RESUMO
Breast cancer is more frequent in human nulliparae, whereas its incidence is reduced by early fullterm pregnancy. Rodent studies suggest that chorionic gonadotropin secretion during pregnancy affords protection by inducing breast structure differentiation. Opposite effects, however, have been observed in cancer prone transgenic mice overexpressing the ß subunit of chorionic gonadotropin or pituitary luteinic hormone (LH). Here we assessed the effect of administration of human chorionic gonadotropin (hCG) for 21 days (corresponding to the duration of a mouse pregnancy) in virgin female mice transgenic for the activated rat (r-) ERBB-2 oncogene (BALB-neuT). In these mice, the onset of atypical mammary duct hyperplasia and its progression towards multiple mammary carcinomas is accelerated by hCG. hCG enhances the in vitro proliferation and in vivo metastatization of tumor cells from a BALB-neuT mammary tumor expressing the hCG/LH as well as the ERBB-2 receptors. These findings suggest that hCG favours the growth and progression of hCG/LH and ERBB-2 receptor-positive breast tumors.
Assuntos
Gonadotropina Coriônica/metabolismo , Gonadotropina Coriônica/farmacologia , Hormônio Luteinizante/metabolismo , Neoplasias Mamárias Experimentais/patologia , Receptor ErbB-2/metabolismo , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Progressão da Doença , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Imuno-Histoquímica , Injeções Intravenosas , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/secundário , Camundongos , Camundongos Endogâmicos BALB C , Ovariectomia , Ovário/efeitos dos fármacos , Ovário/metabolismo , Sais de Tetrazólio , TiazóisRESUMO
Castleman's disease is a lymphoproliferative disorder thought to be related to deregulated production of IL-6. We have previously shown that mice lacking the trans-acting factor C/EBP beta, a transcriptional regulator of IL-6 and a mediator of IL-6 intracellular signaling, develop a pathology nearly identical to multicentric Castleman's disease, together with increasingly high levels of circulating IL-6. We describe here how the simultaneous inactivation of both IL-6 and C/EBP beta genes prevents the development of pathological traits of Castleman's disease observed in C/EBP beta-deficient mice. Histological and phenotypic analysis of lymph nodes and spleen of double mutant mice did not show either the lymphoadenopathy and splenomegaly or the abnormal expansion of myeloid, B and plasma cell compartments observed in C/EBP beta-/- mice, while B cell development, although delayed, was normal. Our data demonstrate that IL-6 is essential for the development of multicentric Castleman's disease in C/EBP beta-/- mice.
Assuntos
Hiperplasia do Linfonodo Gigante/genética , Proteínas de Ligação a DNA/genética , Interleucina-6/genética , Proteínas Nucleares/genética , Animais , Linfócitos B/patologia , Proteínas Estimuladoras de Ligação a CCAAT , Hiperplasia do Linfonodo Gigante/patologia , Hiperplasia do Linfonodo Gigante/prevenção & controle , Proteínas de Ligação a DNA/metabolismo , Citometria de Fluxo , Linfonodos/patologia , Camundongos , Camundongos Mutantes , Proteínas Nucleares/metabolismo , Fenótipo , Plasmócitos/patologia , Baço/patologia , Timo/patologiaRESUMO
Transgenic Balb/c mice expressing the transforming rat HER-2/neu oncogene develop early and multifocal mammary carcinomas. Within the first 5 months of life the tissue-specific expression of HER-2/neu causes a progression in all their 10 mammary glands from atypical hyperplasia to invasive carcinoma. It was previously observed that chronic administration of interleukin (IL)-12 increased tumor latency, but every mouse eventually succumbed to multiple carcinomas. A significant improvement in tumor prevention was sought by administering allogeneic mammary carcinoma cells expressing HER-2/neu combined with systemic IL-12. This treatment reduced tumor incidence by 90% and more than doubled mouse lifetime. For the maximum prevention p185(neu) antigen must be expressed by allogeneic cells. IL-12 treatment strongly increased the cell vaccine efficacy. The mammary glands of mice receiving the combined treatment displayed a markedly reduced epithelial cell proliferation, angiogenesis, and HER-2/neu expression, while the few hyperplastic foci were heavily infiltrated by granulocytes, macrophages, and CD8(+) lymphocytes. Specific anti-HER-2/neu antibodies were produced and a nonpolarized activation of CD4(+) and CD8(+) cells secreting IL-4 and interferon (IFN)-gamma were evident. A central role for IFN-gamma in the preventive effect was proven by the lack of efficacy of vaccination in IFN-gamma gene knockout HER-2/neu transgenic Balb/c mice. A possible requirement for IFN-gamma is related to its effect on antibody production, in particular on IgG2a and IgG2b subclasses, that were not induced in IFN-gamma knockout HER-2/neu mice. In conclusion, our data show that an allogeneic HER-2/neu-expressing cell vaccine combined with IL-12 systemic treatment can prevent the onset of genetically determined tumors.
