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1.
MAbs ; 16(1): 2329321, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38494955

RESUMO

Antibody-mediated delivery of immunogenic viral CD8+ T-cell epitopes to redirect virus-specific T cells toward cancer cells is a promising new therapeutic avenue to increase the immunogenicity of tumors. Multiple strategies for viral epitope delivery have been shown to be effective. So far, most of these have relied on a free C-terminus of the immunogenic epitope for extracellular delivery. Here, we demonstrate that antibody-epitope conjugates (AECs) with genetically fused epitopes to the N-terminus of the antibody can also sensitize tumors for attack by virus-specific CD8+ T cells. AECs carrying epitopes genetically fused at the N-terminus of the light chains of cetuximab and trastuzumab demonstrate an even more efficient delivery of the T-cell epitopes compared to AECs with the epitope fused to the C-terminus of the heavy chain. We demonstrate that this increased efficiency is not caused by the shift in location of the cleavage site from the N- to the C-terminus, but by its increased proximity to the cell surface. We hypothesize that this facilitates more efficient epitope delivery. These findings not only provide additional insights into the mechanism of action of AECs but also broaden the possibilities for genetically fused AECs as an avenue for the redirection of multiple virus-specific T cells toward tumors.


Assuntos
Imunoconjugados , Neoplasias , Humanos , Epitopos de Linfócito T , Linfócitos T CD8-Positivos , Anticorpos , Neoplasias/terapia
2.
Arch Virol ; 148(10): 2023-37, 2003 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-14551822

RESUMO

Bovine herpesvirus 1 (BoHV-1), the cause of infectious bovine rhinotracheitis and infectious pustular vulvovaginitis in cattle, establishes a lifelong infection, despite the presence of antiviral immunity in the host. BoHV-1 has been shown to elude the host immune system, but the viral gene products responsible for this interference have not yet been identified. Studies aiming at the identification of BoHV-1-encoded immune evasion genes have been hampered by the lack of bovine-specific immunological reagents. Some of the immune evasion molecules identified for other herpesviruses are host species specific; others can act across the species barrier. In this study, experiments were performed to investigate whether BoHV-1 can infect human cells and interfere with antigen processing and presentation in these cells. A human melanoma cell line, Mel JuSo, appeared to be permissive for BoHV-1 infection. BoHV-1 induced expression of major viral glycoproteins at the surface of these cells and produced progeny virus up to 10(5) plaque forming units per ml. BoHV-1 infection resulted in impaired intracellular transport of human MHC class I molecules and inhibition of human TAP. These data indicate that the BoHV-1-encoded molecule(s) that block antigen presentation in bovine cells are able to interact with homologous components of the human MHC class I presentation pathway. The fact that immune evasion by BoHV-1 can be studied in human cells will facilitate the identification of the BoHV-1 gene products involved in this process. Moreover, the data presented here suggest that the BoHV-1 encoded inhibitors of antigen presentation represent potential immune suppressive agents for use in humans.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Herpesvirus Bovino 1/patogenicidade , Antígenos de Histocompatibilidade Classe I/metabolismo , Peptídeos/metabolismo , Animais , Bovinos , Linhagem Celular , Humanos , Transporte Proteico , Células Tumorais Cultivadas , Replicação Viral
3.
Int J Cancer ; 91(5): 612-8, 2001 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-11267969

RESUMO

Tumor-specific T-helper (Th) immunity was found to play a pivotal role in the natural and vaccine-induced immune defense against tumors. Since the majority of cervical cancers express human papillomavirus type 16 (HPV16) E7 oncoprotein, it is important to investigate the Th response against this target antigen in detail. By means of PBMC cultures from HLA-typed healthy donors, we identified the central part of HPV16 E7 (E7(41-72)) as the major immunogenic region within this antigen. Furthermore, we mapped 3 distinct Th epitopes within this region (DR15/E7(50-62), DR3/E7(43-77), DQ2/E7(35-50)). In a parallel approach, employing IFN-gamma ELISPOT analysis, we detected Th immunity against HPV16 E7 in subjects with HPV16+ lesions. Several of these responses matched with the 3 Th epitopes defined in our study. A number of other HPV16+ subjects did not display any E7-specific type 1 cytokine-producing T-cell immunity, indicating failure of the immune response. Our combined data argue for more extensive as well as longitudinal analysis of HPV16-specific T-cell immunity using the ELISPOT assay described, as well as for HPV-specific vaccination of individuals with HPV+ lesions.


