RESUMO
Human CD19 antigen is a 95-kDa type I membrane glycoprotein in the immunoglobulin superfamily whose expression is limited to the various stages of B-cell development and differentiation and is maintained in the majority of B-cell malignancies, including leukemias and non-Hodgkin lymphomas of B-cell origin. Coupled with its differential and favorable expression profile, CD19 has rapid internalization kinetics and is not shed into the circulation, making it an ideal target for the development of antibody-drug conjugates (ADCs) to treat B-cell malignancies. ADCT-402 (loncastuximab tesirine) is a novel CD19-targeted ADC delivering SG3199, a highly cytotoxic DNA minor groove interstrand crosslinking pyrrolobenzodiazepine (PDB) dimer warhead. It showed potent and highly targeted in vitro cytotoxicity in CD19-expressing human cell lines. ADCT-402 was specifically bound, internalized, and trafficked to lysosomes in CD19-expressing cells and, following release of the PBD warhead, resulted in formation of DNA crosslinks that persisted for 36 hours. Bystander killing of CD19- cells by ADCT-402 was also observed. In vivo, single doses of ADCT-402 resulted in highly potent, dose-dependent antitumor activity in several subcutaneous and disseminated human tumor models with marked superiority to comparator ADCs delivering tubulin inhibitors. Dose-dependent DNA crosslinks and γ-H2AX DNA damage response were measured in tumors by 24 hours after single dose administration, whereas matched peripheral blood mononuclear cells showed no evidence of DNA damage. Pharmacokinetic analysis in rat and cynomolgus monkey showed excellent stability and tolerability of ADCT-402 in vivo. Together, these impressive data were used to support the clinical testing of this novel ADC in patients with CD19-expressing B-cell malignancies.
Assuntos
Antígenos CD19/biossíntese , Antineoplásicos , Regulação Leucêmica da Expressão Gênica , Imunoconjugados , Leucemia de Células B , Linfoma não Hodgkin , Proteínas de Neoplasias/biossíntese , Antineoplásicos/farmacocinética , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Humanos , Imunoconjugados/farmacocinética , Imunoconjugados/farmacologia , Leucemia de Células B/tratamento farmacológico , Leucemia de Células B/metabolismo , Leucemia de Células B/patologia , Linfoma não Hodgkin/tratamento farmacológico , Linfoma não Hodgkin/metabolismo , Linfoma não Hodgkin/patologia , Lisossomos/metabolismo , Lisossomos/patologiaRESUMO
The normal electron-demand Diels-Alder (DA) cycloaddition is a classic transformation routinely used in synthesis; however, applications in biological systems are limited. Here, we report a spiro[2.4]hepta-4,6-diene-containing noncanonical amino acid (SCpHK) capable of efficient incorporation into antibodies and subsequent coupling with maleimide via a DA reaction. SCpHK was stable throughout protein expression in mammalian cells and enabled covalent attachment of maleimide drug-linkers yielding DA antibody-drug conjugates (DA-ADCs) with nearly quantitative conversion in a one-step process. The uncatalyzed DA reaction between SCpHK and maleimide in aqueous buffer was rapid (1.8-5.4 M-1 s-1), and the antibody-drug adduct was stable in rat serum for at least 1 week at 37 °C. Anti-EphA2 DA-ADCs containing AZ1508 or SG3249 maleimide drug-linkers were potent inhibitors of tumor growth in PC3 tumor models in vivo. The DA bioconjugation strategy described here represents a simple method to produce site-specific and stable ADCs with maleimide drug-linkers.
