Lauren L. Ott

Radiation exposure to the eye during cancer treatment can lead to ocular radiation injury (ORI), a devastating condition that can have a profound and permanent impact on vision-related quality of life. Rodent models do not have adequate ocular anatomy to accurately simulate human ORI, and modeling in non-human primates is limited by logistical and ethical concerns. To improve future translational research investigating ways to treat or prevent ORI, we developed protocols for a tree shrew model of ORI. Northern tree shrews (Tupaia belangeri) were obtained by our laboratory. Custom housing and handling methods were developed, including custom body suits to maintain the tree shrew’s body temperature during procedures. Radiation delivery was optimized to accurately deliver radiation, and imaging was performed to observe fundus changes from ORI. Optimization of tree shrew handling, housing, anesthesia approaches, radiation delivery, and clinically-relevant ocular imaging permitted successful induction and assessment of ORI in tree shrews. With these protocols, tree shrews can be used as a highly relevant model organism with key anatomic features similar to humans to study ORI.

Purpose: Navtemadlin is a potent small-molecule inhibitor of MDM2, which has completed a phase 0 window-of-opportunity study in glioblastoma (GBM). To optimally interpret the clinical data, a detailed analysis of navtemadlin pharmacokinetics (PK), pharmacodynamics, and efficacy was performed in GBM patient-derived xenografts (PDX). Experimental Design: Response to navtemadlin was characterized in vitro and in vivo in GBM PDXs with and without MDM2 amplification. Efficacy in vivo was integrated with measured plasma and intra-tumoral drug levels to develop a translational PK/efficacy model comparing exposure effective in PDX to exposure achieved in phase 0 patient samples. Results: In vitro, navtemadlin showed robust on-target activity in TP53–wild-type GBM. In vivo efficacy strongly correlated with MDM2 amplification status. In subcutaneous PDXs, navtemadlin significantly extended survival when dosed at 25 mg/kg in an MDM2-amplified PDX compared with 100 mg/kg in a non-amplified PDX. Central nervous system distribution was limited by blood–brain barrier efflux (Kp_brain = 0.009). In an MDM2-amplified orthotopic PDX model, navtemadlin was ineffective at 100 mg/kg; when established in mice with deficient blood–brain barrier efflux (Rag−/−Abcb1a−/−Abcg2−/−), 25 mg/kg doubled survival. A tumor PK/efficacy model was built to define target exposure for efficacy in GBM, using the effective 25 mg/kg dose. Modeled exposures exceeded this threshold in three (of 16) tumor samples from phase 0 study patients at the 240 mg dose level. Conclusions: Navtemadlin efficacy was highly dependent on adequate brain penetration. Our translational PK/efficacy model suggests that the minimum effective tumor exposures were achieved only in a minority of patients with GBM.

Top row: Correlation of Normalized Enrichment Scores (NES) and viability of IR+peposertib normalized to IR in Homologous Recombination (HR), Microhomology-mediated End Joining (MMEJ), Non-Homologous End Joining (NHEJ), and mRNA PRKDC rpkm in four PDX GBM lines (GBM 6, 10, 22, and 76). Bottom row: Same data but split into TP53 wildtype lines (TP53wt, GBM10 and GBM76) vs. TP53 mutant lines (TP53mut, GBM 6 and 22) with IR and IR+peposertib data normalized to DMSO.

Standardization curve of peposertib (M3814) concentration as measured by MALDI in normal and tumor tissue.

A, Immunoflourescence of 53BP1 foci in U251 cells 1- and 24-hours after indicated treatments. B, PRKDC methylation scores in indicated GBM PDX lines. C, Somatic mutations and Copy Number Variations in GBM PDX lines tested in vivo in key GBM pathways. D, Western blot confirmation of DNA-PK knock down by shRNA (shPRKDC) or non-targeting shRNA (shNT).

Supplementary Data S2: derivation of modified biologically effective dose model.

Supplementary Figure S2 demonstrates agreement between AUC computed from simulated and measured profiles.

Ataxia Telangiectasia–mutated (ATM) inhibitors are being developed as radiosensitizers to improve the antitumor effects of radiotherapy, but ATM inhibition can also radiosensitize normal tissues. Therefore, understanding the elevated risk of normal tissue toxicities is critical for radiosensitizer development. This study focused on modeling the relationship between acute mucosal toxicity, radiation dose, fractionation schedule, and radiosensitizer exposure. The ATM inhibitor WSD0628 was combined with single or fractionated doses of radiation delivered to the oral cavity or esophagus of Friend Leukemia virus B (FVB) mice. The potentiation by WSD0628 was quantified by a sensitizer enhancement ratio (SER), which describes the changes in radiation tolerance for radiation combined with WSD0628 relative to radiation-only regimens. WSD0628 profoundly enhanced radiation-induced acute oral and esophageal toxicities. For oral mucosal toxicity, the enhancement by WSD0628 with 3 fractions of radiation resulted in an SER ranging from 1.3 (0.25 mg/kg) to 3.1 (7.5 mg/kg). For the 7.5 mg/kg combination, the SER increased with increasing number of fractions from 2.2 (1 fraction) to 4.3 (7 fractions) for oral toxicity and from 2.2 (1 fraction) to 3.6 (3 fractions) for esophageal toxicity, which reflects a loss of the normal tissue sparing benefit of fractionated radiation. These findings were used to develop a modified biologically effective dose model to determine alternative radiation schedules with or without WSD0628 that result in similar levels of toxicity. Successful radiosensitizer dose escalation to a maximally effective therapeutic dose will require careful deliberation of tumor site and reduction of radiation dose volume limits for organs at risk.

Supplementary Figure S1 includes repeats of immunofluorescence images and an example of H&E staining from untreated oral mucosa.