RESUMEN
Vascular endothelial growth factor and its receptors, FLK1/KDR and FLT1, are key regulators of angiogenesis. Unlike FLK1/KDR, the role of FLT1 has remained elusive. FLT1 is produced as soluble (sFLT1) and full-length isoforms. Here, we show that pericytes from multiple tissues produce sFLT1. To define the biologic role of sFLT1, we chose the glomerular microvasculature as a model system. Deletion of Flt1 from specialized glomerular pericytes, known as podocytes, causes reorganization of their cytoskeleton with massive proteinuria and kidney failure, characteristic features of nephrotic syndrome in humans. The kinase-deficient allele of Flt1 rescues this phenotype, demonstrating dispensability of the full-length isoform. Using cell imaging, proteomics, and lipidomics, we show that sFLT1 binds to the glycosphingolipid GM3 in lipid rafts on the surface of podocytes, promoting adhesion and rapid actin reorganization. sFLT1 also regulates pericyte function in vessels outside of the kidney. Our findings demonstrate an autocrine function for sFLT1 to control pericyte behavior.
Asunto(s)
Glomérulos Renales/citología , Glomérulos Renales/metabolismo , Podocitos/metabolismo , Receptor 1 de Factores de Crecimiento Endotelial Vascular/metabolismo , Animales , Gangliósidos/metabolismo , Humanos , Técnicas In Vitro , Metabolismo de los Lípidos , Lípidos/química , Ratones , Ratones Transgénicos , Pericitos/metabolismo , Proteinuria/metabolismo , Transducción de Señal , Sindecanos/metabolismo , Receptor 1 de Factores de Crecimiento Endotelial Vascular/genéticaRESUMEN
Strong evidence exists supporting the important role T cells play in the immune response against tumors. Still, the ability to initiate tumor-specific immune responses remains a challenge. Recent clinical trials suggest that bispecific antibody-mediated retargeted T cells are a promising therapeutic approach to eliminate hematopoietic tumors. However, this approach has not been validated in solid tumors. PF-06671008 is a dual-affinity retargeting (DART®)-bispecific protein engineered with enhanced pharmacokinetic properties to extend in vivo half-life, and designed to engage and activate endogenous polyclonal T cell populations via the CD3 complex in the presence of solid tumors expressing P-cadherin. This bispecific molecule elicited potent P-cadherin expression-dependent cytotoxic T cell activity across a range of tumor indications in vitro, and in vivo in tumor-bearing mice. Regression of established tumors in vivo was observed in both cell line and patient-derived xenograft models engrafted with circulating human T lymphocytes. Measurement of in vivo pharmacodynamic markers demonstrates PF-06671008-mediated T cell activation, infiltration and killing as the mechanism of tumor inhibition.
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Anticuerpos Biespecíficos/inmunología , Anticuerpos Biespecíficos/farmacología , Cadherinas/inmunología , Inmunoterapia/métodos , Neoplasias/inmunología , Neoplasias/terapia , Linfocitos T/inmunología , Animales , Complejo CD3/inmunología , Línea Celular Tumoral , Cricetinae , Cricetulus , Femenino , Células HCT116 , Células HT29 , Humanos , Ratones , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Using an expanded genetic code, antibodies with site-specifically incorporated nonnative amino acids were produced in stable cell lines derived from a CHO cell line with titers over 1 g/L. Using anti-5T4 and anti-Her2 antibodies as model systems, site-specific antibody drug conjugates (NDCs) were produced, via oxime bond formation between ketones on the side chain of the incorporated nonnative amino acid and hydroxylamine functionalized monomethyl auristatin D with either protease-cleavable or noncleavable linkers. When noncleavable linkers were used, these conjugates were highly stable and displayed improved in vitro efficacy as well as in vivo efficacy and pharmacokinetic stability in rodent models relative to conventional antibody drug conjugates conjugated through either engineered surface-exposed or reduced interchain disulfide bond cysteine residues. The advantages of the oxime-bonded, site-specific NDCs were even more apparent when low-antigen-expressing (2+) target cell lines were used in the comparative studies. NDCs generated with protease-cleavable linkers demonstrated that the site of conjugation had a significant impact on the stability of these rationally designed prodrug linkers. In a single-dose rat toxicology study, a site-specific anti-Her2 NDC was well tolerated at dose levels up to 90 mg/kg. These experiments support the notion that chemically defined antibody conjugates can be synthesized in commercially relevant yields and can lead to antibody drug conjugates with improved properties relative to the heterogeneous conjugates formed by nonspecific chemical modification.
