Cancer Research Targets Frontiers in Basic Cancer Research
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Cancer Research Clinical Cancer Research
Cancer Epidemiology Biomarkers & Prevention Molecular Cancer Therapeutics
Molecular Cancer Research Cancer Prevention Research
Cancer Prevention Journals Portal Cancer Reviews Online
Annual Meeting Education Book Meeting Abstracts Online
[Cancer Research 59, 1954-1960, April 1, 1999]
© 1999 American Association for Cancer Research
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Paul, P.
Right arrow Articles by Carosella, E. D.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Paul, P.
Right arrow Articles by Carosella, E. D.
[Cancer Research 59, 1954-1960, April 15, 1999]
© 1999 American Association for Cancer Research

Immunology

Heterogeneity of HLA-G Gene Transcription and Protein Expression in Malignant Melanoma Biopsies1

Pascale Paul2, Francisco Adrián Cabestré, Fréderique-Anne Le Gal, Iman Khalil-Daher, Caroline Le Danff, Michel Schmid, Sylvaine Mercier, Marie-Françoise Avril, Jean Dausset, Jean-Gérard Guillet and Edgardo D. Carosella

DSV-DRM, SRHI/CEA, Commissariat à l’Energie Atomique, Centre Hayem, Hôpital Saint-Louis, 75010 Paris [P. P., F. A. C., I. K-D., C. L. D., E. D. C.]; Laboratoire d’Immunologie des Pathologies Infectieuses et Tumorales, Institut National de la Santé et de la Recherche Médicale U445, Institut Cochin de Génétique Moléculaire, Université René Descartes, 75014 Paris [J-G. G., F-A. L. G.]; Service de Dermatologie, Institut Gustave, 94805 Villejuif cedex [S. M., M-F. A.]; Institut National de la Santé et de la Recherche Médicale U462, Centre Hayem, 75010 Paris [M. S.]; and Fondation Jean Dausset, CEPH, 75010 Paris [J. D.], France


   ABSTRACT
Top
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Nonclassical MHC class I HLA-G antigen expression is tissue specific and is thought to play a role in tolerance of the semiallogeneic fetus by the maternal immune system. Ectopic expression of HLA-G by tumor cells provides them with an additional mechanism of escape from immunosurveillance by host cytotoxic effector mechanisms. The aim of this study was to assess the expression of nonclassical HLA-G antigens in ex vivo human melanoma biopsies.

HLA-G mRNA levels corresponding to both membrane-bound and soluble protein isoforms were analyzed in tumor specimens obtained from primary or metastatic melanomas of 23 patients. High levels of HLA-G transcription were detected in tumor specimens in 5 of 23 patients and found to be comparable in both lymph node and skin metastases. HLA-G mRNA transcript levels at tumor sites in 18 of these patients were compared with those in samples of their own healthy skin and were higher in the tumor tissue in 12 patients. Differential expression of mRNA transcripts corresponding to soluble and membrane-bound HLA-G was also observed in some tumor biopsies. HLA-G protein expression was detected in tumors that exhibited high levels of HLA-G transcription by immunofluorescence of frozen sections and Western blot analysis of both tumor and healthy skin biopsies, using anti-HLA-G-specific monoclonal antibodies. This work provides evidence that HLA-G gene transcription and protein expression can be up-regulated ex vivo in melanoma. Our finding that several of the tumors studied expressed high levels of HLA-G provides additional clues as to how a tumor can be selected in vivo to escape from cytotoxic antitumor responses, constituting a new parameter to be considered in the design of therapeutic approaches aimed at enhancing antitumor immune responses.


   INTRODUCTION
Top
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Several studies have highlighted the importance of an effective T-cell immune response against specific human melanoma-associated antigens. Most tumors are still able to escape cell-mediated immune responses, progressing into metastatic disease. Several explanations have been proposed as to why the T cell-mediated immune response is ineffective in eliminating melanoma cells, including defective helper T-cell function, inadequate costimulation of T cells, and deficient or reduced expression of MHC class I molecules caused by a defect in the class I heavy chain, the ß2-microglobulin light chain, or the TAP peptide transporter (1, 2, 3, 4) . Melanoma therapies aimed at enhancing tumor immunogenicity have recently been the subject of intense investigation (5) , including tumor-specific target antigens, such as MART-1/MelanA, gp100/Pmel17, gp75, MAGE, tyrosinase, and tyrosinase-related proteins 1 and 2, all of which have been identified as targets for MHC class I-restricted recognition by CTLs. Experiments with adoptive transfer of ex vivo-expanded patient T cells specific for these tumor antigens, peptide-based vaccines, and transduction of cytokine genes in tumor cells to enhance immunogenicity have been described (6 , 7) . Despite enhancement of peripheral T-cell immunity in patients undergoing adoptive immunotherapy, clinical effectiveness and tumor regression are not always obtained by these approaches. Ineffective vaccination and adoptive therapy are thought to occur via down-regulation or loss of melanoma-associated antigens or of MHC class I expression by an in situ subset of tumor cells that acquire resistance to CTL-mediated lysis (1 , 6 , 8 , 9) . Delivery of costimulatory signals and presentation of tumor antigens by antigen-presenting cells is also considered to be crucial for the induction of antigen-specific responses, and highly motile peptide-pulsed dendritic cells are now being considered for use in vaccination therapy (10 , 11) . Heterogeneous populations of cells within a tumor acquire different degrees of sensitivity to such therapies and HLA class I loss should be considered a failure factor in tumor eradication because class I-negative cells with high metastatic potential will not respond to specific T-cell therapy (12) . Such cells could be eliminated by NK3 cells (13) , which have the ability to lyse HLA-class I-negative cells, due to the lack of an inhibitory lysis signal delivered by appropriate class I antigens to KIRs expressed by NK (14) . Recent studies suggest that disruption of KIR interactions with their MHC class I ligand on melanoma cells may enhance the antitumor response (15 , 16) . CTL clones that express such inhibitory receptors can also be anergized in their MHC-restricted response to tumor cell clones that exhibit partial loss of HLA class I antigens, a phenomenon frequently observed in tumors (1 , 17 , 18) . Heterogeneity of melanoma cell populations with regard to MHC class I expression is, thus, a key factor to be taken into account when investigating the acquisition of metastatic potential by tumors as well as their capacity to escape from cytotoxic antitumor responses.