Assuntos
Imunoconjugados/química , Maleimidas/química , Animais , Células CHO , Sobrevivência Celular/efeitos dos fármacos , Cricetulus , Reação de Cicloadição , Humanos , Imunoconjugados/farmacologia , Modelos Moleculares , Células PC-3 , Conformação Proteica , Compostos de Espiro/químicaRESUMO
An analysis of Antibody-Drug Conjugate Payload manufacturing has revealed that the majority of the cost is associated with the use of high-containment facilities for the latter stages of the synthesis. To make a significant reduction in the Cost of Goods (CoGs), a new approach to route design has been introduced which focuses on minimizing the number of steps that require high containment. This approach has been exemplified in a new synthesis of tesirine, including the first application of a ring-closing copper(I)/TEMPO aerobic oxidation to the pyrrolobenzodiazepine ring system, affording a 60% reduction in CoGs.
Assuntos
Benzodiazepinas/síntese química , Desenho de Fármacos , Imunoconjugados/química , Pirróis/síntese química , Benzodiazepinas/química , Ciclização , Estrutura Molecular , Pirróis/químicaRESUMO
A novel pyrrolobenzodiazepine dimer payload, SG3227, was rationally designed based on the naturally occurring antitumour compound sibiromycin. SG3227 was synthesized from a dimeric core in an efficient fashion. An unexpected room temperature Diels-Alder reaction occurred during the final step of the synthesis and was circumvented by use of an iodoacetamide conjugation moiety in place of a maleimide. The payload was successfully conjugated to trastuzumab and the resulting ADC exhibited potent activity against a HER2-expressing human cancer cell line in vitro.
Assuntos
Aminoglicosídeos/química , Antineoplásicos/química , Benzodiazepinas/química , Imunoconjugados/química , Linhagem Celular Tumoral , Avaliação Pré-Clínica de Medicamentos , Humanos , Técnicas In VitroRESUMO
Antibody-drug conjugates (ADC) have emerged as potent antitumor drugs that provide increased efficacy, specificity, and tolerability over chemotherapy for the treatment of cancer. ADCs generated by targeting cysteines and lysines on the antibody have shown efficacy, but these products are heterogeneous, and instability may limit their dosing. Here, a novel technology is described that enables site-specific conjugation of toxins to antibodies using chemistry to produce homogeneous, potent, and highly stable conjugates. We have developed a cell-based mammalian expression system capable of site-specific integration of a non-natural amino acid containing an azide moiety. The azide group enables click cycloaddition chemistry that generates a stable heterocyclic triazole linkage. Antibodies to Her2/neu were expressed to contain N6-((2-azidoethoxy)carbonyl)-l-lysine at four different positions. Each site allowed over 95% conjugation efficacy with the toxins auristatin F or a pyrrolobenzodiazepine (PBD) dimer to generate ADCs with a drug to antibody ratio of >1.9. The ADCs were potent and specific in in vitro cytotoxicity assays. An anti Her2/neu conjugate demonstrated stability in vivo and a PBD containing ADC showed potent efficacy in a mouse tumor xenograph model. This technology was extended to generate fully functional ADCs with four toxins per antibody. The high stability of the azide-alkyne linkage, combined with the site-specific nature of the expression system, provides a means for the generation of ADCs with optimized pharmacokinetic, biological, and biophysical properties.
Assuntos
Aminoácidos/química , Azidas/química , Química Click , Reação de Cicloadição , Imunoconjugados/química , Aminoácidos/genética , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/genética , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/uso terapêutico , Azidas/metabolismo , Humanos , Imunoconjugados/genética , Imunoconjugados/imunologia , Imunoconjugados/uso terapêutico , Masculino , Camundongos , Neoplasias/tratamento farmacológico , Engenharia de Proteínas , Ratos Sprague-Dawley , Receptor ErbB-2/imunologiaRESUMO
A highly cytotoxic DNA cross-linking pyrrolobenzodiazepine (PBD) dimer with a valine-alanine dipeptide linker was conjugated to the anti-CD70 h1F6 mAb either through endogenous interchain cysteines or, site-specifically, through engineered cysteines at position 239 of the heavy chains. The h1F6239C-PBD conjugation strategy proved to be superior to interchain cysteine conjugation, affording an antibody-drug conjugate (ADC) with high uniformity in drug-loading and low levels of aggregation. In vitro cytotoxicity experiments demonstrated that the h1F6239C-PBD was potent and immunologically specific on CD70-positive renal cell carcinoma (RCC) and non-Hodgkin lymphoma (NHL) cell lines. The conjugate was resistant to drug loss in plasma and in circulation, and had a pharmacokinetic profile closely matching that of the parental h1F6239C antibody capped with N-ethylmaleimide (NEM). Evaluation in CD70-positive RCC and NHL mouse xenograft models showed pronounced antitumor activities at single or weekly doses as low as 0.1 mg/kg of ADC. The ADC was tolerated at 2.5 mg/kg. These results demonstrate that PBDs can be effectively used for antibody-targeted therapy.
