubMed1993

Defining the extent and nature of cytogenetic events in prostatic adenocarcinoma: paraffin FISH vs. metaphase analysis.

A comparative study of primary prostatic tumors utilizing conventional metaphase analysis of prostate tumor cultures and fluorescence in situ hybridization (FISH) analysis of paraffin-embedded tissue sections revealed significant differences in type and extent of cytogenetic aberrations. Clonal trisomy 7 was identified in two tumors by metaphase analysis of prostate cultures, but not confirmed in either case by FISH analysis. True gain of chromosome 8 was revealed by FISH analysis in malignant epithelium of four tumors but not in adjacent normal or hyperplastic glands. Neither gain nor loss of this chromosome was observed by metaphase analysis in any of the tumors. Significant monosomy and nullisomy of chromosome 10 was identified in one case by FISH, but no cells with gain or loss of chromosome 10 were observed by metaphase analysis. Significant loss of the Y chromosome was revealed in one tumor by FISH, but no cells with -Y were identified by metaphase analysis. Clonal loss of the Y chromosome was identified in two other tumors by metaphase analysis. Paraffin FISH analysis of these tumors revealed overall monosomy in both, although in one tumor there was extensive nodular loss of the Y chromosome. Paraffin FISH analysis permits identification of cytogenetic aberrations in areas identified as carcinoma (CaP), prostatic intraepithelial neoplasia (PIN), and benign prostatic hyperplasia (BPH). This technique appears more informative in defining the true extent and nature of cytogenetic aberrations in prostate cancer than metaphase analysis of prostate tumor cultures.

Adenocarcinoma↗
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

Cytogenetics of primary prostatic adenocarcinoma. Clonality and chromosome instability.

We have examined 62 prostatic adenocarcinomas by conventional cytogenetic analysis. Most were primary cultures harvested in 14 days or less. The most consistent finding was a normal male diploid karyotype, found in 87% of all cells analyzed, and as the exclusive finding in 19 tumors. Nonrandom chromosomal changes included gain of chromosome 7 and loss of the Y chromosome. In addition, clonal gains of chromosomes 8, 12, and 18, and clonal losses of chromosomes 14 and 19 were noted in individual cases. Two structural clonal aberrations, a 9p+ in one case and a t(Y;22) (q11.2;p12) in another, were also seen. Ten of 62 cultures demonstrated chromosome instability, defined herein as nonclonal gain or loss of chromosomes in more than 10% of the metaphases examined from that culture. In those cases with nonclonal numerical aberrations, loss of chromosomes was more common than gain. The distribution of apparently random numeric abnormalities was similar to that of the clonal abnormalities in that the most frequent nonclonal gain was of chromosome 7 and the most frequent nonclonal loss was of the Y chromosome. Apparently random structural aberrations were observed in less than 1% of all analyzed cells. These included a 4p-,del(3)(q13), and t(1;11). The extent of apparently random aneuploidy suggests that chromosome instability characterizes cultured prostatic adenocarcinomas. An increase in the frequency of nonclonal aberrations may be an indicator of tumor origin in a predominantly diploid cell population. The coexistence of clonally aberrant, nonclonally aberrant, and normal diploid cells in culture may reflect heterogeneity of prostate tumors in vivo.

Adenocarcinoma↗
PubMed1992

Clonal chromosomal abnormalities in desmoid tumors. Implications for histopathogenesis.

Desmoid tumors (aggressive fibromatosis) are regarded as lesions of uncertain histopathogenesis. Cytogenetic analyses of 26 desmoid tumor specimens from abdominal or extraabdominal sites of 22 patients with or without Gardner's syndrome (GS) showed clonal karyotypic abnormalities in 7 cases, random abnormalities in 14 cases, and striking telomeric fusion in 5 cases. Loss of chromosome Y, a reported feature of fibromatosis in penile and palmar locations, was detected as a clonal aberration in two patients. Additionally, involvement of 5q was observed in six patients, two of whom had GS. Clonal interstitial deletions of 5q were observed in three patients, one with and two without GS. These findings confirm a clonal and probable neoplastic origin for desmoid tumor and suggest that abnormalities of the Y chromosome and 5q may be important in the genesis of this neoplasm.

Adolescent↗
PubMed1992

Flow cytometric and genetic study of familial gonadal tumors.

Three cases of familial gonadal tumors are reported. Genetic studies were conducted in 2 cases: a brother and sister with embryonal carcinoma and a benign cystic teratoma, respectively, and a father and son with embryonal carcinoma. DNA flow cytometry of paraffin-embedded tumors and constitutional karyotyping were performed. Further genetic studies are emphasized.

