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C Bordignon

Publications and source records attributed to C Bordignon.

At least 19 recordsLinked to original sources
PubMed1997

Functional heterogeneity of HLA-A*02 subtypes revealed by presentation of a MAGE-3-encoded peptide to cytotoxic T cell clones.

The peptide-binding and presentation characteristics of seven naturally occurring HLA-A2 subtypes were studied using M3(271), a peptide derived from the tumor-specific Ag encoded by gene MAGE-3, which has been shown to be processed and presented by A*0201+ melanoma lines. Three independent M3(271)-specific CTL clones were obtained from two unrelated A*0201+ donors. B lymphoblastoid cell lines (BLCLs) expressing A*0201, A*0207, or A*0209 could be sensitized to lysis by all three clones upon incubation with the relevant peptide. Furthermore, the same BLCLs were able to present endogenous M3(271) in IFN-gamma release assays. These findings demonstrate, for the first time, the existence of a functional overlap between A*0207 and other A*02 subtypes. One of the CTL clones also lysed M3(271)-pulsed BLCLs expressing A*0204 and A*0206, while the other two clones recognized M3(271) only in the context of either of these two subtypes. Peptide-pulsed BLCLs expressing A*0202 or A*0205 were not lysed, although A*0205 and, with lower affinity, A*0202 molecules were shown to bind peptide M3(271). These findings have implications for the selection of cancer patients for specific immunotherapy with peptide M3(271).

Antigen Presentation↗
PubMed1997

HSV-TK gene transfer into donor lymphocytes for control of allogeneic graft-versus-leukemia.

In allogeneic bone marrow transplantation (allo-BMT), donor lymphocytes play a central therapeutic role in both graft-versus-leukemia (GvL) and immune reconstitution. However, their use is limited by the risk of severe graft-versus-host disease (GvHD). Eight patients who relapsed or developed Epstein-Barr virus-induced lymphoma after T cell-depleted BMT were then treated with donor lymphocytes transduced with the herpes simplex virus thymidine kinase (HSV-TK) suicide gene. The transduced lymphocytes survived for up to 12 months, resulting in antitumor activity in five patients. Three patients developed GvHD, which could be effectively controlled by ganciclovir-induced elimination of the transduced cells. These data show that genetic manipulation of donor lymphocytes may increase the efficacy and safety of allo-BMT and expand its application to a larger number of patients.

Bone Marrow Transplantation↗
PubMed1996

Characterization of antigenic peptides presented by HLA-B44 molecules on tumor cells expressing the gene MAGE-3.

The amino acid sequence of the protein encoded by the gene MAGE-3 was screened for peptides containing the binding motif for HLA-B44. Nine peptides were synthesized, and their binding affinity for HLA-B*4402 and -B*4403 was analyzed in an HLA class I alpha-chain refolding assay. Four peptides with binding affinity for HLA-B*4403 were chosen for in vitro cytotoxic T-lymphocyte induction assays using as antigen-presenting cells peptide-pulsed, autologous activated B lymphoblasts from a healthy, B*4403+ donor. Peptide-specific effectors could be raised only against one peptide, M3-167. Cytotoxic T lymphocytes specific for this peptide were also able to recognize melanoma cell lines expressing HLA-B44 and the gene MAGE-3, strongly suggesting that M3-167 is a naturally processed MAGE-3-encoded epitope presented by HLA-B44. M3-167 is a I amino acid N-terminal extension of M3-168, a naturally processed epitope MAGE-3-encoded epitope presented by HLA-A1 that has been previously described. TAP binding studies of these 2 peptides revealed that the TAP affinity of M3-167 is about 9-fold higher than that of M3-168. M3-167 or a longer precursor could be transported into the endoplasmatic reticulum, where it could be trimmed for presentation by HLA-A1 or -B44 molecules. Taken together, our data suggest that M3-167 could be an immunodominant peptide encoded by the gene MAGE-3.

Amino Acid Sequence↗
PubMed1996

Multiple HLA-A alleles can present an immunodominant peptide of the human melanoma antigen Melan-A/MART-1 to a peptide-specific HLA-A*0201+ cytotoxic T cell line.

