RESUMO
B-cell maturation antigen (BCMA), a member of the tumor necrosis factor family of receptors, is predominantly expressed on the surface of terminally differentiated B cells. BCMA is highly expressed on plasmablasts and plasma cells from multiple myeloma (MM) patient samples. We developed a BCMAxCD3 bispecific antibody (teclistamab [JNJ-64007957]) to recruit and activate T cells to kill BCMA-expressing MM cells. Teclistamab induced cytotoxicity of BCMA+ MM cell lines in vitro (H929 cells, 50% effective concentration [EC50] = 0.15 nM; MM.1R cells, EC50 = 0.06 nM; RPMI 8226 cells, EC50 = 0.45 nM) with concomitant T-cell activation (H929 cells, EC50 = 0.21 nM; MM.1R cells, EC50 = 0.1 nM; RPMI 8226 cells, EC50 = 0.28 nM) and cytokine release. This activity was further increased in the presence of a γ-secretase inhibitor (LY-411575). Teclistamab also depleted BCMA+ cells in bone marrow samples from MM patients in an ex vivo assay with an average EC50 value of 1.7 nM. Under more physiological conditions using healthy human whole blood, teclistamab mediated dose-dependent lysis of H929 cells and activation of T cells. Antitumor activity of teclistamab was also observed in 2 BCMA+ MM murine xenograft models inoculated with human T cells (tumor inhibition with H929 model and tumor regression with the RPMI 8226 model) compared with vehicle and antibody controls. The specific and potent activity of teclistamab against BCMA-expressing cells from MM cell lines, patient samples, and MM xenograft models warrant further evaluation of this bispecific antibody for the treatment of MM. Phase 1 clinical trials (monotherapy, #NCT03145181; combination therapy, #NCT04108195) are ongoing for patients with relapsed/refractory MM.
Assuntos
Anticorpos Biespecíficos , Mieloma Múltiplo , Animais , Anticorpos Biespecíficos/farmacologia , Antígeno de Maturação de Linfócitos B , Humanos , Ativação Linfocitária , Camundongos , Mieloma Múltiplo/tratamento farmacológico , Linfócitos TRESUMO
Redirecting T cells to specifically kill malignant cells has been validated as an effective anti-cancer strategy in the clinic with the approval of blinatumomab for acute lymphoblastic leukemia. However, the immunosuppressive nature of the tumor microenvironment potentially poses a significant hurdle to T cell therapies. In hematological malignancies, the bone marrow (BM) niche is protective to leukemic stem cells and has minimized the efficacy of several anti-cancer drugs. In this study, we investigated the impact of the BM microenvironment on T cell redirection. Using bispecific antibodies targeting specific tumor antigens (CD123 and BCMA) and CD3, we observed that co-culture of acute myeloid leukemia or multiple myeloma cells with BM stromal cells protected tumor cells from bispecific antibody-T cell-mediated lysis in vitro and in vivo. Impaired CD3 redirection cytotoxicity was correlated with reduced T cell effector responses and cell-cell contact with stromal cells was implicated in reducing T cell activation and conferring protection of cancer cells. Finally, blocking the VLA4 adhesion pathway in combination with CD3 redirection reduced the stromal-mediated inhibition of cytotoxicity and T cell activation. Our results lend support to inhibiting VLA4 interactions along with administering CD3 redirection therapeutics as a novel combinatorial regimen for robust anti-cancer responses.
