RESUMEN
Idiotype vaccination for follicular lymphoma is primarily being developed as remission consolidation after chemotherapy. We investigated idiotype vaccination as primary intervention for treatment-naive indolent B-cell lymphoma and in a separate cohort as remission consolidation after chemotherapy to assess immunization-induced immune responses in relation to progression-free survival (German Clinical Trials Register, DRKS00000227). Twenty-one patients in each cohort received 6 intradermal injections of adjuvanted recombinant idiotype Fab fragment (Fab(Id)); 76% of patients in both groups developed anti-idiotype antibodies and/or cellular immunity as measured by enzyme-linked immunosorbent assay and interferon-γ ELISpot. In treatment-naive patients, only cellular responses correlated with superior progression-free survival (P < .002) and durable objective remissions (P = .04). Immunization-induced T cells recognized hypermutated or complementarity-determining region 3 epitopes. After remission consolidation immunization, induction of anti-idiotype antibodies correlated with progression-free survival. Low B-cell counts after rituximab therapy predicted for failure to develop anti-idiotype antibodies. These results are similar to published trials showing an association of humoral immunity with control of residual lymphoma. In contrast, effective immunity against untreated lymphoma appears to be dependent on idiotype-specific T cells. Sustained remissions in patients with vaccination-induced cellular immunity suggest clinical benefit and warrant a randomized comparison of this vaccine with expectant management for asymptomatic follicular lymphoma.
Asunto(s)
Vacunas contra el Cáncer/uso terapéutico , Fragmentos Fab de Inmunoglobulinas/inmunología , Idiotipos de Inmunoglobulinas/inmunología , Linfoma de Células B/inmunología , Linfoma de Células B/terapia , Proteínas Recombinantes/inmunología , Adulto , Anciano , Linfocitos B/inmunología , Epítopos de Linfocito T/inmunología , Femenino , Citometría de Flujo , Humanos , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Inducción de Remisión , Tasa de Supervivencia , Resultado del Tratamiento , VacunaciónRESUMEN
Active immunization with the idiotype of follicular lymphoma induces tumor-specific immunity. T cells induced in vivo by idiotype vaccination recognize human leukocyte antigen (HLA)--restricted hypervariable but not conserved idiotype peptides. We hypothesized that idiotype-directed T-cell immunity occurs naturally and performed a reverse immunology analysis of idiotype HLA binding in 39 follicular lymphoma patients. For every idiotype, the sum of HLA-A or -B binding scores of the 20 highest-scoring peptides was calculated for all 39 HLA types through the BIMAS algorithm. The idiotype sum score of every patient's lymphoma was compared on the respective patient's HLA type to the mean of the sum scores of the remaining 38 idiotypes. Autologous idiotypes had lower immunogenicity than allogeneic idiotypes. Differential immunogenicity resided predominantly in all 3 complementarity-determining regions rather than in framework peptides. Idiotype immunogenicity was not changed by somatic hypermutation. These findings indicate T cell-mediated immunosurveillance of follicular lymphoma directed specifically against individual idiotype epitopes.
Asunto(s)
Antígenos HLA/inmunología , Idiotipos de Inmunoglobulinas/inmunología , Linfoma Folicular/inmunología , Linfocitos T/inmunología , Adulto , Anciano , Algoritmos , Alelos , Epítopos de Linfocito T/genética , Epítopos de Linfocito T/inmunología , Epítopos de Linfocito T/metabolismo , Femenino , Antígenos HLA/genética , Antígenos HLA/metabolismo , Antígenos HLA-A/genética , Antígenos HLA-A/inmunología , Antígenos HLA-A/metabolismo , Antígenos HLA-B/genética , Antígenos HLA-B/inmunología , Antígenos HLA-B/metabolismo , Humanos , Idiotipos de Inmunoglobulinas/genética , Idiotipos de Inmunoglobulinas/metabolismo , Linfoma Folicular/genética , Linfoma Folicular/metabolismo , Masculino , Persona de Mediana Edad , Monitorización Inmunológica/estadística & datos numéricos , Unión Proteica , Linfocitos T/citología , Linfocitos T/metabolismoRESUMEN