The majority of HLA-A*0201-restricted tumor-infiltrating lymphocytes from melanoma patients recognize a peptide, MT(27-35), derived from the Melan-A/MART-1 Ag. This study reports that six variants of HLA-A2 and the HLA-A28 subtype A*6901 can present peptide MT(27-35). A CTL line specific for peptide MT(27-35) was generated by in vitro stimulation of PBL of an HLA-A*0201+, healthy donor with peptide-pulsed, activated autologous B lymphoblasts. This CTL line was shown to recognize peptide MT(27-35) after endogenous processing on Melan-A/MART-1+/HLA-A2+ tumor cells. Moreover, a panel of B lymphoblastoid cell lines (BLCLs) expressing A*0202, A*0204, A*0205, A*0206, A*0209, and with lower efficiency A*6901, could be sensitized to lysis upon incubation with the relevant peptide. As demonstrated by the levels of ED50 and CD8 dependency of lysis, HLA-A*0204 and HLA-A*0205 presented the peptide as efficiently as HLA-A*0201, while the other four alleles were less efficient. Peptide-binding studies suggest that TCR- rather than peptide-binding affinity determines the T cell recognition levels of peptide-pulsed EBV-BLCLs expressing A*0201, A*0204, A*0206, and A*0209. Peptide-pulsed BLCLs expressing HLA-A*0207 or two additional subtypes of HLA-A28 were not recognized. MT(27-35)-specific CTL could also be raised from donors expressing HLA-A*0205. These findings have implications on the applicability of peptide vaccination with peptide MT(27-35) on melanoma patients.

Alleles↗
PubMed1996

HLA-A*02 subtype distribution in Caucasians from northern Italy: identification of A*0220.

This study describes a comprehensive easy to perform PCR-SSOP typing approach suitable for complete genomic subtyping of HLA-A*02. A single 1.6 kb PCR-amplificate spanning exons 2, 3 and 4 of the HLA-A*02 gene was used for hybridization with a panel of twenty-four SSOPs. This allowed unequivocal assignment of all so far known HLA-A2 subtypes, including A*0209 and A*0215N which differ for nucleotide substitutions in exon 4, without the need for two separate amplifications. Using this approach, HLA-A*02 subtype distribution was analyzed in 218 samples from unrelated, healthy individuals from northern Italy enrolled in the Italian Bone Marrow Registry and typed as HLA-A2 by serology or generic molecular analysis. As expected, A*0201 was found in the majority (92.6%) of samples. However, a significant number (6.8%) of individuals carried A*0205. Furthermore, A*0202, A*0208, A*0209 and A*0217, so far not described in Caucasians, were detected in a low number of samples (frequency ranging from 0.45% to 1.8%). Finally, a novel HLA-A*02 subtype, A*0220, was detected in 0.9% of the samples. As confirmed by DNA sequencing of exons 2 and 3, this allele is identical to A*0201 except for a single nucleotide substitution in codon 66 which changes the predicted amino acid sequence form Lys to Asn. The findings of this study have implications for the selection of HLA-A*02+ donors in unrelated bone marrow transplantation and of patients for specific immuno-therapy with HLA-A*02 restricted peptide vaccines.

Genes↗
PubMed1995

Gene therapy in peripheral blood lymphocytes and bone marrow for ADA- immunodeficient patients.

Adenosine deaminase (ADA) deficiency results in severe combined immunodeficiency, the first genetic disorder treated by gene therapy. Two different retroviral vectors were used to transfer ex vivo the human ADA minigene into bone marrow cells and peripheral blood lymphocytes from two patients undergoing exogenous enzyme replacement therapy. After 2 years of treatment, long-term survival of T and B lymphocytes, marrow cells, and granulocytes expressing the transferred ADA gene was demonstrated and resulted in normalization of the immune repertoire and restoration of cellular and humoral immunity. After discontinuation of treatment, T lymphocytes, derived from transduced peripheral blood lymphocytes, were progressively replaced by marrow-derived T cells in both patients. These results indicate successful gene transfer into long-lasting progenitor cells, producing a functional multilineage progeny.

Adenosine Deaminase↗
PubMed1995

Expression of the MAGE gene family in primary and metastatic human breast cancer: implications for tumor antigen-specific immunotherapy.

Some human tumors express known antigens that can be utilized as targets for specific immunotherapy. An absolute requirement for the efficacy of this therapeutic strategy is an adequate expression of the candidate antigen by all cells of the primary and metastatic tumor. To examine the presence and distribution of tumor-associated antigens in metastatic breast cancer, we used PCR analysis and ethidium bromide staining to test the expression of genes of the MAGE family in 28 primary tumors and related metastatic samples. Overall, samples obtained from 7 of 28 patients revealed positive. However, 2 of 3 primary tumors positive for MAGE-1 and/or MAGE-3 had corresponding negative metastatic lesions. On the contrary, 4 of the 25 MAGE-negative primary tumors gave rise to positive metastatic nodes. Our results confirm in vivo, at the molecular level, the tumor-antigen heterogeneity previously observed at the cellular level by in vitro analysis. Our data strongly suggest that, at least in patients with breast cancer, multiple different antigens would be required to optimize the recognition of neoplastic cells in immunotherapeutic protocols using MAGE products as target antigens.

