Evaluation of tyrosinase mRNA as a tumor marker in the blood of melanoma patients.

Purpose: The value of tyrosinase messenger RNA (mRNA) detection in the peripheral blood by reverse-transcription polymerase chain reaction (RT-PCR) as a melanoma marker remains controversial. The purpose of this study was to compare the sensitivities of two different blood processing techniques for tyrosinase mRNA detection and evaluate its potential clinical value.

Methods: A total of 50 patients with progressive stage IV melanoma was studied.

Recombinant interleukin-2 in combination with recombinant interferon-gamma in patients with advanced malignancy: a phase 1 study.

Because interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) act synergistically in vitro in the generation of lymphokine-activated killer (LAK) cells. we initiated a clinical trial of these lymphokines in combination. Twenty patients with advanced malignancy were treated at fixed dose levels of recombinant IFN-gamma given by intramuscular (i.

Protein tyrosine kinase inhibition blocks granule exocytosis and cytolytic function of lymphokine-activated killer cells.

Short-term pretreatment of human lymphokine-activated killer cells (LAK) with the protein tyrosine kinase-specific inhibitor Herbimycin A (Herb A) blocked cytotoxic function against the NK-resistant (LAK-sensitive) tumor targets, SK-Mel-1 (human melanoma) and Daudi (human lymphoma). Greater than 50% inhibition of LAK activity was observed after a 2.5-h pretreatment with 0.

Predominance of a type 2 intratumoural immune response in fresh tumour-infiltrating lymphocytes from human gliomas.

Increasing evidence suggests the existence of polarized human T cell responses described as Th1-type (promoting cell-mediated immunity) and Th2-type (promoting humoral immunity), characterized by a dominant production of either interferon-gamma (IFN-gamma) or IL-4, respectively. Little is known about the intratumoural activation of infiltrating lymphocytes (TIL) in human gliomas. Therefore, we assessed fresh TIL at cellular and molecular levels to find out if they were activated and polarized into a type 1 or 2 immune response.

Interleukin-6 production and secretion in human melanoma cell lines: regulation by interleukin-1.

These investigations were designed to test the hypothesis that exogenous and/or endogenous interleukin-1 (IL-1) regulates interleukin-6 (IL-6) production in human melanoma cell lines. Ten cell lines were examined for IL-1 and IL-6 expression. Six of these 10 lines constitutively expressed detectable IL-6 mRNA by RT-PCR; three of these six cell lines also produced intracellular and secreted IL-6 as evidenced by positive reaction for IL-6 using immunohistochemistry staining and ELISA methods; three others produced only intracellular IL-6.

Molecular analysis of a silent polymorphism in the PDZ domain of p55, the major palmitoylated erythrocyte membrane protein.

Two independently published cDNA sequences of p55, the X-linked major palmitoylated erythrocyte membrane protein, revealed a discrepancy between G and T at position 358 (Genbank: M64925). This results in codon 85, in exon 3 in the PDZ (PSD-95, discs-large, Z0-1) domain, being either ACG or ACT. As both ACG and ACT code for threonine, this represents a silent polymorphism.

Characterization of the M(r) 65,000 lymphokine-activated killer proteins phosphorylated after tumor target binding: evidence that pp65a and pp65b are phosphorylated forms of L-plastin.

Contact between lymphokine-activated killer (LAK) cells and natural killer-resistant tumor targets SK-Mel-1 (human melanoma) or Raji (human lymphoma) stimulates phosphorylation of two M(r) 65,000 LAK proteins (pp65a and pp65b) with nearly identical isoelectric points. Phosphoamino acid analysis of pp65a and pp65b detected phosphorylation exclusively on serine residues. Phosphotyrosine could not be detected on either substrate after immunoblotting with an antiphosphotyrosine antibody, and herbimycin A treatment failed to inhibit p65 phosphorylation induced by target contact.

Tumor target binding induces phosphorylation of two M(r) 65,000 lymphokine-activated killer proteins.

