RESUMO
Plant-made vaccines have been the subject of intense interest because they can be produced economically in large scale without the use of animal-derived components. Plant-made therapeutic vaccines against challenging chronic diseases, such as cancer, have received little research attention, and no previous human clinical trials have been conducted in this vaccine category. We document the feasibility of using a plant viral expression system to produce personalized (patient-specific) recombinant idiotype vaccines against follicular B cell lymphoma and the results of administering these vaccines to lymphoma patients in a phase I safety and immunogenicity clinical trial. The system allowed rapid production and recovery of idiotypic single-chain antibodies (scFv) derived from each patient's tumor and immunization of patients with their own individual therapeutic antigen. Both low and high doses of vaccines, administered alone or co-administered with the adjuvant GM-CSF, were well tolerated with no serious adverse events. A majority (>70%) of the patients developed cellular or humoral immune responses, and 47% of the patients developed antigen-specific responses. Because 15 of 16 vaccines were glycosylated in plants, this study also shows that variation in patterns of antigen glycosylation do not impair the immunogenicity or affect the safety of the vaccines. Collectively, these findings support the conclusion that plant-produced idiotype vaccines are feasible to produce, safe to administer, and a viable option for idiotype-specific immune therapy in follicular lymphoma patients.
Assuntos
Vacinas Anticâncer/uso terapêutico , Linfoma de Células B/terapia , Linfoma Folicular/terapia , Adulto , Idoso , Anticorpos Antineoplásicos/sangue , Anticorpos Antineoplásicos/química , Anticorpos Antineoplásicos/genética , Anticorpos Antineoplásicos/uso terapêutico , Vacinas Anticâncer/administração & dosagem , Vacinas Anticâncer/efeitos adversos , Vacinas Anticâncer/imunologia , Feminino , Fator Estimulador de Colônias de Granulócitos e Macrófagos/administração & dosagem , Humanos , Imunidade Celular , Idiótipos de Imunoglobulinas/química , Idiótipos de Imunoglobulinas/genética , Idiótipos de Imunoglobulinas/uso terapêutico , Injeções Subcutâneas , Linfoma de Células B/genética , Linfoma de Células B/imunologia , Linfoma Folicular/genética , Linfoma Folicular/imunologia , Masculino , Pessoa de Meia-Idade , Plantas Geneticamente Modificadas , Proteínas Recombinantes , Segurança , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/efeitos adversos , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/uso terapêuticoRESUMO
Squalene synthetase (farnesyl-diphosphate:farnesyl-diphosphate farnesyltransferase, EC 2.5.1.21) catalyzes the first committed step for sterol biosynthesis and is thought to play an important role in the regulation of isoprenoid biosynthesis in eukaryotes. Using degenerate oligonucleotides based on a conserved region found in yeast and human squalene synthetase genes, a cDNA was cloned from the plant Nicotiana benthamiana. The cloned cDNA contained an open reading frame of 1234 bp encoding a polypeptide of 411 amino acids (M(r) 47002). Northern blot analysis of poly(A)+ mRNA from N. benthamiana and N. tabacum cv. MD609 revealed a single band of ca. 1.6 kb in both Nicotiana species. The identity and functionality of the cloned plant squalene synthetase cDNA was further confirmed by expression of the cDNA in Escherichia coli and in a squalene synthetase-deficient erg9 mutant of Saccharomyces cerevisiae. Antibodies raised against a truncated form of the protein recognized an endogenous plant protein of appropriate size as well as the full-length bacterially expressed protein as detected by western analysis. Comparison of the deduced primary amino acid sequences of plant, yeast, rat and human squalene synthetase revealed regions of conservation that may indicate similar functions within each polypeptide.
Assuntos
Farnesil-Difosfato Farnesiltransferase/genética , Nicotiana/genética , Plantas Tóxicas , Sequência de Aminoácidos , Animais , Sequência de Bases , Northern Blotting , Southern Blotting , Clonagem Molecular , DNA Complementar , Escherichia coli/genética , Farnesil-Difosfato Farnesiltransferase/metabolismo , Humanos , Dados de Sequência Molecular , Saccharomyces cerevisiae/genética , Homologia de Sequência de Aminoácidos , Nicotiana/enzimologiaRESUMO
Peak levels of 1-aminocyclopropane-l-carboxylic acid (ACC) in flower parts of ageing carnations (Dianthus caryophyllus L. cv Scanea 3C) were detected 6 to 9 days after flower opening. The ethylene climacteric and the first visible sign of wilting was observed 7 days after opening. The concentration of conjugated ACC in these same tissues peaked at day three with reduction of 70% by day 4. From day 5 to day 9 all parts followed a diurnal pattern of increasing in conjugate levels 1 day and decreasing the next. Concentrations of conjugated ACC were significantly higher than those of ACC in all ageing parts. Preclimacteric petals treated with ACC or 1-(malonylamino)-cycloprane-1-carboxylic acid (MACC), started to senesce 30 to 36 hours after treatment. When petals were treated with MACC plus by 0.1 millimolar aminoethyoxyvinylglycine, premature senescence was induced, while ethylene production was suppressed relative to MACC-treated petals. Petals treated with MACC and silver complex produced ethylene, but did not senesce. The MACC-induced ethylene was inhibited by the addition of 1.0 millimolar CoC1(2). These results demonstrate MACC-induced senescence in preclimacteric petals. The patterns of ACC and MACC detected in the flower parts support the view that an individual part probably does not export an ethylene precursor to the remainder of the flower inducing senescence.