Assuntos
Adjuvantes Imunológicos , Vacinas Anticâncer/imunologia , Interleucina-12/imunologia , Neoplasias Mamárias Experimentais/prevenção & controle , Receptor ErbB-2/fisiologia , Animais , Mama/patologia , Linfócitos T CD4-Positivos/imunologia , Transplante de Células , Feminino , Imunidade Celular/imunologia , Interferon gama/genética , Interferon gama/imunologia , Interleucina-12/administração & dosagem , Neoplasias Mamárias Experimentais/imunologia , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Camundongos Transgênicos , Ratos , Receptor ErbB-2/genética , Transplante Homólogo , Células Tumorais Cultivadas , Vacinação/métodosRESUMO
The ability of interleukin (IL)-12 to prevent tumors when administered to individuals with a genetic risk of cancer was studied in two lines of transgenic mice expressing rat HER-2/neu oncogene in the mammary gland. Female BALB/c (H-2(d)) mice carrying the activated HER-2/ neu oncogene show no morphological abnormalities of the mammary gland until 3 wk of age. They then progress through atypical hyperplasia to in situ lobular carcinoma and at 33 wk of age all 10 mammary glands display invasive carcinomas. Adult FVB mice (H-2(q)) carrying the HER-2/neu protooncogene develop mammary carcinomas with a longer latency (38-49 wk) and a lower multiplicity (mean of 2.6 tumors/mice). Treatment with IL-12 (5 daily intraperitoneal injections, 1 wk on, 3 wk off; the first course with 50 ng IL-12/day, the second with 100 ng IL-12/day) begun at 2 wk of age in BALB/c mice and at 21 wk of age in FVB mice markedly delayed tumor onset and reduced tumor multiplicity. Analogous results were obtained in immunocompetent and permanently CD8(+) T lymphocyte-depleted mice. In both transgenic lines, tumor inhibition was associated with mammary infiltration of reactive cells, production of cytokines and inducible nitric oxide synthase, and reduction in microvessel number, in combination with a high degree of hemorrhagic necrosis.
Assuntos
Antineoplásicos/farmacologia , Carcinoma in Situ/prevenção & controle , Carcinoma Lobular/prevenção & controle , Interleucina-12/farmacologia , Neoplasias Mamárias Experimentais/prevenção & controle , Receptor ErbB-2/fisiologia , Animais , Antineoplásicos/imunologia , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Carcinoma in Situ/genética , Carcinoma in Situ/imunologia , Carcinoma Lobular/genética , Carcinoma Lobular/imunologia , Quimiocina CXCL10 , Quimiocina CXCL9 , Quimiocinas CXC/genética , Feminino , Interferon gama/imunologia , Interleucina-12/imunologia , Depleção Linfocítica , Masculino , Neoplasias Mamárias Experimentais/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Óxido Nítrico Sintase/biossíntese , Óxido Nítrico Sintase Tipo II , Ratos , Receptor ErbB-2/genética , Fator de Necrose Tumoral alfa/imunologia , Molécula 1 de Adesão de Célula Vascular/biossínteseRESUMO
The proliferative responses to phytohemagglutinin and the role of subcellular factors in the modulation of blastogenesis of thymoma lymphocytes from 5 thymoma patients were investigated. The addition of exogenous interleukin 1, a macrophage product, strongly augmented the blastic transformation of cultured thymocytes from both normal and neoplastic glands by influencing the production of interleukin 2 (IL-2) by a well-defined T-cell subset. The magnitudes of the blastic responses were ultimately modulated by the amount of IL-2 released in culture. The higher proliferative responses exhibited by thymocytes from thymoma were effectively sustained by a higher production of IL-2 in culture. In addition to having distinctive surface membrane receptors in common with normal thymocytes, thymoma lymphocytes were also under the influence of the same subcellular factors involved in T-cell blastic activation as thymocytes. These observations imply the presence of functionally distinct subpopulations in the thymoma lymphocyte component and add arguments in favor of their nonneoplastic nature.