Assuntos
Complexo Principal de Histocompatibilidade , Proteínas Oncogênicas Virais/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Linfócitos T Auxiliares-Indutores/metabolismo , Neoplasias do Colo do Útero/virologia , Vacinas Anticâncer , Carcinoma/química , Divisão Celular , Células Cultivadas , Citocinas/metabolismo , Mapeamento de Epitopos , Epitopos/química , Feminino , Genes MHC da Classe II , Antígenos HLA-DQ/química , Antígenos HLA-DR/química , Subtipos Sorológicos de HLA-DR , Antígeno HLA-DR3/química , Humanos , Memória Imunológica , Imunofenotipagem , Concentração Inibidora 50 , Interferon gama/metabolismo , Leucócitos Mononucleares/metabolismo , Proteínas Oncogênicas Virais/química , Proteínas E7 de Papillomavirus , Peptídeos/química , Peptídeos/metabolismo , Ligação Proteica , Estrutura Terciária de Proteína , Proteínas Recombinantes/metabolismo , Recidiva , Linfócitos T Auxiliares-Indutores/química , Neoplasias do Colo do Útero/química
4.
J Immunother ; 23(2): 255-66, 2000.
Artigo em Inglês | MEDLINE | ID: mdl-10746552

RESUMO

Human papillomavirus type 16 (HPV16)-encoded E7 oncoprotein is constitutively expressed in cervical carcinoma cells and is required for cellular transformation to be maintained. The E7 protein, therefore, forms an attractive target for T-cell-mediated immune intervention to prevent or treat HPV16+ tumors. The authors performed a peptide-based phase I/II vaccination trial to induce anti-tumor immune responses in patients with recurrent or residual cervical carcinoma. Fifteen HLA-A*0201+ patients with HPV16+ cervical carcinoma received vaccinations with synthetic peptides representing 2 HPV16 E7-encoded, HLA-A*0201-restricted cytotoxic T lymphocyte epitopes and a pan-HLA-DR-binding T-helper epitope, PADRE, in adjuvant. No signs of toxicity were observed. Two patients had stable disease for more than 1 year after vaccination, 3 patients died of the disease during or shortly after the vaccination period, and 10 patients maintained progressive cervical carcinoma. Specific immune responses directed against the vaccine components were analyzed in peripheral blood samples. No cytotoxic T lymphocyte responses against the HPV16 E7 peptides were detectable. After vaccination, strong PADRE helper peptide-specific proliferation was detected in 4 of 12 patients. In conclusion, peptide vaccination with 2 HPV16 E7 cytotoxic T lymphocyte epitopes and a universal T helper epitope is well tolerated by patients with advanced cervical carcinoma. Despite a reduction of in vitro cytolytic or proliferative recall responses to some, but not all, conventional antigens in this patient group, peptide-specific proliferative responses were induced in 4 patients. Based on the current study, it is now feasible to perform peptide vaccination in earlier stages of HPV16-induced cervical disease.


Assuntos
Vacinas Anticâncer/uso terapêutico , Carcinoma/imunologia , Papillomaviridae/imunologia , Peptídeos/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Neoplasias do Colo do Útero/imunologia , Adulto , Idoso , Carcinoma/terapia , Linhagem Celular Transformada , Epitopos de Linfócito T/imunologia , Feminino , Humanos , Células K562 , Vacinas Antimaláricas/biossíntese , Vacinas Antimaláricas/imunologia , Pessoa de Meia-Idade , Proteínas Oncogênicas Virais/biossíntese , Proteínas Oncogênicas Virais/imunologia , Orthomyxoviridae/imunologia , Proteínas E7 de Papillomavirus , Peptídeos/uso terapêutico , Linfócitos T Citotóxicos/imunologia , Células Tumorais Cultivadas , Neoplasias do Colo do Útero/terapia
5.
Eur J Cancer ; 35(6): 946-52, 1999 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-10533477

