RESUMO
Because apoptosis of infected cells can limit virus production and spread, some viruses have co-opted prosurvival genes from the host. This includes the Epstein-Barr virus (EBV) gene BHRF1, a homolog of human Bcl-2 proteins that block apoptosis and are associated with cancer. Computational design and experimental optimization were used to generate a novel protein called BINDI that binds BHRF1 with picomolar affinity. BINDI recognizes the hydrophobic cleft of BHRF1 in a manner similar to other Bcl-2 protein interactions but makes many additional contacts to achieve exceptional affinity and specificity. BINDI induces apoptosis in EBV-infected cancer lines, and when delivered with an antibody-targeted intracellular delivery carrier, BINDI suppressed tumor growth and extended survival in a xenograft disease model of EBV-positive human lymphoma. High-specificity-designed proteins that selectively kill target cells may provide an advantage over the toxic compounds used in current generation antibody-drug conjugates.
Assuntos
Herpesvirus Humano 4/química , Engenharia de Proteínas , Proteínas/farmacologia , Proteínas Virais/antagonistas & inibidores , Sequência de Aminoácidos , Animais , Apoptose/efeitos dos fármacos , Biologia Computacional , Cristalografia por Raios X , Infecções por Vírus Epstein-Barr/tratamento farmacológico , Herpesvirus Humano 4/fisiologia , Xenoenxertos , Humanos , Linfoma de Células B/tratamento farmacológico , Camundongos , Modelos Moleculares , Dados de Sequência Molecular , Transplante de Neoplasias , Proteínas/química , Proteínas/metabolismo , Alinhamento de Sequência , Proteínas Virais/químicaRESUMO
Fenretinide, a synthetic retinoid, induces apoptotic cell death in B-cell non-Hodgkin lymphoma (B-NHL) and acts synergistically with rituximab in preclinical models. We report results from a phase I-II study of fenretinide with rituximab for B-NHLs. Eligible diagnoses included indolent B-NHL or mantle cell lymphoma. The phase I design de-escalated from fenretinide at 900 mg/m2 PO BID for days 1-5 of a 7-day cycle. The phase II portion added 375 mg/m2 IV rituximab weekly on weeks 5-9 then every 3 months. Fenretinide was continued until progression or intolerance. Thirty-two patients were treated: 7 in phase I, and 25 in phase II of the trial. No dose-limiting toxicities were observed. The phase II component utilized fenretinide 900 mg/m2 twice daily with rituximab. The most common treatment-related adverse events of grade 3 or higher were rash (n = 3) and neutropenia (n = 3). Responses were seen in 6 (24%) patients on the phase II study, with a median duration of response of 47 months (95% confidence interval, 2-56). The combination of fenretinide and rituximab was well tolerated, yielded a modest overall response rate, but with prolonged remission durations. Further study should focus on identifying the responsive subset of B-NHL.
Assuntos
Fenretinida/administração & dosagem , Linfoma de Células B/tratamento farmacológico , Linfoma de Célula do Manto/tratamento farmacológico , Rituximab/administração & dosagem , Adulto , Idoso , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Sinergismo Farmacológico , Exantema/induzido quimicamente , Feminino , Humanos , Linfoma de Células B/complicações , Linfoma de Célula do Manto/complicações , Masculino , Pessoa de Meia-Idade , Neutropenia/induzido quimicamente , Indução de RemissãoRESUMO
α-Emitting radionuclides deposit a large amount of energy within a few cell diameters and may be particularly effective for radioimmunotherapy targeting minimal residual disease (MRD). To evaluate this hypothesis, (211)At-labeled 1F5 monoclonal antibody (mAb) (anti-CD20) was studied in both bulky lymphoma tumor xenograft and MRD animal models. Superior treatment responses to (211)At-labeled 1F5 mAb were evident in the MRD setting. Lymphoma xenograft tumor-bearing animals treated with doses of up to 48 µCi of (211)At-labeled anti-CD20 mAb ([(211)At]1F5-B10) experienced modest responses (0% cures but two- to threefold prolongation of survival compared with negative controls). In contrast, 70% of animals in the MRD lymphoma model demonstrated complete eradication of disease when treated with (211)At-B10-1F5 at a radiation dose that was less than one-third (15 µCi) of the highest dose given to xenograft animals. Tumor progression among untreated control animals in both models was uniformly lethal. After 130 days, no significant renal or hepatic toxicity was observed in the cured animals receiving 15 µCi of [(211)At]1F5-B10. These findings suggest that α-emitters are highly efficacious in MRD settings, where isolated cells and small tumor clusters prevail.
