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T L Whiteside

Publications and source records attributed to T L Whiteside.

At least 19 recordsLinked to original sources
PubMed2003

Role of prolactin receptor and CD25 in protection of circulating T lymphocytes from apoptosis in patients with breast cancer.

Prolactin (PRL) has been reported to inhibit apoptosis in various cell types and to serve as a cofactor in the upregulation of CD25 on T cells during activation. We investigated a possible relation between prolactin receptor (PRL-R) or IL-2 receptor alpha (IL-2Ralpha, CD25) expression on circulating T lymphocytes and their apoptosis in patients with breast cancer. Peripheral blood mononuclear cells obtained from 25 patients, 25 normal controls (NC) and three cord blood samples were evaluated for Annexin V binding and expression of CD95, CD25, and PRL-R on CD3(+) T cells by multicolour flow cytometry. Plasma levels of PRL, sCD95L, and sIL-2R were determined in patients and controls and related to T-cell apoptosis. The ability of PRL to protect T cells from apoptosis induced by various agents was also studied. Expression of PRL-R on the surface of T cells was comparable in patients with breast cancer and NC, but PRL plasma levels in patients were significantly lower (P<0.05). In patients, 18+/-11% (mean+/-s.d.) of CD3(+) cells bound Annexin V, compared to 9+/-6% in NC (P<0.0004). Percentages of CD3(+)Fas(+) and CD3(+)CD25(+) cells were higher in the peripheral circulation of patients than NC (P<0.0001 and <0.04, respectively). Levels of sFasL were lowest in plasma of the patients with the highest proportions of CD3(+)Fas(+) T cells. Most T cells undergoing apoptosis were CD3(+)CD25(-) in patients, and the proportion of CD3(+)CD25(-) Annexin V(+) cells was significantly increased in patients compared to NC (P<0.006). Ex vivo PRL protected T cells from starvation-induced or anti-CD3Ab-induced but not from Fas/FasL-dependent apoptosis. These results indicate that expression of CD25 but not of PRL-R on the surface of activated T lymphocytes appears to be involved in modulating Fas/Fas - ligand interactions, which are, in part, responsible for apoptosis of T lymphocytes and excessive turnover of immune cells in the circulation of patients with breast cancer.

Aged↗
PubMed2003

Effector CD8+CD45RO-CD27-T cells have signalling defects in patients with squamous cell carcinoma of the head and neck.

A subset of circulating T cells (CD8(+)CD45RO(-)CD27(-)) with a naïve phenotype, but mediating effector function, is considered to play an important role in host antitumour defence. To investigate the attributes of these effector T cells in patients with squamous cell carcinoma (SCC) of the head and neck cancer, venous blood was obtained from 39 individuals with cancer and 45 normal controls (NC). Peripheral blood mononuclear cells were isolated, stained with labelled monoclonal antibodies specific for CD8, CD45RO, CD45RA, CD62L, CD27, TCR-zeta as well as isotype controls and examined by multicolour flow cytometry. Annexin V binding to CD8(+) T cells and PMA/ionomycin-induced IFN-gamma expression were also evaluated in patients and NC. The proportions of CD45RA(+)CD45RO(-) (naïve) and CD45RA(-)CD45RO(+) (memory) cells were found to be comparable within the CD8(+) T-cell subset. However, relative to NC, the frequency of effector CD8(+)CD45RO(-)CD27(-) cells was strikingly increased in all SCC patients regardless of the disease status (P=0.0003). The proportion of these cells was found to increase with age in both patients and NC. In NC, stimulated IFN-gamma expression was largely restricted to CD8(+)CD45RO(-)CD27(+) cells, while in patients CD8(+)CD45RO(-)CD27(-) expressed IFN-gamma after ex vivo stimulation. Expression of the TCR-associated zeta chain was decreased or absent in freshly isolated CD8(+)CD45RO(-)CD27(-) T cells in patients (P<0.0001). Annexin V was found to bind to a higher proportion of circulating CD8(+) T cells in patients than NC (P<0.006), and significantly more Annexin V(+) T cells were present in the effector (P<0.0059) than the naïve subset within the CD8(+)CD45RO(-) compartment. The data indicate that the expanded CD8(+)CD45RO(-)CD27(-) T cells, which contain precursors of IFN-gamma-producing T cells, are zeta-negative and sensitive to apoptosis in the circulation of patients with HNC.