RESUMO

A phase I-II clinical trial was performed involving vaccination with HPV16 E7 peptides of patients suffering from HPV16 positive cervical carcinoma which was refractory to conventional treatment. Patients receiving the vaccine were HLA-A*0201 positive with HPV16 positive cervical carcinoma. The clinical trial was designed as a dose-escalation study, in which successive groups of patients received 100 micrograms, 300 micrograms or 1000 micrograms of each peptide, respectively. The vaccine consisted of two HPV16 E7 peptides and one helper peptide emulsified in Montanide ISA 51 adjuvant. 19 patients were included in the study, no adverse side-effects were observed. 2 patients showed stable disease for 1 year after vaccination; 15 patients showed progressive disease of whom 1 died during the vaccination treatment due to progressive disease; and 2 patients showed tumour-regression after chemotherapy following vaccination. A relative low count of lymphocytes before and after vaccination was present in 11/19 patients indicating that these patients were immunocompromised. This study shows that HPV16 E7 peptide vaccination is feasible, even in a group of patients with terminal disease. This paves the way for vaccinating patients with less advanced disease, whose immune system is less compromised by progressive disease.


Assuntos
Vacinas Anticâncer/uso terapêutico , Proteínas Oncogênicas Virais/imunologia , Papillomaviridae/imunologia , Neoplasias do Colo do Útero/tratamento farmacológico , Vacinas Virais/uso terapêutico , Adulto , Idoso , Progressão da Doença , Feminino , Humanos , Imunoterapia , Pessoa de Meia-Idade , Proteínas E7 de Papillomavirus , Resultado do Tratamento , Neoplasias do Colo do Útero/virologia
6.
Eur J Immunol ; 29(4): 1292-303, 1999 04.
Artigo em Inglês | MEDLINE | ID: mdl-10229097

RESUMO

Several cancer immune intervention protocols aim at inducing T cell immunity against antigens presented by HLA-A2, the most common human MHC class I molecule. In the context of HLA-A*0201, we previously identified two cytotoxic T lymphocyte epitopes (E7(11-20) and E7(86-93)) encoded by the human papillomavirus type 16 E7 (HPV16 E7) oncoprotein, which is a tumor-specific antigen for cervical carcinoma. This study reports that the two HPV16 epitopes and a control hepatitis B virus epitope bind equally well to five HLA-A2 alleles (A*0201, A*0202, A*0203, A*0204, and A*0209). These HLA-A2 variants display comparable binding characteristics in accordance with the A2 supertype (M. F. Del Guercio et al., J. Immunol. 1995. 154: 685-693). Cervical carcinoma patients expressing these alleles may benefit from vaccination with the two HPV16 E7 peptides. In contrast, none of the peptides tested bound to A*0207 or A*0208, whereas heterogeneous binding was observed for A*0205 and A*0206. Therefore, the amino acid substitutions that discriminate these HLA-A2 variants from A*0201 affect antigen presentation. Taken together, our findings have implications for application of the A2 supertype concept and for vaccination with A*0201-binding peptides, in particular HPV16 E7 peptides.


Assuntos
Alelos , Antígeno HLA-A2/genética , Proteínas Oncogênicas Virais/imunologia , Neoplasias do Colo do Útero/terapia , Vacinas Virais/imunologia , Linhagem Celular , Epitopos de Linfócito T , Feminino , Antígeno HLA-A2/análise , Humanos , Proteínas Oncogênicas Virais/metabolismo , Proteínas E7 de Papillomavirus , Linfócitos T Citotóxicos/imunologia , Neoplasias do Colo do Útero/imunologia , Vacinação
7.
Cancer Res ; 58(4): 724-31, 1998 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-9485027

RESUMO

Human papillomavirus (HPV) E6 and E7 oncoproteins are attractive targets for T-cell-based immunotherapy of cervical cancer. In this study, we demonstrate that dendritic cells (DCs) pulsed with HPV16 E7 protein are not only recognized in vitro by E7-specific CTLs but also elicit E7-specific CTL responses in vivo, associated with protection against a challenge with syngeneic HPV16-induced tumor cells. Vaccination with soluble E7 protein in incomplete Freund's adjuvant likewise induces E7-specific CTL responses associated with tumor protection. The presence of HPV16 E7-specific CTLs in vivo and the observation that depletion of CD8+ cells completely abolishes tumor protection demonstrate that CTLs are the major effector cells in mediating antitumor activity. The in vivo involvement of DCs in the activation of protective CTLs is suggested by the surface display of E7 peptide-loaded MHC class I molecules on these cells after E7 protein immunization. These data show that HPV16 E7 protein-pulsed DCs, as well as the administration of E7 protein antigen in adjuvant, can effectively stimulate tumor-specific MHC class I-restricted CD8+ T-cell-mediated protective immunity to HPV16-induced cancers.