Asunto(s)
Anticuerpos/metabolismo , Inmunoconjugados/metabolismo , Preparaciones Farmacéuticas/síntesis química , Ingeniería de Proteínas/métodos , Animales , Anticuerpos/sangre , Anticuerpos/química , Anticuerpos/toxicidad , Técnicas de Cultivo Celular por Lotes , Células CHO , Muerte Celular/efectos de los fármacos , Línea Celular , Cricetinae , Cricetulus , Cisteína/metabolismo , Humanos , Inmunoconjugados/química , Inmunoconjugados/farmacocinética , Inmunoconjugados/toxicidad , Preparaciones Farmacéuticas/sangre , Preparaciones Farmacéuticas/química , Estabilidad Proteica/efectos de los fármacos , RatasRESUMEN
There is a considerable ongoing work to identify new cytotoxic payloads that are appropriate for antibody-based delivery, acting via mechanisms beyond DNA damage and microtubule disruption, highlighting their importance to the field of cancer therapeutics. New modes of action will allow a more diverse set of tumor types to be targeted and will allow for possible mechanisms to evade the drug resistance that will invariably develop to existing payloads. Spliceosome inhibitors are known to be potent antiproliferative agents capable of targeting both actively dividing and quiescent cells. A series of thailanstatin-antibody conjugates were prepared in order to evaluate their potential utility in the treatment of cancer. After exploring a variety of linkers, we found that the most potent antibody-drug conjugates (ADCs) were derived from direct conjugation of the carboxylic acid-containing payload to surface lysines of the antibody (a "linker-less" conjugate). Activity of these lysine conjugates was correlated to drug-loading, a feature not typically observed for other payload classes. The thailanstatin-conjugates were potent in high target expressing cells, including multidrug-resistant lines, and inactive in nontarget expressing cells. Moreover, these ADCs were shown to promote altered splicing products in N87 cells in vitro, consistent with their putative mechanism of action. In addition, the exposure of the ADCs was sufficient to result in excellent potency in a gastric cancer xenograft model at doses as low as 1.5 mg/kg that was superior to the clinically approved ADC T-DM1. The results presented herein therefore open the door to further exploring splicing inhibition as a potential new mode-of-action for novel ADCs.
Asunto(s)
Productos Biológicos/química , Inmunoconjugados/química , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Animales , Ácidos Carboxílicos/química , Línea Celular Tumoral , Transformación Celular Neoplásica , Cisteína/química , Humanos , Inmunoconjugados/farmacocinética , Inmunoconjugados/farmacología , Lisina/química , Maleimidas/química , Ratones , Piranos/química , Distribución TisularRESUMEN
The Antibody Drug Conjugate (ADC) is a therapeutic modality consisting of a monoclonal antibody attached to a cytotoxic, small-molecule payload. The antibody portion of the ADC serves as a transport vehicle that recognizes and binds to a protein antigen expressed in tumor tissues. The localized delivery and release of the payload within or near malignant cells allows for targeted delivery of a potent cytotoxic agent to diseased tissue, while reducing damage to antigen-negative, normal tissues. Recent years have witnessed an explosive increase in ADC-based therapies, due mainly to clinical reports of activity in both hematologic and epithelial cancers. Accompanying this upsurge in ADC development is a renewed interest in natural product cytotoxins, which are typically highly potent cell-killing agents, but suffer from poor drug-like properties and narrow safety margins when systemically administered as conventional chemotherapeutics. In this review, we discuss recent advances related to the construction of ADCs, the optimization of ADC safety and efficacy, and the increasingly pivotal roles of natural product payloads in the current and future landscape of ADC therapy.