Adult↗
PubMed2002

Recovery of zeta-chain expression and changes in spontaneous IL-10 production after PSA-based vaccines in patients with prostate cancer.

Circulating T lymphocytes of patients with prostate cancer have been reported to have functional deficits, including low or absent zeta-chain expression. To determine whether these functional impairments could be reversed by prostate specific antigen-based vaccination therapy, 10 patients treated with recombinant human prostate specific antigen plus GM-CSF and eight others receiving prostate specific antigen plus oil emulsion in two pilot clinical trials were evaluated prior to and after vaccination for several immunologic end points, including zeta-chain expression and cytokine production by circulating T cells as well as the frequency of T cells able to respond to prostate specific antigen in ELISPOT assays. The flow cytometry assay for zeta-chain expression was standardized to allow for a reliable comparison of pre- vs post-vaccination samples. Prior to therapy, the patients were found to have significantly lower zeta-chain expression in circulating CD3(+) cells and a higher percentage of zeta-chain negative CD3(+) and CD4(+) cells than normal donors. The patients' peripheral blood mononuclear cells spontaneously produced more IL-10 ex vivo than those of normal controls. After vaccination, recovery of zeta-chain expression was observed in 50% of patients in both clinical trials. Also, spontaneous IL-10 secretion by peripheral blood mononuclear cells decreased following immunotherapy in patients treated with prostate specific antigen and GM-CSF. The frequency of prostate specific antigen-reactive T cells was detectable in 7 out of 18 patients vs 4 out of 18 patients prior to vaccination. Only one of 18 patients was a clinical responder. The vaccine had stimulatory effects on the patients' immune system, but post-vaccine immune recovery could not be correlated to progression-free survival in this small cohort of patients with prostate cancer.

Aged↗
PubMed2001

Identification of cyclin B1 as a shared human epithelial tumor-associated antigen recognized by T cells.

We eluted peptides from class I molecules of HLA-A2.1(+) breast adenocarcinoma and loaded reverse phase high-performance liquid chromatography (HPLC) fractions onto dendritic cells to prime naive CD8(+) T cells. Fractions that supported growth of tumor-specific cytotoxic T lymphocytes were analyzed by nano-HPLC micro-ESI tandem mass spectrometry. Six HLA-A2.1-binding peptides, four 9-mers (P1-P4) differing in the COOH-terminal residue, and two 10-mers (P5 and P6) with an additional COOH-terminal alanine, were identified in one fraction. Peptide sequences were homologous to cyclin B1. We primed CD8(+) T cells from another HLA-A2.1(+) healthy donor with synthetic peptides and generated P4-specific responses. We also detected memory T cells specific for one or more of these peptides in patients with breast cancer and squamous cell carcinomas of the head and neck (SCCHN). T cells from one patient, restimulated once in vitro, could kill the tumor cell line from which the peptides were derived. Immunohistochemical analysis of tumor lines and tissue sections showed cyclin B1 overexpression and aberrant localization in the cytoplasm instead of the nucleus. Sequencing genomic DNA and cDNA corresponding to P1-P6 region showed that differences in COOH-terminal residues were not due to either DNA mutations or errors in transcription, suggesting a high error rate in translation of cyclin B1 protein in tumors.

Antigens, Neoplasm↗
PubMed2001

Modulation of TcR/CD3-zeta chain expression by a circulating factor derived from ovarian cancer patients.

In women with ovarian cancer, suppression of components of the immune system may promote tumour development. Previous studies in ovarian cancer have demonstrated that decreased expression and function of the T-cell receptor (TcR)-associated signal transducing zeta-chain correlates with deficient immune responsiveness of T cells. In this study, sera and ascitic fluids obtained from woman with advanced ovarian cancer were found to suppress the expression of TcR-associated zeta chain. This suppression of zeta chain expression was dose-dependent and was not observed with biologic fluids obtained from healthy women. The factor responsible for the loss of zeta chain was purified from ascites and characterized as a protein with an appropriate molecular weight of 14 kD. Suppression of T-cell TcR-zeta was specific, since neither lck nor ZAP-70 expression was affected, while zeta chain was almost completely suppressed. This selective suppression of TcR-zeta expression by the 14 kD ascites-derived factor was shown to operate at the mRNA level. By defining the mechanism through which this protein modulates TcR-zeta chain levels, it might be possible to ultimately prevent the suppressive influences of the tumour microenvironment and restor immune competence in patients with ovarian carcinoma.