Assuntos
Benzodiazepinas/química , Ligante CD27/química , Imunoconjugados/farmacologia , Animais , Dimerização , Desenho de Fármacos , Feminino , Meia-Vida , Imunoconjugados/química , Camundongos , Camundongos Endogâmicos BALB CRESUMO
Antibody-drug conjugates (ADC) delivering pyrrolobenzodiazepine (PBD) DNA cross-linkers are currently being evaluated in clinical trials, with encouraging results in Hodgkin and non-Hodgkin lymphomas. The first example of an ADC delivering a PBD DNA cross-linker (loncastuximab tesirine) has been recently approved by the FDA for the treatment of relapsed and refractory diffuse large B-cell lymphoma. There has also been considerable interest in mono-alkylating PBD analogs. We conducted a head-to-head comparison of a conventional PBD bis-imine and a novel PBD mono-imine. Key Mitsunobu chemistry allowed clean and convenient access to the mono-imine class. Extensive DNA-binding studies revealed that the mono-imine mediated a type of DNA interaction that is described as "pseudo cross-linking," as well as alkylation. The PBD mono-imine ADC demonstrated robust antitumor activity in mice bearing human tumor xenografts at doses 3-fold higher than those that were efficacious for the PBD bis-imine ADC. A single-dose toxicology study in rats demonstrated that the MTD of the PBD mono-alkylator ADC was approximately 3-fold higher than that of the ADC bearing a bis-imine payload, suggesting a comparable therapeutic index for this molecule. However, although both ADCs caused myelosuppression, renal toxicity was observed only for the bis-imine, indicating possible differences in toxicologic profiles that could influence tolerability and therapeutic index. These data show that mono-amine PBDs have physicochemical and pharmacotoxicologic properties distinct from their cross-linking analogs and support their potential utility as a novel class of ADC payload.
Assuntos
Imunoconjugados , Linfoma não Hodgkin , Humanos , Animais , Camundongos , Ratos , Alquilação , DNA , Iminas , Imunoconjugados/farmacologiaRESUMO
PURPOSE: We evaluated the activity of AZD8205, a B7-H4-directed antibody-drug conjugate (ADC) bearing a novel topoisomerase I inhibitor (TOP1i) payload, alone and in combination with the PARP1-selective inhibitor AZD5305, in preclinical models. EXPERIMENTAL DESIGN: IHC and deep-learning-based image analysis algorithms were used to assess prevalence and intratumoral heterogeneity of B7-H4 expression in human tumors. Several TOP1i-ADCs, prepared with Val-Ala or Gly-Gly-Phe-Gly peptide linkers, with or without a PEG8 spacer, were compared in biophysical, in vivo efficacy, and rat toxicology studies. AZD8205 mechanism of action and efficacy studies were conducted in human cancer cell line and patient-derived xenograft (PDX) models. RESULTS: Evaluation of IHC-staining density on a per-cell basis revealed a range of heterogeneous B7-H4 expression across patient tumors. This informed selection of bystander-capable Val-Ala-PEG8-TOP1i payload AZ14170133 and development of AZD8205, which demonstrated improved stability, efficacy, and safety compared with other linker-payload ADCs. In a study of 26 PDX tumors, single administration of 3.5 mg/kg AZD8205 provided a 69% overall response rate, according to modified RECIST criteria, which correlated with homologous recombination repair (HRR) deficiency (HRD) and elevated levels of B7-H4 in HRR-proficient models. Addition of AZD5305 sensitized very low B7-H4-expressing tumors to AZD8205 treatment, independent of HRD status and in models representing clinically relevant mechanisms of PARPi resistance. CONCLUSIONS: These data provide evidence for the potential utility of AZD8205 for treatment of B7-H4-expressing tumors and support the rationale for an ongoing phase 1 clinical study (NCT05123482). See related commentary by Pommier and Thomas, p. 991.
