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Biomedical subjects

S M Freeman

Publications and source records attributed to S M Freeman.

At least 37 records · Page 2Linked to original sources
PubMed1996

p53 and PCNA coexpression of 81 pleural and peritoneal effusion specimens: an immunohistochemical study.

Several studies have indicated that wild type p53 plays an important role in controlling cell growth and acts as a cyclin modifier. Abnormalities in p53 induce the overexpression of proliferating cell nuclear antigen. The aim of this study is to correlate immunocytochemically the expression of mutant p53 and the proliferative index (PI) as indicated by image analysis of PCNA immunoreactivity in 81 cases of pleural and peritoneal effusions. There was a strong correlation (r = 73%) between p53 immunoreactivity and PCNA PIs. Forty-three (71%) cases indicated p53 immunostaining out of 61 cases with PCNA immunoreactivity, forty of which (93%) proved to have a diagnosis of malignancy using histological or clinical data. Furthermore, 7 malignant cases showed PCNA reactivity but no p53 immunostaining. An additional four malignant cases indicated no reactivity for either p53 or PCNA. Also, there was a significant difference in the PCNA PI between benign and malignant effusions (p < 0.001). These clinical observations confirm the function of wild p53 as a check point during cell cycling, and as a strong negative feedback effect on PCNA expression. Furthermore, such co-expression represents a significant indicator for malignancy.

Antibodies, Monoclonal↗
PubMed1996

Role of Borrelia burgdorferi in the pathogenesis of morphea/scleroderma and lichen sclerosus et atrophicus: a PCR study of thirty-five cases.

Morphea (localized scleroderma), and lichen sclerosus et atrophicus (LSA) share common features with acrodermatitis chronica atrophicans (ACA), a known chronic form of borreliosis. These include similar histologic findings such as diffuse dermal fibrosis. These observations have led several investigators to consider the possibility of Borrelia burgdorferi (Bb) as a common etiologic factor among all of these diseases. The aim of this study is to investigate the role of Bb in the pathogenesis of morphea and LSA, by assaying for its presence in lesional skin biopsies from patients with these diseases. We utilized the nested polymerase chain reaction (PCR) technique to selectively amplify a longer segment of a Bb-specific somatic gene, on DNA from paraffin-embedded, formalin-fixed tissues. The results revealed no Bb-specific DNA sequence in 28 specimens of morphea/scleroderma and 7 of LSA with varying stages of disease. Furthermore, confirmatory Southern blot of the PCR product, resulted in similar findings. These data seriously question the role played by this spirochete in the pathogenesis of morphea and LSA, at least in the southeastern part of the USA.

Adult↗
PubMed1996

An immunohistochemical study of leu 7 and PCNA expression in thyroid neoplasms.

Monoclonal Antibody (MoAb) HNK, or anti-leu-7, is reactive with several neuroendocrine and nonneuroendocrine tumors. The aim of this study is to examine anti-leu-7 reactivity in thyroid neoplasms and its relationship to cellular proliferation as determined by anti-PCNA reactivity. The expression of anti-leu-7 in 56 thyroid neoplasms (24 papillary carcinomas, 14 follicular carcinomas, two medullary carcinomas and 16 follicular adenomas) was examined immunohistochemically. Papillary and follicular thyroid carcinomas reacted with anti-leu-7 in a membranous and cytoplasmic pattern in 88% and 93% of cases, respectively. The adjacent benign tissues were nonreactive. Only eight cases diagnosed as follicular adenomas were reactive with anti-leu-7. Furthermore, the mean proliferative index (PI), as measured by the percentage of nuclei immunoreactive with anti-PCNA, was greater than 30% in all thyroid neoplasms reactive with anti-leu-7. The PI was 58% for papillary carcinomas and 68% and 48% for follicular carcinomas, and follicular adenomas, respectively. Lesions originally classified as follicular adenomas that were nonreactive with anti-leu-7 had a PI of 24% and were reclassified as hyperplastic nodules. These data suggest that anti-leu-7 may be useful for characterizing thyroid neoplasia.