The variety of tumor cell phenotypes that influence the antitumor response may be even more complex. We have previously demonstrated that up-regulation of the nonclassical HLA-G gene may be observed in melanoma cell lines, in which it confers the ability to inhibit the NK cytotoxic function in vitro (19) . HLA-G exists in multiple protein isoforms, including membrane-bound (HLA-G1, -G2, -G3, and -G4) and soluble (HLA-G5 and -G6) molecules, as a result of alternative splicing (20) . Classical class I antigens are expressed at various levels in most tissues throughout the organism (21) , whereas high levels of HLA-G gene transcription and protein expression are physiologically restricted to trophoblast cells during pregnancy. However, HLA-G protein expression has also been reported in thymic epithelial cells (22) , first-trimester placental chorionic blood vessel endothelial cells (23) , and IFN-{gamma}-treated mononuclear phagocytes (24) . The HLA-G molecule exhibits very limited polymorphism and is thought to be able to interact with several described KIRs of the immunoglobulin superfamily including p49 (25) , ILT2 and ILT4 (26 , 27) , expressed at the surface of NK cells but also on T-cell subsets, dendritic cells, B cells, and monocyte lineages. Direct interaction of HLA-G with the widely expressed CD94/NKG2A KIR receptors of the C-type lectin superfamily has been recently refuted (28 , 29) , with the demonstration that the nonclassical HLA-E molecule is the ligand for this receptor. The HLA-G peptide leader, like other class I peptides, allows HLA-E expression at the cell surface and can, thus, indirectly favor such interactions. The capacity of HLA-G to function as an antigen that can inhibit cytotoxic NK effector function of the maternal immune system against its semiallogeneic fetus (30) has led us to investigate how ectopic expression of HLA-G in tumors could favor escape of HLA-G-expressing tumor cells from immunosurveillance (19) . To further evaluate the biological relevance for patients of our previous observation of HLA-G expression in melanoma cell lines and to confirm that HLA-G is, indeed, detectable in vivo, HLA-G gene transcription and protein expression were analyzed in ex vivo primary and metastatic human melanoma biopsies obtained from 23 patients. Our finding that several of the tumors studied expressed high levels of HLA-G may provide further clues as to how a tumor can be selected in vivo to escape from cytotoxic antitumor responses.


   MATERIALS AND METHODS
Top
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Patients and Tissue Samples.
From 1997 through 1998, patients undergoing surgery for melanoma in the Gustave Roussy Institute, Villejuif, France, were asked to participate in a prospective study approved by the local ethics committee (Comité Consultatif de Protection des Personnes pour la Recherche Biomédicale). Patients with primary melanoma were included only if the size of their tumor was large enough to adequately assess Breslow’s thickness and to provide a sample for the study. Small-sized primary tumors were excluded. For skin and regional lymph node metastases, samples were provided by the pathologist if tissue samples were large enough to assess diagnosis and resection quality. After giving informed consent, 23 consecutive patients undergoing surgery at the Institut Gustave Roussy were included in the study. No patient had received chemotherapy or systemic treatment during the 6 months prior to participation in the study. At the time they were included in the study, eight patients had an isolated primary melanoma, two patients had local recurrences within a 5-cm radius of the resected primary lesion, three patients had multiple skin metastases on the legs, eight patients had regional lymph node metastasis (including one who had an unresected primary tumor and another with in-transit metastasis), and two patients had disseminated disease for which they had undergone diagnostic (an untreated patient) or palliative regional lymph node resection. Tissue samples included healthy skin resected at the border of the surgical wound (n = 18), healthy lymph node (n = 1), healthy s.c. tissue (n = 1), skin primary tumor (n = 9), skin metastases (n = 9), lymph node metastases (n = 10), and a tumor regression area within a very large primary tumor (n = 1). Immediately after surgical excision, samples were removed from tumors under aseptic conditions, frozen in liquid nitrogen, and stored until RT-PCR and immunohistochemical staining could be carried out. Frozen blocks were cut into 5-µm sections, mounted on slides, and stained by the H&E method to evaluate the presence of tumor. Immunoperoxidase staining with S-100 and HMB45 antibodies (DAKO, Glostrup, Denmark) was used to confirm diagnosis as described (31) . Patient characteristics at the time of sample inclusion are listed in Table 1Citation .


View this table:
[in this window]
[in a new window]

  		Table 1 Clinical and histopathological characteristics and levels of HLA-G transcripts encoding membrane-bound and soluble isoforms of surgically removed biopsies of 23 melanoma patients

 
RT-PCR.
Total RNA was isolated from frozen samples by homogenization, using an ultraturrax device (IKA Labortechnic, Staufen, Germany) and RNA Now reagent (Biogentex, Seabrook, TX) according to manufacturers’ recommendations and checked by electrophoresis in a 1.5% agarose denaturing gel. One µg of RNA was reverse-transcribed to cDNA using oligo(dT) priming and Moloney murine leukemia virus reverse transcriptase (Life Technologies, Inc.) at 42°C for 1 h. PCR was carried out in 100-µl volumes containing 200 µM each deoxynucleotide, 100 ng of each primer, 10 µl of 10x PCR buffer (Perkin-Elmer Corp.), and 2.5 units of Taq DNA polymerase (Perkin-Elmer Corp.). HLA-G-specific PCR amplifications were carried out using G.257 (5'-GGA AGA GGA GAC ACG GAA CA) and G3.U (5'-GGC TGG TCT CTG CAC AAA GAG A) as primers, detecting all alternatively spliced HLA-G mRNA isoforms (19) . Specific amplification of the soluble HLA-G5 transcript was conducted with G.526 (5'-CCA ATG TGG CTG AAC AAA GG) and G.i4b (5'-AAC GGA GGT GAA GGT GAG GG) primers. The PCR was conducted for 35 cycles, consisting of 1 min at 94°C, 90 s at 65°C (61°C for G5), and 2 min at 72°C, followed by a final extension of 7 min at 72°C. Coamplification of ß-actin cDNA was carried out in each sample, using the ß-actin amplimer set (Clontech) for 16 cycles, to compare RNA levels in samples. PCR specificity was confirmed by Southern blotting onto nylon membranes (Hybond N+; Amersham). Hybridization was carried out with the HLA-G-specific G.R (5'-GGT CTG CAG GTT CAT TCT GTC) and G.i4F (5'-GAG GCA TCA TGT CTG TTA GG) probes, as described previously (19) .

Immunofluorescence Analysis of Tumor Biopsies.
Standard methods were used to carry out immunohistochemistry on frozen sections that had been fixed in acetone for 8 min and then rinsed in PBS and blocked in normal rabbit serum (DAKO) in PBS for 10 min. Samples were incubated with primary antibody for 1 h at room temperature, followed by incubation with a secondary FITC-conjugated antimouse immunoglobulin antibody for 15 min. Sections were counterstained with 4',6-diamidino-2-phenylindole (Sigma Chemical Co., St. Louis, MO) nuclear dye and mounted with antifading mounting medium (DAKO). Fluorescence was analyzed with an MRC 1024 confocal microscope (Bio-Rad, Hempstead, United Kingdom) equipped with a krypton-argon ion laser for fluorescein detection and a UV ion laser for 4',6-diamidino-2-phenylindole detection. A Nikon Diaphot 300 inverted microscope and a Nikon planochromat x60 objective lens with a numerical aperture of 1.4 (Nikon Corp., Melville, NY) were used for fluorescence microscopy. The following antibodies were used: anti-MHC class I W6/32 (DAKO), antihuman melanoma HMB45 (DAKO), and anti-HLA-G 87G (which detects the HLA-G1 isoform; Ref. 32 ), a kind gift of Dan Geraghty, Fred Hutchinson Cancer Research Center (Seattle, WA).