Assuntos
Ativação Linfocitária , Linfócitos/imunologia , Timoma/imunologia , Neoplasias da Glândula Tireoide/imunologia , Adulto , Membrana Celular/imunologia , Feminino , Humanos , Imunoglobulina G , Imunoglobulina M , Interleucina-1/imunologia , Interleucina-2/imunologia , Masculino , Pessoa de Meia-Idade , Fito-Hemaglutininas/farmacologia , Receptores de Antígenos de Linfócitos B , Receptores Fc , Formação de RosetaRESUMO
BACKGROUND: Numerous animal model studies have examined the ability of genetically engineered tumor cells to release cytokines and to elicit an immune memory against the parental tumor. Often only a single cytokine is studied, and few comparative studies have been conducted. PURPOSE: We evaluated the antitumor efficacy of adenocarcinoma cells engineered to release interleukin (IL)-12 in a mouse model system. The efficacy of this cytokine was compared with that of other cytokines released by engineered adenocarcinoma cells and that of exogenous IL-12 administered both locally and intraperitoneally. METHODS: BALB/cAnCr mice were inoculated with syngeneic parental mammary adenocarcinoma (TSA) cells in quantities sufficient to lead to tumors in all inoculated mice. TSA cells engineered to release IL-12 (TSA-IL12) were also injected into normal and selectively immunosuppressed BALB/cAnCr mice. Tumor incidence, growth, and rejection patterns were evaluated by the measurement of neoplastic masses and by the study of the histologic and ultrastructural features of the tumor site. The effects of local or intraperitoneal administration of recombinant IL-12 (rIL-12) on tumor-bearing animals were also studied. RESULTS: Most mice rejected TSA-IL12 cells through a CD8-positive, T-lymphocyte-dependent reaction associated with macrophage infiltration, vessel damage, and necrosis. The systemic immunity of mice that had rejected TSA-IL12 cells to a subsequent challenge with parental TSA cells was less efficient than that elicited by TSA cells engineered to release IL-4 or IL-10 but equivalent to that elicited by TSA cells engineered to release IL-2, IL-7, and interferon alfa. Compared with TSA cells engineered to produce other cytokines, TSA-IL12 cells were the most efficient in curing mice with established TSA tumors; injection of 0.1 million proliferating cells contralaterally to the tumor growth area cured five of 15 mice bearing 1-day-old tumors; injection of the same dose of proliferating cells into the tumor growth area cured two of 20 tumor-bearing mice. However, two 5-day courses with a nontoxic dose (0.1 microgram) of rIL-12 given intraperitoneally cured a similar proportion of these animals (six of 20). Only two of 20 mice with 7-day-old TSA tumors were cured by vaccination with proliferating TSA-IL12 cells, whereas 24 of 30 mice with such tumors were cured by intraperitoneal administration of rIL-12. CONCLUSIONS: TSA cells engineered to release IL-12 are rejected by most mice; the ensuing immune memory for TSA parental cells, however, was less efficient than that elicited by proliferating TSA cells engineered to release other cytokines (e.g., IL-4, IL-10, and possibly interferon gamma). The immune reaction elicited by TSA-IL12 cells was the most efficient in curing mice with established TSA tumors; notably though, the same or a better cure rate was obtained with rIL-12 given intraperitoneally.
Assuntos
Adenocarcinoma/tratamento farmacológico , Interleucina-12/farmacologia , Interleucina-12/fisiologia , Neoplasias Mamárias Experimentais/tratamento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/patologia , Animais , Modelos Animais de Doenças , Feminino , Infusões Parenterais , Injeções Intralesionais , Interleucina-12/administração & dosagem , Interleucina-12/biossíntese , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/farmacologia , Transdução GenéticaRESUMO
BACKGROUND: Defective mismatch repair (MMR) in humans is particularly associated with familial colorectal cancer, but defective repair in mice is generally associated with lymphoma in the absence of experimental exposure to carcinogens. Loss of MMR also confers resistance to the toxic effects of methylating agents. We investigated whether resistance to methylation contributes to increased susceptibility to colorectal cancer in mice by exposing mice with defects in the MMR gene msh2 to a methylating agent. METHODS: Tumor incidence and time of death in msh2(+/+), msh2(+/-), and msh2(-/-) mice were analyzed after weekly exposure (until tumor appearance) to the methylating agent 1,2-dimethylhydrazine (DMH). Chemically induced and spontaneous tumors were characterized by frequency, type, and location. The tumor incidence in untreated and treated mice of each genotype was compared by a Mann-Whitney U test. Carcinogen-induced apoptosis in histologic sections of small and large intestines was also determined. All statistical tests were two-sided. RESULTS: Homozygous inactivation of the msh2 gene statistically significantly accelerated (P<.0001) death due to the development of DMH-induced colorectal tumors and lymphomas. Rates of death from DMH-induced colorectal adenocarcinoma were similar in msh2 heterozygous and wild-type mice, but only msh2 heterozygotes (msh(+/-)) developed additional, noncolorectal malignancies (notably trichofolliculoma [two of 21], angiosarcoma of the kidney capsule [two of 21], and lymphoma [one of 21]), suggesting that heterozygosity for msh2 slightly increases DMH susceptibility. DMH induced apoptosis in small intestinal and colonic epithelial crypts that was dependent on active msh2. CONCLUSIONS: Inactivation of msh2 allows the proliferation of gastrointestinal tract cells damaged by methylating agents. Furthermore, MMR constitutes a powerful defense against colorectal cancer induced by DNA methylation.