Assuntos
Anticorpos Monoclonais/uso terapêutico , Antígenos CD20/imunologia , Astato/uso terapêutico , Imunoconjugados/uso terapêutico , Linfoma de Células B/radioterapia , Animais , Feminino , Humanos , Células Jurkat , Linfoma de Células B/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos NOD , Camundongos Nus , Camundongos SCID , Radioimunoterapia , Resultado do Tratamento , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Antibodies armed with biologic drugs could greatly expand the therapeutic potential of antibody-drug conjugates for cancer therapy, broadening their application to disease targets currently limited by intracellular delivery barriers. Additional selectivity and new therapeutic approaches could be realized with intracellular protein drugs that more specifically target dysregulated pathways in hematologic cancers and other malignancies. A multifunctional polymeric delivery system for enhanced cytosolic delivery of protein drugs has been developed that incorporates endosomal-releasing activity, antibody targeting, and a biocompatible long-chain ethylene glycol component for optimized safety, pharmacokinetics, and tumor biodistribution. The pH-responsive polymeric micelle carrier, with an internalizing anti-CD22 monoclonal targeting antibody, effectively delivered a proapoptotic Bcl-2 interacting mediator (BIM) peptide drug that suppressed tumor growth for the duration of treatment and prolonged survival in a xenograft mouse model of human B-cell lymphoma. Antitumor drug activity was correlated with a mechanistic induction of the Bcl-2 pathway biomarker cleaved caspase-3 and a marked decrease in the Ki-67 proliferation biomarker. Broadening the intracellular target space by more effective delivery of protein/peptide drugs could expand the repertoire of antibody-drug conjugates to currently undruggable disease-specific targets and permit tailored drug strategies to stratified subpopulations and personalized medicines.
Assuntos
Anticorpos Monoclonais , Sistemas de Liberação de Medicamentos , Imunoconjugados/farmacologia , Peptídeos , Animais , Apoptose/efeitos dos fármacos , Disponibilidade Biológica , Biomarcadores , Linhagem Celular Tumoral , Citocromos c/biossíntese , Modelos Animais de Doenças , Estabilidade de Medicamentos , Humanos , Imunoconjugados/química , Imunoconjugados/farmacocinética , Imunoconjugados/toxicidade , Linfoma de Células B/tratamento farmacológico , Linfoma de Células B/metabolismo , Linfoma de Células B/mortalidade , Linfoma de Células B/patologia , Camundongos , Micelas , Polímeros/química , Proteínas Proto-Oncogênicas c-bcl-2/química , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Distribuição Tecidual , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Despite aggressive chemotherapy combined with hematopoietic stem cell transplantation (HSCT), many patients with acute myeloid leukemia (AML) relapse. Radioimmunotherapy (RIT) using monoclonal antibodies labeled with ß-emitting radionuclides has been explored to reduce relapse. ß emitters are limited by lower energies and nonspecific cytotoxicity from longer path lengths compared with α emitters such as (211)At, which has a higher energy profile and shorter path length. We evaluated the efficacy and toxicity of anti-CD45 RIT using (211)At in a disseminated murine AML model. Biodistribution studies in leukemic SJL/J mice showed excellent localization of (211)At-anti-murine CD45 mAb (30F11) to marrow and spleen within 24 hours (18% and 79% injected dose per gram of tissue [ID/g], respectively), with lower kidney and lung uptake (8.4% and 14% ID/g, respectively). In syngeneic HSCT studies, (211)At-B10-30F11 RIT improved the median survival of leukemic mice in a dose-dependent fashion (123, 101, 61, and 37 days given 24, 20, 12, and 0 µCi, respectively). This approach had minimal toxicity with nadir white blood cell counts >2.7 K/µL 2 weeks after HSCT and recovery by 4 weeks. These data suggest that (211)At-anti-CD45 RIT in conjunction with HSCT may be a promising therapeutic option for AML.
Assuntos
Anticorpos Monoclonais/uso terapêutico , Astato/uso terapêutico , Transplante de Medula Óssea , Leucemia/terapia , Antígenos Comuns de Leucócito/imunologia , Radioimunoterapia/métodos , Animais , Terapia Combinada/métodos , Modelos Animais de Doenças , Feminino , Leucemia/mortalidade , Leucemia/patologia , Leucemia/radioterapia , Camundongos , Metástase Neoplásica , Análise de Sobrevida , Distribuição Tecidual , Resultado do Tratamento , Células Tumorais CultivadasRESUMO
Mantle cell lymphoma (MCL) remains incurable for most patients, and proteasome inhibitors like bortezomib induce responses in a minority of patients with relapsed disease. Fenretinide is a retinoid that has shown preclinical activity in B-cell lymphomas. We hypothesized that these agents could yield augmented antitumor activity. MCL lines (Granta-519, Jeko-1, and Rec-1) were treated with escalating concentrations of bortezomib and fenretinide singly and in combination. Cytotoxicity was assessed using the MTT assay. Flow cytometric methods were used to assess apoptosis and necrosis, with annexin V-FITC/propidium iodide staining, and G1 and G2 cell-cycle changes were assessed by DAPI staining. Changes in cyclin D1, cyclin B, IκBα, and IKKα expressions were quantified by western blotting. Cytotoxicity was mediated through apoptosis; both agents showed observed versus expected cytotoxicities of 92.2 versus 55.1% in Granta-519, of 87.6 versus 36.3% in Jeko-1, and of 63.2 versus 29.8% in Rec-1. Isobolographic analysis confirmed synergy in Jeko-1 and Rec-1 cell lines. Bortezomib induced G2-phase arrest, with a 1.7-fold increase compared with control, and fenretinide resulted in G1-phase arrest, with an increase of 1.3-fold compared with control. In the combination, G2-phase arrest predominated, with a 1.4-fold increase compared with control, and there was reduced expression of cyclin D1 to 24%, cyclin B to 52 and 64%, cyclin D3 to 25 and 43%, IκBα to 23 and 46%, and IκBα kinase to 34 and 44%. Bortezomib and fenretinide exhibit synergistic cytotoxicity against MCL cell lines. This activity is mediated by IκBα kinase modulation, decreased cyclin expression, cell cycle dysregulation, and apoptotic cell death.
