RESUMEN
A chromosome 14 inversion was found in a patient who developed bone marrow aplasia following treatment with allogeneic chimeric antigen receptor (CAR) Tcells containing gene edits made with transcription activator-like effector nucleases (TALEN). TALEN editing sites were not involved at either breakpoint. Recombination signal sequences (RSSs) were found suggesting recombination-activating gene (RAG)-mediated activity. The inversion represented a dominant clone detected in the context of decreasing absolute CAR Tcell and overall lymphocyte counts. The inversion was not associated with clinical consequences and wasnot detected in the drug product administered to this patient or in any drug product used in this or other trials using the same manufacturing processes. Neither was the inversion detected in this patient at earlier time points or in any other patient enrolled in this or other trials treated with this or other product lots. This case illustrates that spontaneous, possibly RAG-mediated, recombination events unrelated to gene editing can occur in adoptive cell therapy studies, emphasizes the need for ruling out off-target gene editing sites, and illustrates that other processes, such as spontaneous V(D)J recombination, can lead to chromosomal alterations in infused cells independent of gene editing.
Asunto(s)
Trasplante de Células Madre Hematopoyéticas , Receptores Quiméricos de Antígenos , Humanos , Edición Génica , Nucleasas de los Efectores Tipo Activadores de la Transcripción/genética , Linfocitos T , Receptores Quiméricos de Antígenos/genética , Inmunoterapia Adoptiva/efectos adversosRESUMEN
Patients with relapsed or refractory acute myeloid leukemia (AML) have a dismal prognosis and limited treatment options. Chimeric antigen receptor (CAR) T cells have achieved unprecedented clinical responses in patients with B cell leukemias and lymphomas and could prove highly efficacious in AML. However, a significant number of patients with AML may not receive treatment with an autologous product due to manufacturing failures associated with low lymphocyte counts or rapid disease progression while the therapeutic is being produced. We report the preclinical evaluation of an off-the-shelf CAR T cell therapy targeting Fms-related tyrosine kinase 3 (FLT3) for the treatment of AML. Single-chain variable fragments (scFvs) targeting various epitopes in the extracellular region of FLT3 were inserted into CAR constructs and tested for their ability to redirect T cell specificity and effector function to FLT3+ AML cells. A lead CAR, exhibiting minimal tonic signaling and robust activity in vitro and in vivo, was selected and then modified to incorporate a rituximab-responsive off-switch in cis. We found that allogeneic FLT3 CAR T cells, generated from healthy-donor T cells, eliminate primary AML blasts but are also active against mouse and human hematopoietic stem and progenitor cells, indicating risk of myelotoxicity. By employing a surrogate CAR with affinity to murine FLT3, we show that rituximab-mediated depletion of FLT3 CAR T cells after AML eradication enables bone marrow recovery without compromising leukemia remission. These results support clinical investigation of allogeneic FLT3 CAR T cells in AML and other FLT3+ hematologic malignancies.
Asunto(s)
Inmunoterapia Adoptiva , Leucemia Mieloide Aguda/inmunología , Leucemia Mieloide Aguda/terapia , Receptores Quiméricos de Antígenos/inmunología , Linfocitos T/inmunología , Tirosina Quinasa 3 Similar a fms/inmunología , Animales , Médula Ósea/inmunología , Médula Ósea/metabolismo , Modelos Animales de Enfermedad , Humanos , Inmunoterapia Adoptiva/efectos adversos , Inmunoterapia Adoptiva/métodos , Leucemia Mieloide Aguda/diagnóstico , Ratones , Receptores Quiméricos de Antígenos/genética , Especificidad del Receptor de Antígeno de Linfocitos T , Linfocitos T/metabolismo , Resultado del Tratamiento , Ensayos Antitumor por Modelo de Xenoinjerto , Tirosina Quinasa 3 Similar a fms/antagonistas & inhibidoresRESUMEN
FLT3 (FMS-like tyrosine kinase 3), expressed on the surface of acute myeloid leukemia (AML) blasts, is a promising AML target, given its role in the development and progression of leukemia, and its limited expression in tissues outside the hematopoietic system. Small molecule FLT3 kinase inhibitors have been developed, but despite having clinical efficacy, they are effective only on a subset of patients and associated with high risk of relapse. A durable therapy that can target a wider population of AML patients is needed. Here, we developed an anti-FLT3-CD3 immunoglobulin G (IgG)-based bispecific antibody (7370) with a high affinity for FLT3 and a long half-life, to target FLT3-expressing AML blasts, irrespective of FLT3 mutational status. We demonstrated that 7370 has picomolar potency against AML cell lines in vitro and in vivo. 7370 was also capable of activating T cells from AML patients, redirecting their cytotoxic activity against autologous blasts at low effector-to-target (E:T) ratio. Additionally, under our dosing regimen, 7370 was well tolerated and exhibited potent efficacy in cynomolgus monkeys by inducing complete but reversible depletion of peripheral FLT3+ dendritic cells (DCs) and bone marrow FLT3+ stem cells and progenitors. Overall, our results support further clinical development of 7370 to broadly target AML patients.