Adenoma↗
PubMed1996

In situ use of suicide genes for cancer therapy.

Gene therapy has now become a standard experimental approach for treating cancers that have failed conventional therapies. As the understanding of the molecular nature of carcinogenesis develops, new approaches are being taken to directly target tumor cells, thus bypassing the difficulties of killing cells that are resistant to chemotherapy and radiation. One emerging gene therapy approach has been through the genetic modification of tumor cells with a suicide gene such as the herpes simplex virus thymidine kinase gene (HSV-TK) and ganciclovir (GCV) therapy. Death of tumor cells modified with the HSV-TK gene leads to killing of unmodified in situ tumor cells in a phenomenon termed the "bystander effect." The basis both for this effect and other gene therapy trials underway for the treatment of cancer will be discussed.

Cell Death↗
PubMed1996

In vivo analysis of the 'bystander effect': a cytokine cascade.

The "bystander effect" refers to the death of unmodified tumor cells when in contact with ganciclovir (GCV)-exposed, herpes simplex virus-thymidine kinase (HSV-TK)-modified tumor cells. Although the exact mechanism or mechanisms involved in mediating the bystander effect in vivo are unknown, our findings suggest that an intact host immune system is required for the phenomenon to occur. The present study was designed to establish the effect of HSV-TK-modified tumor cells and GCV on the tumor and its microenvironment in vivo. In sublethally irradiated and immunodeficient Balb/c mice, the bystander effect was observed to be diminished or abrogated. Histopathologic examination of the tumor mass from immunocompetent mice demonstrated centralized hemorrhagic tumor necrosis (38%) after inoculation of the HSV-TK-modified tumor cells and GCV in tumor-bearing mice compared with the control mice (5%), indicating that cytokines such as tumor necrosis factor-alpha (TNF-alpha) were being released locally. This hypothesis was underscored using reverse transcriptase polymerase chain reaction (RT-PCR), by the demonstration of cytokine mRNA expression in mice treated with HSV-TK-expressing tumors and GCV. Semiquantitative PCR analysis for TNF-alpha using PCR-MIMIC on tumor samples from mice treated on days 1 and 4 showed a two-fold increase in the level on mRNA expression. Also, immunohistochemical staining for TNF-alpha showed that mononuclear inflammatory cells infiltrating the tumor were its source. Finally, characterization of tumor-infiltrating lymphocytes (TIL) in experimental animals demonstrated a two- to three-fold increase in the number of macrophages and T cells compared with control animals. These results demonstrate that, in vivo, the bystander effect is mediated in part by an antitumor response through the release of cytokines. Further, the cytokine milieu and tumor microenvironment can be modulated following injection of HSV-TK cells and GCV to enhance the host immune response, which is of potential use in clinical trials.

Animals↗
PubMed1995

The role of cytokines in mediating the bystander effect using HSV-TK xenogeneic cells.

We have previously demonstrated that genetically modified syngeneic murine tumor cells (KBALB) expressing the herpes simplex virus thymidine kinase gene (HSV-STK) can kill nearby unmodified tumor cells in the presence of ganciclovir (GCV). The killing was mediated by a 'bystander effect' as evidenced by the prolonged animal survival when syngeneic HSV-TK gene-modified tumor cells were inoculated into mice with an intraperitoneal tumor. In this study we investigated whether irradiated xenogeneic HSV-TK gene-modified tumor cells, a human colon carcinoma cell line (HCT) transfected with the HSV-TK gene, can mediate the 'bystander effect' when used in vitro and in vivo. In vitro experiments indicate that irradiated HSV-TK gene-modified xenogeneic cells (HCT) can mediate a bystander effect on the adjacent cells when the tumor population consisted of as few as 10% of the HSV-TK expressing HCT tumor cells. In vivo, animal survival experiments demonstrate that the xenogeneic gene-modified tumor cells could generate the 'bystander effect' in mice with intraperitoneal tumors as evidenced by prolonged animal survival. In addition, histologic examination of the tumors from experimental animals showed extensive tumor necrosis 3 days post HSV-TK/GCV treatment in comparison to control animals. To evaluate the cause of necrosis in vivo, we assayed for cytokines, which may be involved in mediating this process, by performing RT-PCR and immunohistochemistry on tumor RNA and tumor cells, respectively. Production of IL-1 alpha and IL-6 mRNA within the experimental tumors was observed by RT-PCR. However, mRNA expression for other cytokines including IFN-gamma, IL-2 and IL-4 was not present. Immunohistochemical analysis for IL-1 alpha protein showed reactivity within the infiltrating mononuclear cells indicating the release of this soluble factor. These results indicate that the bystander effect can be generated using irradiated xenogeneic cells both, in vitro and in vivo. Furthermore, this process is mediated by the release of cytokines such as IL-1 alpha, IL-6 which enhances the bystander effect in vivo by immunostimulation.