Western Blot Analysis of HLA-G Proteins in Tumor Samples.
When enough tumor specimen was available, proteins were extracted from melanoma tissues by homogenization in lysis buffer [25 mM Tris (pH 7.5), 150 mM NaCl, 10 mM EDTA, 1% NP40, and protease inhibitors (Complete; Boehringer Mannheim)], using an ultraturrax device. After removal of insoluble material by centrifugation at 10,000 x g for 20 min, the protein concentration was determined using a Bio-Rad (Richmond, CA) DC protein assay kit. Twenty-five-µg aliquots of total protein were separated by SDS-PAGE (10%) and transferred onto nitrocellulose membranes (Hybond-C extra; Amersham). After blocking, the membrane was probed with a 1:100 dilution of anti HLA-G mAb 4H84 (22) directed against the {alpha}-1 domain of the HLA-G molecule (kindly provided by M. McMaster). Antibody binding was detected using a peroxidase-conjugated secondary antibody (Sigma) and chemiluminescence (ECL+ detection kit; Amersham).


   RESULTS
Top
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
Detection of HLA-G mRNA Levels in Human Melanoma Biopsies.
HLA-G mRNA levels in 28 melanoma specimens (described in Table 1Citation ) were obtained from the primary or metastatic tumors of 23 melanoma patients as well as healthy skin biopsies from 18 of the 23 patients, and tested by RT-PCR. In healthy skin samples, RT-PCR resulted in detection of basal low levels of the most abundant HLA-G1 transcripts, whereas the HLA-G2 transcripts were rarely detected, a pattern that is comparable to that described in peripheral blood lymphocytes (Fig. 1BCitation ; Refs. 33 , 34 ). In 12 of 18 matched healthy and melanoma tissues, we detected a higher level of HLA-G transcription in tumor tissue than in the same patient’s healthy tissue (Figs. 1Citation and 2Citation and Table 1Citation ). Six of the 23 patients exhibited the same basal low level of HLA-G transcription in both tumor and healthy skin biopsies. A high level of HLA-G transcription, comparable to that observed in trophoblast-derived JEG-3 cells, was observed in five melanoma patients (VIG, CHO, ESN, GOU, and ETI; Fig. 1Citation ). The HLA-G transcription level observed at various tumor sites in the same patient were comparable, transcription activation being detected in both lymph node and s.c. skin metastases. Representative cases are shown in Fig. 1Citation , and the results of analysis in other patients are summarized in Table 1Citation .



View larger version (21K):
[in this window]
[in a new window]
[Download PPT slide]
  		Fig. 1. A, RT-PCR analysis of HLA-G transcripts in malignant melanoma biopsies. RT-PCR analysis of HLA-G mRNA isoforms in ex vivo skin melanoma metastasis biopsies. Pan-HLA-G primers were used for RT-PCR amplification of HLA-G transcripts from ex vivo biopsies of healthy skin (HS), skin metastasis (SM), lymph node metastasis (LNM), and skin primary tumors (SPT). JEG-3 choriocarcinoma cells were used as controls for high HLA-G transcription. HLA-G-specific bands were detected by hybridization with the GR-specific probe located in exon 2. Shown are bands corresponding to HLA-G1, G2, G3, G4, and G5 transcripts. PCR products coamplified in the same reaction by ß-actin primers were detected on the same membrane by a ß-actin-specific probe. B, RT-PCR analysis of HLA-G transcription in peripheral blood mononuclear cells (PBMC) and YT2C2 NK cells used as controls for basal and no HLA-G transcription, respectively.

 


View larger version (14K):
[in this window]
[in a new window]
[Download PPT slide]
  		Fig. 2. RT-PCR analysis of HLA-G5 transcripts in malignant melanoma biopsies. Specific RT-PCR detection of HLA-G5 transcripts that correspond to the soluble isoform from melanoma biopsies was carried out by specific RT-PCR amplification of the HLA-G5 transcripts, using G.526 and G.i4b primers in the same patients presented in Fig. 1Citation .

 
Differential Expression of Soluble HLA-G Transcripts in Melanoma Biopsies.
Specific RT-PCR analysis of soluble HLA-G5 RNA transcripts, using primers that amplify only the full-length secreted isoform, allowed us to detect differential transcription of soluble HLA-G5 transcripts in several patients (Fig. 2)Citation . Among the five melanomas that exhibited high levels of transcription, three (ESN, VIG, and ETI) had low or undetectable HLA-G5 transcript levels, whereas the two remaining tumor specimens (CHO and GOU) exhibited overall enhancement of all HLA-G transcripts. The three melanoma biopsies (MEN, PER, and LOR) that had lower HLA-G mRNA levels were also found to express high levels of the soluble HLA-G transcript. Representative cases are shown in Fig. 2Citation , and the results of analysis in other patients are summarized in Table 1Citation .

Detection of HLA-G Proteins in Human Melanoma Biopsies.
HLA-G protein expression in melanoma tissues was evaluated by immunofluorescence, using mAb 87G, which is specific for the full-length HLA-G1 isoform. A correlation between high levels of HLA-G transcription and the expression of HLA-G protein was found in six melanoma biopsies obtained from four (ETI, CHO, ESN, and VIG) of the five patients in whom high levels of HLA-G mRNA were detected. For patient ESN, it was possible to analyze healthy skin as well as skin and lymph node metastases. HLA-G expression was selectively detected in both tumor sites but not observed in healthy skin (Fig. 3A)Citation . Biopsies of patients ETI (Fig. 3A)Citation and CHO (data not shown), in whom a high level of HLA-G transcript was detected, were also positively stained by the anti-HLA-G antibody. Six biopsies of healthy skin obtained from melanoma patients as well as one sample of healthy skin obtained from normal breast plastic surgery, in which low or intermediate levels of HLA-G transcripts were detectable, were negative with the HLA-G antibody. Primary skin tumors of patients MEN and PER and skin metastasis of patient DES, all of which exhibited low levels of HLA-G transcription, were also negative for HLA-G protein expression. One of these tumor specimens (FLI) exhibited only moderate levels of HLA-G transcripts, similar to those found in healthy skin, and low HLA-G expression was reproducibly observed in this tumor by immunofluorescence (data not shown), indicating that low HLA-G transcription can still generate cell surface HLA-G protein expression. No immunofluorescence staining was detected with 87G in the healthy skin of this patient.