Adult↗
PubMed2001

The number of intratumoral dendritic cells and zeta-chain expression in T cells as prognostic and survival biomarkers in patients with oral carcinoma.

BACKGROUND: Dendritic cells (DCs) are antigen-presenting cells with a unique ability to cross prime T cells and generate strong antitumor responses. This study evaluates the presence and prognostic significance of DCs as well as functional T cells, which accumulate in the microenvironment in patients with oral squamous cell carcinoma (OSCC). METHODS: Immunohistochemistry for S-100 positive or p55 positive DCs and for T-cell receptor (TcR)-associated zeta-chain expression in tumor-infiltrating lymphocytes (TILs) was performed in 132 paraffin embedded specimens from patients with primary OSCC. The median clinical follow-up for the patients was 50 months. The numbers of intratumoral DCs or TILs expressing the zeta chain were determined microscopically and compared with clinical and pathohistologic prognostic parameters, including disease stage, T stage or tumor grade, lymph node involvement, as well as disease free survival and overall survival. RESULTS: Immunostaining identified DCs in the epithelial compartment of the tumors (S-100 positive) as well as interdigitating reticular DCs (p55 positive) in peritumoral areas. Based on S-100 staining, intratumoral DC infiltrates were low (<10 DCs per high-power field [HPF]) in 20% of OSCC specimens, intermediate (10-20 DCs per HPF) in 42% of OSCC specimens, and high (>20 DCs per HPF) in 37% of OSCC specimens. The number of S-100 positive DCs was positively and significantly correlated with that of p55 and of TILs with normal zeta-chain expression. A low number of infiltrating S-100 positive DCs was more predictive of poor survival (hazard ratio, 7.95) than lymph node involvement (hazard ratio, 3.36) or late T stage (hazard ratio, 2.92). A significant but weaker association of p55 positive DC infiltration with survival was observed. Low density of DCs and low or absent expression of the zeta chain in TILs correlated with each other and predicted the poorest survival and the greatest risk. CONCLUSIONS: The number of DCs infiltrating the tumor is a highly significant prognostic parameter in patients with OSCC. Furthermore, the absence or paucity of DCs is strongly linked to abnormalities in the TcR-associated zeta chain in TILs. The two biomarkers, zeta-chain expression in TILs and the number of S-100+ DCs in the tumor, independently predict overall survival, disease free survival, and time to disease recurrence in patients with OSCC.

Adult↗
PubMed2001

Immunobiology and immunotherapy of head and neck cancer.

The development of head and neck cancer (HNC) is strongly influenced by the host immune system. Immunoselection of tumors resistant to immune attack and the ability of established tumors to disarm or eliminate immune cells favor tumor progression. Recent evidence for local as well as systemic apoptosis of T lymphocytes, the paucity of dendritic cells (DC) at the tumor site, or the presence of signaling defects in T lymphocytes of patients with HNC emphasizes the fact that their antitumor responses are compromised. The clinical and biologic importance of these immune biomarkers is revealed by the finding that they appear to independently predict 5-year survival in patients with oral carcinoma. Whereas the mechanisms responsible for immune dysfunction in HNC are being investigated, new immunotherapeutic strategies, including antitumor vaccines and DC-based interventions, aim at the restoration of tumor-targeted immune responses. These novel biologic therapies, alone or in combination with conventional therapies, might be expected to protect immune cells from dysfunction or death and to enhance their antitumor activity.

Adjuvants, Immunologic↗
PubMed2001

Therapeutic activity of NK cells against tumors.

While it is generally accepted that natural killer (NK) cells, by killing tumor cells in the circulation, represent a first line of defense against metastases, their therapeutic activity against established tumors has been limited. In this review, we describe studies to improve the therapeutic effectiveness of activated NK cells in both animal models and clinical trials to better understand the biological problems that limit their effectiveness.

Adoptive Transfer↗
PubMed2001

Decreased zeta chain expression and apoptosis in CD3+ peripheral blood T lymphocytes of patients with melanoma.

