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T Hercend

Publications and source records attributed to T Hercend.

At least 37 records · Page 2Linked to original sources
PubMed1994

Evidence for T-cell clonal expansion in a patient with squamous cell carcinoma of the head and neck.

Squamous cell carcinomas of the head and neck (SCCHN) often contain a large mononuclear cell infiltrate, composed mainly of T-lymphocytes which could reflect an in situ immune reaction against the malignant SCCHN cells. We analyzed the molecular structure of the T-cell receptor (TCR) alpha and beta chains expressed by lymphocytes in the tumor, peripheral blood (PBMC), and in the peritumoral tissue in six cases of localized SCCHN. We first determined V alpha and V beta gene segment subfamily usage by polymerase chain reaction using a panel of V subfamily-specific oligonucleotide primers (V alpha 1-w29 and V beta 1-w24). An apparently unrestricted usage of V alpha or V beta gene segment subfamilies was observed for all three samples from the six cases examined. No major difference was observed in T-cell receptor repertoire expression of PBMC between SCCHN and healthy donors, making unlikely the expression of putative tumor-related superantigen(s) in these patients. Intersample comparisons for a given V alpha or V beta T-cell receptor specificity revealed some differences in V gene segment usage in tumor-infiltrating lymphocytes versus PBMC. A detailed analysis of these V segment subfamily specificities (cloning followed by sequencing and CDR3 size distribution analysis) in one patient revealed the presence of recurrent T-cell receptor transcripts (i.e., identical V-N-J sequences) in the tumor (e.g., 40%) and in PBMC (e.g., 75%). These results show that unique T-cell subpopulations are clonally amplified in SCCHN patients, possibly as a consequence of antigen-driven selection.

Adult↗
PubMed1994

Lymphocyte-activation gene 3/major histocompatibility complex class II interaction modulates the antigenic response of CD4+ T lymphocytes.

The activation requirements for antigen-dependent proliferation of CD4+ T cells are well documented, while the events leading to the inactivation phase are poorly understood. Here, we tested the hypothesis that the lymphocyte-activation gene 3 (LAG-3), a second major histocompatibility complex (MHC) class II ligand, plays a regulatory role in CD4+ T lymphocyte activation. CD4+ class II-restricted T cell clones were stimulated by their relevant antigen (hemagglutinin peptide or diphteria toxoid) and antigen-presenting cells with or without anti-LAG-3 monoclonal antibody (mAb). Kinetic studies were performed to monitor different activation parameters, including the measurement of thymidine incorporation, expression of activation antigens and cytokine secretion. Results showed that the time course from the initial time points up to the peak time point was not modified in the presence of anti-LAG-3 mAb. However, addition of these antibodies, either as whole IgG or as Fab fragments, led to increased thymidine incorporation values for late time points and, hence, to a shift in the decreasing proliferation curve. We also showed that expression of activation antigens, such as CD25, was higher in the presence of anti-LAG-3 mAb, and that cytokine concentrations, i.e. of interferon-gamma or interleukin-4, were higher in the corresponding culture supernatants. In addition, we tested whether the effects of anti-LAG-3 mAb were limited to antigen-dependent, MHC class II-restricted responses. The proliferative responses of CD4+ T cell clones following stimulation with either interleukin-2, mitogens, a combination of anti-CD2 mAb, immobilized anti-CD3 or anti-T cell receptor mAb were not altered by anti-LAG-3 mAb. The allogeneic proliferative response of a CD8+ T cell clone was also not affected. Overall, the present analysis reveals a modulating effect of anti-LAG-3 mAb, mediated specifically on antigen-dependent, MHC class II-restricted responses of CD4+ T cell lines. These results support the view that LAG-3/MHC class II interaction down-regulates antigen-dependent stimulation of CD4+ T lymphocytes.

Antigens, CD↗
PubMed1994

TCR gene segments from at least one third of V alpha subfamilies rearrange at the delta locus.

Using PCR and an experimentally validated V alpha subfamily-specific oligonucleotide panel (V alpha 1-w29), we have investigated whether the TCR delta chain may increase its combinatorial diversity by using V genes considered as alpha chain-specific. We show that at least 10 distinct human V alpha segments rearrange at the J delta locus, leading to scrambling of the two V gene repertoires. Fifty-five per cent of the V alpha/J delta transcripts characterized here were in frame. The 17 V alpha/C delta chains analysed included an extended CDR3 region with up to 18 aa encoded by the junctional region. In addition, a new J delta segment (J delta 4) has been characterized. Together, these findings demonstrate that combinatorial diversity in the human delta locus is larger than previously thought.