Assuntos
1,2-Dimetilidrazina/toxicidade , Adenocarcinoma/genética , Alquilantes/toxicidade , Carcinógenos/toxicidade , Neoplasias do Colo/genética , Reparo do DNA/genética , Proteínas de Ligação a DNA , Linfoma não Hodgkin/genética , Proteínas Proto-Oncogênicas/fisiologia , Adenocarcinoma/induzido quimicamente , Animais , Apoptose/efeitos dos fármacos , Pareamento Incorreto de Bases , Divisão Celular/efeitos dos fármacos , Colo/efeitos dos fármacos , Colo/patologia , Neoplasias do Colo/induzido quimicamente , Resistência a Medicamentos , Genótipo , Doenças do Cabelo/induzido quimicamente , Doenças do Cabelo/genética , Folículo Piloso , Hemangiossarcoma/induzido quimicamente , Hemangiossarcoma/genética , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/patologia , Intestino Delgado/efeitos dos fármacos , Intestino Delgado/patologia , Neoplasias Renais/induzido quimicamente , Neoplasias Renais/genética , Linfoma não Hodgkin/induzido quimicamente , Metilação , Camundongos , Camundongos Knockout , Proteína 2 Homóloga a MutS , Neoplasia de Células Basais/induzido quimicamente , Neoplasia de Células Basais/genética , Proteínas Proto-Oncogênicas/deficiência , Proteínas Proto-Oncogênicas/genéticaRESUMO
Immunotherapy applied to patients with established tumors rarely leads to an objective response, whereas patients apparently free from disease after conventional treatment and at risk of recurrence are beginning to receive vaccination. New classes of patients or not-yet patients are those with a high genetic or environmental risk of developing cancer. They may draw benefit from a "soft" treatment such as vaccination. This overview discusses the prospects of immune stimulation as a means of cancer prevention by inducing various forms of nonspecific or even specific immunity. Attainment of this goal provides the rationale and motivation for embarking on such a new and potentially rewarding enterprise.
Assuntos
Imunização , Neoplasias/prevenção & controle , Humanos , Neoplasias/imunologiaRESUMO
The antitumor activity of recombinant murine interleukin-12 (rIL-12) is documented by a large set of data from numerous mouse models. Because the cellular and molecular mechanisms by which rIL-12 impairs tumor growth are still not fully defined, we compared the effects of local and systemic rIL-12 administration in mice harboring an invasive 7-day-old moderately differentiated and spontaneously metastasizing mammary adenocarcinoma (TSA). Whereas the immune events elicited via the two routes of rIL-12 administration seem to be the same, systemic rIL-12 is markedly more effective; tumor destruction is dependent on a prompt antitumor response resulting from the cooperation of several subsets of reactive cells. The reactions that seem to play a key role are: (a) indirect inhibition of angiogenesis by secondary cytokines (mainly IFN-gamma) and third-level chemokines (inducible protein 10 and monokine induced by IFN-gamma); (b) systemic activation of leukocyte subsets capable of producing proinflammatory cytokines, CTLs, and antitumor antibodies; and (c) destruction of tumor vessels by polymorphonuclear cells. The markedly higher efficacy of systemic rIL-12 seems to rest on its ability to recruit these systemic reactions more quickly and efficiently than local rIL-12.