Assuntos
Imunoconjugados , Neoplasias , Ratos , Humanos , Animais , Imunoconjugados/farmacologia , Imunoconjugados/uso terapêutico , Inibidores da Topoisomerase I , Neoplasias/tratamento farmacológico , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Poli(ADP-Ribose) Polimerase-1/genéticaRESUMO
OBJECTIVES: Pyrrolobenzodiazepine (PBD) dimers, tethered through inert propyldioxy or pentyldioxy linkers, possess potent bactericidal activity against a range of Gram-positive bacteria by virtue of their capacity to cross-link duplex DNA in sequence-selective fashion. Here we attempt to improve the antibacterial activity and cytotoxicity profile of PBD-containing conjugates by extension of dimer linkers and replacement of one PBD unit with phenyl-substituted or benzo-fused heterocycles that facilitate non-covalent interactions with duplex DNA. METHODS: DNase I footprinting was used to identify high-affinity DNA binding sites. A staphylococcal gene microarray was used to assess epidemic methicillin-resistant Staphylococcus aureus 16 phenotypes induced by PBD conjugates. Molecular dynamics simulations were employed to investigate the accommodation of compounds within the DNA helix. RESULTS: Increasing the length of the linker in PBD dimers led to a progressive reduction in antibacterial activity, but not in their cytotoxic capacity. Complex patterns of DNA binding were noted for extended PBD dimers. Modelling of DNA strand cross-linking by PBD dimers indicated distortion of the helix. A majority (26 of 43) of PBD-biaryl conjugates possessed potent antibacterial activity with little or no helical distortion and a more favourable cytotoxicity profile. Bactericidal activity of PBD-biaryl conjugates was determined by inability to excise covalently bound drug molecules from bacterial duplex DNA. CONCLUSIONS: PBD-biaryl conjugates have a superior antibacterial profile compared with PBD dimers such as ELB-21. We have identified six PBD-biaryl conjugates as potential drug development candidates.
Assuntos
Antibacterianos/farmacologia , Benzodiazepinas/farmacologia , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Pirróis/farmacologia , Antibacterianos/metabolismo , Benzodiazepinas/metabolismo , Sítios de Ligação , Pegada de DNA , DNA Bacteriano/metabolismo , Perfilação da Expressão Gênica , Análise em Microsséries , Viabilidade Microbiana/efeitos dos fármacos , Simulação de Dinâmica Molecular , Pirróis/metabolismoRESUMO
The pyrrolobenzodiazepines (PBDs) are naturally occurring antitumor antibiotics and a PBD dimer (SJG-136, SG2000) is in Phase II trials. SG2000 is a propyldioxy linked PBD dimer which binds sequence selectively in the minor groove of DNA forming DNA interstrand and intrastrand cross-linked adducts, and also mono-adducts depending on sequence. SG2057 is the corresponding dimer containing a pentyldioxy linkage. SG2057 has multilog differential in vitro cytotoxicity against a panel of human tumour cell lines with a mean GI(50) of 212 pM. The agent is highly efficient at producing DNA interstrand cross-links in cells which form rapidly and persist over a 48 h period. Significant antitumor activity was demonstrated in several human tumor xenograft models. Cures were obtained in a LOX-IMVI melanoma model following a single administration and dose-dependent activity, including regression responses, observed in SKOV-3 ovarian and HL-60 promyelocytic leukemia models following repeat dose schedules. In the advanced stage LS174T model, SG2057 administered either as a single dose, or in two repeat dose schedules, was superior to irinotecan. SG2057 is therefore a highly active antitumor agent, with more potent in vitro activity and superior in vivo activity to SG2000, warranting further development.
