Scott M. Freeman

Study of tumor infiltrating lymphocytes and transforming growth factor- β as prognostic factors in breast carcinoma

Cytokines and growth factors are powerful modulators of the immune response. Their aberrant expression either by the tumor cells or by the tumor infiltrating lymphocytes confers a selective advantage to the tumor to grow and suppress the cytotoxic activity of the infiltrating lymphocytes. Therefore, analysis of these soluble factors in the tumor microenvironment can provide an insight into the understanding of the tumor behavior and may be used as a prognostic factor. In the present study the nature of the tumor infiltrating lymphocytes (TILs) and cytokine profile was examined in 36 and 19 mammary carcinoma tissues, respectively, by immunohistochemistry and PCR.

Chronic corticosterone impairs memory performance in the Barnes maze.

Chronic stress has been reported to impair spatial memory and cause hippocampal impairment in rodents. Glucocorticoids are believed to be the active agent in this impairment. Studies have demonstrated that chronic glucocorticoid administration results in animals being impaired in the Morris water maze (MWM) or eight-arm radial maze. Although both of these methods are well established means of testing spatial memory, neither might be considered optimal for studying the behavioral effects of stress. The Morris maze is itself highly stressful to the animals. The eight-arm maze relies on a food reward to motivate the animals, and glucocorticoids have profound effects on hunger and satiety. We therefore investigated behavioral deficits of corticosterone-treated animals in the two previously used mazes and the Barnes circular platform maze (BCM), a test similar in design to the Morris maze, but one that does not require the animal to perform a highly stressful swim. Consistent with results in other tests, we found that animals that had been treated for 3 months with stress-equivalent concentrations of glucocorticoids showed significantly impaired behavior in the Barnes maze.

Tumor-host interaction: Analysis of cytokines, growth factors, and tumorinfiltrating lymphocytes in ovarian carcinomas

The host-tumor interaction may play an important role in determining tumor progress. Recent studies have shown that this interaction can be influenced by the release of soluble factors by tumor cells and tumor-infiltrating lymphocytes (TIL). The aim of our study is to characterize the nature of cytokines and growth factors and their relationship to the cellular infiltrates in 16 patients with ovarian cancer using reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemistry. Total RNA from 20 malignant and 10 benign specimens were used to assay for expression of 12 cytokines. Additionally, monoclonal antibodies (MAbs) were used to detect T cells, CD4+ helper and CD8+ cytotoxic/suppressor T-cell subtypes, B cells, and macrophages. Our results showed the expression of transforming growth factor-beta1 (TGF-beta1), interleukin-10 (IL-10), and granulocyte-macrophage colony-stimulating factor (GM-CSF) in 19, 17, and 10 malignant specimens, P < .001, .001, and .05, respectively. Other cytokines such as interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), TNF-beta/LT, IL-2, and IL-6 were expressed in a few cases, and IL-1alpha and IL-4 expression were not detected. The benign samples did not express IL-10, but GM-CSF, TGF-beta1, and IL-8 were expressed in one, one, and four specimens, respectively. Interestingly, in four cases in which samples from the primary and relapse tumors were available for analysis, the tumors in relapse showed a significant increase for TGF-beta1 (P < .05) and a decreased trend in IL-10 mRNA levels. The source of these factors was tumor cells as detected immunohistochemically. This combined alteration of TGF-beta1 and IL-10 was associated with a significant reduction in number of TIL in general, and CD8+ and macrophages in particular (P = .036 and .049, respectively). Our findings suggest the important role of certain soluble factors in the complex process of tumor progression. Furthermore, understanding the tumor-host relationship and the factors influencing the interaction may be helpful in developing effective and innovative treatment methods.

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.

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 Protocol The Treatment of Malignant Mesothelioma with a Gene Modified Cancer Cell Line: A Phase I Study

1.1 To evaluate the safety and side effects of treatment with the gene-modified ovarian cancer cell line PA1-STK, which is administered intrapleurally and activated with ganciclovir. 1.2 To determine a maximum treatment dose (MTD) and the dose limiting side effects of this treatment. 1.3 To evaluate the immunologic response to this treatment. 1.4 To measure intrapleural pharmakokinetics of the drug ganciclovir. 1.5 To observe for clinical effects on the residual malignant mesothelioma.

Aging in the hippocampus: Interrelated actions of neurotrophins and glucocorticoids

Over the past two decades, evidence has been accumulating that diffusible molecules, such as growth factors and steroids hormones, play an important part in neural senescence, particularly in the hippocampus. There is also evidence that these molecules do not act as independent signals, but show interrelated regulation and cooperative control over the aging process. Here, we review some of the changes that occur in the hippocampus with age, and the influence of two classes of signaling substances: glucocorticoids and neurotrophins. We also examine the interactions between these substances and how this could influence the aging process.