Amino Acid Sequence↗
PubMed1994

Immunotherapy with interleukin-2 (IL2) and lymphokine-activated natural killer cells: improvement of clinical responses in metastatic renal cell carcinoma patients previously treated with IL2.

Treatment with interleukin-2 (IL2) induces clinical responses in 15-30% of metastatic renal cell carcinoma (MRCC) patients, with mainly partial responses. In order to improve clinical response, we decided to treat partial response patients from a previous IL2 treatment with a second course of IL2 associated with lymphokine-activated natural killer (LANAK) cells. 10 patients who underwent PR after an IL2 protocol (24 x 10(6) U/m2/day, 2 days a week for 5 weeks, either alone or with interferon-gamma) subsequently received a combination of high-dose IL2 (16-20 x 10(6) U/m2/day, 2 days a week) and LANAK cell infusions. Four complete responses were obtained, and 2 additional patients whose tumour mass was further reduced achieved complete response following surgery. These results support the view that initial responses obtained with primary IL2 courses can be improved by complementary treatments. The potential role of cellular immunotherapy and, more particularly, of LANAK cells as an effective procedure to further reduce tumour burden in patients responsive to IL2 will have to be assessed in randomised studies.

Adult↗
PubMed1994

Influence of human leukocyte antigen genes on TCR V gene segment frequencies.

Human leukocyte antigen (HLA)-dependent selection mechanisms exerted during thymic maturation are supposed to be main contributing factors to the genetic predetermination of the TCR repertoire and may have a detectable effect on adult peripheral blood lymphocyte V segment frequencies. Here, we analyzed whether polymorphic or non-polymorphic HLA determinants are associated with selected expression of some V gene segment specificities. We first examined the reactivity of 17 V segment specific mAb on purified CD4+ and CD8+ cell fractions in 10 unrelated people. We found a significant overexpression of only three V segment products (V beta 2, V beta 5.1 and V beta 6.7) in CD4+ and none in CD8+ cell fractions in most individuals. Skewing of certain V beta segments by non-polymorphic HLA determinants (i.e. class II for CD4+ and class I for CD8+ cells) is therefore more limited (3/17) than previously thought. Considering the effects of polymorphic HLA determinants, we compared TCR V segment frequencies in HLA-identical siblings to sibling pairs who differ at one or both HLA haplotypes, using 13 V beta specific mAb. In pairwise comparisons, we found that the HLA complex had no detectable effect on TCR repertoire in five large families with multiple siblings. Together, these observations suggest that HLA-predicted selection mechanisms exerted during thymic maturation might not have a predominant influence shaping the TCR repertoire of normal adults.

Antibodies, Monoclonal↗
PubMed1994

T-cell receptor repertoire in colorectal adenocarcinoma patients with hepatic metastases and its changes induced by preoperative adjuvant interleukin-2 therapy.

The liver is the primary site for colorectal metastases and hepatic resection often fails to cure these patients. Little is known about T-cell immune response in these patients or its changes after interleukin-2 (IL-2) infusion. Using polymerase chain reaction methodology, we investigated T-cell receptor (TCR) V alpha and V beta gene segment subfamily usage in tumor, hepatic tissue, and blood of six patients undergoing hepatic resection and, in addition, in six patients receiving preoperative adjuvant IL-2 therapy (randomized phase I trial). The objectives were (a) to analyze the TCR repertoire in patients undergoing hepatic resection (without IL-2), (b) to analyze the TCR repertoire in patients undergoing hepatic resection after IL-2 therapy, (c) to analyze the effects of preoperative IL-2 infusion on the tumor-infiltrating lymphocyte (TIL) TCR repertoire by comparing TCR V gene segment usage in IL-2-treated versus nontreated patients, and (d) to analyze the effect of IL-2 infusion on the peripheral blood mononuclear cell (PBMC) TCR repertoire by comparing TCR V gene segment expression in pre- versus posttreatment PBMC of IL-2-treated patients. With regard to the first objective, we observed an unrestricted use of V alpha and V beta gene segment subfamily specificities in tumor hepatic tissue and blood of patients undergoing hepatic resection. In addition, we found that some V alpha and V beta specificities were overexpressed in tumor compared with hepatic tissue and blood, suggesting that the corresponding T-cell subpopulations were expanded at the tumor site. In IL-2-treated patients, in whom the tumor and liver were heavily infiltrated by T lymphocytes, the TIL TCR repertoire was also highly diverse and roughly similar to that detected in hepatic tissue and blood, except for a few overexpressions. To analyze the effect of IL-2 on TIL, we next compared the data obtained in IL-2-treated versus nontreated patients. No significant difference between the two groups was observed. Finally, no major changes in the PBMC TCR repertoire were found after IL-2 infusion. Further studies are needed to identify discrete T-cell subpopulations in these patients that may participate in either tumor immune surveillance or active immunotherapy mechanisms triggered by systemic IL-2 infusion.