Assuntos
Vacinas Anticâncer/imunologia , Células Dendríticas/imunologia , Adjuvante de Freund/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Proteínas Oncogênicas Virais/imunologia , Papillomaviridae/patogenicidade , Infecções Tumorais por Vírus/prevenção & controle , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Citotoxicidade Imunológica , Camundongos , Camundongos Endogâmicos BALB C , Proteínas E7 de Papillomavirus
8.
Proc Natl Acad Sci U S A ; 94(26): 14660-5, 1997 Dec 23.
Artigo em Inglês | MEDLINE | ID: mdl-9405669

RESUMO

Vaccines harboring genes that encode functional oncoproteins are intrinsically hazardous, as their application may lead to introduction of these genes into normal cells and thereby to tumorigenesis. On the other hand, oncoproteins are especially attractive targets for immunotherapy of cancer, as their expression is generally required for tumor growth, making the arisal of tumor variants lacking these antigens unlikely. Using murine tumor models, we investigated the efficacy of polyepitope recombinant adenovirus (rAd) vaccines, which encode only the immunogenic T cell epitopes derived from several oncogenes, for the induction of protective anti-tumor immunity. We chose to employ rAd, as these are safe vectors that do not induce the side effects associated with, for example, vaccinia virus vaccines. A single polyepitope rAd was shown to give rise to presentation of both H-2 and human leukocyte antigen-restricted cytotoxic T lymphocyte (CTL) epitopes. Moreover, vaccination with a rAd encoding H-2-restricted CTL epitopes, derived from human adenovirus type 5 early region 1 and human papilloma virus type 16-induced tumors, elicited strong tumor-reactive CTL and protected the vaccinated animals against an otherwise lethal challenge with either of these tumors. The protection induced was superior compared with that obtained by vaccination with irradiated tumor cells. Thus, vaccination with polyepitope rAd is a powerful approach for the induction of protective anti-tumor immunity that allows simultaneous immunization against multiple tumor-associated T cell epitopes, restricted by various major histocompatibility complex haplotypes.


Assuntos
Citotoxicidade Imunológica , Epitopos de Linfócito T/imunologia , Neoplasias Experimentais/imunologia , Linfócitos T/imunologia , Adenoviridae , Animais , Sequência de Bases , DNA Recombinante , Humanos , Camundongos , Dados de Sequência Molecular , Neoplasias Experimentais/prevenção & controle , Oligonucleotídeos , Vacinação
9.
Ann Med ; 28(6): 471-7, 1996 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-9017105

RESUMO

Cervical carcinoma is the second most common cause of cancer-related deaths in women worldwide. Recurrences occur in 15% of the patients after optimal treatment of low-risk early-stage disease. Treatment results of recurrent disease are relatively poor and for this reason new therapeutic strategies are warranted. Viral infection with human papillomavirus seems to have an essential part in the aetiology of cervical carcinoma. Evidence for the assumption that cervical carcinoma, among other malignancies such as melanomas, renal malignancies and Kaposi sarcoma, are immunogenic is provided by the fact that these malignancies grow more rapidly in the presence of systemic immunosuppression. Spontaneous regression for these tumour types is also described and immunohistochemical studies show extensive infiltrates in the tumour, consisting of immunocompetent cells. It is thus postulated that cellular immunity, and mainly the T-cell system plays an important role in the antitumour defence in cervical carcinoma. This review describes the rationale for the use of immunotherapy as treatment for cervical carcinoma as well as the results of recent developments in tumour immunology and its implications for the clinical use of immunotherapeutical approaches.