Expression of T-cell receptor- or Fcgamma receptor III-associated signal-transducing zeta chain is important for the functional integrity of immune cells. We found that significantly higher proportions of circulating CD3+ T cells as well as natural killer cells had low or absent expression of the zeta chain in patients with advanced melanoma than in normal donors (P < 0.0005). Decreased zeta expression was always observed in a small subset of circulating CD3+ T cells that were in the process of apoptosis, i.e., bound Annexin V or were terminal deoxynucleotidyl transferase-mediated nick end labeling positive. Up to 80% of T cells in the peripheral blood of patients with melanoma were Fas+, with the mean percentage of Fas+CD3+ cells significantly higher in patients (P < 0.004) than normal controls. These Fas+CD3+ T cells were found to preferentially undergo apoptosis. Annexin V binding, the loss of Fas expression from the cell surface as well as zeta down-regulation, which are associated with early apoptosis, were detected in a proportion of circulating Fas+CD3+. In Jurkat cells incubated with agonistic anti-Fas antibody (CH-11), a rapid loss of Fas expression from the cell surface coincided with Annexin V binding and preceded the loss of zeta chain during early apoptosis. In a subset of Jurkat cells coincubated with human melanoma cells, Annexin V binding and zeta degradation as well as DNA fragmentation were observed, indicating that the tumor induced T-cell death. Triggering of death receptors expressed on activated T lymphocytes was accompanied by the loss of zeta expression. On the other hand, soluble factors secreted by melanoma cells induced down-regulation but no apoptosis in activated normal T cells. In the circulation of patients with melanoma, apoptosis of immune effector cells may be related to the state of chronic activation, resulting in the up-regulation of death receptors and increased susceptibility to apoptosis.

Annexin A5↗
PubMed2000

Competition of peptide-MHC class I tetrameric complexes with anti-CD3 provides evidence for specificity of peptide binding to the TCR complex.

BACKGROUND: Major histocompatibility complex (MHC)-peptide tetrameric complexes (tetramers) are valuable tools for detecting and characterizing peptide-specific T cells. Because the frequency of these cells is generally very low, it may be difficult to discriminate between nonspecific and specific tetramer binding. METHODS: A four-color flow cytometric assay that simultaneously measures tetramer, CD3, CD8, and CD14 was used to investigate the sensitivity and specificity of MHC class I tetramer staining. This was accomplished by using the influenza virus matrix protein peptide, GILGFVFTL (FLU), as a model recall antigen and the human immunodeficiency virus (HIV) reverse transcriptase peptide, ILKEPVHGV (HIV), as a model novel antigen. Peripheral blood mononuclear cells (PBMC) from 31 HLA-A2.1(+) and 10 HLA-A2.1(-) healthy individuals were stained with the tetramers. RESULTS: The lower limit of detection was established at approximately 1/8,000. In HLA-A2(+) PMBC, frequencies of tetramer-positive CD8(+) T cells were log normally distributed and were high for FLU (1/910) but low for HIV (1/6,067). A novel competition assay, in which tetramer binding was shown to diminish subsequent staining with anti-CD3 antibody, was used to confirm the specificity of tetramer binding to the T-cell receptor (TCR) complex. The competition assay was validated by evaluating several anti-CD3 antibodies and showing that in PBMC from HLA-A2(-) subjects, spurious tetramer-positive events (1/20,000) failed to compete with CD3 binding. For the "recall" FLU tetramer, the degree of competition was proportional to the frequency, suggesting a selection of high avidity cells. Although CD3 competition was also highly correlated with the intensity of tetramer staining, competition allowed the identification of false positive cases with relatively high tetramer staining intensity. CONCLUSION: The data indicate that competition of CD3 binding allows confirmation of the specificity of tetramer binding to the TCR, extending the usefulness of tetramers in the frequency analysis of peptide-specific T lymphocytes.

Amino Acid Sequence↗
PubMed2000

Generation of T cells specific for the wild-type sequence p53(264-272) peptide in cancer patients: implications for immunoselection of epitope loss variants.