Assuntos
Antineoplásicos/uso terapêutico , Benzodiazepinonas/uso terapêutico , Reagentes de Ligações Cruzadas/uso terapêutico , Neoplasias/tratamento farmacológico , Animais , Antineoplásicos/farmacologia , Benzodiazepinonas/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Reagentes de Ligações Cruzadas/farmacologia , DNA/efeitos dos fármacos , DNA/metabolismo , Feminino , Humanos , Camundongos , Camundongos Nus , Camundongos SCID , Neoplasias/patologia , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Antibody-drug conjugate (ADC) research has typically focused on the release of highly potent cytotoxic agents to achieve antitumor efficacy. However, recently approved ADCs trastuzumab deruxtecan and sacituzumab govitecan release lower-potency topoisomerase inhibitors. This has prompted interest in ADCs that release lower-potency cytotoxic drugs to potentially enhance therapeutic index and reduce unwanted toxicity. Pyrrolobenzodiazepine (PBD) dimer ADCs have been widely investigated in human clinical trials, which have focused on high-potency PBDs. In this study, we evaluated five ADCs that release the low-potency PBD dimer SG3650. The relatively low clogD for this agent facilitated higher drug-to-antibody ratio (DAR) conjugation without the need for antibody engineering or functionalization of the drug. The rank order of potency for DAR 2 site-specific ADCs (conjugated at the C239i position) matched the order for the corresponding free drugs in vitro. Despite free drug SG3650 being inactive in vivo, the DAR 2 ADCs derived from the corresponding drug-linker SG3584 showed antitumor efficacy in solid (anti-HER2) and hematologic (anti-CD22) xenograft models. Antitumor activity could be enhanced by conjugating SG3584 to trastuzumab at higher DARs of 4 and 8 and by adjusting dosing and schedule. Higher-DAR conjugates were stable and displayed good rat pharmacokinetic profiles as measured by ELISA and LC/MS-MS. A single intravenous dose of isotype control SG3584 DAR 2 ADC resulted in no mortality in rats or monkeys at doses of up to 25 and 30 mg/kg, respectively. These findings suggest that further investigations of low-potency PBD dimers in ADCs that target hematologic and solid tumors are warranted.
Assuntos
Antineoplásicos , Imunoconjugados , Animais , Antineoplásicos/farmacologia , Benzodiazepinas/farmacologia , Linhagem Celular Tumoral , Humanos , Imunoconjugados/uso terapêutico , Pirróis , Ratos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
A series of novel DNA-interactive C8-linked pyrrolobenzodiazepine (PBD)-heterocycle polyamide conjugates has been synthesised to explore structure/sequence-selectivity relationships. One conjugate (2d) has a greater selectivity and DNA binding affinity for inverted CCAAT sequences within the Topoisomerase IIα promoter than the known C8-bis-pyrrole PBD conjugate GWL-78 (1b).