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Antineoplásicos/uso terapéutico , Productos Biológicos/uso terapéutico , Inmunoconjugados/uso terapéutico , Anticuerpos Monoclonales/administración & dosificación , Anticuerpos Monoclonales/uso terapéutico , Antineoplásicos/farmacología , Productos Biológicos/farmacología , Humanos , Inmunoconjugados/farmacología , Estructura Molecular , Neoplasias/tratamiento farmacológicoRESUMEN
Objectives of the present investigation were: (1) to compare three literature reported tumor growth inhibition (TGI) pharmacodynamic (PD) models and propose an optimal new model that best describes the xenograft TGI data for antibody drug conjugates (ADC), (2) to translate efficacy of the ADC Trastuzumab-emtansine (T-DM1) from mice to patients using the optimized PD model, and (3) to apply the translational strategy to predict clinically efficacious concentrations of a novel in-house anti-5T4 ADC, A1mcMMAF. First, the performance of all four of the PD models (i.e. 3 literature reported + 1 proposed) was evaluated using TGI data of T-DM1 obtained from four different xenografts. Based on the estimates of the pharmacodynamic/pharmacokinetic (PK/PD) modeling, a secondary parameter representing the efficacy index of the drug was calculated, which is termed as the tumor static concentration (TSC). TSC values derived from all four of the models were compared with each other, and with literature reported values, to assess the performance of these models. Subsequently, using the optimized PK/PD model, PD parameters obtained from different cell lines, human PK, and the proposed translational strategy, clinically efficacious doses of T-DM1 were projected. The accuracy of projected efficacious dose range for T-DM1 was verified by comparison with the clinical doses. Aforementioned strategy was then applied to A1mcMMAF for projecting its efficacious concentrations in clinic. TSC values for A1mcMMAF, obtained by fitting TGI data from 4 different xenografts with the proposed PK/PD model, were estimated to range from 0.6 to 11.5 µg mL⻹. Accordingly, the clinically efficacious doses for A1mcMMAF were projected retrospectively. All in all, the improved PD model and proposed translational strategy presented here suggest that appropriate correction for the clinical exposure and employing the TSC criterion can help translate mouse TGI data to predict first in human doses of ADCs.
Asunto(s)
Anticuerpos/farmacología , Antineoplásicos/farmacología , Antineoplásicos/farmacocinética , Inmunoconjugados/farmacología , Inmunoconjugados/farmacocinética , Neoplasias Experimentales/tratamiento farmacológico , Ado-Trastuzumab Emtansina , Animales , Anticuerpos Monoclonales Humanizados/farmacocinética , Anticuerpos Monoclonales Humanizados/farmacología , Línea Celular Tumoral , Femenino , Humanos , Maitansina/análogos & derivados , Maitansina/farmacocinética , Maitansina/farmacología , Ratones , Ratones Desnudos , Trastuzumab , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
The commentary by Colombo and Rich recently published in Cancer Cell provides a timely and comprehensive review of the clinical maximum tolerated doses (MTDs) of antibody-drug conjugates (ADCs) and their corresponding small molecules/chemotherapies. The authors identified similarities between their MTDs and therefore question the historic assumptions made for ADCs, namely, that they increase the MTDs of their corresponding cytotoxic molecules. However, the authors did not address the superior anti-tumor responses of ADCs compared to their corresponding chemotherapies, as reported in clinical trials. In this point of view, we propose a revised model wherein the anti-tumor activities of ADCs and consequently their therapeutic indexes (TIs) are not solely associated with changes not only in their MTDs but also in their minimal effective doses (MEDs). In addition, when using an exposure-based TI calculation method, the superior anti-tumor activities of ADCs relative to their corresponding chemotherapy can readily be explained. We discussed the clinical and preclinical data in support of lower MEDs of ADCs and generated a revised graph illustrating the TI improvements of ADCs vs chemotherapy more accurately. We believe that our revised model can provide a blueprint for future improvements in protein engineering and chemical engineering of toxins to further advance ADC research and development.
Asunto(s)
Antineoplásicos , Inmunoconjugados , Neoplasias , Humanos , Inmunoconjugados/uso terapéutico , Inmunoconjugados/química , Antineoplásicos/uso terapéutico , Antineoplásicos/química , Neoplasias/tratamiento farmacológico , Ingeniería de Proteínas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
In the RIP1-Tag2 mouse model of pancreatic islet carcinoma, angiogenesis is switched on in a discrete premalignant stage of tumor development, persisting thereafter. Signaling through VEGF receptor tyrosine kinases is a well-established component of angiogenic regulation. We show that five VEGF ligand genes are expressed in normal islets and throughout islet tumorigenesis. To begin dissecting their contributions, we produced an islet beta cell specific knockout of VEGF-A, resulting in islets with reduced vascularity but largely normal physiology. In RIP1-Tag2 mice wherein most oncogene-expressing cells had deleted the VEGF-A gene, both angiogenic switching and tumor growth were severely disrupted, as was the neovasculature. Thus, VEGF-A is crucial for angiogenesis in a prototypical model of carcinogenesis, whose loss is not readily compensated.