Adult↗
PubMed1992

Emergence of myeloid stem cell line from T-lymphoid blastic phase of chronic myeloid leukemia in culture.

A new cell line designated JA-CML was derived from the peripheral blood of a patient with blastic phase CML. Sequential evolution of phenotypic and genetic markers was demonstrated during adaptation from primary to continuous culture in vitro. In the primary sample the majority of blast cells displayed the early T-cell markers, CD7, HLA-DR, and TdT, but were negative for the common ALL antigen (CALLA), CD4 and CD8. Simultaneously, unstimulated metaphase cells showed great karyotypic variation with a range of 43-46 chromosomes per cell. Clonal changes included the Ph chromosome t(9;22), loss of the Y and gain of several altered chromosomes. The cells grew slowly in suspension during the first 10 weeks of culture. During that time, cells still expressed the CD7 and HLA-DR antigens. Karyotypic analysis at ten weeks showed a pattern of 46,X,-Y,t(9;22),+8 in more than 90% of metaphases with disappearance of all other abnormal chromosomes noted in the original sample. A tetraploid subline exhibiting duplication of most chromosomes, including the Ph, comprised the remaining metaphases. Upon further cultivation in vitro, the cells transformed spontaneously over a period of several weeks, from T-lymphoid into myeloid cells. Expression of CD7 was lost, but reactivities with monoclonal antibodies to CD34, CD33 and CD13 were newly acquired. The karyotype was hypertriploid and all cells carried two copies of t(9;22) and lacked normal copies of No. 9 or Y. The cells have since maintained stable cytogenetic and phenotypic profiles. Molecular rearrangement of the breakpoint cluster region was identified in the primary blasts and the established line and T-cell receptor gene rearrangements were not found. These observations suggest that the leukemic blast arose from primitive stem cells, not irreversibly committed T cells, and that these stem cells retained the capacity to differentiate along the myeloid pathway.

Antigens, CD↗
PubMed1992

An approach to definition of genetic alterations in prostate cancer.

Growth patterns in prostatic cancer can reduce detectability of genetic alterations. Tumors show histologic grade heterogeneity, multifocality, interdigitation of benign and malignant glands, and varying amounts of stroma. These characteristics introduce sampling errors when one uses traditional methods for genetic analysis that depend on disaggregated cells [metaphase or interphase chromosome studies] or on tissue extracts [Southern blotting or polymerase chain reaction (PCR)] to detect molecular events. To circumvent these problems, we used two approaches to study paraffin-embedded tumors, which permit focused analysis of critical tissue components. Serial 4- to 5-microns sections are applied to slides in groups of three. Every second slide is hematoxylin and eosin stained to visualize areas of carcinoma, dysplasia, hyperplasia, and stroma; tumor-rich areas are circled with ink and used as templates to examine or excise the same areas from adjacent nonstained sections. PCR methods for quantitative and qualitative gene assay are effective in evaluating samples when alteration at a particular locus is suspected. Fluorescence in situ hybridization with chromosome-specific paracentromeric probes for detection of copy number of the relevant chromosome is applied to the adjacent section. Normal chromosome controls for both methods were demonstrated. This protocol enables us to correlate genetic alterations precisely with tumor extent and morphology.

Chromosomes, Human↗
PubMed1992

Immunocytochemical characterization of male breast cancer.

Biologic properties of breast cancer in men that might reflect alterations in pathogenesis from the disease in women were examined. We studied 22 tumors from males, 18 invasive carcinomas, three of which were papillary, and three in situ tumors of which one was papillary, and one papilloma. Our data support the previously reported high incidence of papillary carcinoma in men. Estrogen receptor status and the expression of cancer-associated antigens recognized by antibodies DF3, B73.2, SP-1, and c-erbB-2 were compared to matched tumors from females. Immunocytochemistry was performed on formalin-fixed, paraffin-embedded sections using standard avidin-biotin techniques; anti-PSA was used to exclude the possibility of metatastic prostate cancer, and 12 cases of gynecomastia were included as nonmalignant controls. The incidence of estrogen receptor positivity was higher in tumors from males (73%) than from females (54%), as has been reported previously. The range of expression of all breast cancer antigens tested in male tumors was similar to that observed in females, but some interesting differences were noted. With the exception of the anti-mucin DF3, all the antibodies reacted only with neoplastic tissues. Expression of the oncoprotein c-erbB-2 was lower (17%) in males than in females (33%), despite the preponderance in men of the large-cell type carcinomas that have been associated with c-erbB-2 expression. Unexpectedly, the pregnancy-associated hormone detected by SP-1 was expressed in 33% of tumors from males and, in contrast to females, was found in less differentiated tumors.

Adult↗