Asunto(s)
Anticuerpos Biespecíficos/farmacología , Antineoplásicos Inmunológicos/farmacología , Complejo CD3/antagonistas & inhibidores , Hematopoyesis/efectos de los fármacos , Tirosina Quinasa 3 Similar a fms/antagonistas & inhibidores , Animales , Anticuerpos Biespecíficos/química , Anticuerpos Biespecíficos/uso terapéutico , Antineoplásicos Inmunológicos/química , Antineoplásicos Inmunológicos/uso terapéutico , Médula Ósea/efectos de los fármacos , Médula Ósea/metabolismo , Médula Ósea/patología , Complejo CD3/química , Línea Celular Tumoral , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Humanos , Inmunoglobulina G/farmacología , Inmunofenotipificación , Leucemia Mieloide Aguda , Depleción Linfocítica , Macaca fascicularis , Ratones , Modelos Moleculares , Dominios Proteicos/efectos de los fármacos , Relación Estructura-Actividad , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Linfocitos T/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto , Tirosina Quinasa 3 Similar a fms/químicaRESUMEN
Clinical success of autologous CD19-directed chimeric antigen receptor T cells (CAR Ts) in acute lymphoblastic leukemia and non-Hodgkin lymphoma suggests that CAR Ts may be a promising therapy for hematological malignancies, including multiple myeloma. However, autologous CAR T therapies have limitations that may impact clinical use, including lengthy vein-to-vein time and manufacturing constraints. Allogeneic CAR T (AlloCAR T) therapies may overcome these innate limitations of autologous CAR T therapies. Unlike autologous cell therapies, AlloCAR T therapies employ healthy donor T cells that are isolated in a manufacturing facility, engineered to express CARs with specificity for a tumor-associated antigen, and modified using gene-editing technology to limit T cell receptor (TCR)-mediated immune responses. Here, transcription activator-like effector nuclease (TALEN) gene editing of B cell maturation antigen (BCMA) CAR Ts was used to confer lymphodepletion resistance and reduced graft-versus-host disease (GvHD) potential. The safety profile of allogeneic BCMA CAR Ts was further enhanced by incorporating a CD20 mimotope-based intra-CAR off switch enabling effective CAR T elimination in the presence of rituximab. Allogeneic BCMA CAR Ts induced sustained antitumor responses in mice supplemented with human cytokines, and, most importantly, maintained their phenotype and potency after scale-up manufacturing. This novel off-the-shelf allogeneic BCMA CAR T product is a promising candidate for clinical evaluation.