View larger version (107K):
[in this window]
[in a new window]
[Download PPT slide]
  		Fig. 3. Confocal microscopy analysis of HLA-G expression in melanoma biopsies and Western blot detection of the HLA-G protein in melanoma protein extracts. A, 87G positive staining of a skin metastasis section from ESN (b), lymph node metastasis from ESN (c), and lymph node metastasis from ETI (d), whereas the healthy skin sample from ESN remains unstained with the 87G antibody (a), displaying good correlation with results obtained at the transcription level. B, the Mr 39,000 protein, corresponding to the HLA-G1 isoform, detected by the anti-HLA-G 4H84 antibody in the JEG-3 choriocarcinoma cell line used as the positive control, and in lymph node metastasis protein lysates from VIG, ETI, and ESN. Healthy skin from MAL (HS) was tested as a negative control, and no Mr 39,000 band was detected.

 
Western blot analysis, using the anti-HLA-G 4H84 mAb, revealed the presence of a Mr 39,000 band in protein extracts from lymph node metastastic tumor samples obtained from patients ESN, VIG, and ETI. This band is similar to one observed in the JEG-3 choriocarcinoma cell line, known to express the HLA-G1 protein. No signal corresponding to HLA-G was detected in healthy skin (Fig. 3B)Citation .

Levels of HLA-G Transcription and Protein Expression Measured in Healthy Skin, Skin Primary Tumor, Tumor Regression Site, and Melanoma Metastasis in the Same Patient Were Found to Correlate with the Proportion of Tumor Cells That Expressed Melanoma-associated Antigens.
In one patient (VIG), it was possible to test healthy skin, primary cutaneous tumor, lymph node metastasis, and tumor regression within the skin primary tumor site specimen. Activation of HLA-G transcription was high in this patient’s primary and metastatic tumor specimens but low and comparable in both healthy skin and tumor regression sites (Fig. 4)Citation . No transcripts corresponding to HLA-G5 were detected in these samples. HLA-G, melanoma antigen, and classical class I protein expression were analyzed in primary skin tumor, lymph node metastases, healthy skin, and a tumor regression site in this patient. Healthy skin from this patient was negative for 87G (Fig. 5A)Citation . Both the nodular area of primary melanoma and lymph node metastases exhibited strong homogeneous staining with anti-HLA-G, anti-classical HLA class I, and anti-melanoma-associated antigen antibodies (Fig. 5, B and C)Citation . No staining could be detected in a regressive area of the primary tumor in the same patient (Fig. 5D)Citation .



View larger version (75K):
[in this window]
[in a new window]
[Download PPT slide]
  		Fig. 4. RT-PCR analysis of HLA-G transcripts in biopsies from patient VIG. Analysis of HLA-G transcription in patient VIG: healthy skin (HS), skin primary tumor (SPT), lymph node metastasis (LNM), and tumor regression (R). HLA-G transcript levels were analyzed in the same patient, using pan-HLA-G or HLA-G5-specific primer sets.

 


View larger version (133K):
[in this window]
[in a new window]
[Download PPT slide]
  		Fig. 5. HLA class I, HLA-G and melanoma antigen expression in healthy skin, skin primary tumor, lymph node metastasis, and tumor regression tissues of patient VIG. A–D, immunohistochemical analysis of HLA-G expression in biopsies from patient VIG. The 87G antibody (c) binds only to skin primary tumor (B) and lymph node metastasis (C) sections that were stained with the HMB45 anti-melanoma-associated antigen antibody (d). Healthy skin (A) and tumor regression (D) biopsies were not stained by either 87G (c) or HMB45 (d) antibodies. Positive staining with the antibody W6/32 can be seen in the four specimens (b), whereas the negative control (a) remains unstained.

 

   DISCUSSION
Top
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
Constitutive and inducible HLA-G expression has been demonstrated in cultured tumor cell lines (19 , 35 , 36) . HLA-G protein expression in surgically removed tumor biopsies has also been analyzed in a limited series of colorectal cancer, laryngeal carcinoma, kidney, thyroid, and breast cancer. Such expression remains controversial (36, 37, 38) and is probably underestimated because of the paucity of reagents able to detect HLA-G isoforms other than the full-length HLA-G1 membrane-bound isoform. The results obtained in this study are new findings demonstrating that primary and metastatic melanoma cells can express HLA-G in vivo. Various mechanisms can account for the escape of melanoma cells from immune destruction, including down-regulation of MHC class I molecules, FasL expression (39) , and secretion of inhibitory factors such as interleukin 10 and transforming growth factor-ß by tumor cells (40) . Peptide-based vaccine and adoptive transfer clinical approaches do not always counterbalance the selection capacity of escape mechanisms provided by tumor cell phenotype heterogeneity. Apart from the commonly held opinion that tumors are poor stimulators of immune responses, recent evidence favors an active role of tumors in inducing T-cell tolerance (41) .

Some phenotypical features, such as expression of the Fas ligand, are common to immune-privileged sites in which tolerance is established, such as trophoblast and the anterior chamber of the eye, where HLA-G expression is also physiologically restricted. We demonstrate here that ectopic HLA-G gene transcription and protein expression can be detected in melanoma cells ex vivo. Several genes, including MAGE, that exhibit testis- and placenta-restricted distribution are activated in melanoma. These genes are thought to be activated through demethylation processes or by release from stringent genetic control, which could also trigger the expression of other genes, including HLA-G (42) . We have shown that activation of HLA-G gene transcription and protein expression can occur in melanoma cells and could be considered a way the immune system is rendered ineffective against HLA-G-positive tumor subsets. This could occur by inhibiting the cytotoxic function of immune effector cells (19) through interaction with KIRs of both NK and T cells. Recent works show that the expression of immunoglobulin superfamily KIR receptors that interact with HLA-G is not restricted to NK cells. In addition to NK cells, the ILT2 receptor is expressed in T-cell subsets, B lymphocytes, and myelomonocytic cells (27 , 43) and ILT4 is selectively expressed in monocytes, macrophages, and dendritic cells (26) . Other as yet unidentified immunological functions of HLA-G expression in melanoma, such as enhancement of HLA-E expression (28 , 44) and interaction with other activating or inhibitory receptors expressed on various immunocompetent cells, may, thus, become evident in the near future.