Alterations in the p53 gene occur frequently and can lead to accumulation of p53 protein in squamous cell carcinomas of the head and neck (SCCHN). Since accumulation of p53 is associated with enhanced presentation of wild-type sequence (wt) p53 peptides to immune cells, the development of pan vaccines against SCCHN has focused on wt p53 epitopes. We used the HLA-A2.1-restricted wt p53(264-272) epitope to generate CTL from circulating precursor T cells of HLA-A2.1(+) healthy donors and patients with SCCHN. Autologous peptide-pulsed dendritic cells were used for in vitro sensitization. CTL specific for the wt p53(264-272) peptide were generated from PBMC obtained from two of seven normal donors and three of seven patients with SCCHN. These CTL were HLA class I restricted and responded to T2 cells pulsed with p53(264-272) peptide as well as HLA-A2-matched SCCHN cell lines naturally presenting the epitope. Paradoxically, none of the tumors in the three patients who generated CTL could adequately present the epitope; two had a wt p53 genotype and no p53 protein accumulation, while the third tumor expressed a point mutation (R to H) in codon 273 that prevents presentation of the p53(264-272) epitope. In contrast, patients who did not generate CTL had tumors that accumulated altered p53 and potentially could present the p53(264-272) epitope. These findings suggest that in vivo, CTL specific for the wt p53(264-272) peptide might play a role in the elimination of tumor cells expressing this epitope and in immunoselection of epitope-loss tumor cells. Immunoselection of tumors that become resistant to anti-p53 immune responses has important implications for future p53-based vaccination strategies.

Autoantibodies↗
PubMed2000

Generation of tumor-specific T-lymphocytes by cross-priming with human dendritic cells ingesting apoptotic tumor cells.

It has been suggested that dendritic cells (DCs) are capable of ingesting apoptotic tumor cells (ATCs) and presenting tumor-associated antigens to immune cells. We evaluated the potential of human DCs, which have ingested ATCs, to serve as a source of antigenic epitopes for presentation to T cells specific for PCI-13, a squamous cell carcinoma of the head and neck cell line. Immature DCs (DCimm) generated in the presence of interleukin 4 and granulocyte machrophage colony-stimulating factor from peripheral blood monocytes of HLA-A2+ healthy donors were incubated in the presence of ATCs. Uptake of ATCs by DCs was monitored by flow cytometry and confocal microscopy after 2-18 h of coincubation. When DCs were matured (DCmat) in the presence of proinflammatory cytokines, their capacity to uptake ATCs was reduced. Responses of PCI-13-specific CD8+ T cells to unmodified PCI-13 cells and to DCimm or DCmat +/- ATCs or +/- tumor lysates were tested in gamma-IFN enzyme-linked immunospot and cytotoxicity assays. Unmodified tumor cells were found to be the best stimulators of antitumor activity of the established T-cell line, and ATCs alone were minimally stimulatory. However, DCs that ingested ATCs were able to present tumor antigens to CTLs, and DCimm and DCmat were almost equally stimulatory. When DCs plus various tumor-derived preparations were used as antigen-presenting cells with autologous HLA-A2+ T cells obtained from normal donors, DCs that had ingested ATCs were more effective in generating CD8+ CTLs than tumor cells alone or DCs pulsed with tumor lysates. The results indicate that human DCs fed with ATCs and then matured effectively generated T cell-mediated antitumor responses in vitro.

Antigens, CD↗
PubMed2000

Analysis of therapeutic and immunologic effects of R(24) anti-GD3 monoclonal antibody in 37 patients with metastatic melanoma.

BACKGROUND: Antitumor effects of antibodies against ganglioside antigens of melanoma have been reported, but neither optimal doses nor mechanisms have been established. METHODS: This Phase IB trial of the murine immunoglobulin IgG(3) monoclonal antibody R(24) against disialoganglioside GD3 was conducted with 37 patients to define better the dose-response relation and mechanism of action of R(24) in patients with metastatic melanoma. RESULTS: Dose-limiting toxicity consisted of a pulmonary capillary leak syndrome in 3 of 5 patients in the 80 mg/M(2)/day dosage tier. Serial blood and tumor biopsy samples were obtained prior to therapy and on Days 5, 9, and 22 following R(24) infusion. Tumor biopsy-infiltrating lymphocytes were enumerated in peritumoral, endotumoral, and perivascular compartments: endotumoral CD4(+) and CD8(+) T cells and HLA-DR(+) T cells increased over time on R(24) antibody. Endotumoral CD4 lymphoid infiltrate activation (DR expression) and antibody-dependent cytotoxicity were the greatest in the one patient who achieved a complete response. CONCLUSIONS: Clinical response was associated with depression in natural killer (CD56(+) and CD56(+)DR(+)) blood cells (P = 0.03) and was associated with R(24) dosage (P = 0.01). A complete response that lasted 2 years and a partial response that lasted 2 months occurred at a dose of 1 mg/M(2)/day. The limited number of clinical responses observed in this trial hampered the correlation of antitumor and immune parameters but provided a rational foundation for the future evaluation of antiganglioside antibodies.

