RESUMEN
Targeted immune agonist (TIA) comprising a TLR7 agonist conjugated to tumor-targeting antibodies have been shown to induce potent anti-tumor responses in various preclinical models. However, the clinical proof-of-concept of a TIA has been hampered by systemic dose-limiting immune-related toxicities, including rapid induction of anti-drug antibodies in patients. We have developed ELISPOT-based assay to measure activation of antibody-secreting cells (ASCs), intended to simulate the interaction between TIA and peripheral B cells as a tool to pre-clinically de-risk tumor target-independent peripheral B-cell activation by TIA. This method has proven to be robust and has fast turn-around time to evaluate the induction of spontaneous B-cell activation by TIA in a tumor target- and FcγR-independent manner. This novel ASC assay platform may serve as a preclinical tool to de-risk TIAs that can potentially induce immune-related adverse effects in the clinic.
Asunto(s)
Neoplasias , Receptor Toll-Like 7 , Humanos , Adyuvantes Inmunológicos , Anticuerpos , Activación de LinfocitosRESUMEN
Janus kinase (JAK) inhibitors, such as tofacitinib (Xeljanz) and filgotinib (Jyseleca), have been approved for treatment of ulcerative colitis with several other JAK inhibitors in late-stage clinical trials for inflammatory bowel disease (IBD). Despite their impressive efficacy, the risk of adverse effects accompanying the use of JAK inhibitors has brought the entire class under scrutiny, leading to them receiving an FDA black box warning. In this study we investigated whether ileocolonic-targeted delivery of a pan-JAK inhibitor, tofacitinib, can lead to increased tissue exposure and reduced systemic exposure compared to untargeted formulations. The stability of tofacitinib in the presence of rat colonic microbiota was first confirmed. Next, in vivo computed tomography imaging was performed in rats to determine the transit time and disintegration site of ileocolonic-targeted capsules compared to gastric release capsules. Pharmacokinetic studies demonstrated that systemic drug exposure was significantly decreased, and colonic tissue exposure increased at 10 mg/kg tofacitinib dosed in ileocolonic-targeted capsules compared to gastric release capsules and an oral solution. Finally, in a rat model of LPS-induced colonic inflammation, targeted tofacitinib capsules significantly reduced concentrations of proinflammatory interleukin 6 in colonic tissue compared to a vehicle-treated control (p = 0.0408), unlike gastric release tofacitinib capsules and orally administered dexamethasone. Overall, these results support further development of ileocolonic-targeted tofacitinib, and potentially other specific JAK inhibitors in pre-clinical and clinical development, for the treatment of IBD.
RESUMEN
Peggy Sue is a capillary-based western/immunoassay platform that can separate and characterize proteins by size or charge. A quick and automated immunogenicity assay was developed on Peggy Sue based on charge separation and compared with a traditional bridging method using preclinical samples from non-human primate studies. The results generated with the Peggy Sue assay were comparable to those of the bridging assays. The Peggy Sue platform has several advantages, including time efficiency, low sample consumption, and easy automation. The platform is especially ideal for further characterization of anti-drug antibody (ADA) specificity against complex biologics such as bispecific or multi-specific biotherapeutics as it is easy to conduct domain specificity assessment of observed ADA responses. Our evaluation suggests that the Peggy Sue platform is a promising tool for preclinical ADA analysis.
Asunto(s)
Anticuerpos/aislamiento & purificación , Productos Biológicos/uso terapéutico , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos/diagnóstico , Inmunoensayo/métodos , Anticuerpos de Dominio Único/aislamiento & purificación , Animales , Automatización , Camélidos del Nuevo Mundo , Electroforesis Capilar , Humanos , Macaca mulattaRESUMEN
We have synthesized and evaluated a series of triaryl sulfonamide-based PTP1B inhibitors in which a difluoro-methylenephosphonate group of a potent lead has been replaced by potential bioisosteric replacements. Several mono- or di-charged compounds (8a, 8b, and 15a) were shown exhibit inhibitory activity in the low micromolar range, demonstrating the feasibility of using this approach in identifying non-phosphonate pTyr mimetics in a small molecular scaffold. These results also provide a useful indication of the relative effectiveness of these pTyr mimetics.