Effector-target cell conjugate formation is an essential step during lymphokine-activated killer (LAK) cell-mediated cytotoxicity. Protein phosphorylation changes in human LAKs after contact with NK-resistant (LAK-sensitive) tumors were examined by two-dimensional SDS-PAGE. Exposure to either SK-Mel-1 (melanoma) or Raji (lymphoma) targets led to increased phosphorylation of two M(r) 65,000 LAK proteins, pp65a and pp65b, with isoelectric points of 5.

High expression of adhesion molecules/activation markers with little interleukin-2, interferon gamma, and tumor necrosis factor beta gene activation in fresh tumor-infiltrating lymphocytes from lung adenocarcinoma.

Little is known about the activation level of tumor-infiltrating lymphocytes (TIL) in human lung adenocarcinoma. We investigated the activation of fresh TIL at cellular and molecular levels and compared it with autologous and healthy normal peripheral blood lymphocytes (PBL) for baseline level. TIL were extracted from 12 primary lung adenocarcinomas by mechanical disruption without enzyme use and isolated by double-density Ficoll gradients.

Interleukin-1 production in tumor cells of human melanoma surgical specimens.

To determine whether IL-1 alpha and/or IL-1 beta protein is expressed by human melanoma tumor in vivo, we first analyzed nine human melanoma cell lines and optimized the in situ detection of these proteins. Three of the melanoma cell lines stained positively for both IL-1 alpha and IL-1 beta using immunohistochemistry (IHC). THe specificity of IHC was confirmed by the ability of purified recombinant IL-1 alpha and IL-1 beta protein to abolish the staining after being adsorbed by their respective antibodies before use in IHC.

Differential inhibition of lymphokine-activated killing, proliferation, and cytokine secretion by humanized antibodies against the low- and intermediate-affinity interleukin-2 receptors. A novel model for activation of human peripheral blood mononuclear cells by interleukin 2.

Human PBMCs generate LAK, proliferate, and secrete secondary cytokines in response to IL-2. We assessed the roles of the various receptor complexes in these responses to IL-2 using antibodies that block the interaction of IL-2 with the intermediate-affinity IL-2R (humanized Mik beta 1) and high- and low-affinity interactions (HTac). The HTac antibody produced dramatic reductions in proliferation (27%) and in the secretion of cytokines (TNF)-alpha and -beta and IFN-gamma inhibited 68%, 72%, and 80%, respectively) with no decrease in LAK.

Therapeutic effect of a retroviral wild-type p53 expression vector in an orthotopic lung cancer model.

Background: Mutations in the p53 tumor suppressor gene (also known as TP53) are common in human lung cancers. The wild-type form of p53 is dominant over the mutant; thus, restoration of wild-type p53 function in lung cancer cells may suppress their growth as tumors.

Purpose: We investigated the therapeutic efficacy of direct administration of a retroviral wild-type p53 (wt-p53) expression vector (LNp53B) in an orthotopic human lung cancer model in nu/nu mice.

Mechanism of the anti-tumour effect of biochemotherapy in melanoma: preliminary results.

During the conduct of a biochemotherapy trial in which cisplatin, vinblastine and dacarbazine (CVD) were administered concurrently with interleukin-2 (IL-2) plus interferon-alpha 2a (IFN-alpha 2a) (biochemotherapy) in advanced melanoma, we performed a series of laboratory studies in an attempt to understand better the mechanism of anti-tumour effect of the regimen. We initially hypothesized that CVD enhanced the anti-tumour effect of the biotherapy. However, in the first 10 patients studied, of whom eight were responders, we observed no lymphokine-associated killer cell (LAK) and minimal natural killer (NK) cell activities.

Human papillomavirus in laryngeal and hypopharyngeal carcinomas. Relationship to survival.

Objective: To determine the relationship between human papillomavirus (HPV) DNA detection in upper aerodigestive tract malignancies and patient outcome.

Design: Archival paraffin-embedded specimens from 78 previously untreated patients with squamous carcinomas of the larynx and hypopharynx were pathologically verified and analyzed by polymerase chain reaction for detection of HPV DNA. Charts were independently reviewed and coded until final analysis.