Asunto(s)
Factores de Crecimiento Endotelial/metabolismo , Islotes Pancreáticos/irrigación sanguínea , Islotes Pancreáticos/metabolismo , Neovascularización Patológica/metabolismo , Neoplasias Pancreáticas/irrigación sanguínea , Neoplasias Pancreáticas/metabolismo , Animales , División Celular , Factores de Crecimiento Endotelial/genética , Proteínas Activadoras de GTPasa/metabolismo , Eliminación de Gen , Perfilación de la Expresión Génica , Humanos , Islotes Pancreáticos/patología , Linfocinas/genética , Linfocinas/metabolismo , Ratones , Ratones Transgénicos , Neovascularización Patológica/genética , Especificidad de Órganos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factor A de Crecimiento Endotelial Vascular , Factores de Crecimiento Endotelial VascularRESUMEN
To reveal the functional significance of hypoxia and angiogenesis in astrocytoma progression, we created genetically engineered transformed astrocytes from murine primary astrocytes and deleted the hypoxia-responsive transcription factor HIF-1alpha or its target gene, the angiogenic factor VEGF. Growth of HIF-1alpha- and VEGF-deficient transformed astrocytes in the vessel-poor subcutaneous environment results in severe necrosis, reduced growth, and vessel density, whereas when the same cells are placed in the vascular-rich brain parenchyma, the growth of HIF-1alpha knockout, but not VEGF knockout tumors, is reversed: tumors deficient in HIF-1alpha grow faster, and penetrate the brain more rapidly and extensively. These results demonstrate that HIF-1alpha has differential roles in tumor progression, which are greatly dependent on the extant microenvironment of the tumor.
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Astrocitoma/irrigación sanguínea , Astrocitoma/metabolismo , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Hipoxia/metabolismo , Hipoxia/patología , Neovascularización Patológica , Animales , Astrocitos/metabolismo , Astrocitos/patología , Astrocitos/trasplante , Astrocitoma/patología , Neoplasias Encefálicas/irrigación sanguínea , División Celular , Transformación Celular Neoplásica , Progresión de la Enfermedad , Eliminación de Gen , Subunidad alfa del Factor 1 Inducible por Hipoxia , Ratones , Invasividad Neoplásica , Trasplante de Neoplasias , Tamaño de los Órganos , Tasa de Supervivencia , Factores de Transcripción/deficiencia , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Células Tumorales Cultivadas , Factor A de Crecimiento Endotelial Vascular/genética , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
We deleted the hypoxia-responsive transcription factor HIF-1alpha in endothelial cells (EC) to determine its role during neovascularization. We found that loss of HIF-1alpha inhibits a number of important parameters of EC behavior during angiogenesis: these include proliferation, chemotaxis, extracellular matrix penetration, and wound healing. Most strikingly, loss of HIF-1alpha in EC results in a profound inhibition of blood vessel growth in solid tumors. These phenomena are all linked to a decreased level of VEGF expression and loss of autocrine response of VEGFR-2 in HIF-1alpha null EC. We thus show that a HIF-1alpha-driven, VEGF-mediated autocrine loop in EC is an essential component of solid tumor angiogenesis.
Asunto(s)
Neoplasias/irrigación sanguínea , Neoplasias/metabolismo , Neovascularización Patológica , Factores de Transcripción/fisiología , Factor A de Crecimiento Endotelial Vascular/metabolismo , Animales , Hipoxia de la Célula , Proliferación Celular , Células Cultivadas , Quimiotaxis , Células Endoteliales/patología , Regulación de la Expresión Génica , Subunidad alfa del Factor 1 Inducible por Hipoxia , Ratones , Neoplasias/patología , Factores de Transcripción/genética , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Cicatrización de HeridasRESUMEN
Corneal avascularity-the absence of blood vessels in the cornea-is required for optical clarity and optimal vision, and has led to the cornea being widely used for validating pro- and anti-angiogenic therapeutic strategies for many disorders. But the molecular underpinnings of the avascular phenotype have until now remained obscure and are all the more remarkable given the presence in the cornea of vascular endothelial growth factor (VEGF)-A, a potent stimulator of angiogenesis, and the proximity of the cornea to vascularized tissues. Here we show that the cornea expresses soluble VEGF receptor-1 (sVEGFR-1; also known as sflt-1) and that suppression of this endogenous VEGF-A trap by neutralizing antibodies, RNA interference or Cre-lox-mediated gene disruption abolishes corneal avascularity in mice. The spontaneously vascularized corneas of corn1 and Pax6+/- mice and Pax6+/- patients with aniridia are deficient in sflt-1, and recombinant sflt-1 administration restores corneal avascularity in corn1 and Pax6+/- mice. Manatees, the only known creatures uniformly to have vascularized corneas, do not express sflt-1, whereas the avascular corneas of dugongs, also members of the order Sirenia, elephants, the closest extant terrestrial phylogenetic relatives of manatees, and other marine mammals (dolphins and whales) contain sflt-1, indicating that it has a crucial, evolutionarily conserved role. The recognition that sflt-1 is essential for preserving the avascular ambit of the cornea can rationally guide its use as a platform for angiogenic modulators, supports its use in treating neovascular diseases, and might provide insight into the immunological privilege of the cornea.