Enhancement of tumor killing using a combination of tumor immunization and HSV-tk suicide gene therapy

Tumor cells genetically modified with the herpes simplex virus thymidine kinase (HSV‐tk) gene in combination with ganciclovir (GCV) demonstrate a “bystander effect”. Previous attempts to enhance the bystander tumor killing by combining cytokine genes with HSV‐tk/GCV have met with varying results. The present study was designed to determine the effects of tumor immunization in combination with HSV‐tk gene‐modified tumor cells and GCV on tumor killing and to determine if the bystander tumor killing could be enhanced. Tumor‐bearing mice immunized with syngeneic tumor (KBALB) prior to treatment with an i.p. injection of xenogeneic HSV‐tk gene‐modified tumor cells (PA‐1STK) had prolonged animal survival (group 4, 56.4 days). In contrast, unimmunized tumor‐bearing mice (group 2) or tumor‐bearing mice immunized to the xenogeneic PA‐1STK tumor cells (group 5) showed a mean survival of about 27 days after receiving an i.p. injection of PA‐1STK cells and GCV. Control groups, which were either not immunized and did not receive HSV‐tk cells (group 1) or immunized but treated only with GCV (group 3) showed short survival (16–18 days). Analysis of tumors for cytokine mRNA expression revealed increased TNF‐α and IL‐1α mRNA expression in group 4 mice. Furthermore, IL‐2 mRNA expression was detectable on days 2 and 4 only in group 4 mice. Immunophenotypic analysis for tumor‐infiltrating lymphocytes demonstrated an increase in macrophage (4%, p = 0.0001) and T cells (1.8%, p < 0.001) in group 4 mice with an enhanced T‐cell response as compared with mice from groups 1, 2 and 3. Our results demonstrate that tumor immunization combined with HSV‐tk/GCV treatment results in increased animal survival with enhanced immune response. Furthermore, the cytokine milieu observed in the present study can modulate the tumor micro‐environment in vivo from one that is immunosuppressive to one that is immune‐stimulatory. Int. J. Cancer 80:380–386, 1999.

p53, c-erbB2, and PCNA Status in Benign, Proliferative, and Malignant Ovarian Surface Epithelial Neoplasms A Study of 75 Cases

Low malignant potential tumors of the ovary are believed to behave in a manner intermediate to their benign and malignant counterparts. However, recent evidence suggests these lesions are in fact benign and better classified as proliferative. Based on our previous work and evaluating p53, c-erbB2, and PCNA status in a full spectrum of ovarian surface epithelial tumors, with emphasis on low malignant potential tumors, we tested this hypothesis. Immunohistochemical stains with monoclonal antibodies were used on 75 archival ovarian neoplasms. The results demonstrated anti-p53 reactivity in 30 carcinomas (40%), 2 of which were proliferative, and no reactivity in the benign tumors. Overexpression of c-erbB2 was seen in 31 malignant neoplasms (64.5%), 4 of which were proliferative (22.1%), and none in benign tumors. The PCNA proliferative index showed means of 42.8%, 22.8%, and 14.9% with benign, low malignant potential, and malignant tumors, respectively. Predicting immunoreactivity in carcinomas for anti-PCNA (Student t test), anti-p53, and anti-c-erbB2 (Pearson chi2 test) versus a lack of immunoreactivity in proliferative tumors indicate P values of .001, <.001, and <.001, respectively. These data show significant differences in the expression of these markers in ovarian tumors and suggest a possible role for these oncogenes as supplemental tools in diagnostic pathology. Further, our findings also support the designation of proliferative as opposed to the current nomenclature of low malignant potential tumors.

Gene therapy : clinical potential and relationships to drug treatment

Gene therapy has been defined as the alteration of the genetic material of a cell with resultant benefit to a patient. Since its first clinical application in 1989, gene therapy has become a standard experimental approach for a number of diseases that have no alternative treatment. Gene therapy trials have been separated into 2 broad categories: therapeutic trials, in which the goal is to treat a disease; and marking trials, in which the goal is to transfer a gene to label a cell type to determine the fate of a cell or the marker gene. Clinical trials are ongoing in 5 different general areas, which will be discussed. Before a clinical trial begins in the US, the protocol must undergo review by both the National Institutes of Health Recombinant DNA Advisory Committee (RAC), if the protocol has received federal funding, and the US Food and Drug Administration. For each section, a general overview of the basic science background for each application is outlined, along with the current clinical protocoL The focus of this article is on the utility of gene transfer as a drug delivery system, and both therapeutic and marking trials are discussed.