Assuntos
Antineoplásicos/farmacologia , Ácidos Borônicos/farmacologia , Fenretinida/farmacologia , Linfoma de Célula do Manto/patologia , Pirazinas/farmacologia , Apoptose/efeitos dos fármacos , Bortezomib , Linhagem Celular Tumoral/efeitos dos fármacos , Ciclina B/metabolismo , Ciclina D1/metabolismo , Ciclina D3/metabolismo , Sinergismo Farmacológico , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Humanos , Proteínas I-kappa B/metabolismo , Inibidor de NF-kappaB alfaRESUMO
Given the poor outcomes of relapsed aggressive lymphomas and preclinical data suggesting that ≥2·5 µmol/l concentrations of vorinostat synergize with both etoposide and platinums, we hypothesized that pulse high-dose vorinostat could safely augment the anti-tumour activity of (R)ICE [(rituximab), ifosphamide, carboplatin, etoposide] chemotherapy. We conducted a phase I dose escalation study using a schedule with oral vorinostat ranging from 400 mg/d to 700 mg bid for 5 d in combination with the standard (R)ICE regimen (days 3, 4 and 5). Twenty-nine patients [median age 56 years, median 2 prior therapies, 14 chemoresistant (of 27 evaluable), 2 prior transplants] were enrolled and treated. The maximally tolerated vorinostat dose was defined as 500 mg twice daily × 5 d. Common dose limiting toxicities included infection (n = 2), hypokalaemia (n = 2), and transaminitis (n = 2). Grade 3 related gastrointestinal toxicity was seen in 9 patients. The median vorinostat concentration on day 3 was 4·5 µmol/l (range 4·2-6·0 µmol/l) and in vitro data confirmed the augmented antitumour and histone acetylation activity at these levels. Responses were observed in 19 of 27 evaluable patients (70%) including 8 complete response/unconfirmed complete response. High-dose vorinostat can be delivered safely with (R)ICE, achieves potentially synergistic drug levels, and warrants further study, although adequate gastrointestinal prophylaxis is warranted.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Linfoma/tratamento farmacológico , Adulto , Idoso , Anticorpos Monoclonais Murinos/administração & dosagem , Anticorpos Monoclonais Murinos/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Carboplatina/administração & dosagem , Carboplatina/efeitos adversos , Sinergismo Farmacológico , Etoposídeo/administração & dosagem , Etoposídeo/efeitos adversos , Feminino , Humanos , Ácidos Hidroxâmicos/administração & dosagem , Ácidos Hidroxâmicos/efeitos adversos , Ifosfamida/administração & dosagem , Ifosfamida/efeitos adversos , Masculino , Pessoa de Meia-Idade , Rituximab , VorinostatRESUMO
Pretargeted radioimmunotherapy (PRIT) using an anti-CD45 antibody (Ab)-streptavidin (SA) conjugate and DOTA-biotin labeled with ß-emitting radionuclides has been explored as a strategy to decrease relapse and toxicity. α-emitting radionuclides exhibit high cytotoxicity coupled with a short path length, potentially increasing the therapeutic index and making them an attractive alternative to ß-emitting radionuclides for patients with acute myeloid leukemia. Accordingly, we have used (213)Bi in mice with human leukemia xenografts. Results demonstrated excellent localization of (213)Bi-DOTA-biotin to tumors with minimal uptake into normal organs. After 10 minutes, 4.5% ± 1.1% of the injected dose of (213)Bi was delivered per gram of tumor. α-imaging demonstrated uniform radionuclide distribution within tumor tissue 45 minutes after (213)Bi-DOTA-biotin injection. Radiation absorbed doses were similar to those observed using a ß-emitting radionuclide ((90)Y) in the same model. We conducted therapy experiments in a xenograft model using a single-dose of (213)Bi-DOTA-biotin given 24 hours after anti-CD45 Ab-SA conjugate. Among mice treated with anti-CD45 Ab-SA conjugate followed by 800 µCi of (213)Bi- or (90)Y-DOTA-biotin, 80% and 20%, respectively, survived leukemia-free for more than 100 days with minimal toxicity. These data suggest that anti-CD45 PRIT using an α-emitting radionuclide may be highly effective and minimally toxic for treatment of acute myeloid leukemia.