Assuntos
Antineoplásicos Imunológicos/farmacologia , Medula Óssea/efeitos dos fármacos , Complexo CD3/imunologia , Integrina alfa4beta1/antagonistas & inibidores , Leucemia Mieloide Aguda/tratamento farmacológico , Mieloma Múltiplo/tratamento farmacológico , Animais , Anticorpos Biespecíficos/farmacologia , Anticorpos Biespecíficos/uso terapêutico , Antineoplásicos Imunológicos/uso terapêutico , Antígeno de Maturação de Linfócitos B/antagonistas & inibidores , Antígeno de Maturação de Linfócitos B/imunologia , Medula Óssea/imunologia , Medula Óssea/patologia , Complexo CD3/antagonistas & inibidores , Linhagem Celular Tumoral , Feminino , Humanos , Integrina alfa4beta1/imunologia , Subunidade alfa de Receptor de Interleucina-3/antagonistas & inibidores , Subunidade alfa de Receptor de Interleucina-3/imunologia , Leucemia Mieloide Aguda/imunologia , Leucemia Mieloide Aguda/patologia , Camundongos , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/patologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/patologia , Microambiente Tumoral/efeitos dos fármacosRESUMO
CD33 is expressed in 90% of patients with acute myeloid leukemia (AML), and its extracellular portion consists of a V domain and a C2 domain. A recent study showed that a single nucleotide polymorphism (SNP), rs12459419 (C > T), results in the reduced expression of V domain-containing CD33 and limited efficacy of V domain-binding anti-CD33 antibodies. We developed JNJ-67571244, a novel human bispecific antibody capable of binding to the C2 domain of CD33 and to CD3, to induce T-cell recruitment and CD33+ tumor cell cytotoxicity independently of their SNP genotype status. JNJ-67571244 specifically binds to CD33-expressing target cells and induces cytotoxicity of CD33+ AML cell lines in vitro along with T-cell activation and cytokine release. JNJ-67571244 also exhibited statistically significant antitumor activity in vivo in established disseminated and subcutaneous mouse models of human AML. Furthermore, this antibody depletes CD33+ blasts in AML patient blood samples with concurrent T-cell activation. JNJ-67571244 also cross-reacts with cynomolgus monkey CD33 and CD3, and dosing of JNJ-67571244 in cynomolgus monkeys resulted in T-cell activation, transient cytokine release, and sustained reduction in CD33+ leukocyte populations. JNJ-67571244 was well tolerated in cynomolgus monkeys up to 30 mg/kg. Lastly, JNJ-67571244 mediated efficient cytotoxicity of cell lines and primary samples regardless of their SNP genotype status, suggesting a potential therapeutic benefit over other V-binding antibodies. JNJ-67571244 is currently in phase 1 clinical trials in patients with relapsed/refractory AML and high-risk myelodysplastic syndrome.
Assuntos
Leucemia Mieloide Aguda , Linfócitos T , Animais , Domínios C2 , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Macaca fascicularis , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico/genética , Linfócitos T/metabolismoRESUMO
T-cell-mediated approaches have shown promise in myeloma treatment. However, there are currently a limited number of specific myeloma antigens that can be targeted, and multiple myeloma (MM) remains an incurable disease. G-protein-coupled receptor class 5 member D (GPRC5D) is expressed in MM and smoldering MM patient plasma cells. Here, we demonstrate that GPRC5D protein is present on the surface of MM cells and describe JNJ-64407564, a GPRC5DxCD3 bispecific antibody that recruits CD3+ T cells to GPRC5D+ MM cells and induces killing of GPRC5D+ cells. In vitro, JNJ-64407564 induced specific cytotoxicity of GPRC5D+ cells with concomitant T-cell activation and also killed plasma cells in MM patient samples ex vivo. JNJ-64407564 can recruit T cells and induce tumor regression in GPRC5D+ MM murine models, which coincide with T-cell infiltration at the tumor site. This antibody is also able to induce cytotoxicity of patient primary MM cells from bone marrow, which is the natural site of this disease. GPRC5D is a promising surface antigen for MM immunotherapy, and JNJ-64407564 is currently being evaluated in a phase 1 clinical trial in patients with relapsed or refractory MM (NCT03399799).