Asunto(s)
Antígeno de Maduración de Linfocitos B/inmunología , Trasplante de Células/métodos , Inmunoterapia Adoptiva/métodos , Mieloma Múltiple/terapia , Receptores Quiméricos de Antígenos/inmunología , Linfocitos T/inmunología , Linfocitos T/trasplante , Animales , Antineoplásicos Inmunológicos/uso terapéutico , Antígeno de Maduración de Linfocitos B/genética , Donantes de Sangre , Línea Celular Tumoral , Trasplante de Células/efectos adversos , Citotoxicidad Inmunológica/genética , Edición Génica , Vectores Genéticos , Enfermedad Injerto contra Huésped/terapia , Humanos , Inmunoterapia Adoptiva/efectos adversos , Ratones , Ratones Endogámicos NOD , Ratones SCID , Mieloma Múltiple/patología , Supervivencia sin Progresión , Receptores Quiméricos de Antígenos/genética , Receptores Quiméricos de Antígenos/metabolismo , Rituximab/uso terapéutico , Linfocitos T/metabolismo , Nucleasas de los Efectores Tipo Activadores de la Transcripción/genética , Transducción Genética , Trasplante Homólogo/métodosRESUMEN
Anemia of inflammation (AI) is commonly observed in chronic inflammatory states and may hinder patient recovery and survival. Induction of hepcidin, mediated by interleukin 6, leads to iron-restricted erythropoiesis and anemia. Several translational studies have been directed at neutralizing hepcidin overexpression as a therapeutic strategy against AI. However, additional hepcidin-independent mechanisms contribute to AI, which are likely mediated by a direct effect of inflammatory cytokines on erythropoiesis. In this study, we used wild-type, hepcidin knockout (Hamp-KO) and interleukin 6 knockout (IL-6-KO) mice as models of AI. AI was induced with heat-killed Brucella abortus (BA). The distinct roles of iron metabolism and inflammation triggered by interleukin 6 and hepcidin were investigated. BA-treated wild-type mice showed increased expression of hepcidin and inflammatory cytokines, as well as transitory suppression of erythropoiesis and shortened red blood cell lifespan, all of which contributed to the severe anemia of these mice. In contrast, BA-treated Hamp-KO or IL-6-KO mice showed milder anemia and faster recovery compared with normal mice. Moreover, they exhibited different patterns in the development and resolution of anemia, supporting the notion that interleukin 6 and hepcidin play distinct roles in modulating erythropoiesis in AI.
Asunto(s)
Anemia/inmunología , Brucella abortus , Brucelosis/inmunología , Hepcidinas/inmunología , Interleucina-6/inmunología , Anemia/genética , Anemia/microbiología , Animales , Médula Ósea/inmunología , Brucelosis/complicaciones , Modelos Animales de Enfermedad , Eritropoyesis/inmunología , Femenino , Hepcidinas/genética , Calor , Interleucina-6/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Recuperación de la Función/inmunologíaRESUMEN
Iron maldistribution has been implicated in the etiology of many diseases including the anemia of inflammation (AI), atherosclerosis, diabetes, and neurodegenerative disorders. Iron metabolism is controlled by hepcidin, a 25-amino-acid peptide. Hepcidin is induced by inflammation and causes iron to be sequestered within cells of the reticuloendothelial system, suppressing erythropoiesis and blunting the activity of erythropoiesis stimulating agents (ESAs). For this reason, neutralization of hepcidin has been proposed as a therapeutic treatment of AI. The aim of the current work was to generate fully human anti-hepcidin antibodies (Abs) as a potential human therapeutic for the treatment of AI and other iron maldistribution disorders. An enzyme-linked immunosorbent assay was established using these Abs to identify patients likely to benefit from either ESAs or anti-hepcidin agents. Using human hepcidin knock-in mice, the mechanism of action of the Abs was shown to be due to an increase in available serum iron leading to enhanced red cell hemoglobinization. One of the Abs, 12B9m, was validated in a mouse model of AI and demonstrated to modulate serum iron in cynomolgus monkeys. The 12B9m Ab was deemed to be an appropriate candidate for use as a potential therapeutic to treat AI in patients with kidney disease or cancer.
Asunto(s)
Anemia Ferropénica/tratamiento farmacológico , Anticuerpos Neutralizantes/farmacología , Eritrocitos/efectos de los fármacos , Hemoglobinas/biosíntesis , Hierro/sangre , Anemia Ferropénica/sangre , Anemia Ferropénica/patología , Animales , Anticuerpos Neutralizantes/biosíntesis , Modelos Animales de Enfermedad , Evaluación Preclínica de Medicamentos , Ensayo de Inmunoadsorción Enzimática , Eritrocitos/metabolismo , Eritrocitos/patología , Eritropoyesis/efectos de los fármacos , Femenino , Hematínicos/farmacología , Humanos , Inflamación/prevención & control , Macaca fascicularis , Masculino , RatonesRESUMEN
The magnitude of CAR T cell expansion has been associated with clinical efficacy. Although cytokines can augment CAR T cell proliferation, systemically administered cytokines can result in toxicities. To gain the benefits of cytokine signaling while mitigating toxicities, we designed constitutively active synthetic cytokine receptor chimeras (constitutive Turbodomains) that signal in a CAR T cell-specific manner. The modular design of Turbodomains enables diverse cytokine signaling outputs from a single homodimeric receptor chimera and allows multiplexing of different cytokine signals. Turbodomains containing an IL-2/15Rß-derived signaling domain closely mimicked IL-15 signaling and enhanced CAR T cell potency. Allogeneic TurboCAR T cells targeting BCMA showed no evidence of aberrant proliferation yet displayed enhanced expansion and antitumor activity, prolonging survival and preventing extramedullary relapses in mouse models. These results illustrate the potential of constitutive Turbodomains to achieve selective potentiation of CAR T cells and demonstrate the safety and efficacy of allogeneic BCMA TurboCAR T cells, supporting clinical evaluation in multiple myeloma.