Adult↗
PubMed1995

Evaluation of recombinant human erythropoietin to facilitate autologous blood donation before surgery in anaemic patients with cancer of the gastrointestinal tract.

The aim of this study was to determine whether the preoperative administration of recombinant human erythropoietin (rHuEPO) could increase the rate of autologous blood donation and reduce the perioperative need for homologous blood in anaemic patients with cancer. Twenty-two anaemic (haematocrit less than 34 per cent), iron-deficient (iron less than 700 micrograms/l) patients, with gastric or colorectal cancer scheduled for elective surgery, were allocated randomly to two groups. The first (n = 11) received iron saccharate 200 mg/day intravenously for 12 consecutive days. The second (n = 11) received rHuEPO subcutaneously (300 units/kg as first administration, and 100 units/kg 4, 8 and 12 days later) with supplemental iron. On days 4, 8 and 12, if the haematocrit was greater than 34 per cent, patients donated one unit (350 ml) of autologous blood. In the iron group the mean haematocrit did not change from admission (31 per cent) to day 12 of treatment (31 per cent), and no patient could donate autologous blood. In the rHuEPO group, eight patients donated two units of autologous blood and three donated one unit. Four patients in the iron group received perioperative transfusion of homologous blood compared with none in the rHuEPO group. Administration of rHuEPO facilitated the donation of autologous blood and reduced perioperative homologous blood transfusion in anaemic patients with cancer.

Adolescent↗
PubMed1995

A family of rapidly evolving genes from the sex reversal critical region in Xp21.

Patients with an intact SRY gene and duplications of portions of Xp21 develop as phenotypic females. We have recently mapped this sex reversal locus, DSS, to a 160-kb region of Xp21 that includes the adrenal hypoplasia congenita locus. To clone the gene(s) underlying DSS and AHC, we isolated expressed sequences from the region. Here we describe the characterization of two related genes. DAM10 and DAM6, expressed in adult testis and lung tumors. The predicted DAM10 and DAM6 proteins are 66% identical and are both highly similar to the MAGE family of tumor-associated antigens and to mouse necdin. Genes belonging to the MAGE superfamily, DAMs, MAGEs, and necdin, are likely to have originated from a common ancestor and to be subject to an unusually rapid evolution. The tumor-restricted expression of DAM proteins and their structural similarity to MAGE genes suggest that DAM peptides may be targets for active immunotherapy in lung cancer patients.

Adult↗
PubMed1995

Transfer of the HSV-tk gene into donor peripheral blood lymphocytes for in vivo modulation of donor anti-tumor immunity after allogeneic bone marrow transplantation.

The infusion of donor lymphocytes after allogeneic bone marrow transplantation is a promising therapeutic tool for achieving a graft versus leukemia (GvL) effect in case of leukemic relapse (1-7), and for the treatment of other complications related to the severe immunosuppressive status of transplanted patients, such as Epstein Barr virus-induced lymphoproliferative disorders (EBV-BLPD) (8) or reactivation of CMV infection (9). Although the delay in the administration of T lymphocytes is expected to reduce the risk of severe GvHD, this risk is still present at higher doses of donor T-cells. The transfer of a suicide gene into donor lymphocytes could allow the in vivo selective elimination of cells responsible for severe GvHD. Additionally, under appropriate conditions, it may allow in vivo modulation of donor anti-tumor responses, and to separate GvL from GvHD. Finally, crucial questions concerning survival and function of donor lymphocytes could be answered by their gene marking. Previous studies documented that T lymphocytes are suitable targets for gene transfer through retroviral vectors (10, 11). This protocol has been designed to evaluate in the contest of allogeneic BMT: 1--the safety of increasing doses of donor lymphocytes transduced with a suicide retroviral vector; 2--the efficacy in terms of survival and immunologic potential of donor lymphocytes after in vitro activation, gene transduction, and immunoselection; 3--the possibility of in vivo down regulation of GvHD by the administration of ganciclovir to patients treated by tk-transduced donor lymphocytes.(ABSTRACT TRUNCATED AT 250 WORDS)

Bone Marrow Transplantation↗
PubMed1995

Complete generic and extensive fine-specificity typing of the HLA-B locus by the PCR-SSOP method.