Adult↗
PubMed2000

Generation of PSA-reactive effector cells after vaccination with a PSA-based vaccine in patients with prostate cancer.

BACKGROUND: JBT 1001 is a vaccine used for therapy of prostate cancer (CA), which consists of recombinant prostate-specific antigen (PSA) with lipid A formulated in liposomes. Patients with prostate CA were vaccinated with JBT 1001 emulsified in mineral oil (n = 5) or with the vaccine in combination with granulocyte-macrophage colony-stimulating factor (GM-CSF) administered locally at the site of vaccination (n = 5). Frequency of PSA-reactive T cells was measured in peripheral blood mononuclear cells (PBMC) before and after immunization, using an interferon-gamma (IFN-gamma) enzyme-linked immunospot (ELISPOT) assay with autologous dendritic cells (DC) as antigen-presenting cells. The hypothesis tested was that PSA-based vaccines induce T cell responses to human PSA. METHODS: In order to expand precursor cells, in vitro sensitization (IVS) was performed. Microcultures of peripheral blood lymphocytes (PBL) (1 x 10(5)/well) in medium supplemented with interleukin-2 (IL-2) (10 IU/ml) and interleukin-7 (IL-7) (10 ng/ml) were stimulated twice (day 0 and day 7) with monocyte-derived autologous DC, generated by culture with interleukin-4 (IL-4) and GM-CSF and pulsed with PSA (10 microg/ml) at an effector to stimulator ratio of 10:1. ELISPOT assays were performed on day 14 of culture. In addition, PBMC were separated on immunobeads into CD4(+) and CD8(+) subsets for ELISPOT assays performed without IVS. RESULTS: Two patients had PSA-reactive responses before vaccination (frequency range, 1/700-1/4,400). After vaccination, 8/10 patients had measurable PSA-reactive T-cell frequencies, ranging from 1/200-1/1900, using IVS. In contrast, without IVS, but after immunoselection to enrich in CD8(+) and CD4(+) T cells, only 2/10 patients had detectable PSA-reactive T cells after vaccination, at a frequency ranging from 1/2,600-1/4,000. CONCLUSIONS: Vaccination with PSA formulated into liposomes induced T-cell responses in 8/10 patients with prostate carcinoma. The frequency of PSA-reactive precursor T cells was relatively low in the blood of these patients, and IVS, leading to amplification of the precursor cells prior to ELISPOT, was necessary for quantification of the PSA-responding T cells. Cellular responses to PSA were predominantly mediated by CD4(+) T lymphocytes.

Aged↗
PubMed2000

B7.1 on human carcinomas: costimulation of T cells and enhanced tumor-induced T-cell death.

Human squamous cell carcinomas of the head and neck (SCCHN) do not express the costimulatory molecules B7.1 or B7.2 in situ or in culture. Transduction of B7.1(-) SCCHN cells with the retroviral B7. 1 and neo(r) genes resulted in the expression of high levels of the transgene in these tumor cells. When B7.1(+) SCCHN cells were used as stimulators of autologous or allogeneic PBL in mixed lymphocyte-tumor cultures (MLTC), T-cell proliferation and generation of antitumor effector T cells as well as levels of their lytic activity were significantly increased. At the same time, a proportion of activated T cells seen to undergo apoptosis was found to be significantly higher upon coincubation with B7.1(+) SCCHN than with B7.1(-) SCCHN. Both B7.1(+) and B7.1(-) SCCHN cells were found to express FasL on the cell surface and in the cytoplasm, as well as mRNA for FasL and mRNA for TRAIL. However, expression of the B7.1 transgene did not lead to increased expression of FasL protein on tumor cells. Yet, up to 50% of activated CD28(+) allogeneic T cells, which were CD95(+), showed evidence of DNA fragmentation in JAM and TUNEL assays upon incubation with an excess of B7.1(+) SCCHN for 24 h. Tumor-induced T-cell death was equally and only in part blocked by anti-Fas antibodies in both B7.1(+) and B7.1(-) MLTC. While surface expression of B7.1 molecules on SCCHN cells enhanced T-cell costimulation via B7.1-CD28 interactions, it did not rescue activated T cells from tumor-induced apoptosis. The outcome of MLTC under these conditions was dependent on the ratio of tumor to T cells. Thus, in the presence of an excess of B7.1(+) tumor cells, activated T cells showed increased sensitivity to apoptosis which did not appear to be Fas/FasL mediated. These data are important for the development of B7.1 gene therapy and efforts directed at the generation of effector cells in MLTC.

