RESUMEN
The outcome for metastatic pediatric osteosarcoma (OS) remains poor. Thus, there is an urgent need to develop novel therapies, and immunotherapy with CAR T cells has the potential to meet this challenge. However, there is a lack of preclinical models that mimic salient features of human disease including reliable development of metastatic disease post orthotopic OS cell injection. To overcome this roadblock, and also enable real-time imaging of metastatic disease, we took advantage of LM7 OS cells expressing firefly luciferase (LM7.ffLuc). LM7.ffLuc were implanted in a collagen mesh into the tibia of mice, and mice reliably developed orthotopic tumors and lung metastases as judged by bioluminescence imaging and histopathological analysis. Intratibial implantation also enabled surgical removal by lower leg amputation and monitoring for metastases development post-surgery. We then used this model to evaluate the antitumor activity of CAR T cells targeting B7-H3, an antigen that is expressed in a broad range of solid tumors including OS. B7-H3-CAR T cells had potent antitumor activity in a dose-dependent manner and inhibited the development of pulmonary metastases resulting in a significant survival advantage. In contrast T cells expressing an inactive B7-H3-CAR had no antitumor activity. Using unmodified LM7 cells also enabled us to demonstrate that B7-H3-CAR T cells traffic to orthotopic tumor sites. Hence, we have developed an orthotopic, spontaneously metastasizing OS model. This model may improve our ability not only to predict the safety and efficacy of current and next generation CAR T cell therapies but also other treatment modalities for metastatic OS.
Asunto(s)
Antígenos B7/inmunología , Neoplasias Óseas/terapia , Inmunoterapia Adoptiva , Osteosarcoma/terapia , Receptores Quiméricos de Antígenos/inmunología , Animales , Neoplasias Óseas/patología , Línea Celular Tumoral , Femenino , Humanos , Neoplasias Pulmonares/secundario , Neoplasias Pulmonares/terapia , Ratones , Osteosarcoma/patología , Tibia/patología , Resultado del Tratamiento , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: We report pharmacokinetic (PK) data, evaluation of modifications for increased stability, evaluation for cellular uptake, and mediation of regression of breast cancer for the aptamer OPN-R3. METHODS: The OPN-R3 aptamer was assessed for PK data in vivo with additional comparison of IV and subcutaneous dosing. Five aptamer variants were generated by differential 2'-O-methylation for comparison with parent. OPN-R3-Cy3 was incubated with MDA-MB231 cells and cellular uptake evaluated under confocal microscopy. Mice were treated with OPN-R3, mutant, or saline 3 weeks after inoculation with MDA-MB231 cells and tumor size was evaluated. RESULTS: OPN-R3 PK data were: t(1/2) 7.76 hours, T(max) 3 hours, C(max) 13.2 mmol/L, mean residence time 9 hours, AUC (0-t) 161.9 mmol/hr/L, and K(d) 57.2 nmol/L. The half-life was higher when given intravenously versus subcutaneously (E(1/2) 7.93 vs 0.74 hours). The 2' methylation of all available bases increased unmodified aptamer stability and affinity (t(1/2) 6.2 hours; K(d) 520 nmol/L), but this did not improve on parent aptamer (t(1/2) 7.78 hours, K(d) 18 nmol/L). The aptamer remained extracellular. OPN-R3 caused regression of tumor to levels seen at 1 week after tumor inoculation. CONCLUSION: We show the efficacy of OPN-R3 for reversing growth of breast cancer cells with adequate PK stability for clinical application.