Assuntos
Imunoterapia , Papillomaviridae/imunologia , Infecções por Papillomavirus/terapia , Vacinas contra Papillomavirus , Infecções Tumorais por Vírus/terapia , Neoplasias do Colo do Útero/terapia , Vacinas Virais/administração & dosagem , Animais , Feminino , Humanos , Infecções por Papillomavirus/complicações , Infecções Tumorais por Vírus/complicações , Neoplasias do Colo do Útero/etiologia , Neoplasias do Colo do Útero/imunologia
10.
Clin Cancer Res ; 2(11): 1825-8, 1996 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-9816136

RESUMO

An impaired immune response is frequently observed in patients and experimental animals with advanced cancer. We and others have shown alterations in CD3-associated signal-transducing zeta molecules in tumor-infiltrating T cells and peripheral blood lymphocytes (PBLs) of patients with advanced cancer. By using flow cytometric analysis of permeabilized cells with a monoclonal antibody (TIA-2) that reacts with the cytoplasmic domain of the zeta chain, here we demonstrate a marked decrease (P < 0.01) in the expression of the signal-transducing CD3 zeta chain of PBLs in patients with cervical cancer (n = 22) as compared to PBLs from healthy donors (n = 21). In addition, PBLs isolated from patients (n = 23) with cervical intraepithelial neoplasia (CIN), to a lesser but significant (P < 0. 01) extent, expressed reduced CD3 zeta levels as compared to those from healthy donors. This decreased expression of zeta chains was also observed on CD16(+) natural killer cells in PBLs from patients with cervical cancer. Surface expression of CD3 epsilon on PBLs was also decreased in cervical cancer patients as compared to healthy donors, but not on PBLs from patients with CIN. CD3 zeta chain expression significantly (r = 0.53, P < 0.01) correlated with the ability of the PBLs to produce tumor necrosis factor in response to anti-CD3 stimulation. These findings suggest that alterations of signal-transducing zeta molecules commonly occur in patients with cervical cancer and to a lesser extent with CIN, and that they are associated with reduced cellular functions such as production of tumor necrosis factor.


Assuntos
Complexo CD3/metabolismo , Células Matadoras Naturais/metabolismo , Linfócitos T/metabolismo , Neoplasias do Colo do Útero/imunologia , Anticorpos Monoclonais/imunologia , Complexo CD3/imunologia , Feminino , Humanos , Linfócitos do Interstício Tumoral/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores de IgG/metabolismo , Transdução de Sinais , Fator de Necrose Tumoral alfa/metabolismo
11.
Immunotechnology ; 2(4): 241-51, 1996 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-9373306

RESUMO

Recent progress in defining the molecular nature of antigens and in finding ways to manipulate T cell-mediated immune responses may provide new modalities for cancer treatment. In this report, we review preclinical studies as well as the first clinical trials with vaccination strategies aiming at the induction of anti-tumor immunity. In particular, we focus on the development of a vaccine against human papillomavirus-induced cervical carcinoma.


Assuntos
Vacinas Anticâncer/uso terapêutico , Imunoterapia Ativa , Neoplasias/imunologia , Neoplasias/prevenção & controle , Linfócitos T/imunologia , Animais , Ensaios Clínicos como Assunto , Humanos
12.
Cancer Res ; 56(3): 582-8, 1996 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-8564975

RESUMO

Most cervical carcinoma (Cxca) cells constitutively express human papillomavirus type 16 (HPV16) E6 and E7 oncoproteins. These proteins are, therefore, attractive targets for T cell-based immunotherapy. Previously, we identified HVP16 E7-encoded CTL epitopes. In patients with cervical intraepithelial neoplasia or Cxca, little is known concerning T-cell activity against viruses in general and against HPV16 in particular. Here, we have screened the blood of 10 healthy donor controls and of 22 patients with HPV16+ cervical lesions for the presence of CTLs directed against HPV16 E7- and control influenza virus matrix-derived epitopes presented by HLA-A *0201. We detected influenza virus-specific CTLs in all donors and in the majority of patients, indicating that most patients have functioning T-cell responses despite their lesions or therapeutic interventions. Moreover, we show that patients with HPV16+ lesions occasionally have memory CTLs against a HPV16 E7-encoded epitope (sequence YMLD-LQPETT), providing evidence for natural CTL immunity against HPV16 in patients with cervical lesions. Combined, these findings raise possibilities for vaccination with HPV16 E7-encoded peptides to induce or augment CTL responses for treatment or prevention of Cxca.