Animals↗
PubMed1995

Gene-modified cells for the treatment of cancer.

Gene therapy involves the insertion of a gene into an organism to treat a disease. Since its early development in the 1970s, gene therapy has expanded rapidly both in terms of the methods available and the number of candidate diseases for treatment. This report reviews gene therapy for cancer, including methodology, pre-clinical studies and experimental clinical trials.

Clinical Trials as Topic↗
PubMed1995

Tumor oncogenic expression in malignant effusions as a possible method to enhance cytologic diagnostic sensitivity. An immunocytochemical study of 87 cases.

The diagnostic ability of cytological preparations can be hampered by specimen inadequacy and the presence of representative cells, which may result in a diagnostic accuracy of only 70%. An immunocytochemical battery (ICC), which included anti-p53, anti-c-erbB-2, and B72.3 MoAbs, was used to enhance sensitivity in 87 specimens of body effusions. Thirty-six cases were positive for malignancy using conventional cytology. Forty cases were negative and 11 cases were inconclusive or had an equivocal diagnosis. Sensitivity was 65%, and there was a negative predictive value (NPV) of 62%. p53 was expressed in 50 cases (56%, sensitivity = 83%, NPV = 73%), and B72.3 MoAb was positive in 36 cases (37%, sensitivity = 66%, NPV = 64%). Forty-eight cases (56%) displayed reactivity with anti-c-erbB-2 (sensitivity = 75%, NPV = 63%). The authors concluded that application of an ICC panel of anti-p53, B72.3 and c-erbB-2 to complement conventional cytology increases sensitivity to 98% (P < .0005) with an NPV of 96% (P = .001).

Adult↗
PubMed1995

Gastric carcinoma: recent issues in prognostic factors.

The prognosis of gastric carcinoma remains unfavorable despite a greater understanding of its molecular pathology. This retrospective study of primary gastric carcinomas was collected from one of the highest risk regions of China and examined for the oncogenetic expression of p53, c-erbB-2, and PCNA using immunohistochemistry and DNA contents by flow cytometry and image analysis. These products are reported to influence the tumor behavior. The p53 nuclear and c-erbB-2 membrane-bound stainings were seen in 58% and 34% of cases, respectively. A high PCNA index was found in 90% of the tumors. The p53 expression did not correlate with the histological differentiation, gross morphology, and depth of tumor invasion. Additionally, p53 and c-erbB-2 reactivity did not correlate with the proliferative index (PI) or S-phase DNA content. However, the mutant p53 expression was detected in the dysplastic cells adjacent to the tumor, suggesting a possible role of the oncogene in tumor pathogenesis. Mutant p53 expression can also be helpful in early detection of cases with dysplasia in well-differentiated adenocarcinomas.

Adenocarcinoma↗
PubMed1993

The "bystander effect": tumor regression when a fraction of the tumor mass is genetically modified.