The capacity of tumors to secrete the soluble HLA-G isoform could also favor their spread. Soluble HLA-G5, similar in structure to full-length HLA-G1, lacks the transmembrane domain and includes a specific protein sequence translated from "intronic" mRNA. The soluble form of the HLA-G protein binds the same endogenous peptide motifs as HLA-G1 but differs with respect to TAP association (32) .The soluble HLA-G5 antigen is secreted by cytotrophoblasts and activated placental macrophages (45) and is also detected in serum and amniotic fluid (46) . However, to the best of our knowledge, no data are available concerning secretion of sHLA-G by malignant cells and its possible functional implications. The release of soluble molecules, such as CD59/ICAM-1 (47) , GD3 gangliosides (48) , or HLA antigens (49 , 50) has been described as a factor favoring in vivo escape from constitutive or treatment-induced immune control. Soluble class I molecules have been shown to inhibit the NK cytotoxic effector function (51) and may regulate interactions with cellular and humoral immune responses in the host, thus contributing to and facilitating the establishment of tolerance to tumor cells. Soluble classical class I molecules that were shed from cell membranes have been detected in the serum of patients with autoimmune and inflammatory diseases and after allogeneic organ transplantation (52) . Although the biological significance of soluble class I molecules remains enigmatic, their capacity to induce apoptosis in alloreactive cytotoxic lymphocytes has been demonstrated (53) to enable down-regulation of T-cell responses. The soluble HLA molecules that present self-peptides are also thought to play a role in maintaining self-tolerance. When expressed in tumors, HLA-G secretion could also have such an inhibitory effect on T-cell responses (54) . The observed discrepancy between membrane-bound and soluble HLA-G isoforms at the transcription level is reported here for the first time and could represent adaptive mechanisms of tumor clones enabling them to selectively express membrane-bound isoforms. Further studies are needed to investigate the secretion of HLA-G protein and its potential correlation with disease severity. Whether the absence of HLA-G secretion would lead to a stronger immune response and to a better prognosis is also an interesting question. The roles of HLA-G isoforms that lack one or two extracellular domains also remain to be elucidated in the context of tumor escape mechanisms. Considering the accumulation of recent data related to HLA-G function in the establishment of immune tolerance, it is tempting to postulate that HLA-G expression could favor the appearance of metastatic clones. Although the limited number of patients included in this study did not allow us to correlate HLA-G expression in tumor cells or the secretion of soluble HLA-G with the severity of disease or with metastatic potential, this work clearly establishes the fact that such HLA-G expression in melanoma is, indeed, observed in vivo. Additional follow-up studies of HLA-G expression status in primary tumors regarding their capacity to metastasize will permit evaluation of this parameter as a facilitator of tumor progression.

Analysis of the HLA-G status of tumors before and during treatment and the design and development of pharmacological tools that selectively down-regulate HLA-G expression at the surface of HLA-G-positive tumor cells should help to provide complementary therapeutic approaches in the treatment and follow-up of human melanoma.


   ACKNOWLEDGMENTS
 
We thank Daniel Geraghty, Susan Fisher, and Michael McMaster for providing us with HLA-G antibodies. We also thank Christelle Dolliger for her technical assistance. We are grateful to Pierre Duvillard and Alvaro Margulis (Department of Pathology and Surgery, Institut Gustave Roussy, Villejuif, France) for providing us with histopathology reports and tumor sample biopsies. We thank Noah Hardy, Nathalie Rouas-Freiss, and Julie-Ann Gavigan for reading and correcting the manuscript. We express our thanks to Prof. R. Gómez-Lus for his interest in this work.


   FOOTNOTES
 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1 This study was supported by grants from the French Commissariat à l’Energie Atomique and the Association pour la Recherche sur le Cancer. F. A. was the recipient of a grant from the MENRT in the context of a French-Spanish scientific collaboration web. Back

2 To whom requests for reprints should be addressed, at DSV-DRM, SRHI/CEA, Commissariat à l’Energie Atomique, Centre Hayem, Hôpital Saint-Louis, 1, avenue Claude-Vellefaux, 75010 Paris, France. Phone: 33 (0)1 53 72 21 42; Fax: 33 (0)1 48 03 19 60; E-mail: paul{at}dsvidf.cea.fr Back

3 The abbreviations used are: NK, natural killer; KIR, killing inhibitory receptor; RT-PCR, reverse transcriptase-PCR; mAb, monoclonal antibody. Back

Received 9/14/98. Accepted 2/18/99.