Assuntos
Interleucina-12/uso terapêutico , Neoplasias Mamárias Experimentais/terapia , Animais , Anticorpos Antineoplásicos/sangue , Citocinas/genética , Citocinas/metabolismo , Feminino , Imuno-Histoquímica , Interleucina-12/administração & dosagem , Neoplasias Mamárias Experimentais/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Metástase Neoplásica , Neutrófilos/fisiologia , RNA Mensageiro/análise , Proteínas Recombinantes/uso terapêutico , Linfócitos T Citotóxicos/imunologiaRESUMO
T-lymphocytes (LYs) from normal and IFN-gamma knockout mice were activated by anti-CD3 and anti-CD28 antibodies and cultured in inserts in the presence of interleukin (IL)-12 (IL-12-activated LYs) or not (activated LYs). Their ability to modulate the genetic programs of two tumor lines growing at the bottom of transwells was evaluated. cDNA gene expression array, reverse transcription-PCR, and protein expression showed that LPS, transcription termination factor 1, transforming growth factor, and fibroblast growth factor genes were up-modulated by factors other than IFN-gamma released by activated LYS: The high levels of IFN-gamma released by normal IL-12-activated LYs up-modulated the expression of STAT1, IRF-1, LMP2, LMP7, monokine induced by IFN-gamma, monocyte chemoattractant protein 1, and angiopoietin 2 genes but down-modulated the expression of vascular endothelial growth factor. PA28, IFN-inducible protein 10, inducible NO synthetase, and macrophage-inhibitory protein 2 genes were up-modulated by factors released only by IL-12-activated LYs apart from IFN-gamma. The opposite modulations of vascular endothelial growth factor expression and of angiopoietin 2, monokine induced by IFN-gamma, IFN-inducible protein 10, and inducible NO synthetase by IL-12-activated LYs fit in well with the inhibition of angiogenesis that characterizes the antitumor activity of IL-12. T-LYs thus modify a tumor's behavior so that it becomes a party to its own inhibition.
Assuntos
Adenocarcinoma/genética , Adenocarcinoma/imunologia , Interleucina-12/imunologia , Ativação Linfocitária/imunologia , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/imunologia , Linfócitos T/imunologia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacologia , Antígenos CD28/imunologia , Complexo CD3/imunologia , Técnicas de Cocultura , Citocinas/genética , Citocinas/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Interferon gama/genética , Interferon gama/imunologia , Interleucina-12/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Linfócitos T/efeitos dos fármacos , Células Tumorais CultivadasRESUMO
With a slight asynchronous but consistent progression, all of the mammary glands of female BALB/c mice transgenic for the transforming rat HER-2/neu oncogene progress to atypical hyperplasia and to invasive carcinoma. Previous studies have shown that chronic administration of interleukin (IL) 12 started at the 2nd week of age hampers this progression because of its ability to inhibit tumor angiogenesis and activate a nonspecific immune response. Here we show that a similar inhibition is achieved when 7-week-old mice with fully blown atypical hyperplasia receive a weekly injection of 100 ng IL-12 for 16 times. This lower-dose and later IL-12 administration induces high and sustained levels of serum IFN-gamma equivalent to those elicited by more frequent administrations. A lower-dose and less toxic treatment may thus be envisaged as a possible option in the management of preneoplastic mammary lesions.
Assuntos
Genes erbB-2/genética , Interleucina-12/farmacologia , Glândulas Mamárias Animais/patologia , Neoplasias Mamárias Experimentais/prevenção & controle , Animais , Relação Dose-Resposta a Droga , Feminino , Hiperplasia/patologia , Interferon gama/biossíntese , Glândulas Mamárias Animais/efeitos dos fármacos , Glândulas Mamárias Animais/metabolismo , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/metabolismo , Camundongos , Camundongos TransgênicosRESUMO
Previous studies in mice have shown that chronic administration of recombinant interleukin-12 (IL-12) hampers the progression of both chemical- and oncogene-dependent carcinogenesis. This suggests that a new preventive strategy may be envisaged for individuals with a genetic risk of cancer or carrying preneoplastic lesions. Starting at progressive stages of mammary carcinogenesis, female BALB/c and FVB mice carrying the activated rat HER2/neu oncogene (BALB-neuT) or the proto-oncogene (FVB-neuN) under the mouse mammary tumor virus promoter received multiple 5-day courses of different doses of IL-12. The times of tumor appearance, multiplicity, and histopathological features of the neoplastic lesions were evaluated. In both BALB-neuT and FVB-neuN mice, 5-day i.p. courses of 50/100 ng of IL-12/day inhibited mammary carcinogenesis when they coincided with the progression of early preneoplastic lesions. Inhibition appears to depend primarily on the ability of IL-12 to interfere with early tumor angiogenesis. Later treatments are much less effective, and daily doses of 10 and 2 ng are useless. The efficacy of early IL-12 courses suggests that they could be used to prevent mammary tumors in individuals at risk, whereas their lower efficacy in later stages of carcinogenesis and the dose range required pose some constraints on their use in the management of overt preneoplastic lesions. Precise understanding of tumor progression means that effective treatments can be commenced relatively late in the life of individuals at risk and that no lifetime administration is required.