Assuntos
Anticorpos/farmacologia , Bismuto/farmacologia , Leucemia Mieloide Aguda/radioterapia , Antígenos Comuns de Leucócito/antagonistas & inibidores , Radioimunoterapia/métodos , Radioisótopos/farmacologia , Animais , Biotina/análogos & derivados , Biotina/farmacologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Relação Dose-Resposta Imunológica , Relação Dose-Resposta à Radiação , Feminino , Humanos , Leucemia Mieloide Aguda/imunologia , Antígenos Comuns de Leucócito/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Compostos Organometálicos/farmacologia , Indução de Remissão , Estreptavidina/farmacologia , Taxa de Sobrevida , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Radioimmunotherapy (RIT) with α-emitting radionuclides is an attractive approach for the treatment of minimal residual disease because the short path lengths and high energies of α-particles produce optimal cytotoxicity at small target sites while minimizing damage to surrounding normal tissues. Pretargeted RIT (PRIT) using antibody-streptavidin (Ab-SA) constructs and radiolabeled biotin allows rapid, specific localization of radioactivity at tumor sites, making it an optimal method to target α-emitters with short half-lives, such as bismuth-213 (²¹³Bi). Athymic mice bearing Ramos lymphoma xenografts received anti-CD20 1F5(scFv)(4)SA fusion protein (FP), followed by a dendrimeric clearing agent and [²¹³Bi]DOTA-biotin. After 90 minutes, tumor uptake for 1F5(scFv)4SA was 16.5% ± 7.0% injected dose per gram compared with 2.3% ± .9% injected dose per gram for the control FP. Mice treated with anti-CD20 PRIT and 600 µ Ci [²¹³Bi]DOTA-biotin exhibited marked tumor growth delays compared with controls (mean tumor volume .01 ± .02 vs. 203.38 ± 83.03 mm³ after 19 days, respectively). The median survival for the 1F5(scFv)4SA group was 90 days compared with 23 days for the control FP (P < .0001). Treatment was well tolerated, with no treatment-related mortalities. This study demonstrates the favorable biodistribution profile and excellent therapeutic efficacy attainable with ²¹³Bi-labeled anti-CD20 PRIT.
Assuntos
Antígenos CD20/metabolismo , Bismuto/uso terapêutico , Linfoma não Hodgkin/imunologia , Linfoma não Hodgkin/radioterapia , Neoplasia Residual/tratamento farmacológico , Radioimunoterapia/métodos , Ensaios Antitumorais Modelo de Xenoenxerto , Animais , Anticorpos Antineoplásicos/imunologia , Biotina/efeitos adversos , Biotina/análogos & derivados , Biotina/farmacocinética , Biotina/farmacologia , Biotina/uso terapêutico , Bismuto/efeitos adversos , Bismuto/farmacocinética , Bismuto/farmacologia , Contagem de Células Sanguíneas , Linhagem Celular Tumoral , Humanos , Região Variável de Imunoglobulina/imunologia , Rim/efeitos dos fármacos , Rim/metabolismo , Rim/fisiopatologia , Testes de Função Hepática , Linfoma não Hodgkin/patologia , Linfoma não Hodgkin/fisiopatologia , Camundongos , Neoplasia Residual/imunologia , Compostos Organometálicos/efeitos adversos , Compostos Organometálicos/farmacocinética , Compostos Organometálicos/farmacologia , Compostos Organometálicos/uso terapêutico , Radiometria , Proteínas Recombinantes de Fusão/metabolismo , Análise de Sobrevida , Distribuição Tecidual/efeitos dos fármacosRESUMO
Mantle cell lymphoma (MCL) affects approximately 4500 patients/year in the US and demonstrates a male to female ratio of approximately 4:1. While the pathobiology underlying this ratio is unknown, the hematopoietic system is characterized by sex-related differences in androgen receptor (AR) expression, leading us to hypothesize that the male-biased incidence of MCL may reflect sex-related differences in AR signaling during MCL lymphomagenesis. To explore the AR axis in MCL, we evaluated AR expression in MCL cell lines and human tumors, and tested the impact of androgen pathway inhibition on MCL proliferation. AR transcript levels ranged up to ~26 fold higher in MCL lines vs non-MCL NHL lines (p = 0.006) and were correlated with expression of the canonical AR-regulated gene, prostate-specific antigen (PSA; r = 0.715, p = 0.001), consistent with functional AR activity. Patient-derived MCL samples demonstrated a range of AR expression. Treatment of four different MCL lines with the potent AR antagonist enzalutamide demonstrated suppression of proliferation across both male and female-derived cell lines. These data suggest androgen-axis blockade may represent a novel therapeutic modality in MCL. This novel treatment approach is currently under investigation in a phase II clinical trial of AR inhibition in patients with relapsed/refractory MCL.