Asunto(s)
Trasplante de Células Madre Hematopoyéticas , Receptores Quiméricos de Antígenos , Animales , Ratones , Receptores Quiméricos de Antígenos/genética , Inmunoterapia Adoptiva/métodos , Antígeno de Maduración de Linfocitos B , Recurrencia Local de Neoplasia , Linfocitos T , CitocinasRESUMEN
PURPOSE: Small cell lung cancer (SCLC) is an aggressive disease with limited treatment options. Delta-like ligand 3 (DLL3) is highly expressed on SCLC and several other types of neuroendocrine cancers, with limited normal tissue RNA expression in brain, pituitary, and testis, making it a promising CAR T-cell target for SCLC and other solid tumor indications. EXPERIMENTAL DESIGN: A large panel of anti-DLL3 scFv-based CARs were characterized for both in vitro and in vivo activity. To understand the potential for pituitary and brain toxicity, subcutaneous or intracranial tumors expressing DLL3 were implanted in mice and treated with mouse cross-reactive DLL3 CAR T cells. RESULTS: A subset of CARs demonstrated high sensitivity for targets with low DLL3 density and long-term killing potential in vitro. Infusion of DLL3 CAR T cells led to robust antitumor efficacy, including complete responses, in subcutaneous and systemic SCLC in vivo models. CAR T-cell infiltration into intermediate and posterior pituitary was detected, but no tissue damage in brain or pituitary was observed, and the hormone-secretion function of the pituitary was not ablated. CONCLUSIONS: In summary, the preclinical efficacy and safety data presented here support further evaluation of DLL3 CAR T cells as potential clinical candidates for the treatment of SCLC.
Asunto(s)
Trasplante de Células Madre Hematopoyéticas , Neoplasias Pulmonares , Carcinoma Pulmonar de Células Pequeñas , Animales , Masculino , Ratones , Ligandos , Neoplasias Pulmonares/tratamiento farmacológico , Carcinoma Pulmonar de Células Pequeñas/genética , Linfocitos T/metabolismoAsunto(s)
Anemia , Eritropoyetina/análogos & derivados , Hematínicos/uso terapéutico , Hepcidinas/sangre , Anemia/sangre , Anemia/inducido químicamente , Anemia/tratamiento farmacológico , Antineoplásicos/efectos adversos , Antineoplásicos/uso terapéutico , Darbepoetina alfa , Eritropoyetina/uso terapéutico , Femenino , Humanos , Masculino , Estudios Multicéntricos como Asunto , Neoplasias/sangre , Neoplasias/tratamiento farmacológico , Ensayos Clínicos Controlados Aleatorios como AsuntoRESUMEN
Iron maldistribution has been implicated in multiple diseases, including the anemia of inflammation (AI), atherosclerosis, diabetes, and neurodegenerative disorders. Iron metabolism is controlled by hepcidin, a 25-amino acid peptide. Hepcidin is induced by inflammation, causes iron to be sequestered, and thus, potentially contributes to AI. Human hepcidin (hHepc) overexpression in mice caused an iron-deficient phenotype, including stunted growth, hair loss, and iron-deficient erythropoiesis. It also caused resistance to supraphysiologic levels of erythropoiesis-stimulating agent, supporting the hypothesis that hepcidin may influence response to treatment in AI. To explore the role of hepcidin in inflammatory anemia, a mouse AI model was developed with heat-killed Brucella abortus treatment. Suppression of hepcidin mRNA was a successful anemia treatment in this model. High-affinity antibodies specific for hHepc were generated, and hHepc knock-in mice were produced to enable antibody testing. Antibody treatment neutralized hHepc in vitro and in vivo and facilitated anemia treatment in hHepc knock-in mice with AI. These data indicate that antihepcidin antibodies may be an effective treatment for patients with inflammatory anemia. The ability to manipulate iron metabolism in vivo may also allow investigation of the role of iron in a number of other pathologic conditions.