Assuntos
Genes erbB-2 , Interleucina-12/uso terapêutico , Neoplasias Mamárias Experimentais/tratamento farmacológico , Neoplasias Mamárias Experimentais/patologia , Receptor ErbB-2/genética , Animais , Progressão da Doença , Feminino , Neoplasias Mamárias Experimentais/prevenção & controle , Vírus do Tumor Mamário do Camundongo/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Regiões Promotoras Genéticas , Ratos , Fatores de TempoRESUMO
Prevention of the progression of precancerous lesions by vaccines is virtually uncharted territory. Their potential, however, is being assessed in transgenic mice which develop autochthonous tumors with defined stages of progression. In this paper we show that the DNA micro-array technology significantly helps assessment of the preventive efficacy of a combined DNA and cell vaccine. All female rat Her-2/neu transgenic BALB/c (BALB-neuT) mice develop an invasive carcinoma in each of their mammary glands within 25 weeks of age. This is elicited by the activated transforming rat Her-2/neu oncogene embedded in their genome. We have previously shown that vaccination of mice bearing multiple in situ carcinomas with DNA plasmids which code for the extracellular and transmembrane domain of rat p185neu, the product of the rat Her-2/neu oncogene, followed by a boost with rat p185neu+ allogeneic cells engineered to secrete interferon-gamma, keeps 48% of mice tumor free until week 32. We have now extended our follow-up until mice reach one year of age and show that protection vanishes as time progresses. This observation suggests that the accuracy of the results studying immunotherapy against life-threatening tumors is a function of the length of the follow-up. The application of microarrays, and the concordance of morphologic and gene expression data led us to identify antibody as the main mechanism induced by vaccination. Protection is associated with a break of tolerance and a limited autoimmunity against the endogenous mouse p185neu.
Assuntos
Autoimunidade/efeitos dos fármacos , Glicoproteínas/genética , Neoplasias Mamárias Experimentais/prevenção & controle , Lesões Pré-Cancerosas/terapia , Receptor ErbB-2/genética , Vacinas de DNA/uso terapêutico , Animais , Linhagem Celular Tumoral , Clonagem Molecular , Feminino , Neoplasias Mamárias Experimentais/imunologia , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Transgênicos , Lesões Pré-Cancerosas/imunologia , Lesões Pré-Cancerosas/patologia , Ratos , TransgenesRESUMO
Because BALB/c mice transgenic for the rat Her-2/neu oncogene develop multifocal carcinomas in all mammary glands by week 33, they constitute an aggressive model for investigation of treatments designed to oppose mammary carcinogenesis. Nonspecific immune reaction elicited by systemic interleukin (IL)-12 both delayed the appearance of the first tumor and reduced the number of glands affected. However, only 5% of mice were tumor free at week 33. On the other hand, specific vaccination with plasmids encoding for the rat p185neu resulted in a further delay, so much so that 58% of mice were tumor free at week 33. No CTL response was evoked in either IL-12-treated or DNA-vaccinated mice, whereas an anti-rat p185neu antibody response was evident in the latter. Pathological examinations showed that in both IL-12-treated and DNA-vaccinated mice, the tumor growth area was infiltrated by reactive cells associated with expression of endothelial adhesion molecules and antiangiogenic proinflammatory cytokines. In the vaccinated mice, reduction of the number of cells expressing rat p185neu was combined with down-regulation of its membrane expression and even a marked inhibition in development of the terminal ductal lobular units. The reactive infiltrate in vaccinated mice contained numerous granulocytes that likely played an antiangiogenic and angiodestructive role and also joined other cells in the antibody-mediated killing of the r-p185neu+ cells. These results suggest that the elicitation of nonspecific and specific immunity could be beneficially used in individuals with a high risk of developing tumors.