Asunto(s)
Córnea/irrigación sanguínea , Córnea/metabolismo , Receptor 1 de Factores de Crecimiento Endotelial Vascular/metabolismo , Animales , Eliminación de Gen , Ratones , Neovascularización Fisiológica , ARN Mensajero/genética , ARN Mensajero/metabolismo , Solubilidad , Trichechus , Factor A de Crecimiento Endotelial Vascular/metabolismo , Receptor 1 de Factores de Crecimiento Endotelial Vascular/deficiencia , Receptor 1 de Factores de Crecimiento Endotelial Vascular/genéticaRESUMEN
pHLA complexes represent the largest class of cell surface markers on cancer cells, making them attractive for targeted cancer therapies. Adoptive cell therapies expressing TCRs that recognize tumor specific pHLAs take advantage of the unique selectivity and avidity of TCR: pHLA interactions. More recently, additional protein binding domains binding to pHLAs, known as TCR mimics (TCRm), were developed for tumor targeting of high potency therapeutic modalities, including bispecifics, ADCs, CAR T and -NK cells. TCRm compounds take advantage of the exquisite tumor specificity of certain pHLA targets, including cell lineage commitment markers and cancer testis antigens (CTAs). To achieve meaningful anti-tumor responses, it is critical that TCRm compounds integrate both, high target binding affinities and a high degree of target specificity. In this review, we describe the most advanced approaches to achieve both criteria, including affinity- and specificity engineering of TCRs, antibodies and alternative protein scaffolds. We also discuss the status of current TCRm based therapeutics developed in the clinic, key challenges, and emerging trends to improve treatment options for cancer patients treated with TCRm based therapeutics in Oncology.
RESUMEN
Extra domain B splice variant of fibronectin (EDB+FN) is an extracellular matrix protein (ECM) deposited by tumor-associated fibroblasts, and is associated with tumor growth, angiogenesis, and invasion. We hypothesized that EDB+FN is a safe and abundant target for therapeutic intervention with an antibody-drug conjugate (ADC). We describe the generation, pharmacology, mechanism of action, and safety profile of an ADC specific for EDB+FN (EDB-ADC). EDB+FN is broadly expressed in the stroma of pancreatic, non-small cell lung (NSCLC), breast, ovarian, head and neck cancers, whereas restricted in normal tissues. In patient-derived xenograft (PDX), cell-line xenograft (CLX), and mouse syngeneic tumor models, EDB-ADC, conjugated to auristatin Aur0101 through site-specific technology, demonstrated potent antitumor growth inhibition. Increased phospho-histone H3, a pharmacodynamic biomarker of response, was observed in tumor cells distal to the target site of tumor ECM after EDB-ADC treatment. EDB-ADC potentiated infiltration of immune cells, including CD3+ T lymphocytes into the tumor, providing rationale for the combination of EDB-ADC with immune checkpoint therapy. EDB-ADC and anti-PD-L1 combination in a syngeneic breast tumor model led to enhanced antitumor activity with sustained tumor regressions. In nonclinical safety studies in nonhuman primates, EDB-ADC had a well-tolerated safety profile without signs of either on-target toxicity or the off-target effects typically observed with ADCs that are conjugated through conventional conjugation methods. These data highlight the potential for EDB-ADC to specifically target the tumor microenvironment, provide robust therapeutic benefits against multiple tumor types, and enhance activity antitumor in combination with checkpoint blockade.