Asunto(s)
Anemia Ferropénica/tratamiento farmacológico , Anticuerpos Neutralizantes/farmacología , Péptidos Catiónicos Antimicrobianos/antagonistas & inhibidores , Hierro/metabolismo , Anemia Ferropénica/genética , Anemia Ferropénica/inmunología , Anemia Ferropénica/metabolismo , Animales , Anticuerpos Neutralizantes/inmunología , Péptidos Catiónicos Antimicrobianos/biosíntesis , Péptidos Catiónicos Antimicrobianos/genética , Péptidos Catiónicos Antimicrobianos/inmunología , Brucella abortus , Modelos Animales de Enfermedad , Eritropoyesis/efectos de los fármacos , Eritropoyesis/genética , Hepcidinas , Humanos , Inflamación/complicaciones , Inflamación/tratamiento farmacológico , Inflamación/genética , Inflamación/inmunología , Inflamación/metabolismo , Ratones , Ratones TransgénicosRESUMEN
Although cytokine support can enhance CAR T-cell function, coadministering cytokines or engineering CAR T cells to secrete cytokines can result in toxicities. To mitigate these safety risks, we engineered iTurboCAR T cells that coexpress a novel inducible Turbo (iTurbo) cytokine signaling domain. iTurbo domains consist of modular components that are customizable to a variety of activating inputs, as well as cytokine signaling outputs multiplexable for combinatorial signaling outcomes. Unlike most canonical cytokine receptors that are heterodimeric, iTurbo domains leverage a compact, homodimeric design that minimizes viral vector cargo. Using an iTurbo domain activated by the clinically validated dimerizer, AP1903, homodimeric iTurbo domains instigated signaling that mimicked the endogenous heterodimeric cytokine receptor. Different iTurbo domains programmed iTurboCAR T cells toward divergent phenotypes and resulted in improved antitumor efficacy. iTurbo domains, therefore, offer the flexibility for user-programmable signaling outputs, permitting control over cellular phenotype and function while minimizing viral cargo footprint.
Asunto(s)
Inmunoterapia Adoptiva , Receptores de Antígenos de Linfocitos T , Citocinas , Inmunoterapia Adoptiva/métodos , Receptores de Antígenos de Linfocitos T/genética , Transducción de Señal , Linfocitos TRESUMEN
Multiple myeloma remains an incurable plasma cell malignancy despite the rapidly evolving treatment landscape. Chimeric antigen receptor T cells targeted against BCMA have recently shown great promise in relapsed refractory multiple myeloma; however, all patients ultimately still progress from their disease. Lack of CAR T-cell persistence, impaired T-cell fitness in autologous CAR T-cell products and the presence of an immunosuppressive bone marrow (BM) microenvironment are contributory factors to treatment failure. We generated anti-BCMA CAR T cells from healthy donors (HD) and patients with multiple myeloma at different stages of disease to compare their T-cell profile, fitness, and cytotoxic activity in preclinical studies. We also used an ex vivo assay with multiple myeloma BM biopsies from distinct genomic subgroups to test the efficacy of HD-derived CAR T cells in a clinically relevant model. HD volunteers showed increased T-cell counts, higher CD4/CD8 ratio, and expanded naïve T-cell population compared with patients with multiple myeloma. After anti-BCMA CAR T-cell production, patients with relapsed multiple myeloma had lower frequencies of CAR+ T cells, decreased central memory phenotype, and increased checkpoint inhibitory markers compared with HD-derived products, which compromised their expansion and cytotoxicity against multiple myeloma cells in vitro. Importantly, HD-derived CAR T cells efficiently killed primary multiple myeloma cells within the BM microenvironment of different multiple myeloma genomic subgroups and their cytotoxic activity could be boosted with gamma secretase inhibitors. In conclusion, allogeneic anti-BCMA CAR T cells are a potential therapeutic strategy for patients with relapsed multiple myeloma and should be further developed in the clinic. Significance: Multiple myeloma is an incurable cancer of the plasma cells. A new therapy with anti-BCMA CAR T cells - the patient's own T cells genetically engineered to find and kill myeloma cancer cells - has shown encouraging results. Unfortunately, patients still relapse. In this study, we propose to use T cells from HD volunteers, which have a stronger T-cell fitness, higher cancer killing capacity, and are ready to be administered when needed.