Assuntos
Vacinas Anticâncer , Genes erbB-2/genética , Interleucina-12/uso terapêutico , Neoplasias Mamárias Animais/prevenção & controle , Receptor ErbB-2/uso terapêutico , Animais , DNA/metabolismo , Regulação para Baixo , Feminino , Citometria de Fluxo , Imuno-Histoquímica , Interleucina-12/biossíntese , Interleucina-12/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Plasmídeos/metabolismo , Ratos , Linfócitos T Citotóxicos/metabolismo , Fatores de TempoRESUMO
Transduction of the murine interferon-alpha (IFN-alpha) gene into various malignant mouse tumor cells has resulted in the loss of tumorigenicity and an acquired capacity to induce long-lasting antitumor immunity following their injection into immunocompetent syngeneic mice. In the present study, we investigated the effectiveness of IFN-alpha-producing tumor cells in the therapy of mice with established mouse tumors. In DBA/2 mice bearing subcutaneous (s.c.) Friend erythroleukemia cell (FLC) tumors, we found that to achieve some antitumor response (i) it was necessary to inject high numbers of IFN-alpha-producing FLC, which occasionally lead to the formation of slowly growing tumors; and, that (ii) repeated injections of irradiated IFN-alpha-FLC did not result in any antitumor effect. The therapeutic potential of IFN-alpha-producing FLC rendered sensitive to ganciclovir (GCV), by transfer of the herpes simplex virus thymidine kinase (tk) gene, was investigated. Complete tumor rejection and cure was observed in > or = 70% of the animals after injection of high numbers (10(7)) of IFN-alpha-producing tk-expressing tumor cells followed 4 days later by repeated GCV treatments, whereas only a slight increase in survival time was obtained after administration of control tk-expressing tumor cells (not producing IFN) and GCV. Tumor rejection was associated with a dramatic destruction of tumor tissue and with the subsequent development of a potent and long-lasting antitumor immunity. No therapeutic effect was observed in immunosuppressed nude mice. These data indicate that this approach may represent an effective and safe therapeutic strategy for antitumor cytokine gene therapy.
Assuntos
Vírus da Leucemia Murina de Friend , Ganciclovir/farmacologia , Interferon-alfa/genética , Leucemia Eritroblástica Aguda/terapia , Simplexvirus/enzimologia , Timidina Quinase/genética , Animais , Linhagem Celular , Células Clonais , Expressão Gênica , Sistema Imunitário , Interferon-alfa/metabolismo , Leucemia Eritroblástica Aguda/patologia , Masculino , Camundongos , Camundongos Endogâmicos DBA , Camundongos Nus , Metástase Neoplásica , Simplexvirus/genética , Timidina Quinase/metabolismo , Transfecção , Células Tumorais CultivadasRESUMO
The parental cells of the TSA murine mammary adenocarcinoma (TSA-pc) were transfected with both the interferon-gamma (IFN-y) gene and the cytosine deaminase (CD) suicide gene to obtain a therapeutic vaccine active against TSA-pc lung metastases. Even in the absence of treatment with the prodrug 5-fluorocytosine (5-FC), the local growth of double transfectants (CD-y clones) was inhibited by a marked recruitment of granulocytes and macrophages. In mice harboring TSA-pc micrometastases, therapeutic vaccination with either IFN-gamma or CD single transfectants reduced the number of lung nodules, whereas CD-gamma double transfectants abrogated metastasis growth in up to 80% of mice. Treatment of mice with 5-FC did not alter the curative efficacy of CD-gamma double-transfectant cells. By contrast, in mice vaccinated with CD single-transfectant cells, 5-FC treatment caused a significant loss of their curative activity. Host T cells played an active role in the cure of lung metastases, because vaccination of nude mice with CD-gamma cells was uneffective.
Assuntos
Adenocarcinoma/genética , Interferon gama/genética , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/terapia , Neoplasias Mamárias Experimentais/genética , Nucleosídeo Desaminases/genética , Transfecção , Adenocarcinoma/imunologia , Adenocarcinoma/patologia , Animais , Citosina Desaminase , Citotoxicidade Imunológica , Feminino , Terapia Genética , Imunoterapia Ativa , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/secundário , Neoplasias Mamárias Experimentais/imunologia , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Transplante de Neoplasias , Linfócitos T Citotóxicos/imunologiaRESUMO
The onco-suppressor gene TP53 has potential use in the gene therapy of many human cancers including leukemias. The latter indication derived from numerous experimental reports of p53-mediated suppressing effects on human and murine leukemia cells in vitro. However, few in vivo experiments have been performed, and those that have used a subcutaneous injection of p53-transduced leukemia cells. Thus, we developed an orthotopic leukemia model in adult, syngenic mice to evaluate the feasibility of TP53-mediated therapeutic approaches. We found that among other cells, v-src-transformed 32D myeloid progenitors induce leukemia when injected intravenously in syngenic mice. The resulting malignancy resembles the clinical manifestations of human acute myeloid leukemia because it is characterized by a massive invasion of bone marrow compartments, splenomegaly, generalized lymphadenopathy, and a macroscopic or microscopic infiltration of the kidneys, liver, and lungs. When these 32Dv-src cells were infected with a TP53-recombinant retrovirus before intravenous injection, we found a decreased mortality and, in those animals that develop leukemia, a drastic reduction of the generalized organ infiltration, suggesting that exogenous TP53 expression might be used for ex vivo bone marrow purging from leukemia cells.