Assuntos
Anticorpos Biespecíficos/uso terapêutico , Antineoplásicos Imunológicos/uso terapêutico , Mieloma Múltiplo/terapia , Receptores Acoplados a Proteínas G/imunologia , Linfócitos T/efeitos dos fármacos , Animais , Anticorpos Biespecíficos/imunologia , Antineoplásicos Imunológicos/imunologia , Linhagem Celular Tumoral , Citotoxicidade Imunológica/efeitos dos fármacos , Feminino , Humanos , Imunoterapia , Camundongos Endogâmicos BALB C , Mieloma Múltiplo/imunologia , Linfócitos T/imunologiaRESUMO
PURPOSE: Multiple myeloma (MM) patients with disease refractory to all available drugs have a poor outcome, indicating the need for new agents with novel mechanisms of action. EXPERIMENTAL DESIGN: We evaluated the anti-MM activity of the fully human BCMA×CD3 bispecific antibody JNJ-7957 in cell lines and bone marrow (BM) samples. The impact of several tumor- and host-related factors on sensitivity to JNJ-7957 therapy was also evaluated. RESULTS: We show that JNJ-7957 has potent activity against 4 MM cell lines, against tumor cells in 48 of 49 BM samples obtained from MM patients, and in 5 of 6 BM samples obtained from primary plasma cell leukemia patients. JNJ-7957 activity was significantly enhanced in patients with prior daratumumab treatment, which was partially due to enhanced killing capacity of daratumumab-exposed effector cells. BCMA expression did not affect activity of JNJ-7957. High T-cell frequencies and high effector:target ratios were associated with improved JNJ-7957-mediated lysis of MM cells. The PD-1/PD-L1 axis had a modest negative impact on JNJ-7957 activity against tumor cells from daratumumab-naïve MM patients. Soluble BCMA impaired the ability of JNJ-7957 to kill MM cells, although higher concentrations were able to overcome this negative effect. CONCLUSIONS: JNJ-7957 effectively kills MM cells ex vivo, including those from heavily pretreated MM patients, whereby several components of the immunosuppressive BM microenvironment had only modest effects on its killing capacity. Our findings support the ongoing trial with JNJ-7957 as single agent and provide the preclinical rationale for evaluating JNJ-7957 in combination with daratumumab in MM.
Assuntos
Anticorpos Biespecíficos/farmacologia , Anticorpos Monoclonais/farmacologia , Antígeno de Maturação de Linfócitos B/imunologia , Complexo CD3/imunologia , Mieloma Múltiplo/tratamento farmacológico , Anticorpos Biespecíficos/imunologia , Citotoxicidade Celular Dependente de Anticorpos , Antineoplásicos/farmacologia , Medula Óssea/patologia , Avaliação Pré-Clínica de Medicamentos , Sinergismo Farmacológico , Quimioterapia Combinada , Humanos , Imunoterapia/métodos , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/metabolismo , Mieloma Múltiplo/patologia , Células Tumorais CultivadasRESUMO
Development of targeted cancer therapy requires a thorough understanding of mechanisms of tumorigenesis as well as mechanisms of action of therapeutics. This is challenging because by the time patients are diagnosed with cancer, early events of tumorigenesis have already taken place. Similarly, development of cancer immunotherapies is hampered by a lack of appropriate small animal models with autologous human tumor and immune system. In this article, we report the development of a mouse model of human acute myeloid leukemia (AML) with autologous immune system for studying early events of human leukemogenesis and testing the efficacy of immunotherapeutics. To develop such a model, human hematopoietic stem/progenitor cells (HSPC) are transduced with lentiviruses expressing a mutated form of nucleophosmin (NPM1), referred to as NPM1c. Following engraftment into immunodeficient mice, transduced HSPCs give rise to human myeloid leukemia, whereas untransduced HSPCs give rise to human immune cells in the same mice. The de novo AML, with CD123+ leukemic stem or initiating cells (LSC), resembles NPM1c+ AML from patients. Transcriptional analysis of LSC and leukemic cells confirms similarity of the de novo leukemia generated in mice with patient leukemia and suggests Myc as a co-operating factor in NPM1c-driven leukemogenesis. We show that a bispecific conjugate that binds both CD3 and CD123 eliminates CD123+ LSCs in a T cell-dependent manner both in vivo and in vitro. These results demonstrate the utility of the NPM1c+ AML model with an autologous immune system for studying early events of human leukemogenesis and for evaluating efficacy and mechanism of immunotherapeutics.
Assuntos
Carcinogênese , Leucemia Mieloide , Proteínas Nucleares , Ensaios Antitumorais Modelo de Xenoenxerto , Animais , Células-Tronco Hematopoéticas , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , NucleofosminaRESUMO
The protein methyltransferase Set7/9 was recently shown to regulate p53 activity in cancer cells. However, the impact of Set7/9 on p53 function in vivo is unclear. To explore these issues, we created a null allele of Set7/9 in mice. Cells from Set7/9 mutant mice fail to methylate p53 K369, are unable to induce p53 downstream targets upon DNA damage, and are predisposed to oncogenic transformation. Importantly, we find that methylation of p53 by Set7/9 is required for the binding of the acetyltransferase Tip60 to p53 and for the subsequent acetylation of p53. We provide the first genetic evidence demonstrating that lysine methylation of p53 by Set7/9 is important for p53 activation in vivo and suggest a mechanistic link between methylation and acetylation of p53 through Tip60.