Induction of chemosensitivity in human lung cancer cells in vivo by adenovirus-mediated transfer of the wild-type p53 gene.

Recombinant adenovirus-mediated transfer of the wild-type p53 gene into monolayer cultures or multicellular tumor spheroids of human non-small cell lung cancer cell line H358, which has a homozygous deletion of p53, markedly increased the cellular sensitivity of these cells to the chemotherapeutic drug cisplatin. Treated cells underwent apoptosis with specific DNA fragmentation. Direct injection of the p53-adenovirus construct into H358 tumors s.

Induction of lymphokine-activated killing with reduced secretion of interleukin-1 beta, tumor necrosis factor-alpha, and interferon-gamma by interleukin-2 analogs.

Background: Interleukin-2 (IL-2)-based immunotherapy has been shown to effect clinical responses in 15-35% of patients with metastatic renal cell carcinoma or melanoma. Despite its clinical efficacy, many clinicians refrain from using IL-2 because of the associated toxicity. This toxicity is believed to be mediated by such secondary cytokines as IL-1, tumor necrosis factor (TNF), and interferon (IFN)-gamma, which are produced by the patient's IL-2-stimulated peripheral blood mononuclear cells (PBMCs).

Sequential TNF and TGF-beta regulation of expansion and induction of cytotoxicity in long-term cultures of lymphokine-activated killer cells.

Therapeutic use of lymphokine-activated killer (LAK) cells frequently requires higher numbers of effector cells than can be produced in short-term cultures. Extended stimulation of these cells by interleukin-2 (IL-2) past 2 weeks leads to a decrease in cytolytic activity and cell proliferation. To determine how known regulatory cytokines are involved in the mechanism responsible for loss lf IL-2 responsiveness, adherent LAK (A-LAK), nonadherent LAK (NA-LAK), monocyte-depleted (MD-LAK), and unmanipulated LAK (UN-LAK) cells derived from human peripheral blood were stimulated with high-dose IL-2 for 4 weeks, and cytokine production was measured serially.

Retroviral-mediated transduction of p53 gene increases TGF-beta expression in a human glioblastoma cell line.

Transforming growth factor-beta (TGF-beta) has been implicated as a potent growth regulator; the degree of responses to it, whether positive or negative, generally correlates with the stage of cell differentiation in various cell types. We examined the effect of the p53 gene, which participates in the control of cell-cycle progression, on the expression of human TGF-beta. The human glioblastoma cell line SNB-19, which expresses the latent form of TGF-beta, was transfected with a retroviral vector containing wild-type p53 (wt-p53) or p53 with a mutation (mut-p53) at codon 273.

Gene therapeutics and gene therapy for cancer.

Recent advances in molecular biology have opened new avenues of basic genetic engineering technology and have made possible the application of this technology in clinical human gene therapy. Replication-defective viral vectors and biocompatible materials, eg, liposomes, have been developed as vehicles to introduce potentially therapeutic genes into mammalian cells. Over the past 2 years, this technology has increased the possibilities for therapy in numerous genetic diseases.

A retroviral wild-type p53 expression vector penetrates human lung cancer spheroids and inhibits growth by inducing apoptosis.

Multicellular tumor spheroids approximate the three-dimensional configuration of primary and metastatic tumors. The effects of retrovirus-mediated transduction of wild-type p53 (wt-p53) were studied on multicellular tumor spheroids of human non-small cell lung cancer cell lines H322a, the p53 gene of which is homozygously mutated at codon 248, and WT226b, which has endogenous wt-p53. The growth of WT226b spheroids was not affected by exogenous wt-p53 transduction; the growth of H322a spheroids, however, was significantly inhibited by the addition of wt-p53 virus stocks.

Cytokine combinations in immunotherapy for solid tumors: a review.

The use of cytokines alone or in combination with other cytokines or cytotoxic drugs has had a profound effect upon widely metastatic disease in many cases. However, despite the encouraging results in early trials, there is much room for improvement. Few responses to these combinations are complete, and toxicity has in some cases been quite severe.