Assuntos
Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Linfoma de Célula do Manto/tratamento farmacológico , Linfoma de Célula do Manto/metabolismo , Proteínas de Neoplasias/biossíntese , Feniltioidantoína/análogos & derivados , Receptores Androgênicos/biossíntese , Benzamidas , Linhagem Celular Tumoral , Feminino , Humanos , Masculino , Nitrilas , Feniltioidantoína/farmacologia , Antígeno Prostático Específico/biossíntese , Fatores SexuaisRESUMO
Streptavidin (SA)-biotin pretargeted radioimmunotherapy (PRIT) that targets CD20 in non-Hodgkin lymphoma (NHL) exhibits remarkable efficacy in model systems, but SA immunogenicity and interference by endogenous biotin may complicate clinical translation of this approach. In this study, we engineered a bispecific fusion protein (FP) that evades the limitations imposed by this system. Briefly, one arm of the FP was an anti-human CD20 antibody (2H7), with the other arm of the FP an anti-chelated radiometal trap for a radiolabeled ligand (yttrium[Y]-DOTA) captured by a very high-affinity anti-Y-DOTA scFv antibody (C825). Head-to-head biodistribution experiments comparing SA-biotin and bispecific FP (2H7-Fc-C825) PRIT in murine subjects bearing human lymphoma xenografts demonstrated nearly identical tumor targeting by each modality at 24 hours. However, residual radioactivity in the blood and normal organs was consistently higher following administration of 1F5-SA compared with 2H7-Fc-C825. Consequently, tumor-to-normal tissue ratios of distribution were superior for 2H7-Fc-C825 (P < 0.0001). Therapy studies in subjects bearing either Ramos or Granta subcutaneous lymphomas demonstrated that 2H7-Fc-C825 PRIT is highly effective and significantly less myelosuppressive than 1F5-SA (P < 0.0001). All animals receiving optimal doses of 2H7-Fc-C825 followed by 90Y-DOTA were cured by 150 days, whereas the growth of tumors in control animals progressed rapidly with complete morbidity by 25 days. In addition to demonstrating reduced risk of immunogenicity and an absence of endogenous biotin interference, our findings offer a preclinical proof of concept for the preferred use of bispecific PRIT in future clinical trials, due to a slightly superior biodistribution profile, less myelosuppression, and superior efficacy. Cancer Res; 76(22); 6669-79. ©2016 AACR.
Assuntos
Anticorpos Biespecíficos/metabolismo , Linfoma de Células B/tratamento farmacológico , Linfoma de Células B/radioterapia , Radioimunoterapia/métodos , Estreptavidina/uso terapêutico , Animais , Anticorpos Biespecíficos/análise , Feminino , Humanos , Linfoma de Células B/patologia , Camundongos , Estreptavidina/farmacologiaRESUMO
PURPOSE: Alpha-emitting radionuclides exhibit a potential advantage for cancer treatments because they release large amounts of ionizing energy over a few cell diameters (50-80 µm), causing localized, irreparable double-strand DNA breaks that lead to cell death. Radioimmunotherapy (RIT) approaches using monoclonal antibodies labeled with α emitters may thus inactivate targeted cells with minimal radiation damage to surrounding tissues. Tools are needed to visualize and quantify the radioactivity distribution and absorbed doses to targeted and nontargeted cells for accurate dosimetry of all treatment regimens utilizing α particles, including RIT and others (e.g., Ra-223), especially for organs and tumors with heterogeneous radionuclide distributions. The aim of this study was to evaluate and characterize a novel single-particle digital autoradiography imager, the ionizing-radiation quantum imaging detector (iQID) camera, for use in α-RIT experiments. METHODS: The iQID camera is a scintillator-based radiation detection system that images and identifies charged-particle and gamma-ray/x-ray emissions spatially and temporally on an event-by-event basis. It employs CCD-CMOS cameras and high-performance computing hardware for real-time imaging and activity quantification of tissue sections, approaching cellular resolutions. In this work, the authors evaluated its characteristics for α-particle imaging, including measurements of intrinsic detector spatial resolutions and background count rates at various detector configurations and quantification of activity distributions. The technique was assessed for quantitative imaging of astatine-211 ((211)At) activity distributions in cryosections of murine and canine tissue samples. RESULTS: The highest spatial resolution was measured at â¼20 µm full width at half maximum and the α-particle background was measured at a rate as low as (2.6 ± 0.5) × 10(-4) cpm/cm(2) (40 mm diameter detector area). Simultaneous imaging of multiple tissue sections was performed using a large-area iQID configuration (ø 11.5 cm). Estimation of the (211)At activity distribution was demonstrated at mBq/µg-levels. CONCLUSIONS: Single-particle digital autoradiography of α emitters has advantages over traditional film-based autoradiographic techniques that use phosphor screens, in terms of spatial resolution, sensitivity, and activity quantification capability. The system features and characterization results presented in this study show that the iQID is a promising technology for microdosimetry, because it provides necessary information for interpreting alpha-RIT outcomes and for predicting the therapeutic efficacy of cell-targeted approaches using α emitters.