Asunto(s)
Trasplante de Células Madre Hematopoyéticas , Mieloma Múltiple , Receptores Quiméricos de Antígenos , Humanos , Mieloma Múltiple/terapia , Receptores Quiméricos de Antígenos/genética , Inhibidores y Moduladores de Gamma Secretasa , Recurrencia Local de Neoplasia , Linfocitos T , Microambiente TumoralRESUMEN
CD70 is highly expressed in renal cell carcinoma (RCC), with limited expression in normal tissue, making it an attractive CAR T target for an immunogenic solid tumor indication. Here we generated and characterized a panel of anti-CD70 single-chain fragment variable (scFv)-based CAR T cells. Despite the expression of CD70 on T cells, production of CAR T cells from a subset of scFvs with potent in vitro activity was achieved. Expression of CD70 CARs masked CD70 detection in cis and provided protection from CD70 CAR T cell-mediated fratricide. Two distinct classes of CAR T cells were identified with differing memory phenotype, activation status, and cytotoxic activity. Epitope mapping revealed that the two classes of CARs bind unique regions of CD70. CD70 CAR T cells displayed robust antitumor activity against RCC cell lines and patient-derived xenograft mouse models. Tissue cross-reactivity studies identified membrane staining in lymphocytes, thus matching the known expression pattern of CD70. In a cynomolgus monkey CD3-CD70 bispecific toxicity study, expected findings related to T-cell activation and elimination of CD70-expressing cells were observed, including cytokine release and loss of cellularity in lymphoid tissues. Finally, highly functional CD70 allogeneic CAR T cells were produced at large scale through elimination of the T-cell receptor by TALEN-based gene editing. Taken together, these efficacy and safety data support the evaluation of CD70 CAR T cells for the treatment of RCC and has led to the advancement of an allogeneic CD70 CAR T-cell candidate into phase I clinical trials. SIGNIFICANCE: These findings demonstrate the efficacy and safety of fratricide-resistant, allogeneic anti-CD70 CAR T cells targeting renal cell carcinoma and the impact of CAR epitope on functional activity. See related commentary by Adotévi and Galaine, p. 2517.
Asunto(s)
Carcinoma de Células Renales , Trasplante de Células Madre Hematopoyéticas , Neoplasias Renales , Animales , Ligando CD27 , Carcinoma de Células Renales/patología , Línea Celular Tumoral , Humanos , Inmunoterapia Adoptiva , Neoplasias Renales/patología , Macaca fascicularis , Ratones , Linfocitos T/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Antibody-based therapeutics have experienced a rapid growth in recent years and are now utilized in various modalities spanning from conventional antibodies, antibody-drug conjugates, bispecific antibodies to chimeric antigen receptor (CAR) T cells. Many next generation antibody therapeutics achieve enhanced potency but often increase the risk of adverse events. Antibody scaffolds capable of exhibiting inducible affinities could reduce the risk of adverse events by enabling a transient suspension of antibody activity. To demonstrate this, we develop conditionally activated, single-module CARs, in which tumor antigen recognition is directly modulated by an FDA-approved small molecule drug. The resulting CAR T cells demonstrate specific cytotoxicity of tumor cells comparable to that of traditional CARs, but the cytotoxicity is reversibly attenuated by the addition of the small molecule. The exogenous control of conditional CAR T cell activity allows continual modulation of therapeutic activity to improve the safety profile of CAR T cells across all disease indications.