   REFERENCES
Top
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 

  1. Ferrone S., Marincola F. M. Loss of HLA class I antigens by melanoma cells: molecular mechanisms, functional significance and clinical relevance. Immunol. Today, 16: 487-494, 1995.[Medline]
  2. Maeurer M., Gollin S. M., Martin D., Swaney W., Bryant J., Castelli C., Robbins P., Parmiani G., Storkus W. J., Lotze M. T. Tumor escape from immune recognition. J. Clin. Invest., 98: 1633-1641, 1996.[Medline]
  3. Seliger B., Maeurer M. J., Ferrone S. TAP off-tumors on. Immunol. Today, 18: 292-298, 1997.[Medline]
  4. Hicklin D., Wang Z., Arienti F., Rivoltini L., Parmiani G., Ferrone S. ß2-microglobulin mutations, HLA class I antigen loss, and tumor progression in melanoma. J. Clin. Invest., 15: 2720-2729, 1998.
  5. Rosenberg S. A. Cancer vaccines based on the identification of genes encoding cancer regression antigens. Immunol. Today, 18: 175-182, 1997.[Medline]
  6. Pardoll D. M. Cancer vaccines. Nat. Med., 4: 525-531, 1998.[Medline]
  7. Rosenberg S. A., Yang J. C., Schwartzentruber D. J., Hwu P., Marincola F., Topalian S. L., Restifo N. P., Dudley M. E., Schwarz S. L., Spiess P. J., Wunderlich J. R., Parkhurst M. R., Kawakami Y., Seipp C. A., Einhorn J. H., White D. E. Immunologic and therapeutic evaluation of a synthetic peptide vaccine for the treatment of patients with metastatic melanoma. Nat. Med., 4: 321-327, 1998.[Medline]
  8. Alexandroff A. B., Robins R. A., Murray A., James K. Tumour immunology: false hopes, new horizons. Immunol. Today, 19: 247-252, 1998.[Medline]
  9. Rivoltini L., Loftus D., Squarcina P., Castelli C., Rini F., Arienti F., Belli F., Marincola F., Geisler C., Borsatti A., Appella E., Parmiani G. Recognition of melanoma-derived antigens by CTL: possible mechanisms involved in down-regulating anti-tumor T-cell reactivity. Crit. Rev. Immunol., 18: 55-63, 1998.[Medline]
  10. Nestle F. O., Aligajic S., Gilliet M., Sun Y., Grabbe S., Dummer R., Burg G., Schadendorf D. Vaccination of melanoma patients with peptide- or tumor lysate-pulsed dendritic cells. Nat. Med., 4: 328-332, 1998.[Medline]
  11. Tüting T., Wilson C. C., Martin D. M., Kasamon Y. L., Rowles J., Ma D. I., Slingluff C. L., Wagner S. N., van der Bruggen P., Baar J., Lotze M. T., Storkus W. J. Autologous human monocyte-derived dendritic cells genetically modified to express melanoma antigens elicit primary cytotoxic T cell responses in vitro: enhancement by cotransfection of genes encoding the Th1-biasing cytokines IL-12 and IFN-{alpha}. J. Immunol., 160: 1139-1147, 1998.[Abstract/Free Full Text]
  12. Ruiz Cabello F., Garrido F. HLA and cancer: from research to clinical impact. Immunol. Today, 19: 539-542, 1998.[Medline]
  13. Porgador A., Mandelboim O., Restifo N., Strominger J. L. Natural killer cell lines kill autologous ß2-microglobulin-deficient melanoma cells: implications for cancer immunotherapy. Proc. Natl. Acad. Sci. USA, 94: 13140-13145, 1997.[Abstract/Free Full Text]
  14. Klein E., Mantovani A. Action of natural killer cells and macrophages in cancer. Curr. Opin. Immunol., 5: 714-718, 1993.[Medline]
  15. Bakker A. B. H., Phillips J. H., Figdor C. G., Lanier L. L. Killer cell inhibitory receptors for MHC class I molecules regulate lysis of melanoma cells mediated by NK cells, {gamma}{delta} T cells, and antigen-specific CTL. J. Immunol., 160: 5239-5245, 1998.[Abstract/Free Full Text]
  16. Pende D., Accame L., Pareti L., Mazzochi A., Moretta A., Parmiani G., Moretta L. The susceptibility to natural killer cell-mediated lysis of HLA class I positive melanomas reflects the insufficient amounts of different HLA class I alleles. Eur. J. Immunol., 28: 2384-2394, 1998.[Medline]
  17. Ikeda H., Lethe B., Lehmann F., van Baren N., Baurain J. F., de Smet C., Chambost H., Vitale M., Moretta A., Boon T., Coulie P. G. Characterization of an antigen that is recognized on a melanoma showing partial HLA loss by CTL expressing an NK inhibitory receptor. Immunity, 6: 199-208, 1997.[Medline]
  18. Garrido F., Ruiz-Cabello F., Cabrera T., Pérez-Villar J. J., Lopez-Botet M., Duggan-Keen M., Stern P. L. Implications for immunosurveillance of altered HLA class I phenotypes in human tumours. Immunol. Today, 18: 89-95, 1997.[Medline]
  19. Paul P., Rouas-Freiss N., Khalil-Daher I., Moreau P., Riteau B., Le Gal F. A., Avril M. F., Dausset J., Guillet J. G., Carosella E. D. HLA-G expression in melanoma: a way for tumor cells to escape from immunosurveillance. Proc. Natl. Acad. Sci. USA, 95: 4510-4515, 1998.[Abstract/Free Full Text]
  20. Carosella E. D., Dausset J., Kirszenbaum M. HLA-G revisited. Immunol. Today, 17: 407-409, 1996.[Medline]
  21. Natali P., Bigotti A., Nicotra M., Viora M., Manfredi D., Ferrone S. Distribution of human class I (HLA-A, B, C) histocompatibility antigens in normal and malignant tissues of nonlymphoid origin. Cancer Res., 44: 4679-4687, 1984.[Abstract/Free Full Text]
  22. Crisa L., McMaster M. T., Ishii J. K., Fisher S. J., Salomon D. R. Identification of a thymic epithelial cell subset sharing expression of the class Ib HLA-G molecule with fetal trophoblasts. J. Exp. Med., 186: 289-298, 1997.[Abstract/Free Full Text]
  23. Blaschitz A., Lenfant F., Mallet V., Hartmann M., Bensussan A., Geraghty D. E., Le Bouteiller P., Dohr G. Endothelial cells in chorionic fetal vessels of first trimester placenta express HLA-G. Eur. J. Immunol., 27: 3380-3388, 1997.[Medline]
  24. Yang Y., Chu W., Geraghty D. E., Hunt J. S. Expression of HLA-G in human mononuclear phagocytes and selective induction by IFN-{gamma}. J. Immunol., 156: 4224-4231, 1996.[Abstract]
  25. Cantoni C., Verdiani S., Falco M., Pessino A., Cilli M., Conte R., Pende D., Ponte M., Mikaelsson M. S., Moretta L., Biassoni R. p49, a putative HLA class I-specific inhibitory NK receptor belonging to the Ig superfamily. Eur. J. Immunol., 28: 1980-1990, 1998.[Medline]
  26. Colonna M., Samaridis J., Cella M., Angman L., Allen R. L., O’Callaghan C., Dunbar R., Ogg G. S., Cerundolo V., Rolink A. Human myelomonocytic cells express an inhibitory receptor for classical and nonclassical MHC class I molecules. J. Immunol., 160: 3096-3100, 1998.[Abstract/Free Full Text]
  27. Colonna M., Navarro F., Bellon T., Llano M., Garcia P., Samaridis J., Angman L., Cella M., Lopez-Botet M. A common inhibitory receptor for major histocompatibility complex class I molecules on human lymphoid and myelomonocytic cells. J. Exp. Med., 186: 1809-1818, 1997.[Abstract/Free Full Text]
  28. Braud V. M., Allan D. S. J., O’Callaghan C. A., Söderström K., D’Andrea A., Ogg G. S., Lazetic S., Young N. T., Bell J. I., Phillips J. H., Lanier L. L., McMichael A. J. HLA-E binds to natural killer cell receptors CD94/NKG2A, B and C. Nature (Lond.), 391: 795-799, 1998.[Medline]
  29. Lee N., Llano M., Carretero M., Ishitani A., Navarro F., Lopez-Botet M., Geraghty D. E. HLA-E is a major ligand for the natural killer inhibitory receptor CD94/NKG2A. Proc. Natl. Acad. Sci. USA, 95: 5199-5204, 1998.[Abstract/Free Full Text]
  30. O’Callaghan C. A., Bell J. I. Structure and function of the human MHC class Ib molecules HLA-E, HLA-F and HLA-G. Immunol. Rev., 163: 129-138, 1998.[Medline]
  31. Skelton H. d., Smith K., Barrett T., Lupton G. P., Graham J. H. HMB-45 staining in benign and malignant melanocytic lesions. A reflection of cellular activation. Am. J. Dermatopathol., 13: 543-550, 1991.[Medline]
  32. Lee N., Malacko A. R., Ishitani A., Chen M. C., Bajorath J., Marquardt H., Geraghty D. E. The membrane-bound and soluble forms of HLA-G bind identical sets of endogenous peptides but differ with respect to TAP association. Immunity, 3: 591-600, 1995.[Medline]
  33. Amiot L., Onno M., Drénou B., Lemarchand B., Lamy T., Semana G., Fauchet R. Distribution of HLA-G alternative mRNAs including soluble forms in normal lymphocytes and in lymphoid cell-derived leukemia. Eur. J. Immunogenet., 23: 311-320, 1996.[Medline]
  34. Kirszenbaum M., Moreau P., Gluckman E., Dausset J., Carosella E. D. An alternatively spliced form of HLA-G mRNA in human trophoblasts and evidence for the presence of HLA-G transcript in adult lymphocytes. Proc. Natl. Acad. Sci. USA, 91: 4209-4213, 1994.[Abstract/Free Full Text]
  35. Amiot L., Onno M., Drénou B., Monvoisin C., Fauchet R. HLA-G class I gene expression in normal and malignant hematopoietic cells. Hum. Immunol., 59: 524-528, 1998.[Medline]
  36. Carosella E. D., Rouas-Freiss N., Paul P., Dausset J. HLA-G: a tolerance molecule from the major histocompatibility complex. Immunol. Today, 20: 60-62, 1999.[Medline]
  37. Fukushima Y., Oshika Y., Nakamura M., Tokunaga T., Hatanaka H., Abe Y., Yamazaki H., Kijima H., Ueyama Y., Tamaoki N. Increased expression of human histocompatibility leukocyte antigen-G in colorectal cancer cells. Int. J. Mol. Med., 2: 349-351, 1998.[Medline]
  38. Amiot L., Onno M., Renard I., Drénou B., Guillaudeux T., Lebouteiller P., Fauchet R. HLA-G transcription studies during the different stages of normal and malignant hematopoiesis. Tissue Antigens, 47: 408-413, 1996.[Medline]
  39. Hahne M., Rimoldi D., Schröter M., Romero P., Schreier M., French L. E., Schneider P., Bornand T., Fontana A., Lienard D., Cerottini J. C., Tschopp J. Melanoma cell expression of Fas (Apo-1/CD95) ligand: implications for tumor escape. Science, 274: 1363 1996.[Abstract/Free Full Text]
  40. Chouaib S., Asselin-Paturel C., Mami-Chouaib F., Caignard A., Blay J. I. The host-tumor immune conflict: from immunosuppression to resistance and destruction. Immunol. Today, 18: 493-497, 1997.[Medline]
  41. Staveley-O’Carroll K., Sotomayor E., Montgomery J., Borrelo I., Hwang L., Fein S., Pardoll D., Levitsky H. Induction of antigen-specific T cell anergy: an early event in the course of tumor progression. Proc. Natl. Acad. Sci. USA, 95: 1178-1183, 1998.[Abstract/Free Full Text]
  42. De Smet C., De Backer O., Faraoni I., Lurquin C., Brasseur F., Boon T. The activation of human gene MAGE-1 in tumor cells is correlated with genome-wide demethylation. Proc. Natl. Acad. Sci. USA, 93: 7149-7153, 1996.[Abstract/Free Full Text]
  43. Cosman D., Fanger N., Borges L., Kubin M., Chin W., Peterson L., Hsu M-l. A novel Ig superfamily receptor for cellular and viral MHC class I molecules. Immunity, 7: 273-283, 1997.[Medline]
  44. Yokoyama W. M. HLA class I specificity for natural killer cell receptor CD94/NKG2A: two for one in more ways than one. Proc. Natl. Acad. Sci. USA, 95: 4791-4794, 1998.[Free Full Text]
  45. Chu W., Fant M. E., Geraghty D. E., Hunt J. S. Soluble HLA-G in human placentas: synthesis in trophoblasts and interferon-{gamma}-activated macrophages but not placental fibroblasts. Hum. Immunol., 59: 435-442, 1998.[Medline]
  46. Houlihan J. M., Biro P. A., Harper H. M., Jenkinson H. J., Holmes C. H. The human amnion is a site of MHC class Ib expression: evidence for the expression of HLA-E and HLA-G. J. Immunol., 154: 5665-5674, 1995.[Abstract]
  47. Brasoveanu L. I., Fonsatti E., Vasintin A., Pavlovic M., Cattarossi I., Colizzi F., Gasparollo A., Coral S., Horejsi V., Altomonte M., Maio M. Melanoma cells constitutively release an anchor-positive soluble form of protein (sCD59) that retains functional activities in homologous complement-mediated cytotoxicity. J. Clin. Invest., 100: 1248-1255, 1997.[Medline]
  48. Bernhard H., Meyer zum Buschenfelde K., Dippold W. G. Ganglioside GD3 shedding by human malignant melanoma cells. Int. J. Cancer, 44: 155-160, 1989.[Medline]
  49. Westhoff U., Fox C., Otto F. Soluble HLA class I antigens in plasma of patients with malignant melanoma. Anticancer Res., 18: 3789-3792, 1998.[Medline]
  50. Becker J., Brocker E. Lymphocyte-melanoma interaction: role of surface molecules. Rec. Results Cancer Res., 139: 105-114, 1995.[Medline]
  51. Carbone E., Terrazzano G., Colonna M., Tuosto L., Piccolella E., Franksson L., Pérez-Villar J. J., Fontana S., Kärre K., Zappacosta S. Natural killer clones recognize specific soluble HLA class I molecules. Eur. J. Immunol., 26: 683-689, 1996.[Medline]
  52. Mc Donald J., Adamashvili I. Soluble HLA: a review of the literature. Hum. Immunol., 59: 387-403, 1998.[Medline]
  53. Zavazava N., Krönke M. Soluble HLA class I molecules induce apoptosis in alloreactive cytotoxic T lymphocytes. Nat. Med., 2: 1005-1010, 1996.[Medline]
  54. Puppo F., Indiveri F., Scudeletti M., Ferrone S. Soluble HLA-G antigens: new roles and uses. Immunol. Today, 18: 154-155, 1997.[Medline]