Assuntos
Autorradiografia/instrumentação , Autorradiografia/métodos , Câmaras gama , Radioimunoterapia/instrumentação , Radioimunoterapia/métodos , Animais , Antígenos CD20/administração & dosagem , Astato , Cães , Desenho de Equipamento , Feminino , Antígenos Comuns de Leucócito/administração & dosagem , Linfonodos/diagnóstico por imagem , Linfonodos/imunologia , Linfonodos/efeitos da radiação , Linfoma não Hodgkin/diagnóstico por imagem , Linfoma não Hodgkin/imunologia , Linfoma não Hodgkin/radioterapia , Camundongos , Camundongos Nus , Transplante de Neoplasias , Imagens de Fantasmas , Radiografia , SoftwareRESUMO
PURPOSE: Pretargeted radioimmunotherapy (PRIT) is a multi-step method of selectively delivering high doses of radiotherapy to tumor cells while minimizing exposure to surrounding tissues. Yttrium-90 (90Y) and lutetium-177 (177Lu) are two of the most promising beta-particle emitting radionuclides used for radioimmunotherapy, which despite having similar chemistries differ distinctly in terms of radiophysical features. These differences may have important consequences for the absorbed dose to tumors and normal organs. Whereas 90Y has been successfully applied in a number of preclinical and clinical radioimmunotherapy settings, there have been few published pretargeting studies with 177Lu. We therefore compared the therapeutic potential of targeting either 90Y or 177Lu to human B-cell lymphoma xenografts in mice. METHODS: Parallel experiments evaluating the biodistribution, imaging, dosimetry, therapeutic efficacy, and toxicity were performed in female athymic nude mice bearing either Ramos (Burkitt lymphoma) or Granta (mantle cell lymphoma) xenografts, utilizing an anti-CD20 antibody-streptavidin conjugate (1F5-SA) and an 90Y- or 177Lu-labeled 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA)-biotin second step reagent. RESULTS: The two radionuclides displayed comparable biodistributions in tumors and normal organs; however, the absorbed radiation dose delivered to tumor was more than twice as high for 90Y (1.3 Gy/MBq) as for 177Lu (0.6 Gy/MBq). More importantly, therapy with 90Y-DOTA-biotin was dramatically more effective than with 177Lu-DOTA-biotin, with 100% of Ramos xenograft-bearing mice cured with 37 MBq 90Y, whereas 0% were cured using identical amounts of 177Lu-DOTA-biotin. Similar results were observed in mice bearing Granta xenografts, with 80% of the mice cured with 90Y-PRIT and 0% cured with 177Lu-PRIT. Toxicities were comparable with both isotopes. CONCLUSION: 90Y was therapeutically superior to 177Lu for streptavidin-biotin PRIT approaches in these human lymphoma xenograft models.