Tumor cells expressing the herpes simplex virus thymidine kinase (HSV-TK) gene are sensitive to the drug ganciclovir (GCV). We demonstrate here that HSV-TK-positive cells exposed to GCV were lethal to HSV-TK-negative cells as a result of a "bystander effect." HSV-TK-negative cells were killed in vitro when the population of cultured cells contained only 10% HSV-TK-positive cells. The mechanism of this "bystander effect" on HSV-TK-negative cells appeared to be related to the process of apoptotic cell death when HSV-TK-positive cells were exposed to GCV. Flow cytometric and electron microscopic analyses suggested that apoptotic vesicles generated from the dying gene-modified cells were phagocytized by nearby, unmodified tumor cells. Prevention of apoptotic vesicle transfer prevented the bystander effect. The toxic effect of HSV-TK-positive cells on HSV-TK-negative cells was reproduced in an in vivo model. A mixed population of tumor cells consisting of HSV-TK-positive and HSV-TK-negative cells was inoculated s.c. into mice. Regression of the tumor mass occurred when the inoculum consisted of as few as 10% HSV-TK-expressing tumor cells. The bystander effect was also demonstrated in i.p. tumor studies. Initial experiments demonstrated that prolonged survival (> 70 days) occurred when a mixture containing 50% HSV-TK-positive and 50% HSV-TK-negative cells was injected i.p. followed by GCV treatment. Further, survival was prolonged for mice with a preexisting HSV-TK-negative i.p. tumor burden by injecting HSV-TK-positive cells and GCV. These results suggest that genetic modification of tumor cells may be useful for developing cancer therapies.

Animals↗
PubMed1993

Gene therapy of cancer.

Retroviral-mediated gene transfer has permitted the development of clinical protocols for the study and treatment of cancer. These protocols can be divided into gene-labeling and gene therapy proposals. Labeling studies include the tracking of tumor infiltrating lymphocytes (TIL) following the administration of those cells, and the detection, at the time of relapse, of tumor cells from transplanted autologous bone marrow. Most gene therapy protocols are designed to induce an immune attack against the tumor by inserting genes into tumor cells themselves. Although uncertainty about the safety of the procedure still exists, gene therapy of cancer holds much promise as an effective treatment modality.

Clinical Trials as Topic↗
PubMed1992

Retroviral expression of recombinant p47phox protein by Epstein-Barr virus-transformed B lymphocytes from a patient with autosomal chronic granulomatous disease.

Patients with chronic granulomatous disease (CGD) have recurrent infections resulting from a failure of phagocytic cells to produce superoxide. One third of CGD patients have an autosomal gene defect resulting in absence of p47phox protein, a cytoplasmic component critical to superoxide production by phagocytic cells. cDNA encoding p47phox has been cloned and recombinant p47phox (rp47phox) restores superoxide-generating activity to a cell-free assay containing cell membranes and cytosol from p47phox-deficient CGD neutrophils. The goal of the present study was to determine the feasibility of retrovirus mediated expression of rp47phox in the HL60 and U937 human hematopoietic cell lines, and in an Epstein-Barr virus transformed B-lymphocyte cell line (EBV-BCL) derived from a p47phox-deficient CGD patient. Normal EBV-BCL contain p47phox and generate small amounts of superoxide, while this CGD EBV-BCL lacks any detectable p47phox protein. Defective amphotropic retrovirus containing p47phox sequence inserted in the LXSN vector in sense and antisense orientations were used to transduce HL60, U937, and CGD EBV-BCL. p47phox mRNA sequence was detected in cells transduced with either sense or antisense retroviral constructs while rp47phox protein was detected only with the sense construct. The amount of rp47phox protein produced within these cells was greater than the native p47phox present in uninduced HL60 or U937 cells, but substantially less than that present in normal neutrophils, induced HL60 cells, or even normal EBV-BCL. Differentiation of transduced HL60 cells and the associated production of native p47phox in response to dimethyl sulfoxide was not affected. These studies demonstrate that retrovirus constructs can be used to mediate stable expression of rp47phox protein in human hematopoietic cell lines and can restore rp47phox protein within the cytosol of p47phox-deficient EBV-BCL from patients with CGD.

B-Lymphocytes↗