Assuntos
Amidas/química , Benzodiazepinas/química , Regiões Promotoras Genéticas , Pirróis/química , Amidas/síntese química , Amidas/metabolismo , Antígenos de Neoplasias/genética , Sequência de Bases , Benzodiazepinas/síntese química , Benzodiazepinas/metabolismo , DNA Topoisomerases Tipo II/genética , Proteínas de Ligação a DNA/genética , Modelos Moleculares , Regiões Promotoras Genéticas/genética , Ligação Proteica , Pirróis/síntese química , Pirróis/metabolismo , Relação Estrutura-AtividadeRESUMO
Antibody-drug conjugates (ADCs) are an emerging class of oncology treatments combining the unique specificity of monoclonal antibodies with the highly cytotoxic properties of small molecule compounds. Pyrrolobenzodiazepines (PBDs) are highly potent agents capable of inhibiting cellular DNA replication which leads to apoptosis. To ensure efficacy and patient safety upon administration of such toxic and heterogeneous molecules, their structure and quality attributes must be closely monitored. Size exclusion chromatography (SEC) is a powerful, fast and robust tool for the separation of compounds varying in molecular weight. When using volatile components in the chromatographic mobile phase, SEC has also been shown to be amenable for interfacing to mass spectrometry, providing potential for reliable identification of protein isoforms across the size variants present. Here, we present a SEC-MS method developed for the characterisation of PBD-based ADCs on the intact molecular level. We demonstrate that information on ADC monomers such as the glycoform distribution and the average drug-antibody ratio (DAR) can be obtained in 15 minutes of analysis time. Qualitative and quantitative information on low and high molecular weight impurities such as aggregates and fragments, fundamental for critical quality attribute analysis of biopharmaceuticals, can be generated simultaneously. SEC-MS enables the characterisation of multiple product quality attributes of complex biotherapeutics at the same time.
Assuntos
Imunoconjugados , Anticorpos Monoclonais , Benzodiazepinas , Cromatografia em Gel , Humanos , Imunoconjugados/análise , Espectrometria de Massas , PirróisRESUMO
Resistance to antibody-drug conjugates (ADCs) has been observed in both preclinical models and clinical studies. However, mechanisms of resistance to pyrrolobenzodiazepine (PBD)-conjugated ADCs have not been well characterized and thus, this study was designed to investigate development of resistance to PBD dimer warheads and PBD-conjugated ADCs. We established a PBD-resistant cell line, 361-PBDr, by treating human breast cancer MDA-MB-361 cells with gradually increasing concentrations of SG3199, the PBD dimer released from the PBD drug-linker tesirine. 361-PBDr cells were over 20-fold less sensitive to SG3199 compared with parental cells and were cross-resistant to other PBD warhead and ADCs conjugated with PBDs. Proteomic profiling revealed that downregulation of Schlafen family member 11 (SLFN11), a putative DNA/RNA helicase, sensitizing cancer cells to DNA-damaging agents, was associated with PBD resistance. Confirmatory studies demonstrated that siRNA knockdown of SLFN11 in multiple tumor cell lines conferred reduced sensitivity to SG3199 and PBD-conjugated ADCs. Treatment with EPZ011989, an EZH2 inhibitor, derepressed SLFN11 expression in 361-PBDr and other SLFN11-deficient tumor cells, and increased sensitivity to PBD and PBD-conjugated ADCs, indicating that the suppression of SLFN11 expression is associated with histone methylation as reported. Moreover, we demonstrated that combining an ataxia telangiectasia and Rad3-related protein (ATR) inhibitor, AZD6738, with SG3199 or PBD-based ADCs led to synergistic cytotoxicity in either resistant 361-PBDr cells or cells that SLFN11 was knocked down via siRNA. Collectively, these data provide insights into potential development of resistance to PBDs and PBD-conjugated ADCs, and more importantly, inform strategy development to overcome such resistance.