Assuntos
Antígenos CD20/imunologia , Imunoconjugados/uso terapêutico , Lutécio/uso terapêutico , Linfoma/radioterapia , Radioimunoterapia/métodos , Radioisótopos de Ítrio/uso terapêutico , Animais , Partículas beta/uso terapêutico , Linhagem Celular Tumoral , Transformação Celular Neoplásica , Feminino , Humanos , Imunoconjugados/efeitos adversos , Imunoconjugados/imunologia , Lutécio/efeitos adversos , Lutécio/farmacocinética , Linfoma/imunologia , Linfoma/patologia , Camundongos , Camundongos Nus , Radioimunoterapia/efeitos adversos , Distribuição Tecidual , Ensaios Antitumorais Modelo de Xenoenxerto , Radioisótopos de Ítrio/efeitos adversos , Radioisótopos de Ítrio/farmacocinéticaRESUMO
UNLABELLED: α-radioimmunotherapy targeting CD45 may substitute for total-body irradiation in hematopoietic cell transplantation (HCT) preparative regimens for lymphoma. Our goal was to optimize the anti-CD45 monoclonal antibody (mAb; CA12.10C12) protein dose for (211)At-radioimmunotherapy, extending the analysis to include intraorgan (211)At activity distribution and α-imaging-based small-scale dosimetry, along with immunohistochemical staining. METHODS: Eight normal dogs were injected with either a 0.75 (n = 5) or 1.00 (n = 3) mg/kg dose of (211)At-B10-CA12.10C12 (11.5-27.6 MBq/kg). Two were euthanized and necropsied 19-22 h after injection, and 6 received autologous HCT 3 d after (211)At-radioimmunotherapy, after lymph node and bone marrow biopsies at 2-4 and/or 19 h after injection. Blood was sampled to study toxicity and clearance; CD45 targeting was evaluated by flow cytometry. (211)At localization and small-scale dosimetry were assessed using two α-imaging systems: an α-camera and an ionizing-radiation quantum imaging detector (iQID) camera. RESULTS: (211)At uptake was highest in the spleen (0.31-0.61% injected activity [%IA]/g), lymph nodes (0.02-0.16 %IA/g), liver (0.11-0.12 %IA/g), and marrow (0.06-0.08 %IA/g). Lymphocytes in blood and marrow were efficiently targeted using either mAb dose. Lymph nodes remained unsaturated but displayed targeted (211)At localization in T lymphocyte-rich areas. Absorbed doses to blood, marrow, and lymph nodes were estimated at 3.1, 2.4, and 3.4 Gy/166 MBq, respectively. All transplanted dogs experienced transient hepatic toxicity. Liver enzyme levels were temporarily elevated in 5 of 6 dogs; one treated with 1.00 mg mAb/kg developed ascites and was euthanized 136 d after HCT. CONCLUSION: (211)At-anti-CD45 radioimmunotherapy with 0.75 mg mAb/kg efficiently targeted blood and marrow without severe toxicity. Dosimetry calculations and observed radiation-induced effects indicated that sufficient (211)At-B10-CA12.10C12 localization was achieved for efficient conditioning for HCT.
Assuntos
Astato/farmacocinética , Transplante de Células-Tronco Hematopoéticas/métodos , Antígenos Comuns de Leucócito , Radioimunoterapia/métodos , Compostos Radiofarmacêuticos/farmacocinética , Partículas alfa , Animais , Ascite/diagnóstico por imagem , Astato/efeitos adversos , Biópsia , Medula Óssea/diagnóstico por imagem , Cães , Sistemas de Liberação de Medicamentos , Imuno-Histoquímica , Linfonodos/diagnóstico por imagem , Radiometria , Cintilografia , Compostos Radiofarmacêuticos/efeitos adversos , Baço/diagnóstico por imagem , Linfócitos T/diagnóstico por imagem , Distribuição TecidualRESUMO
The vast majority of patients with plasma cell neoplasms die of progressive disease despite high response rates to novel agents. Malignant plasma cells are very radiosensitive, but the potential role of radioimmunotherapy (RIT) in the management of plasmacytomas and multiple myeloma has undergone only limited evaluation. Furthermore, CD38 has not been explored as a RIT target despite its uniform high expression on malignant plasma cells. In this report, both conventional RIT (directly radiolabeled antibody) and streptavidin-biotin pretargeted RIT (PRIT) directed against the CD38 antigen were assessed as approaches to deliver radiation doses sufficient for multiple myeloma cell eradication. PRIT demonstrated biodistributions that were markedly superior to conventional RIT. Tumor-to-blood ratios as high as 638:1 were seen 24 hours after PRIT, whereas ratios never exceeded 1:1 with conventional RIT. (90)Yttrium absorbed dose estimates demonstrated excellent target-to-normal organ ratios (6:1 for the kidney, lung, liver; 10:1 for the whole body). Objective remissions were observed within 7 days in 100% of the mice treated with doses ranging from 800 to 1,200 µCi of anti-CD38 pretargeted (90)Y-DOTA-biotin, including 100% complete remissions (no detectable tumor in treated mice compared with tumors that were 2,982% ± 2,834% of initial tumor volume in control animals) by day 23. Furthermore, 100% of animals bearing NCI-H929 multiple myeloma tumor xenografts treated with 800 µCi of anti-CD38 pretargeted (90)Y-DOTA-biotin achieved long-term myeloma-free survival (>70 days) compared with none (0%) of the control animals.