Assuntos
Epitopos/imunologia , Antígenos HLA-A/imunologia , Memória Imunológica/imunologia , Proteínas Oncogênicas Virais/imunologia , Papillomaviridae/imunologia , Linfócitos T Citotóxicos/imunologia , Neoplasias do Colo do Útero/imunologia , Neoplasias do Colo do Útero/virologia , Sequência de Aminoácidos , Feminino , Humanos , Vírus da Influenza A/imunologia , Dados de Sequência Molecular , Proteínas E7 de Papillomavirus , Displasia do Colo do Útero/imunologia , Displasia do Colo do Útero/virologia
14.
J Immunol ; 154(11): 5934-43, 1995 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-7538538

RESUMO

Human papillomavirus type 16 (HPV16) is strongly associated with cervical carcinogenesis. The HPV16 E6 and E7 oncoproteins are constitutively expressed in the majority of cervical tumor cells and are, therefore, attractive targets for CTL-mediated immunotherapy. In mice, the outgrowth of a lethal dose of HPV16-induced tumor cells has been prevented by vaccination with a CTL epitope encoded by HPV16 E7, indicating the feasibility of peptide immunization to obtain antitumor CTL responses. In the present study, the immunogenicity of 9 HLA-A*0201-binding peptides encoded by HPV16 E6 and E7 was analyzed in vivo in HLA-A*0201Kb transgenic mice and in vitro in CTL cultures induced from PBMC of HLA-A*0201+ healthy donors. Four peptides with a good binding affinity were immunogenic in HLA-A*0201Kb transgenic mice, and three of them were also highly immunogenic in CTL induction experiments with PBMC of HLA-A*0201+ healthy donors. Human CTL clones specific for these three peptides were capable of lysing the HPV16 E7-containing HLA-A*0201+ cervical carcinoma cell line CaSki. These E7-derived peptides (11-20, YMLDLQPETT; 82-90, LLMGTLGIV; 86-93, TLGIVCPI), therefore, are likely to represent naturally processed human CTL epitopes of HPV16. Additionally, these three HPV16-encoded peptides have the highest affinity of binding to the HLA-A*0201 molecule. In this study, peptides with a lower binding affinity were less immunogenic. Therefore, our data illustrate that the HLA-binding affinity of a peptide has a major impact on its immunogenicity. In conclusion, we have identified immunogenic peptides encoded by HPV16 E6 and E7 that could be used in vaccines for the prevention and treatment of cervical carcinoma.


Assuntos
Epitopos/imunologia , Antígenos HLA-A/imunologia , Proteínas Oncogênicas Virais/imunologia , Papillomaviridae/imunologia , Proteínas Repressoras , Linfócitos T Citotóxicos/imunologia , Sequência de Aminoácidos , Animais , Células Cultivadas , Testes Imunológicos de Citotoxicidade , Humanos , Camundongos , Camundongos Transgênicos , Dados de Sequência Molecular , Proteínas E7 de Papillomavirus , Ligação Proteica/imunologia , Vacinas Virais/imunologia
15.
J Immunol ; 154(8): 3991-8, 1995 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-7706738

RESUMO

CTL that lyse melanoma cells were previously isolated from several melanoma patients. Such CTL recognize autologous proteins, indicating the occurrence of autoreactive T cells in melanoma patients. We have now raised CTL, using responding T lymphocytes from healthy donor blood, that lysed not only cells incubated with an HLA-A*0201-binding tyrosinase peptide but also melanoma cells endogenously processing and presenting the epitope. Our results suggest that autoreactive CTL precursors are present in healthy donor blood and can be activated in vitro with synthetic peptides presented on appropriate APCs. Therefore, tissue-specific, autoantigen-derived peptides might be useful in immunotherapy of both poorly and nonimmunogenic tumors.