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Neoplasias de la Mama , Inmunoconjugados , Animales , Neoplasias de la Mama/patología , Línea Celular Tumoral , Femenino , Fibronectinas/metabolismo , Humanos , Inmunoconjugados/farmacología , Inmunoconjugados/uso terapéutico , Ratones , Neovascularización Patológica/metabolismo , Microambiente Tumoral , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Endocrine pancreatic beta cells require endothelial signals for their differentiation and function. However, the molecular basis for such signals remains unknown. Here, we show that beta cells, in contrast to the exocrine pancreatic cells, do not form a basement membrane. Instead, by using VEGF-A, they attract endothelial cells, which form capillaries with a vascular basement membrane next to the beta cells. We have identified laminins, among other vascular basement membrane proteins, as endothelial signals, which promote insulin gene expression and proliferation in beta cells. We further demonstrate that beta1-integrin is required for the beta cell response to the laminins. The proposed mechanism explains why beta cells must interact with endothelial cells, and it may apply to other cellular processes in which endothelial signals are required.
Asunto(s)
Membrana Basal/metabolismo , Vasos Sanguíneos/anatomía & histología , Regulación de la Expresión Génica , Células Secretoras de Insulina/fisiología , Insulina/metabolismo , Transducción de Señal/fisiología , Animales , Membrana Basal/citología , Línea Celular , Proliferación Celular , Células Endoteliales/citología , Células Endoteliales/metabolismo , Insulina/genética , Células Secretoras de Insulina/citología , Integrina beta1/metabolismo , Laminina/metabolismo , Ratones , Páncreas/citología , Páncreas/metabolismo , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
The glomerular microvasculature is particularly susceptible to injury in thrombotic microangiopathy, but the mechanisms by which this occurs are unclear. We report the cases of six patients who were treated with bevacizumab, a humanized monoclonal antibody against vascular endothelial growth factor (VEGF), in whom glomerular disease characteristic of thrombotic microangiopathy developed. To show that local reduction of VEGF within the kidney is sufficient to trigger the pathogenesis of thrombotic microangiopathy, we used conditional gene targeting to delete VEGF from renal podocytes in adult mice; this resulted in a profound thrombotic glomerular injury. These observations provide evidence that glomerular injury in patients who are treated with bevacizumab is probably due to direct targeting of VEGF by antiangiogenic therapy.
Asunto(s)
Inhibidores de la Angiogénesis/efectos adversos , Anticuerpos Monoclonales/efectos adversos , Glomérulos Renales/efectos de los fármacos , Podocitos/metabolismo , Trombosis/inducido químicamente , Factor A de Crecimiento Endotelial Vascular/antagonistas & inhibidores , Anciano , Inhibidores de la Angiogénesis/uso terapéutico , Animales , Anticuerpos Monoclonales/uso terapéutico , Anticuerpos Monoclonales Humanizados , Bevacizumab , Femenino , Marcación de Gen , Humanos , Glomérulos Renales/irrigación sanguínea , Glomérulos Renales/patología , Masculino , Ratones , Ratones Noqueados , Microcirculación/efectos de los fármacos , Persona de Mediana Edad , Neoplasias/tratamiento farmacológico , Proteinuria/inducido químicamente , ARN Mensajero/metabolismo , Circulación Renal , Transducción de Señal , Factor A de Crecimiento Endotelial Vascular/genética , Factor A de Crecimiento Endotelial Vascular/inmunología , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Despite major advances in the treatment of non-Hodgkin lymphoma (NHL), including the use of chemotherapeutic agents and the anti-CD20 antibody rituximab, the majority of patients eventually relapse, and salvage treatments with non-cross-resistant compounds are needed to further improve patient survival. Here, we evaluated the antitumor effects of the microtubule destabilizing agent monomethyl auristatin E (MMAE) conjugated to the humanized anti-CD19 antibody hBU12 via a protease-sensitive valine-citrulline (vc) dipeptide linker. hBU12-vcMMAE induced potent tumor cell killing against rituximab-sensitive and -resistant NHL cell lines. CD19 can form heterodimers with CD21, and high levels of CD21 were reported to interfere negatively with the activity of CD19-targeted therapeutics. However, we observed comparable internalization, intracellular trafficking, and drug release in CD21(low) and CD21(high), rituximab-sensitive and -refractory lymphomas treated with hBU12-vcMMAE. Furthermore, high rates of durable regressions in mice implanted with these tumors were observed, suggesting that both rituximab resistance and CD21 expression levels do not impact on the activity of hBU12-vcMMAE. Combined, our data suggest that hBU12-vcMMAE may represent a promising addition to the treatment options for rituximab refractory NHL and other hematologic malignancies, including acute lymphoblastic leukemia.