Apoptosis↗
PubMed2000

Tumor-induced apoptosis of T lymphocytes: elucidation of intracellular apoptotic events.

Our recent studies suggest that human squamous cell carcinoma of the head and neck (SCCHN) is capable of activating an intrinsic mechanism of programmed-cell death in interacting lymphocytes in situ and in vitro. The current study used Jurkat T-cell line as a model to investigate intracellular apoptotic events in T cells interacting with SCCHN. Apoptosis induced in T lymphocytes by tumor cells was in part Fas-mediated, since it was partially, but significantly, inhibited in the presence of anti-Fas ligand Ab or in Fas-resistant Jurkat cells. The synthetic caspase inhibitors, N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (Z-VAD-FMK) and N-benzyloxycarbonyl-Asp-glu-Val-Asp-fluoromethyl ketone (Z-DEVD-FMK), effectively blocked apoptosis of Jurkat cells co-incubated with SCCHN cell lines, suggesting the involvement of caspases in tumor-induced apoptosis of lymphocytes. Overexpression of CrmA, an inhibitor of caspase-1 and caspase-8, partially inhibited tumor-induced T-cell death. Caspase-8 and caspase-3 were identified as effector molecules in the execution of tumor-induced T-cell death, since the proform enzymes were processed into active subunits during co-incubation of T cells with tumor cells. Furthermore, co-incubation with tumor cells resulted in cleavage of poly(ADP-ribose) polymerase (PARP), a common caspase-3 substrate, and in cleavage of TcR-zeta chain, shown by us to be a T-cell specific caspase-3 substrate. Overexpression of Bcl-2 did not provide protection of T cells from SCCHN-induced DNA degradation. Instead, the Bcl-2 protein was cleaved in the target T cells during their co-incubation with tumor cells. These findings demonstrate that tumor cells can trigger in T lymphocytes caspase-dependent apoptotic cascades, which are not effectively protected by Bcl-2. (Blood. 2000;95:2015-2023)

Apoptosis↗
PubMed2000

Identification of a novel tumor necrosis factor-alpha-inducible gene, SCC-S2, containing the consensus sequence of a death effector domain of fas-associated death domain-like interleukin- 1beta-converting enzyme-inhibitory protein.

We report here the isolation and characterization of a novel tumor necrosis factor-alpha (TNF-alpha)-inducible gene, SCC-S2. Based on the nucleotide sequence, the SCC-S2 open reading frame contains a sequence in the amino terminus that shows a significant homology to death effector domain II of cell death regulatory protein, Fas-associated death domain-like interleukin-1beta-converting enzyme-inhibitory protein (FLIP). Unlike FLIP, the SCC-S2 open reading frame contains only one death effector domain and lacks the carboxyl-terminal caspase-like homology domain, raising the possibility that SCC-S2 may be a novel member of the FLIP family. SCC-S2 mRNA expression is found in most normal tissues and malignant cells. The steady state level of SCC-S2 mRNA is significantly induced by TNF-alpha in different tumor cells (TNF-alpha at 20 ng/ml for 3 h: A549, approximately 2-9-fold; SKOV-3, approximately 3-fold; PCI-04A, approximately 3-6-fold). TNF-alpha treatment (100 ng/ml, 4 h) of HeLa cells transiently transfected with FLAG epitope-tagged SCC-S2 cDNA or expression vector alone led to an increase in the number of apoptotic cells as compared with the untreated counterpart. Interestingly, however, SCC-S2 transfectants revealed a significant decrease in the number of apoptotic cells as compared with the vector transfectants (p < 0.001). These data implicate a role of SCC-S2 as a negative mediator of apoptosis in certain cell types.

Adaptor Proteins, Signal Transducing↗