Assuntos
Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Benzodiazepinas/metabolismo , Proteínas Nucleares/metabolismo , Pirróis/metabolismo , Regulação para Baixo , Resistencia a Medicamentos Antineoplásicos , Feminino , Humanos , TransfecçãoRESUMO
Antibody-drug conjugates (ADC) containing pyrrolobenzodiazepine (PBD) dimers are being evaluated clinically in both hematologic and solid tumors. These include ADCT-301 (camidanlumab tesirine) and ADCT-402 (loncastuximab tesirine) in pivotal phase II trials that contain the payload tesirine, which releases the PBD dimer warhead SG3199. An important consideration in future clinical development is acquired resistance. The aim was to generate and characterize PBD acquired resistant cell lines in both hematologic and solid tumor settings. Human Karpas-299 (ALCL) and NCI-N87 (gastric cancer) cells were incubated with increasing IC50 doses of ADC (targeting CD25 and HER2, respectively) or SG3199 in a pulsed manner until stable acquired resistance was established. The level of resistance achieved was approximately 3,000-fold for ADCT-301 and 3-fold for SG3199 in Karpas-299, and 8-fold for ADCT-502 and 4-fold for SG3199 in NCI-N87. Cross-resistance between ADC and SG3199, and with an alternative PBD-containing ADC or PBD dimer was observed. The acquired resistant lines produced fewer DNA interstrand cross-links, indicating an upstream mechanism of resistance. Loss of antibody binding or internalization was not observed. A human drug transporter PCR Array revealed several genes upregulated in all the resistant cell lines, including ABCG2 and ABCC2, but not ABCB1(MDR1). These findings were confirmed by RT-PCR and Western blot, and inhibitors and siRNA knockdown of ABCG2 and ABCC2 recovered drug sensitivity. These data show that acquired resistance to PBD-ADCs and SG3199 can involve specific ATP-binding cassette drug transporters. This has clinical implications as potential biomarkers of resistance and for the rational design of drug combinations.
Assuntos
Benzodiazepinas/química , Resistencia a Medicamentos Antineoplásicos , Imunoconjugados/farmacologia , Linfoma Anaplásico de Células Grandes/genética , Pirróis/química , Neoplasias Gástricas/genética , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/genética , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Anticorpos Monoclonais Humanizados/química , Anticorpos Monoclonais Humanizados/farmacologia , Benzodiazepinas/farmacologia , Linhagem Celular Tumoral , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Imunoconjugados/química , Linfoma Anaplásico de Células Grandes/tratamento farmacológico , Linfoma Anaplásico de Células Grandes/metabolismo , Proteína 2 Associada à Farmacorresistência Múltipla , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/metabolismoRESUMO
cMet is a well-characterized oncogene that is the target of many drugs including small molecule and biologic pathway inhibitors, and, more recently, antibody-drug conjugates (ADCs). However, the clinical benefit from cMet-targeted therapy has been limited. We developed a novel cMet-targeted 'third-generation' ADC, TR1801-ADC, that was optimized at different levels including specificity, stability, toxin-linker, conjugation site, and in vivo efficacy. Our nonagonistic cMet antibody was site-specifically conjugated to the pyrrolobenzodiazepine (PBD) toxin-linker tesirine and has picomolar activity in cancer cell lines derived from different solid tumors including lung, colorectal, and gastric cancers. The potency of our cMet ADC is independent of MET gene copy number, and its antitumor activity was high not only in high cMet-expressing cell lines but also in medium-to-low cMet cell lines (40 000-90 000 cMet/cell) in which a cMet ADC with tubulin inhibitor payload was considerably less potent. In vivo xenografts with low-medium cMet expression were also very responsive to TR1801-ADC at a single dose, while a cMet ADC using a tubulin inhibitor showed a substantially reduced efficacy. Furthermore, TR1801-ADC had excellent efficacy with significant antitumor activity in 90% of tested patient-derived xenograft models of gastric, colorectal, and head and neck cancers: 7 of 10 gastric models, 4 of 10 colorectal cancer models, and 3 of 10 head and neck cancer models showed complete tumor regression after a single-dose administration. Altogether, TR1801-ADC is a new generation cMet ADC with best-in-class preclinical efficacy and good tolerability in rats.