This article has been cited by other articles:


Home page
 Ann OncolHome page
A. Lin, W.-H. Yan, H.-H. Xu, M.-F. Gan, J.-F. Cai, M. Zhu, and M.-Y. Zhou
HLA-G expression in human ovarian carcinoma counteracts NK cell function
Ann. Onc., November 1, 2007; 18(11): 1804 - 1809.
[Abstract] [Full Text] [PDF]

Home page
 Cancer Res.Home page
N. Rouas-Freiss, P. Moreau, S. Ferrone, and E. D. Carosella
HLA-G Proteins in Cancer: Do They Provide Tumor Cells with an Escape Mechanism?
Cancer Res., November 15, 2005; 65(22): 10139 - 10144.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
A. Tajima, T. Tanaka, T. Ebata, K. Takeda, A. Kawasaki, J. M. Kelly, P. K. Darcy, R. E. Vance, D. H. Raulet, K. Kinoshita, et al.
Blastocyst MHC, a Putative Murine Homologue of HLA-G, Protects TAP-Deficient Tumor Cells from Natural Killer Cell-Mediated Rejection In Vivo
J. Immunol., August 15, 2003; 171(4): 1715 - 1721.
[Abstract] [Full Text] [PDF]

Home page
 Cancer Res.Home page
J. Bukur, V. Rebmann, H. Grosse-Wilde, H. Luboldt, H. Ruebben, I. Drexler, G. Sutter, C. Huber, and B. Seliger
Functional Role of Human Leukocyte Antigen-G Up-Regulation in Renal Cell Carcinoma
Cancer Res., July 15, 2003; 63(14): 4107 - 4111.
[Abstract] [Full Text] [PDF]

Home page
 J. Clin. Pathol.Home page
E H Kemp, R A Metcalfe, P F Watson, and A P Weetman
HLA-G does not have a pathophysiological role in Graves' disease
J. Clin. Pathol., June 1, 2003; 56(6): 475 - 477.
[Abstract] [Full Text] [PDF]