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Anticuerpos Monoclonales/uso terapéutico , Antígenos CD19/inmunología , Antineoplásicos/uso terapéutico , Resistencia a Antineoplásicos , Inmunoconjugados/uso terapéutico , Linfoma no Hodgkin/tratamiento farmacológico , Oligopéptidos/uso terapéutico , Animales , Anticuerpos Monoclonales de Origen Murino , Western Blotting , Línea Celular Tumoral , Supervivencia Celular , Citrulina/química , Citrulina/metabolismo , Dimerización , Dipéptidos/metabolismo , Femenino , Citometría de Flujo , Dosificación de Gen , Humanos , Técnicas para Inmunoenzimas , Linfoma no Hodgkin/inmunología , Linfoma no Hodgkin/metabolismo , Lisosomas , Ratones , Ratones SCID , Oligopéptidos/metabolismo , Receptores de Complemento 3d/genética , Receptores de Complemento 3d/inmunología , Receptores de Complemento 3d/metabolismo , Rituximab , Valina/química , Valina/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Rheumatoid arthritis (RA) is characterized by inflammation and cellular proliferation in the synovial lining of joints that result in cartilage and bone destruction. Although the etiology of RA is unclear, activated lymphocytes and proinflammatory molecules, in particular TNF superfamily members, have been implicated in the disease pathology. A TNF superfamily member, CD70, is found on activated lymphocytes and shown to be important in memory and effector responses of lymphocytes. CD70 is expressed at high levels on chronically activated T cells in patients with autoimmune disorders, including RA. The involvement of CD70 in the progression of RA, however, remains unknown. In this study, we report effects of targeting CD70 on disease pathogenesis by using an anti-mouse CD70 Ab in a murine model of collagen-induced arthritis (CIA). In addition to blocking CD70 binding to its receptor CD27, the anti-CD70 Ab used also engages Fc-dependent effector functions including Ab-dependent cellular cytotoxicity, phagocytosis, and complement fixation. Treatment of mice with anti-CD70 Ab both before the onset or after the established disease in CIA model resulted in marked improvements in disease severity and significant reduction in the production of autoantibodies. Histopathological analyses of the joints of mice revealed a substantial reduction of inflammation, and bone and cartilage destruction in response to the anti-CD70 Ab treatment. These results uncover a novel role for CD27-CD70 interactions in the regulation of in vivo inflammatory response leading to arthritis, and provide a molecular basis to support the rationale for anti-CD70 therapy for autoimmune and inflammatory diseases.
Asunto(s)
Anticuerpos Monoclonales/farmacología , Artritis Experimental/tratamiento farmacológico , Ligando CD27/antagonistas & inhibidores , Miembro 7 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/antagonistas & inhibidores , Animales , Artritis Experimental/patología , Autoanticuerpos/análisis , Huesos/efectos de los fármacos , Huesos/patología , Cartílago/efectos de los fármacos , Cartílago/patología , Progresión de la Enfermedad , Inflamación/prevención & control , Artropatías , Masculino , RatonesRESUMEN
Vascular endothelial growth factor (VEGF) is a key regulator of physiological angiogenesis during embryogenesis, skeletal growth and reproductive functions. VEGF has also been implicated in pathological angiogenesis associated with tumors, intraocular neovascular disorders and other conditions. The biological effects of VEGF are mediated by two receptor tyrosine kinases (RTKs), VEGFR-1 and VEGFR-2, which differ considerably in signaling properties. Non-signaling co-receptors also modulate VEGF RTK signaling. Currently, several VEGF inhibitors are undergoing clinical testing in several malignancies. VEGF inhibition is also being tested as a strategy for the prevention of angiogenesis, vascular leakage and visual loss in age-related macular degeneration.