Adenocarcinoma↗
PubMed1994

Direct evidence to support the immunosurveillance concept in a human regressive melanoma.

The concept of immunosurveillance against cancer has been an extensively debated question over the last decades. Multiple indirect arguments have supported the view that the immune system may control, at least in certain cases, malignant cell growth while direct demonstration is still lacking in the human. In an attempt to address this issue, we have selected a study model, namely spontaneously regressive melanoma. In previous series of experiments, the variability of T cell receptors (TCRs) in the lymphocytes infiltrating a regressive tumor lesion was investigated. Results demonstrated that clonal T cell populations, precisely defined through their V-D-J junctional sequences, were amplified in situ. One clone was predominant, expressing the V beta 16 variable gene segment. A specific anti-V beta 16 TCR mAb was generated here to purify and functionally characterize the corresponding cells. A tumor-infiltrating lymphocyte-derived V beta 16+ T cell line was developed using this reagent. These in vitro cultured cells were found to express the in vivo predominant TCR sequence exclusively and to display an HLA-B14-restricted cytotoxic activity against the autologous tumor cells. Immunohistochemical experiments, performed with the anti-V beta 16 mAb, showed that the corresponding CTLs are present in the tumor area, some of them being closely opposed to the melanoma cells. Together, these studies demonstrate the existence of a local adaptive immune response clinically associated to tumor regression, thus strongly supporting the validity of the immunosurveillance concept in certain human tumors.

Aged↗
PubMed1993

Evidence for in situ amplification of cytotoxic T-lymphocytes with antitumor activity in a human regressive melanoma.

We have derived from lymphocytes infiltrating a human regressive melanoma lesion a series of T-cell receptor alpha/beta-dependent, HLA-B14-restricted cytotoxic T-lymphocyte clones reactive against the autologous tumor. Analysis of the T-cell receptor gene expression revealed that all the clones represented a unique cell expressing a V beta 13.1/J beta 1.1 gene segment. T-cell receptor transcripts expressed in the cloned cells were compared to those present in the uncultured tumor tissue. This analysis demonstrated that the specific cytotoxic T-lymphocyte clones characterized in vitro was actually selected and amplified in vivo at the lesion site. These results provide strong evidence that effector T-cells have contributed to tumor regression.

Aged↗
PubMed1993

Influence of interleukin-2 administration on the expression of T-cell receptor V gene segments in patients with renal-cell carcinoma.

Tumor regression induced by IL-2 in a fraction of patients with metastatic renal-cell carcinoma (MRCC) could not be predicted by immunological monitoring. In addition, the general mechanisms leading to tumor regression or even the distinct cell subsets (e.g., T vs. NK cells) involved are poorly identified. To evaluate the influence of IL-2 administration on circulating T-cell subpopulations, TCR V alpha and V beta gene segment usage was analysed by PCR in 7 MRCC patients using a panel of V gene segment subfamily-specific oligonucleotide primers (V alpha I-w29/V beta I-w24). Three samples were examined in each patient: (i) peripheral blood cells (PBMCs) before therapy (day I); (ii) PBMC 2 days after the interruption of IL-2, at day 36 (i.e., at the lymphocytic rebound), (iii) the CD25-enriched cell fraction at day 36. Virtually all V alpha and V beta subfamily specificities were found in pre- as well as in post-treatment PBMCs and CD25+ cell fractions. These results support the view that circulating T-cell subpopulations are highly polyclonal after IL-2 therapy without any major alteration in the TCR V alpha and V beta repertoire. In addition, the results of quantificative densitometric analysis of V alpha and V beta amplified products suggest that a unique V beta 18-expressing T-cell subpopulation may be expanded in the CD25+ cell fraction after IL-2 therapy. Further characterization of these T cells may contribute to a better understanding of in vivo effects of IL-2 on renal-cell carcinoma metastases.