This study describes sequence specific oligonucleotide probe (SSOP) typing of hypervariable regions in exons 2 and 3 of HLA-B locus genes. A single HLA-B specific PCR-product spanning from bp 84 in exon 2 to bp 241 in exon 3 was used for dot blot hybridization to forty-seven chemiluminescent labeled oligonucleotide probes. Thirty-one of these probes were derived from four hypervariability zones in exon 3 of HLA-B genes and covered most known sequence polymorphisms within these regions. In addition, sixteen probes derived from polymorphic regions in exon 2 were used to discriminate alleles not unequivocally characterized by the exon 3 based probes. This SSOP panel gave rise to eighty-six distinct hybridization patterns that could be used to unequivocally define all WHO-designated serological HLA-B specificities except for HLA-B54 in all homo- and heterozygous combinations. Furthermore, sixty-six out of ninety-seven molecularly defined HLA-B subtypes were characterized by unique hybridization patterns in all homozygous and most (possibly all) heterozygous combinations. The reproducibility of these results was confirmed by analysis of forty-four Workshop reference cell lines and of seventy-eight randomly chosen samples (one-hundred forty-seven alleles) from unrelated individuals serologically typed in the laboratory. For sixty-five samples (one-hundred-thirty-three alleles), molecular typing confirmed the results obtained by serology and allowed molecular subtype assignment for ninety-one alleles tested. A serologically blank allele could be defined by molecular analysis in three cases. The method presented here for molecular typing of the HLA-B locus can be used as an alternative to biochemical methods such as one-dimensional isoelectric focusing for assignment of serologically cross-reacting HLA-B molecules as well as for subtype characterization of a large variety of HLA-B alleles.

Alleles↗
PubMed1994

Peripheral blood lymphocytes as target cells of retroviral vector-mediated gene transfer.

Peripheral blood lymphocytes (PBLs) are key target cells for gene therapy of a number of inherited and acquired blood disorders. We have systematically compared four retroviral vectors, designed according to different strategies, for their efficiency in transfer and expression in human PBLs of the same reporter gene. The receptor gene used in the study codes for the human low-affinity nerve growth factor receptor (LNGFR), and is not expressed on the majority of human hematopoietic cells, thus allowing quantitative analysis of the transduced gene expression by immunofluorescence, with single cell resolution. Peripheral blood mononuclear cells (PBMCs), as well as human hematopoietic cell lines of myeloid and lymphoid origin, were transduced with the four vectors and analyzed for efficiency of gene transfer, integration and stability of vector proviruses, and LNGFR expression at both RNA and protein level. Fluorescence-activated cell sorter analysis of coexpression of LNGFR and lineage-specific cell surface markers was performed in transduced cell lines, PBLs, and T-cell clones to study gene expression on specific cell subpopulations. Although crucial differences were observed among different constructs, all retroviral vectors could transduce, under appropriate infection conditions, T-cell populations representative of the normal immune repertoire. Gene transfer and expression could be demonstrated also in circulating progenitors of mature T cells. Expression of the transduced gene was heterogeneous among cell populations infected with the different vectors, with optimal results obtained by two of the four constructs. Finally, we have devised a simple protocol based on vector-mediated gene transfer and positive immunoselection of the transduced cells that produces virtually 100% gene-modified cells. This may represent a crucial improvement in the way of designing efficacious protocols involving the use of gene-modified T lymphocytes in clinical studies.

Cell Line↗
PubMed1994

Characterization of natural peptide ligands for HLA-B*4402 and -B*4403: implications for peptide involvement in allorecognition of a single amino acid change in the HLA-B44 heavy chain.

This study describes the characterization of endogenous peptides associated with the two major subtypes of HLA-B44. The two subtypes differ for a single amino acid substitution from Asp (HLA-B*4402) to Leu (HLA-B*4403) in position 156 of the alpha 2 domain, causing strong alloreactivity in vivo. In order to study the involvement of peptides in this phenomenon, the peptide motifs of the two subtypes were determined from natural peptide pools using Edman degradation. The motif was found to be essentially identical for HLA-B*4402 and -B*4403, with a strong predominance for Glu at position 2, Tyr or Phe at positions 9 and 10 and hydrophobic residues, especially Met, at position 3. Two individual naturally processed ligands of HLA-B*4403 were sequenced and shown to be derived from intracellularly expressed proteins found in protein sequence databases. The sequence of these natural peptide ligands conform well to the determined motif. These data will allow the prediction of HLA-B44 restricted peptide epitopes from viral and tumor antigens of known amino acid sequences. Moreover, they indicate that the peptide repertoire presented by HLA-B*4402 and -B*4403 is very similar, suggesting that the strong alloresponse between these two subtypes is not due to presentation of a different set of self peptides.

Amino Acid Sequence↗