Coexpression of normally incompatible developmental pathways in retinoblastoma genesis.

It is widely believed that the molecular and cellular features of a tumor reflect its cell of origin and can thus provide clues about treatment targets. The retinoblastoma cell of origin has been debated for over a century. Here, we report that human and mouse retinoblastomas have molecular, cellular, and neurochemical features of multiple cell classes, principally amacrine/horizontal interneurons, retinal progenitor cells, and photoreceptors. Importantly, single-cell gene expression array analysis showed that these multiple cell type-specific developmental programs are coexpressed in individual retinoblastoma cells, which creates a progenitor/neuronal hybrid cell. Furthermore, neurotransmitter receptors, transporters, and biosynthetic enzymes are expressed in human retinoblastoma, and targeted disruption of these pathways reduces retinoblastoma growth in vivo and in vitro.

Subconjunctival carboplatin and systemic topotecan treatment in preclinical models of retinoblastoma.

BACKGROUND: The authors demonstrated previously that the combination of topotecan (TPT) and carboplatin (CBP) was more effective than current chemotherapeutic combinations used to treat retinoblastoma in an orthotopic xenograft model. However, systemic coadministration of these agents is not ideal, because both agents cause dose-limiting myelosuppression in children. METHODS: To overcome the toxicity associated with systemic TPT and CBP, the authors explored subconjunctival delivery of TPT or CBP in an orthotopic xenograft model and in a genetic mouse model of retinoblastoma (Chx10-Cre;Rb(lox/lox);p107(-/-);p53(lox/lox)). The effects of combined subconjunctival CBP (CBP(subcon)) and systemic TPT (TPT(syst)) were compared with the effects of combined TPT(subcon) and CBP(syst.) at clinically relevant dosages. RESULTS: Pharmacokinetic and tumor-response studies, including analyses of ocular and hematopoietic toxicity, revealed that CBP(subcon)/TPT(syst) was more effective and had fewer side effects than TPT(subcon)/CBP(syst). CONCLUSIONS: For the first time, retinoblastoma was ablated and long-term vision was preserved in a mouse model by using a clinically relevant chemotherapy regimen. These results eventually may be translated into a clinical trial for children with this debilitating cancer.

Whole-body physiologically based pharmacokinetic model for nutlin-3a in mice after intravenous and oral administration.

Nutlin-3a is an MDM2 inhibitor that is under investigation in preclinical models for a variety of pediatric malignancies, including retinoblastoma, rhabdomyosarcoma, neuroblastoma, and leukemia. We used physiologically based pharmacokinetic (PBPK) modeling to characterize the disposition of nutlin-3a in the mouse. Plasma protein binding and blood partitioning were assessed by in vitro studies. After intravenous (10 and 20 mg/kg) and oral (50, 100, and 200 mg/kg) dosing, tissue concentrations of nutlin-3a were determined in plasma, liver, spleen, intestine, muscle, lung, adipose, bone marrow, adrenal gland, brain, retina, and vitreous fluid. The PBPK model was simultaneously fit to all pharmacokinetic data using NONMEM. Nutlin-3a exhibited nonlinear binding to murine plasma proteins, with the unbound fraction ranging from 0.7 to 11.8%. Nutlin-3a disposition was characterized by rapid absorption with peak plasma concentrations at approximately 2 h and biphasic elimination consistent with a saturable clearance process. The final PBPK model successfully described the plasma and tissue disposition of nutlin-3a. Simulations suggested high bioavailability, rapid attainment of steady state, and little accumulation when administered once or twice daily at dosages up to 400 mg/kg. The final model was used to perform simulations of unbound tissue concentrations to determine which dosing regimens are appropriate for preclinical models of several pediatric malignancies.

MDM2-A, a common Mdm2 splice variant, causes perinatal lethality, reduced longevity and enhanced senescence.

MDM2 is the predominant negative regulator of p53 that functions to maintain the appropriate level of expression and activity of this central tumor suppressor. Mdm2-a is a commonly identified splice variant of Mdm2; however, its physiological function is unclear. To gain insight into the activity of MDM2-A and its potential impact on p53, an Mdm2-a transgenic mouse model was generated. Mdm2-a transgenic mice displayed a homozygous-lethal phenotype that could be rescued by a reduction in p53 expression, demonstrating a dependence upon p53. Mdm2-a hemizygous mice exhibited reduced longevity, and enhanced senescence was observed in their salivary glands. In addition, the transgenic mice lacked typical, accelerated aging phenotypes. Growth of transgenic mouse embryonic fibroblasts (MEFs) was inhibited relative to wild-type MEFs, and MDM2-A was shown to bind to full-length MDM2 in an interaction that could increase p53 activity via reduced MDM2 inhibition. Evidence of p53 activation was shown in the Mdm2-a transgenic MEFs, including p53-dependent growth inhibition and elevated expression of the p53 target protein p21. In addition, MDM2-A increased senescence in a p21-independent manner. In conclusion, unexpected roles for MDM2-A in longevity and senescence were identified in a transgenic mouse model, suggesting that Mdm2 splice variants might be determinants of these phenotypes in vivo.