Asunto(s)
Antígenos de Neoplasias/inmunología , Inmunoterapia Adoptiva/métodos , Metotrexato/administración & dosificación , Neoplasias/terapia , Receptores Quiméricos de Antígenos/metabolismo , Linfocitos T/efectos de los fármacos , Animales , Línea Celular Tumoral , Terapia Combinada/métodos , Femenino , Células HEK293 , Humanos , Inmunoterapia Adoptiva/efectos adversos , Ratones , Neoplasias/inmunología , Cultivo Primario de Células , Receptores Quiméricos de Antígenos/inmunología , Anticuerpos de Dominio Único/inmunología , Anticuerpos de Dominio Único/metabolismo , Especificidad del Receptor de Antígeno de Linfocitos T/efectos de los fármacos , Especificidad del Receptor de Antígeno de Linfocitos T/inmunología , Linfocitos T/inmunología , Linfocitos T/metabolismo , Linfocitos T/trasplante , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Anemia in cancer patients can result from a complex interaction of numerous factors including iron deficiency, inflammation, toxicity related to therapy and effect of cancer on the marrow. Determining effective anemia treatment can therefore be complex, requiring a combination of diagnostic tests. Research on iron metabolism has highlighted the importance of hepcidin and its potential role in development of anemia of inflammation (AI). Hepcidin is a peptide that controls iron flow, is induced by inflammation and is speculated to cause the sequestration of iron in patients with inflammation. In the present study, serum hepcidin concentration determined by LC-MS/MS was shown to correlate with inflammatory markers in patients with anemia of cancer (AoC). In the absence of a widely-available serum hepcidin detection assay, detection of prohepcidin using a commercial assay has been used for several years as a surrogate for measuring serum hepcidin concentration. Analysis of prohepcidin concentration did not reveal any correlation with hepcidin or with inflammatory markers in patient samples and our data suggest that prohepcidin may not be stable in serum. Algorithms to sub-classify AoC patients showed that hepcidin was strongly associated with the population subset with inflammation and without iron deficiency. Serum hepcidin concentrations may therefore be a good predictor of AI, useful in diagnosis of anemia etiology and in treatment determination.
Asunto(s)
Algoritmos , Anemia/sangre , Péptidos Catiónicos Antimicrobianos/sangre , Inflamación/sangre , Hierro/sangre , Precursores de Proteínas/sangre , Anemia/complicaciones , Anemia/diagnóstico , Biomarcadores/sangre , Femenino , Hepcidinas , Humanos , Inflamación/complicaciones , Inflamación/diagnóstico , Masculino , Neoplasias/sangre , Neoplasias/diagnósticoRESUMEN
The restricted expression pattern of B-cell maturation antigen (BCMA) makes it an ideal tumor-associated antigen (TAA) for the treatment of myeloma. BCMA has been targeted by both CD3 bispecific antibody and antibody-drug conjugate (ADC) modalities, but a true comparison of modalities has yet to be performed. Here we utilized a single BCMA antibody to develop and characterize both a CD3 bispecific and 2 ADC formats (cleavable and noncleavable) and compared activity both in vitro and in vivo with the aim of generating an optimal therapeutic. Antibody affinity, but not epitope was influential in drug activity and hence a high-affinity BCMA antibody was selected. Both the bispecific and ADCs were potent in vitro and in vivo, causing dose-dependent cell killing of myeloma cell lines and tumor regression in orthotopic myeloma xenograft models. Primary patient cells were effectively lysed by both CD3 bispecific and ADCs, with the bispecific demonstrating improved potency, maximal cell killing, and consistency across patients. Safety was evaluated in cynomolgus monkey toxicity studies and both modalities were active based on on-target elimination of B lineage cells. Distinct nonclinical toxicity profiles were seen for the bispecific and ADC modalities. When taken together, results from this comparison of BCMA CD3 bispecific and ADC modalities suggest better efficacy and an improved toxicity profile might be achieved with the bispecific modality. This led to the advancement of a bispecific candidate into phase I clinical trials.
Asunto(s)
Anticuerpos Biespecíficos/administración & dosificación , Antígeno de Maduración de Linfocitos B/metabolismo , Complejo CD3/inmunología , Inmunoconjugados/administración & dosificación , Mieloma Múltiple/tratamiento farmacológico , Animales , Anticuerpos Biespecíficos/efectos adversos , Anticuerpos Biespecíficos/farmacología , Afinidad de Anticuerpos , Antígeno de Maduración de Linfocitos B/antagonistas & inhibidores , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Inmunoconjugados/efectos adversos , Inmunoconjugados/farmacología , Ratones , Mieloma Múltiple/metabolismo , Transducción de Señal/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
CAR T-cell therapies hold great promise for treating a range of malignancies but are however challenged by the complexity of their production and by the adverse events related to their activity. Here we report the development of the CubiCAR, a tri-functional CAR architecture that enables CAR T-cell detection, purification and on-demand depletion by the FDA-approved antibody Rituximab. This novel architecture has the potential to streamline the manufacturing of CAR T-cells, allow their tracking and improve their overall safety.