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Factores de Crecimiento Endotelial/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Linfocinas/metabolismo , Neovascularización Patológica , Neovascularización Fisiológica , Receptores de Factores de Crecimiento Endotelial Vascular/metabolismo , Transducción de Señal/fisiología , Animales , Ensayos Clínicos como Asunto , Factores de Crecimiento Endotelial/genética , Factores de Crecimiento Endotelial/uso terapéutico , Endotelio Vascular/citología , Endotelio Vascular/metabolismo , Sustancias de Crecimiento/metabolismo , Humanos , Péptidos y Proteínas de Señalización Intercelular/genética , Péptidos y Proteínas de Señalización Intercelular/uso terapéutico , Linfocinas/genética , Linfocinas/uso terapéutico , Oxígeno/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Factor A de Crecimiento Endotelial Vascular , Factores de Crecimiento Endotelial VascularRESUMEN
PURPOSE: Mortality due to acute myeloid leukemia (AML) remains high, and the management of relapsed or refractory AML continues to be therapeutically challenging. The reapproval of Mylotarg, an anti-CD33-calicheamicin antibody-drug conjugate (ADC), has provided a proof of concept for an ADC-based therapeutic for AML. Several other ADCs have since entered clinical development of AML, but have met with limited success. We sought to develop a next-generation ADC for AML with a wide therapeutic index (TI) that overcomes the shortcomings of previous generations of ADCs. EXPERIMENTAL DESIGN: We compared the TI of our novel CD33-targeted ADC platform with other currently available CD33-targeted ADCs in preclinical models of AML. Next, using this next-generation ADC platform, we performed a head-to-head comparison of two attractive AML antigens, CD33 and CD123. RESULTS: Our novel ADC platform offered improved safety and TI when compared with certain currently available ADC platforms in preclinical models of AML. Differentiation between the CD33- and CD123-targeted ADCs was observed in safety studies conducted in cynomolgus monkeys. The CD33-targeted ADC produced severe hematologic toxicity, whereas minimal hematologic toxicity was observed with the CD123-targeted ADC at the same doses and exposures. The improved toxicity profile of an ADC targeting CD123 over CD33 was consistent with the more restricted expression of CD123 in normal tissues. CONCLUSIONS: We optimized all components of ADC design (i.e., leukemia antigen, antibody, and linker-payload) to develop an ADC that has the potential to translate into an effective new therapy against AML.
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Gemtuzumab/uso terapéutico , Inmunoconjugados/uso terapéutico , Subunidad alfa del Receptor de Interleucina-3/antagonistas & inhibidores , Leucemia Mieloide Aguda/tratamiento farmacológico , Lectina 3 Similar a Ig de Unión al Ácido Siálico/antagonistas & inhibidores , Animales , Antineoplásicos Inmunológicos/inmunología , Antineoplásicos Inmunológicos/farmacocinética , Antineoplásicos Inmunológicos/uso terapéutico , Área Bajo la Curva , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Gemtuzumab/inmunología , Gemtuzumab/farmacocinética , Células HL-60 , Humanos , Inmunoconjugados/inmunología , Inmunoconjugados/farmacocinética , Subunidad alfa del Receptor de Interleucina-3/inmunología , Leucemia Mieloide Aguda/inmunología , Leucemia Mieloide Aguda/metabolismo , Macaca fascicularis , Ratones , Lectina 3 Similar a Ig de Unión al Ácido Siálico/inmunología , Carga Tumoral/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
Aberrant NOTCH3 signaling and overexpression is oncogenic, associated with cancer stem cells and drug resistance, yet therapeutic targeting remains elusive. Here, we develop NOTCH3-targeted antibody drug conjugates (NOTCH3-ADCs) by bioconjugation of an auristatin microtubule inhibitor through a protease cleavable linker to two antibodies with differential abilities to inhibit signaling. The signaling inhibitory antibody rapidly induces ligand-independent receptor clustering and internalization through both caveolin and clathrin-mediated pathways. The non-inhibitory antibody also efficiently endocytoses via clathrin without inducing receptor clustering but with slower lysosomal co-localization kinetics. In addition, DLL4 ligand binding to the NOTCH3 receptor mediates transendocytosis of NOTCH3-ADCs into ligand-expressing cells. NOTCH3-ADCs internalize into receptor and ligand cells independent of signaling and induce cell death in both cell types representing an atypical mechanism of ADC cytotoxicity. Treatment of xenografts with NOTCH3-ADCs leads to sustained tumor regressions, outperforms standard-of-care chemotherapy, and allows targeting of tumors that overexpress NOTCH3 independent of signaling inhibition.