Assuntos
Citotoxicidade Imunológica , Melanoma/imunologia , Monofenol Mono-Oxigenase/imunologia , Linfócitos T Citotóxicos/imunologia , Sequência de Aminoácidos , Autoantígenos/imunologia , Células Cultivadas , Células Clonais , Reações Cruzadas , Relação Dose-Resposta Imunológica , Antígenos HLA-A/imunologia , Humanos , Imunidade Celular , Técnicas In Vitro , Dados de Sequência Molecular , Peptídeos/química , Peptídeos/imunologia
16.
J Virol ; 68(9): 6038-46, 1994 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-7520098

RESUMO

Upon infection with the Moloney murine sarcoma virus-murine leukemia virus (MuLV) complex, H-2b C57BL/6 (B6) mice respond with a class I Db-restricted cytotoxic T-lymphocyte (CTL) response, which protects against virus-induced tumorigenesis. In the B6-derived Db mutant B6.CH-2bm13 (bm13) strain, part of the class I Db antigen-presenting groove is shaped by a class I Kb-encoded sequence. Like B6 mice, bm13 mice reject Moloney virus-induced tumors, but the protective CTL response is Kb restricted. In this study we show enhanced levels of Moloney MuLV-specific CTLp with a restriction for Kb in bm13 mice. Through the use of CTL clones from Moloney virus-immunized bm13 mice, the class I Kb-presented CTL epitope was identified. The epitope is located in the Moloney virus gp70 envelope protein region (Moloney envelope, amino acids 189 to 196 [Mol env (189-196)]), SSWDFITV and has the Kb allele-specific binding motif. The Dbm13 molecule does not present the env(189 to 196) epitope to Kb-restricted bm13 CTL. In B6 mice, Mol env(189-196)-specific CTL could be induced by peptide vaccination. B6 mice thus have CTL precursors specific for this epitope but at considerably lower levels than do bm13 mice. We hypothesize that additional positive selection of Kb-restricted CTL on the Dbm13 molecule in bm13 mice explains this difference in precursor frequencies. We examined related strains of MuLV for the presence of Mol env(189-196) sequence equivalents. Rauscher, Friend, and AKV MuLV-encoded Mol env(189-196) epitope equivalents were properly recognized in cytotoxicity assays, both as synthetic and as endogenously expressed (Rauscher MuLV) peptides. In contrast, the mink cell focus-forming virus MuLV-encoded epitope equivalent, lacking a Kb anchor residue, was not presented for CTL recognition and hence can be excluded as an important CTL epitope for mink cell focus-forming viruses.


Assuntos
Antígenos Virais/imunologia , Antígenos H-2/imunologia , Vírus da Leucemia Murina de Moloney/imunologia , Linfócitos T Citotóxicos/imunologia , Sequência de Aminoácidos , Animais , Células Apresentadoras de Antígenos/imunologia , Epitopos , Produtos do Gene env/imunologia , Antígenos H-2/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Dados de Sequência Molecular , Peptídeos/imunologia
17.
Pharm Res ; 9(2): 266-70, 1992 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-1372732

RESUMO

The influence of lyophilization on the stability of a monoclonal antibody (MN12) was investigated. MN12 was freeze-dried in different formulations [without lyoprotectant or in the presence of sucrose, dextran, or hydroxypropyl-beta-cyclodextrin (HP beta CD)] and under varying conditions (with or without secondary drying). Subsequently, the monoclonal antibody was stored for 18 or 32 days at various temperatures (4, 37, or 56 degrees C). For comparison, solutions of MN12 were stored under the same conditions. Regardless of the lyoprotectant used, precipitation and a concomitant reduction of the antigen-binding capacity by about 10% were observed upon reconstitution of lyophilized MN12. HP beta CD proved to be the most effective stabilizer to prevent degradation of lyophilized MN12 during storage. Compared with MN12 solutions, HP beta CD-containing lyophilized MN12 cakes were more resistant to heat-induced charge alterations and loss of antigen-binding capacity.


Assuntos
Anticorpos Monoclonais/química , Ciclodextrinas/química , Dextranos/química , Imunoglobulina G/química , Sacarose/química , beta-Ciclodextrinas , 2-Hidroxipropil-beta-Ciclodextrina , Cromatografia em Gel , Estabilidade de Medicamentos , Armazenamento de Medicamentos , Eletroforese em Gel de Poliacrilamida , Ensaio de Imunoadsorção Enzimática , Liofilização , Peso Molecular , Temperatura
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