Asunto(s)
Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Compuestos de Flúor/síntesis química , Compuestos Organofosforados/síntesis química , Compuestos Organofosforados/farmacología , Proteína Tirosina Fosfatasa no Receptora Tipo 1/antagonistas & inhibidores , Sulfonamidas/química , Inhibidores Enzimáticos/química , Compuestos de Flúor/química , Compuestos de Flúor/farmacología , Estructura Molecular , Compuestos Organofosforados/química , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Relación Estructura-ActividadRESUMEN
The mammalian target of rapamycin (mTOR) is a central node in a complex signaling network that is regulated by several pathways deregulated in human cancers, including the PI3K/Akt and MAPK pathways. Targeting mTOR therefore presents an opportunity for therapeutic intervention. However, mTOR inhibition with rapamycin analogs or kinase inhibitors reduces cell growth but does not induce apoptosis, and the clinical benefit of rapamycin analogs has been modest. In this study we show that mTOR kinase inhibitors can potentiate apoptosis when used in combination with upstream targeted agents such as PI3K and MEK inhibitors. This increased apoptosis is dependent on genetic background, and correlates with active growth factor survival pathways. In PI3K mutant tumors, mTOR inhibition leads to partial reactivation of Akt which allows cells to survive, whereas in KRAS mutant tumors, this same reactivation of Akt occurs but is not required for cell survival. These data suggest the use of selected rational combinations of mTOR kinase inhibitors with other targeted inhibitors in specific tumor genotypes to achieve the maximal cytotoxic response by inhibiting two nodes in the activated signaling network.
Asunto(s)
Apoptosis/efectos de los fármacos , Quinasas Quinasa Quinasa PAM/antagonistas & inhibidores , Inhibidores de las Quinasa Fosfoinosítidos-3 , Inhibidores de Proteínas Quinasas/farmacología , Serina-Treonina Quinasas TOR/antagonistas & inhibidores , Western Blotting , Línea Celular Tumoral , Ensayo de Inmunoadsorción Enzimática , HumanosRESUMEN
BACKGROUND: DNA-damaging agents are widely used for the treatment of human malignancies. Agents containing the multifunctional alkylating moiety tetrakis(2-chloroethyl)phosphorodiamidic acid are currently under development as cancer therapeutics. MATERIALS AND METHODS: TLK58747, a phophorodiamidate-based prodrug, was tested in vivo for antitumor efficacy and safety. The in vitro responses of tumor cells to TLK58747 were examined by cytotoxicity assays, cell cycle analysis, immunoblots and microscopy. RESULTS: TLK58747 was efficacious in xenograft models of human breast, pancreas, and prostate cancer, as well as in leukemia and glioma. It caused less bone marrow suppression in rats than did cyclophosphamide. In vitro, TLK58747 inhibited the growth of a wide variety of cancer cells and activated the DNA damage-response pathway, leading to G(2)/M cell cycle arrest and subsequent premature senescence or apoptosis. CONCLUSION: TLK58747 is a promising new alkylating agent with broad antitumor activity and superior safety that warrants further development.
Asunto(s)
Antineoplásicos Alquilantes/farmacología , Daño del ADN , ADN de Neoplasias/efectos de los fármacos , Compuestos Organofosforados/farmacología , Profármacos/farmacología , Animales , Antineoplásicos Alquilantes/toxicidad , División Celular/efectos de los fármacos , Procesos de Crecimiento Celular/efectos de los fármacos , Línea Celular Tumoral , ADN de Neoplasias/metabolismo , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Fase G2/efectos de los fármacos , Células HL-60 , Humanos , Masculino , Ratones , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Neoplasias/patología , Compuestos Organofosforados/toxicidad , Profármacos/toxicidad , Ratas , Ratas Sprague-Dawley , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
A series of novel sulfonamides containing a single difluoromethylene-phosphonate group were discovered to be potent inhibitors of protein tyrosine phosphatase 1B. Structure-activity relationships around the scaffold were investigated, leading to the identification of compounds with IC50 or Ki values in the low nanomolar range. These sulfonamide-based inhibitors exhibit 100 and 30 times higher inhibitory activity than the corresponding tertiary amines and carboxamides, respectively.