Asunto(s)
Inmunoterapia Adoptiva , Neoplasias Experimentales/inmunología , Neoplasias Experimentales/cirugía , Receptores Quiméricos de Antígenos/inmunología , Rituximab/farmacología , Animales , Línea Celular Tumoral , Humanos , Ratones , Ratones Endogámicos BALB C , Neoplasias Experimentales/patologíaRESUMEN
T cell activity has been stimulated to enhance anti-tumor activity for several decades. Tumor infiltrating lymphocytes have been expanded and reinfused to treat melanoma and other solid tumor cancers, and adaptive immune modulators have been used to increase endogenous T cell activity against cancer. Both of these strategies rely on the pre-existence of adequate recognition of cancer cells by the T cells which are being activated. Redirected T cell therapies represent a paradigm shift in treatment. They do not rely on endogenous T cell recognition but instead focus the T cells onto a defined tumor antigen. This review summarizes both molecular and cellular T cell redirecting therapies, compares their success in the clinic and highlights both their limitations and some potential future solutions.
Asunto(s)
Neoplasias/terapia , Linfocitos T/inmunología , Traslado Adoptivo , HumanosRESUMEN
Hepcidin (H25) is a hormone peptide synthesized by the liver that binds to ferroportin and blocks iron export. In this study, H25 was inhibited by administration of single and multiple doses of an anti-H25 monoclonal antibody Ab 12B9m in cynomolgus monkeys. The objective of this analysis was to develop a pharmacodynamic model describing the role of H25 in regulating iron homeostasis and the impact of hepcidin inhibition by Ab 12B9m. Total serum H25 and Ab 12B9m were determined in each animal. Corresponding measurements of serum iron and hemoglobin (Hb) were obtained. The PD model consisted of iron pools in serum (FeS), reticuloendothelial macrophages (FeM), hemoglobin (FeHb), and liver (FeL). The iron was assumed to be transported between the FeS, FeHb, and FeM unidirectionally at rates k S, k Hb, and k M. H25 serum concentrations were described by the previously developed PK model with the parameters fixed at their estimates. The serum iron and Hb data were fitted simultaneously. The corresponding estimates of the rate constants were k S/Fe0 = 0.113 h(-1), k M = 0.00191 h(-1), and k Hb = 0.00817 h(-1). The model-based IC50 value for the H25 inhibitory effect on ferroportin activity was 0.398 nM. The PD model predicted a negligible effect of Ab 12B9m on Hb levels for the tested doses. The presented PD model adequately described the serum iron time courses following single and multiple doses of Ab 12B9m. Ab 12B9m-induced inhibition of H25 resulted in a temporal increase in serum and liver iron and a decrease in the iron stored in reticuloendothelial macrophages.
Asunto(s)
Hepcidinas/sangre , Homeostasis/fisiología , Hierro/sangre , Modelos Biológicos , Animales , Anticuerpos Monoclonales/farmacología , Hepcidinas/antagonistas & inhibidores , Homeostasis/efectos de los fármacos , Macaca fascicularisRESUMEN
Chemotherapy-induced thrombocytopenia can lead to chemotherapy treatment delays or dose reductions. The ability of romiplostim, a thrombopoietin (TPO) mimetic, to promote platelet recovery in a mouse model of multicycle chemotherapy/radiation therapy (CRT)-induced thrombocytopenia was examined. In humans, an inverse relationship between platelet counts and endogenous TPO (eTPO) concentration exists. In a CRT mouse model, eTPO was not elevated during the first 5 days after CRT treatment (the "eTPO gap"), then increased to a peak 10 days after each CRT treatment in an inverse relationship to platelet counts seen in humans. To bridge the eTPO gap, mice were treated with 10-1,000 µg/kg of romiplostim on day 0, 1, or 2 after CRT. In some mice, the romiplostim dose was approximately divided over 3 days. Platelet recovery occurred faster with romiplostim in most conditions tested. Romiplostim doses of ≥100 µg/kg given on day 0 significantly lessened the platelet nadir. Fractionating the dose over 3 days did not appear to confer a large advantage. These data may provide a rationale for clinical studies of romiplostim in chemotherapy-induced thrombocytopenia.