Adolescent↗
PubMed1993

Analysis of T-cell-receptor variable gene segment usage in peripheral-blood lymphocytes of advanced cancer patients.

Advanced cancer patients generally display impaired T-cell immune functions. The underlying mechanisms are not well understood. The aim of this study was to analyze whether major alterations of TCR variable gene segment usage could be detected in the blood of these patients. Seventeen individuals with various malignancies were tested using PCR and a panel of V-gene-segment-sub-family-specific (V alpha 1-w29/V beta 1-w24) oligonucleotide primers. The results indicate that these cancer patient lymphocytes expressed most V alpha and V beta sub-family specificities, similarly to the lymphocytes of healthy donors (n = 10). This suggests that immunodepression in advanced cancer patients is not related to major deletions in their T-cell repertoires. We also compared the mean relative expression of each V-sub-family specificity of patients and healthy donors by quantitative densitometric analysis. We found significant differences in 4 V beta specificities (and no V alpha). Our analysis identified unique T-cell sub-sets putatively involved in the mechanisms leading to immunodepression in advanced cancer patients. Alternatively, the observed differences in terms of V beta specificity expression may reflect the host response against the tumor.

Adolescent↗
PubMed1993

Fine specificity of monoclonal antibodies directed at human T cell receptor variable regions: comparison with oligonucleotide-driven amplification for evaluation of V beta expression.

Seven distinct anti-human T cell receptor (TcR) V region monoclonal antibodies (mAb) were generated by immunizing mice with either human T cell lines or transfected murine cells expressing human TcR V beta genes. The specificity of these reagents was determined as follows: T cells recognized by each mAb were purified from the peripheral blood of healthy donors and TcR transcripts expressed in these cells were analyzed using oligonucleotide-driven amplification and cDNA sequencing. Four mAb were found to delineate the V beta 3, V beta 8, V beta 17 and V beta 19 subfamilies, respectively. The remaining reagents recognize subsets within the V beta 2, V beta 5 and V beta 13 subfamilies. Reactivity of the mAb with circulating T cells from 18 unrelated healthy individuals was determined. Limited variability was found from an individual to another. In four donors, mAb staining was compared to oligonucleotide-driven amplification for evaluation of V beta 3, V beta 8, V beta 17 and V beta 19 subfamily expression in the peripheral blood. Although the V gene subfamily-specific oligonucleotides used in this study belong to a carefully controlled series, our results show that this method does not give an accurate estimate of the percentage of peripheral T cells expressing a given TcR beta chain. The present data confirm the necessity to establish a complete set of well-characterized monoclonal reagents to study human T cell responses.

Animals↗
PubMed1993

Limited T-cell receptor diversity in liver-infiltrating lymphocytes from patients with primary biliary cirrhosis.

Primary biliary cirrhosis is associated with the presence of high-titer anti-mitochondrial autoantibodies as well as T-cell infiltration of the liver, suggesting the involvement of autoimmune mechanisms. We have studied here the sequences of T-cell receptor alpha and beta chains expressed by T-cell clones derived from liver-infiltrating lymphocytes of two patients with primary biliary cirrhosis. Among the eight clones studied from the first patient, four expressed the same member of the V beta 6 subfamily, associated with either V alpha 4 (three clones) or V alpha 21 (one clone) gene segment. Two other clones expressed an identical V beta 12 transcript, and two in-frame alpha chain transcripts, involving V alpha 2 and V alpha 7 gene segments. From the second patient, eight out of the nine clones were found to rearrange V beta 17-J beta 2.1 and V alpha 3 gene segments. The remaining clone expressed distinct T-cell receptor chains, involving V beta 9 and V alpha 11 gene segments. As deduced from the analysis of their junctional regions, the eight T-cell clones expressing V beta 17/V alpha 3 gene segments derived from only three different T cells. Furthermore, conserved amino acid motifs were found to be encoded in both the alpha and the beta-chain junctional regions. Together, these data show a local amplification of unique T lymphocytes in both patients. The use of identical V beta J beta and V alpha gene segments with similar junctional sequences by three different cells, evidenced in one of the two cases, strengthens the view that liver-infiltrating T lymphocytes are selected locally by autoantigens in PBC.

Aged↗
PubMed1993

In-situ preferential usage of V alpha 8 T-cell receptor gene segments in a patient with bullous pemphigoid.