Home page
 Proc. Natl. Acad. Sci. USAHome page
P. Moreau, G. Mouillot, P. Rousseau, C. Marcou, J. Dausset, and E. D. Carosella
HLA-G gene repression is reversed by demethylation
PNAS, February 4, 2003; 100(3): 1191 - 1196.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Pathol.Home page
E. C. Ibrahim, Y. Allory, F. Commo, B. Gattegno, P. Callard, and P. Paul
Altered Pattern of Major Histocompatibility Complex Expression in Renal Carcinoma: Tumor-Specific Expression of the Nonclassical Human Leukocyte Antigen-G Molecule Is Restricted to Clear Cell Carcinoma While Up-Regulation of Other Major Histocompatibility Complex Antigens Is Primarily Distributed in All Subtypes of Renal Carcinoma
Am. J. Pathol., February 1, 2003; 162(2): 501 - 508.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
H. Wiendl, M. Mitsdoerffer, V. Hofmeister, J. Wischhusen, A. Bornemann, R. Meyermann, E. H. Weiss, A. Melms, and M. Weller
A Functional Role of HLA-G Expression in Human Gliomas: An Alternative Strategy of Immune Escape
J. Immunol., May 1, 2002; 168(9): 4772 - 4780.
[Abstract] [Full Text] [PDF]

Home page
 BloodHome page
M. Urosevic, J. Willers, B. Mueller, W. Kempf, G. Burg, and R. Dummer
HLA-G protein up-regulation in primary cutaneous lymphomas is associated with interleukin-10 expression in large cell T-cell lymphomas and indolent B-cell lymphomas
Blood, January 15, 2002; 99(2): 609 - 617.
[Abstract] [Full Text] [PDF]

Home page
 IOVSHome page
H. M. H. Hurks, M. M. Valter, L. Wilson, I. Hilgert, P. J. van den Elsen, and M. J. Jager
Uveal Melanoma: No Expression of HLA-G
Invest. Ophthalmol. Vis. Sci., December 1, 2001; 42(13): 3081 - 3084.
[Abstract] [Full Text] [PDF]

Home page
 Proc. Natl. Acad. Sci. USAHome page
N. Lila, N. Rouas-Freiss, J. Dausset, A. Carpentier, and E. D. Carosella
Soluble HLA-G protein secreted by allo-specific CD4+ T cells suppresses the allo-proliferative response: A CD4+ T cell regulatory mechanism
PNAS, September 19, 2001; (2001) 201407398.
[Abstract] [Full Text] [PDF]

Home page
 Cancer Res.Home page
E. C. Ibrahim, N. Guerra, M.-J. T. Lacombe, E. Angevin, S. Chouaib, E. D. Carosella, A. Caignard, and P. Paul
Tumor-specific Up-Regulation of the Nonclassical Class I HLA-G Antigen Expression in Renal Carcinoma
Cancer Res., September 1, 2001; 61(18): 6838 - 6845.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Pathol.Home page
M. Urosevic, M. O. Kurrer, J. Kamarashev, B. Mueller, W. Weder, G. Burg, R. A. Stahel, R. Dummer, and A. Trojan
Human Leukocyte Antigen G Up-Regulation in Lung Cancer Associates with High-Grade Histology, Human Leukocyte Antigen Class I Loss and Interleukin-10 Production
Am. J. Pathol., September 1, 2001; 159(3): 817 - 824.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Pathol.Home page
S. Aractingi, N. Briand, C. Le Danff, M. Viguier, H. Bachelez, L. Michel, L. Dubertret, and E. D. Carosella
HLA-G and NK Receptor Are Expressed in Psoriatic Skin : A Possible Pathway for Regulating Infiltrating T Cells?
Am. J. Pathol., July 1, 2001; 159(1): 71 - 77.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
B. Riteau, N. Rouas-Freiss, C. Menier, P. Paul, J. Dausset, and E. D. Carosella
HLA-G2, -G3, and -G4 Isoforms Expressed as Nonmature Cell Surface Glycoproteins Inhibit NK and Antigen-Specific CTL Cytolysis
J. Immunol., April 15, 2001; 166(8): 5018 - 5026.
[Abstract] [Full Text] [PDF]

Home page
 Int ImmunolHome page
B. Riteau, C. Menier, I. Khalil-Daher, S. Martinozzi, M. Pla, J. Dausset, E. D. Carosella, and N. Rouas-Freiss
HLA-G1 co-expression boosts the HLA class I-mediated NK lysis inhibition
Int. Immunol., February 1, 2001; 13(2): 193 - 201.
[Abstract] [Full Text] [PDF]

Home page
 Biol. Reprod.Home page
J. Copeman, R. N.N. Han, I. Caniggia, M. McMaster, S. J. Fisher, and J. C. Cross
Posttranscriptional Regulation of Human Leukocyte Antigen G During Human Extravillous Cytotrophoblast Differentiation
Biol Reprod, June 1, 2000; 62(6): 1543 - 1550.
[Abstract] [Full Text]

Home page
 NEJMHome page
E. D. Carosella, J. Dausset, and N. Rouas-Freiss
Transplacental Transmission of Natural-Killer-Cell Lymphoma
N. Engl. J. Med., December 16, 1999; 341(25): 1937 - 1937.
[Full Text]

Home page
 J. Biol. Chem.Home page
S. Lefebvre, S. Berrih-Aknin, F. Adrian, P. Moreau, S. Poea, L. Gourand, J. Dausset, E. D. Carosella, and P. Paul
A Specific Interferon (IFN)-stimulated Response Element of the Distal HLA-G Promoter Binds IFN-regulatory Factor 1 and Mediates Enhancement of This Nonclassical Class I Gene by IFN-beta
J. Biol. Chem., February 23, 2001; 276(9): 6133 - 6139.
[Abstract] [Full Text] [PDF]

Home page
 Proc. Natl. Acad. Sci. USAHome page
N. Lila, N. Rouas-Freiss, J. Dausset, A. Carpentier, and E. D. Carosella
Soluble HLA-G protein secreted by allo-specific CD4+ T cells suppresses the allo-proliferative response: A CD4+ T cell regulatory mechanism
PNAS, October 9, 2001; 98(21): 12150 - 12155.
[Abstract] [Full Text] [PDF]

Home page
 CirculationHome page
N. Lila, C. Amrein, R. Guillemain, P. Chevalier, C. Latremouille, J.-N. Fabiani, J. Dausset, E. D. Carosella, and A. Carpentier
Human Leukocyte Antigen-G Expression After Heart Transplantation Is Associated With a Reduced Incidence of Rejection
Circulation, April 23, 2002; 105(16): 1949 - 1954.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Paul, P.
Right arrow Articles by Carosella, E. D.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Paul, P.
Right arrow Articles by Carosella, E. D.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Cancer Research Clinical Cancer Research
Cancer Epidemiology Biomarkers & Prevention Molecular Cancer Therapeutics
Molecular Cancer Research Cancer Prevention Research
Cancer Prevention Journals Portal Cancer Reviews Online
Annual Meeting Education Book Meeting Abstracts Online