Assuntos
Antineoplásicos/farmacologia , Benzodiazepinas/farmacologia , Imunoconjugados/farmacologia , Neoplasias/tratamento farmacológico , Oncogenes/imunologia , Proteínas Proto-Oncogênicas c-met/imunologia , Pirróis/farmacologia , Animais , Antineoplásicos/uso terapêutico , Antineoplásicos/toxicidade , Neoplasias do Sistema Biliar/metabolismo , Linhagem Celular Tumoral , Neoplasias do Colo/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Neoplasias de Cabeça e Pescoço/metabolismo , Humanos , Imunoconjugados/uso terapêutico , Imunoconjugados/toxicidade , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias/imunologia , Proteínas Proto-Oncogênicas c-met/metabolismo , Ratos , Ratos Sprague-Dawley , Neoplasias Gástricas/metabolismo , Análise Serial de Tecidos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
A dynamic equilibrium between covalent 1:1 hairpin and 2:1 duplex DNA adducts of a pyrrolobenzodiazepine (PBD) minor groove binding agent () has been observed for the first time. The equilibrium, which establishes over 1 hour and must require unfolding of both types of adducts, is surprising given that PBDs normally require DNA minor groove structure for binding and take 24 hours for complete reaction with duplex DNA. The equilibrium is interesting from an energetics perspective due to the well known DNA stabilizing effect of PBDs. This observation could have significance for the in vitro and in vivo biological activity of PBDs, as DNA hairpin and loop structures are known to be important in cellular processes such as transcription and replication.
Assuntos
Benzodiazepinas/farmacologia , Adutos de DNA/química , Adutos de DNA/efeitos dos fármacos , DNA/química , DNA/efeitos dos fármacos , Conformação de Ácido Nucleico/efeitos dos fármacos , Pirróis/farmacologia , Sequência de Bases , Benzodiazepinas/química , Sítios de Ligação , Cromatografia Líquida de Alta Pressão , Pirróis/química , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
We report a novel class of biaryl polyamides highly selective for G-quadruplex DNA, and with significant cytotoxicity in several cancer cell lines; they form planar U-shaped structures that match the surface area dimensions of a terminal G-quartet in quadruplex structures rather than the grooves of duplex DNA.
Assuntos
Antineoplásicos/farmacologia , Quadruplex G/efeitos dos fármacos , Nylons/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Dicroísmo Circular , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Concentração Inibidora 50 , Ligantes , Nylons/síntese química , Nylons/químicaRESUMO
A prodrug form (17) of a novel C2/C2'-aryl-substituted pyrrolobenzodiazepine (PBD) dimer (16) has been synthesized by introducing sodium bisulfite groups to the C11/C11'-positions of the parent bis-imine. The prodrug form is highly water soluble, stable in aqueous conditions, and the rate of DNA cross-link formation is much slower compared to the parent bis-imine.
Assuntos
Benzodiazepinonas/síntese química , DNA/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Pró-Fármacos , Pirróis/síntese química , Animais , Benzodiazepinas/farmacologia , Benzodiazepinonas/química , Benzodiazepinonas/farmacologia , Linhagem Celular Tumoral , Simulação por Computador , DNA/metabolismo , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Compostos de Epóxi/farmacologia , Humanos , Modelos Moleculares , Pirróis/química , Pirróis/farmacologia , Solubilidade/efeitos dos fármacos , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
Distamycin is a naturally occurring antibiotic that binds to AT-rich sequences in the minor groove of DNA in a noncovalent manner. It continues to be of interest as a "building block" for more-complex small-molecule ligands capable of targeting specific DNA sequences for gene regulation purposes (i.e., transcription factor inhibitors). We report here a convenient and efficient synthesis of a library of 72 novel analogs (3a-f) of the parent distamycin on SynPhase Lanterns. To investigate structure-activity relationships including DNA-binding affinity and sequence-selectivity, two previously unexplored points of diversification have been introduced into the distamycin structure by replacing one of its pyrrole rings with novel biaryl motifs. The key aspects of the synthetic approach include the development of an efficient protocol for preparation of the heterocyclic polyamide chain, optimization of the Suzuki-Miyaura cross-coupling reaction and application of a split-and-mix technique based on radiofrequency encoding. In addition, a series of biaryl carboxamide derivatives (4a-f) has been synthesized utilizing the title library diversity reagents.