Assuntos
ADP-Ribosil Ciclase 1/imunologia , Anticorpos Monoclonais/uso terapêutico , Compostos Heterocíclicos/uso terapêutico , Terapia de Alvo Molecular/métodos , Neoplasias de Plasmócitos/radioterapia , Compostos Organometálicos/uso terapêutico , Radioimunoterapia/métodos , Animais , Células Cultivadas , Feminino , Humanos , Camundongos , Camundongos Nus , Camundongos Transgênicos , Ensaios Antitumorais Modelo de Xenoenxerto , Radioisótopos de Ítrio/uso terapêuticoRESUMO
Modification of T cells with chimeric antigen receptors (CAR) has emerged as a promising treatment modality for human malignancies. Integration of co-stimulatory domains into CARs can augment the activation and function of genetically targeted T cells against tumors. However, the potential for insertional mutagenesis and toxicities due to the infused cells have made development of safe methods for removing transferred cells an important consideration. We have genetically modified human T cells with a lentiviral vector to express a CD20-CAR containing both CD28 and CD137 co-stimulatory domains, a "suicide gene" relying on inducible activation of caspase 9 (iC9), and a truncated CD19 selectable marker. Rapid expansion (2000 fold) of the transduced T cells was achieved in 28 days after stimulation with artificial antigen presenting cells. Transduced T cells exhibited effective CD20-specific cytotoxic activity in vitro and in a mouse xenograft tumor model. Activation of the iC9 suicide switch resulted in efficient removal of transduced T cells both in vitro and in vivo. Our work demonstrates the feasibility and promise of this approach for treating CD20(+) malignancies in a safe and more efficient manner. A phase I clinical trial using this approach in patients with relapsed indolent B-NHL is planned.
Assuntos
Antígenos CD20/genética , Caspase 9/genética , Genes Transgênicos Suicidas , Imunoterapia Adotiva/métodos , Linfoma/terapia , Receptores de Antígenos de Linfócitos T/genética , Linfócitos T/transplante , Animais , Células Cultivadas , Humanos , Células Jurkat , Linfoma/genética , Linfoma/imunologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Células NIH 3T3 , Proteínas Recombinantes de Fusão/genética , Linfócitos T/metabolismo , Resultado do Tratamento , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
PURPOSE: Pretargeted radioimmunotherapy (PRIT) using streptavidin (SAv)-biotin technology can deliver higher therapeutic doses of radioactivity to tumors than conventional RIT. However, "endogenous" biotin can interfere with the effectiveness of this approach by blocking binding of radiolabeled biotin to SAv. We engineered a series of SAv FPs that downmodulate the affinity of SAv for biotin, while retaining high avidity for divalent DOTA-bis-biotin to circumvent this problem. EXPERIMENTAL DESIGN: The single-chain variable region gene of the murine 1F5 anti-CD20 antibody was fused to the wild-type (WT) SAv gene and to mutant SAv genes, Y43A-SAv and S45A-SAv. FPs were expressed, purified, and compared in studies using athymic mice bearing Ramos lymphoma xenografts. RESULTS: Biodistribution studies showed delivery of more radioactivity to tumors of mice pretargeted with mutant SAv FPs followed by (111)In-DOTA-bis-biotin [6.2 ± 1.7% of the injected dose per gram (%ID/gm) of tumor 24 hours after Y43A-SAv FP and 5.6 ± 2.2%ID/g with S45A-SAv FP] than in mice on normal diets pretargeted with WT-SAv FP (2.5 ± 1.6%ID/g; P = 0.01). These superior biodistributions translated into superior antitumor efficacy in mice treated with mutant FPs and (90)Y-DOTA-bis-biotin [tumor volumes after 11 days: 237 ± 66 mm(3) with Y43A-SAv, 543 ± 320 mm(3) with S45A-SAv, 1129 ± 322 mm(3) with WT-SAv, and 1435 ± 212 mm(3) with control FP (P < 0.0001)]. CONCLUSIONS: Genetically engineered mutant-SAv FPs and bis-biotin reagents provide an attractive alternative to current SAv-biotin PRIT methods in settings where endogenous biotin levels are high.
Assuntos
Linfoma não Hodgkin/radioterapia , Radioimunoterapia/métodos , Animais , Anticorpos Monoclonais/genética , Antígenos CD20/imunologia , Linhagem Celular Tumoral , Linfoma não Hodgkin/imunologia , Camundongos , Camundongos Nus , Engenharia de Proteínas , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Anticorpos de Cadeia Única/genética , Estreptavidina/genética , Estreptavidina/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The structural maintenance of chromosome 1 (Smc1) protein is a member of the highly conserved cohesin complex and is involved in sister chromatid cohesion. In response to ionizing radiation, Smc1 is phosphorylated at two sites, Ser-957 and Ser-966, and these phosphorylation events are dependent on the ATM protein kinase. In this study, we describe the generation of two novel ELISAs for quantifying phospho-Smc1(Ser-957) and phospho-Smc1(Ser-966). Using these novel assays, we quantify the kinetic and biodosimetric responses of human cells of hematological origin, including immortalized cells, as well as both quiescent and cycling primary human PBMC. Additionally, we demonstrate a robust in vivo response for phospho-Smc1(Ser-957) and phospho-Smc1(Ser-966) in lymphocytes of human patients after therapeutic exposure to ionizing radiation, including total-body irradiation, partial-body irradiation, and internal exposure to (131)I. These assays are useful for quantifying the DNA damage response in experimental systems and potentially for the identification of individuals exposed to radiation after a radiological incident.