Assuntos
Histona-Lisina N-Metiltransferase/metabolismo , Proteínas Metiltransferases/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Acetilação , Acetiltransferases/metabolismo , Animais , Ciclo Celular/fisiologia , Linhagem Celular , Linhagem Celular Transformada , Transformação Celular Viral , Dano ao DNA/genética , Embrião de Mamíferos/citologia , Fibroblastos/metabolismo , Dosagem de Genes , Glutationa Transferase/metabolismo , Células HCT116 , Histona Metiltransferases , Histona-Lisina N-Metiltransferase/genética , Humanos , Rim/citologia , Lisina/metabolismo , Metilação , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mutação , Proteínas Metiltransferases/genética , Proteína Supressora de Tumor p53/genéticaAssuntos
Cromossomos de Mamíferos/genética , Metilação de DNA , Perda de Heterozigosidade , Sarcoma/genética , Neoplasias de Tecidos Moles/genética , Animais , Cromossomos de Mamíferos/fisiologia , DNA (Citosina-5-)-Metiltransferase 1 , DNA (Citosina-5-)-Metiltransferases/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , Fibroblastos/metabolismo , Genes da Neurofibromatose 1 , Genes p53 , Humanos , Camundongos , Mutação , Neoplasias/genética , Recombinação GenéticaRESUMO
Genome-wide DNA hypomethylation occurs in many human cancers, but whether this epigenetic change is a cause or consequence of tumorigenesis has been unclear. To explore this phenomenon, we generated mice carrying a hypomorphic DNA methyltransferase 1 (Dnmt1) allele, which reduces Dnmt1 expression to 10% of wild-type levels and results in substantial genome-wide hypomethylation in all tissues. The mutant mice were runted at birth, and at 4 to 8 months of age they developed aggressive T cell lymphomas that displayed a high frequency of chromosome 15 trisomy. These results indicate that DNA hypomethylation plays a causal role in tumor formation, possibly by promoting chromosomal instability.
Assuntos
Aberrações Cromossômicas , Cromossomos de Mamíferos/genética , Metilação de DNA , Linfoma de Células T/genética , Alelos , Animais , Peso ao Nascer , Transformação Celular Neoplásica , Cromossomos de Mamíferos/fisiologia , DNA (Citosina-5-)-Metiltransferase 1 , DNA (Citosina-5-)-Metiltransferases/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , Retrovirus Endógenos/genética , Retrovirus Endógenos/fisiologia , Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Rearranjo Gênico , Rearranjo Gênico da Cadeia beta dos Receptores de Antígenos dos Linfócitos T , Genes myc , Heterozigoto , Perda de Heterozigosidade , Linfoma de Células T/patologia , Camundongos , Camundongos Transgênicos , Vírus da Leucemia Murina de Moloney/genética , Vírus da Leucemia Murina de Moloney/fisiologia , Trissomia , Ativação ViralRESUMO
Biallelic expression of Igf2 is frequently seen in cancers because Igf2 functions as a survival factor. In many tumors the activation of Igf2 expression has been correlated with de novo methylation of the imprinted region. We have compared the intrinsic susceptibilities of the imprinted region of Igf2 and H19, other imprinted genes, bulk genomic DNA, and repetitive retroviral sequences to Dnmt1 overexpression. At low Dnmt1 methyltransferase levels repetitive retroviral elements were methylated and silenced. The nonmethylated imprinted region of Igf2 and H19 was resistant to methylation at low Dnmt1 levels but became fully methylated when Dnmt1 was overexpressed from a bacterial artificial chromosome transgene. Methylation caused the activation of the silent Igf2 allele in wild-type and Dnmt1 knockout cells, leading to biallelic Igf2 expression. In contrast, the imprinted genes Igf2r, Peg3, Snrpn, and Grf1 were completely resistant to de novo methylation, even when Dnmt1 was overexpressed. Therefore, the intrinsic difference between the imprinted region of Igf2 and H19 and of other imprinted genes to postzygotic de novo methylation may be the molecular basis for the frequently observed de novo methylation and upregulation of Igf2 in neoplastic cells and tumors. Injection of Dnmt1-overexpressing embryonic stem cells in diploid or tetraploid blastocysts resulted in lethality of the embryo, which resembled embryonic lethality caused by Dnmt1 deficiency.