Bullous pemphigoid (BP) is an autoimmune blistering skin disease associated with the occurrence of autoantibodies directed to a limited series of antigens located at the basement membrane zone of the dermo-epidermal junction. Previous studies have demonstrated the presence of activated T cells in BP lesions although their actual contribution to the pathogenesis of the disease has remained unclear. One approach to better understanding the significance of these T-cell infiltrates is to study the diversity of the recognition receptors (TCRs) expressed at the lesion sites. We report here an extensive analysis, performed in one patient with typical BP, where 187 TCR in frame transcripts from a lesional area, from clinically normal skin or from peripheral blood lymphocytes have been sequenced and compared. The data show preferential usage of the V alpha 8 subfamily gene segments in the lesion. It is therefore suggested that T-cell infiltration in BP may not simply reflect a non-specific inflammatory process but include antigen-specific responses.

Amino Acid Sequence↗
PubMed1993

Alternatively spliced T cell receptor transcripts expressed in human T lymphocytes.

We used the anchored-polymerase chain reaction (A-PCR) procedure to study human TCR transcripts derived from a variety of polyclonal T cell populations. In this series of experiments, 31 'unusual' cDNAs, which do not include exclusively V-J-C, J-C or 5'C genomic sequences, were identified. Ten of these were found to represent distinct types of alternatively spliced TCR alpha transcripts whose structure is derived from unusual splicing of one, two or even three intervening intronic sequences. The splicing events led to either conservation of a novel exon in the mRNA structure (designated aE1 alpha-aE5 alpha) between the V-J and C segments or to deletion of the 3' V region-J segment. In three cases, the alternatively spliced exons (aE1 alpha-aE3 alpha) interrupt the open translational reading frame of the corresponding V-J alpha segment. Nineteen and two cDNA represent sterile C beta or C delta transcripts, respectively. Their structures are derived from the conservation of a non-translatable exon, aE1 beta or aE1 delta, which is precisely spliced at the 5' end of the corresponding C exon sequences. Interestingly, the 3' region of the aE1 beta sequence is homologous to the murine C beta 0 exon. Together, these results led to the characterization of nine novel exons in the TCR alpha, beta and delta loci.

Alternative Splicing↗
PubMed1993

The use of anchored polymerase chain reaction for the study of large numbers of human T-cell receptor transcripts.

Anchored-PCR (A-PCR) is an approach designed to amplify and clone sequences with unknown 5' or 3' extremities. A-PCR is therefore appropriate for studying variable region of T-cell receptors (TCRs) expressed in polyclonal T-cell populations since it does not prejudge which variable gene segments are actually being used. We report here some critical modifications in the initial procedure to make it easy to clone and sequence large series of TCR transcripts. They have been introduced to improve both the yield and specificity of TCR amplified products and include re-amplification, size selections of the material combined with the successive use of nested TCR constant region specific primers. This procedure has been successfully applied to the study of the repertoire of both TCR alpha/beta+ and gamma/delta+ human T-cells. The efficiency of the present A-PCR protocol will help to precisely analyze TCR usage in normal and pathological situations.

Base Sequence↗
PubMed1993

Combination of interleukin-2 and gamma interferon in metastatic renal cell carcinoma.

The use of high-dose interleukin-2 (IL2), alone or in association with lymphokine activated killer cells in patients with metastatic renal cell carcinoma (MRCC) results in a 20-25% response rate. However, the toxicity of IL2 is substantial and despite many clinical trials, response rates initially reported have not been improved. The aim of this study was to evaluate a combination of IL2 and gamma interferon (IFN) in MRCC with respect to both efficacy and tolerance. IL2 was given by continuous intravenous infusion at a daily dose of 24 x 10(6) U/m2 for 2 consecutive days during 5 consecutive weeks. Gamma IFN was given subcutaneously at a daily dose of 5 x 10(6) U/m2 on the same days as IL2. 33 patients with MRCC entered the study. Clinical responses were comparable with other published series: 7 patients (21%) achieved partial response, 13 (39%) were stable and 13 had progression, despite therapy. Immunological profile observed with this regimen showed a major increase in natural killer cells which became the predominant lymphocyte population at the end of the therapy. Tolerance was good with 92.5% of the planned doses actually received by the patients. This was reflected by an early discharge from the hospital in 95% of the cycles, increasing acceptability of the regimen by the patients.

Adult↗