OBJECTIVES: Individual immunoglobulins expressed by B-cell lymphomas represent tumor-specific antigens ('idiotypes'). Immunization with idiotype in follicular lymphoma patients may induce specific immune responses, sustained progression-free survival, and disappearance of minimal residual disease. Manufacturing of idiotype vaccines has mostly relied on heterohybridomas established from viable lymphoma cells. This paper describes the feasibility of production of GMP-grade idiotype vaccines as recombinant Fab fragments in Escherichia coli. METHODS: IgH and IgL transcripts were analyzed by anchored PCR from 106 lymphoma and nine control biopsies. Lymphoma-derived V segments were inserted into prokaryotic expression plasmids. Recombinant idiotype Fab fragments were expressed in E. coli in a fermentation system. RESULTS: Idiotype IgH and IgL transcripts were identified in 95% of 106 lymphoma biopsies according to stringent clonality criteria. Large-scale idiotype expression was successful in 69 of 78 cases (89%) and yielded a median of 17 mg (range: 1.2-250 mg) recombinant Fab protein. After affinity chromatography, median vaccine purity was 99% heterodimeric Fab protein (range: 72-100%). Bacterial protein contamination was detectable in one vaccine only. Fab proteins with IgL lambda chains had a tendency for inferior yield and lesser purity than kappa-type Fabs. Among other structural idiotype features (isotype, V family usage, somatic hypermutation pattern, novel glycosylation sites, CDR III net charge), no consistent influences on Fab yield or purity were detected. CONCLUSIONS: Anchored PCR cloning and subsequent protein expression in E. coli provides a reliable technological basis for clinical idiotype vaccination trials.
Asunto(s)
Vacunas contra el Cáncer/genética , Fragmentos Fab de Inmunoglobulinas/genética , Cadenas Pesadas de Inmunoglobulina/genética , Idiotipos de Inmunoglobulinas/genética , Cadenas Ligeras de Inmunoglobulina/genética , Linfoma de Células B/genética , Linfoma Folicular/genética , Vacunas contra el Cáncer/aislamiento & purificación , Vacunas contra el Cáncer/uso terapéutico , Escherichia coli/genética , Humanos , Fragmentos Fab de Inmunoglobulinas/aislamiento & purificación , Fragmentos Fab de Inmunoglobulinas/uso terapéutico , Cadenas Pesadas de Inmunoglobulina/aislamiento & purificación , Cadenas Pesadas de Inmunoglobulina/uso terapéutico , Idiotipos de Inmunoglobulinas/aislamiento & purificación , Idiotipos de Inmunoglobulinas/uso terapéutico , Cadenas Ligeras de Inmunoglobulina/aislamiento & purificación , Cadenas Ligeras de Inmunoglobulina/uso terapéutico , Linfoma de Células B/terapia , Linfoma Folicular/terapia , Reacción en Cadena de la Polimerasa/métodos , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/uso terapéutico , VacunaciónRESUMEN
The immunoglobulin receptor of B-cell lymphomas constitutes a specific tumor antigen (idiotype) and a target for active immunotherapy. Encouraging results have been reported in phase II trials after s.c. vaccination of follicular lymphoma patients during clinical remission with idiotype produced from eukaryotic cell lines and coupled to an immunogenic carrier macromolecule. We have developed a good manufacturing protocol for rapid expression of idiotype vaccines as recombinant Fab fragments in Escherichia coli. The objectives of this trial were to show safety and feasibility of intradermal immunization with this vaccine and to investigate whether immune responses were induced by this immunization route. Patients (n = 18) with advanced B-cell malignancies received repetitive intradermal vaccinations with 0.5 to 1.65 mg recombinant idiotype Fab fragment mixed with lipid-based adjuvant in combination with 150 mug granulocyte macrophage colony-stimulating factor s.c. at the same location. The patients' immune status was assessed by flow cytometry of peripheral blood lymphocytes and concomitant hepatitis B vaccination. Cellular and humoral immune responses to the vaccine were assessed by enzyme-linked immunospot and ELISA. Side effects of a total of 65 vaccinations were mild and did not affect the immunization schedule. No patient developed hepatitis B surface antibodies (anti-HBs) after two hepatitis B immunizations. Of 17 evaluable patients, five developed specific anti-vaccine antibodies, and eight developed anti-Fab T-cell responses. T-cell reactivity was independent of the cellular immune status and was idiotype specific as shown by statistical regression analysis (P = 0.0024) and epitope mapping studies. Intradermal administration of uncoupled recombinant idiotype with appropriate adjuvants may overcome profound clinical immunosuppression and induce specific immune responses.