Assuntos
Terapia Genética , Leucemia/terapia , Proteína Supressora de Tumor p53/genética , Animais , Modelos Animais de Doenças , Estudos de Viabilidade , Vetores Genéticos , Leucemia/genética , Leucemia/patologia , Masculino , Camundongos , Camundongos Endogâmicos C3H , Transplante de Neoplasias , Retroviridae/genética , Proteína Supressora de Tumor p53/metabolismo , Proteína Supressora de Tumor p53/uso terapêuticoRESUMO
Immunization of cancer patients with cytokine-engineered tumor cells is being currently tested in several trials. To test the feasibility of this approach in neuroblastoma (NB) patients we investigated the functional consequences of interleukin-2 (IL-2) gene transfer into NB cell lines. Two human NB cell lines were transfected with the plasmid expression vector RSV.5neo containing the human IL-2 cDNA, and their tumorigenicity was evaluated in a nude mice xenograft model after characterization of the growth patterns and phenotypic features in vitro. The combination of IL-2 gene transfection and the xenograft model in nude mice was chosen on the basis of the low or absent expression of HLA class I antigen in human NB tumors. Our aim was to evaluate the effectiveness of an immunization protocol that could elicit a nonspecific antitumor response. The IL-2 stable transfectants were morphologically identical to parental or vector-transfected cells but completely lost tumorigenicity and inhibited, through a bystander effect, the growth of parental cells injected simultaneously at the same site. Histologic and immunohistochemical analysis of the nodules showed extensive necrosis with severe endothelial damage. The infiltrating cells were mainly macrophages, while natural killer (NK) cells were scarce. However, depletion of NK cells by anti-CD122 monoclonal antibody indicated that the rejection process required NK cell activity. The relevance of these data for the development of therapeutic approaches using cytokine-engineered NB cell lines is discussed.
Assuntos
Terapia Genética , Interleucina-2/genética , Interleucina-2/imunologia , Neuroblastoma/terapia , Animais , Feminino , Humanos , Imuno-Histoquímica , Células Matadoras Naturais/imunologia , Camundongos , Camundongos Nus , Transplante de Neoplasias , Neuroblastoma/imunologia , Neuroblastoma/patologia , Linfócitos T Citotóxicos/imunologia , Transfecção , Transplante Heterólogo , Células Tumorais CultivadasRESUMO
A spontaneously metastatic murine mammary adenocarcinoma, TSA, has been transduced with the gene for interferon alpha1 (IFN-alpha). Transfectants were used for the immunotherapy of mice bearing lung colonies induced by the intravenous inoculation of non-transduced parental cells. A significant reduction in the number of tumor colonies was obtained when repeated subcutaneous administrations of mitomycin C-blocked transfectant cells were given, commencing 3 days after an intravenous challenge with TSA cells. Intraperitoneal vaccination induced a stronger anti-tumor response than subcutaneous vaccination, and the proportion of tumor-free mice reached 50%. The potency of IFN-alpha transfectants was similar to that of IFN-gamma transfectants previously obtained from TSA. Admixture of IFN-alpha and IFN-gamma transfectant cells in the same vaccine did not increase the curative effect over that of single vaccines. In nude mice vaccination with IFN-alpha or IFN-gamma transfectants did not lead to a reduction in the number of lung colonies, indicating that an intact T cell response was required for the therapeutic effect observed in immunocompetent mice.
Assuntos
Adenocarcinoma/terapia , Vacinas Anticâncer/uso terapêutico , Interferon-alfa/imunologia , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Neoplasias Mamárias Experimentais/terapia , Mitomicina/farmacologia , Adenocarcinoma/patologia , Animais , Vacinas Anticâncer/administração & dosagem , Feminino , Injeções Intraperitoneais , Injeções Intravenosas , Injeções Subcutâneas , Interferon gama/imunologia , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Linfócitos T/imunologia , Transdução GenéticaRESUMO
This paper is concerned with technical standardization in detecting Fc-IgM receptors on human T lymphocytes. We have investigated a number of factors of critical importance in obtaining easily reproducible and reliable estimates of the numbers of TM cells among human peripheral T lymphocytes. A point of major importance is optimal coating of erythrocytes by IgM molecules. For this condition to be met, particular attention is required when erythrocytes from animals other than the one used for obtaining antiserum are used to prepare EA-IgM. Determination of the agglutinating titer of IgM preparation is useful in determing optimal sensitizing dilutions. Full expression of Fc receptors is favoured when human cord serum is added to the medium. The influence of incubation period of EA-lymphocytes mixtures on TM counts has also been investigated.