An In Vivo CRISPR Screening Platform for Prioritizing Therapeutic Targets in AML.

CRISPR-Cas9-based genetic screens have successfully identified cell type-dependent liabilities in cancer, including acute myeloid leukemia (AML), a devastating hematologic malignancy with poor overall survival. Because most of these screens have been performed in vitro using established cell lines, evaluating the physiologic relevance of these targets is critical. We have established a CRISPR screening approach using orthotopic xenograft models to validate and prioritize AML-enriched dependencies in vivo, including in CRISPR-competent AML patient-derived xenograft (PDX) models tractable for genome editing. Our integrated pipeline has revealed several targets with translational value, including SLC5A3 as a metabolic vulnerability for AML addicted to exogenous myo-inositol and MARCH5 as a critical guardian to prevent apoptosis in AML. MARCH5 repression enhanced the efficacy of BCL2 inhibitors such as venetoclax, further highlighting the clinical potential of targeting MARCH5 in AML. Our study provides a valuable strategy for discovery and prioritization of new candidate AML therapeutic targets. SIGNIFICANCE: There is an unmet need to improve the clinical outcome of AML. We developed an integrated in vivo screening approach to prioritize and validate AML dependencies with high translational potential. We identified SLC5A3 as a metabolic vulnerability and MARCH5 as a critical apoptosis regulator in AML, both of which represent novel therapeutic opportunities.This article is highlighted in the In This Issue feature, p. 275.

A first-generation pediatric cancer dependency map.

Exciting therapeutic targets are emerging from CRISPR-based screens of high mutational-burden adult cancers. A key question, however, is whether functional genomic approaches will yield new targets in pediatric cancers, known for remarkably few mutations, which often encode proteins considered challenging drug targets. To address this, we created a first-generation pediatric cancer dependency map representing 13 pediatric solid and brain tumor types. Eighty-two pediatric cancer cell lines were subjected to genome-scale CRISPR-Cas9 loss-of-function screening to identify genes required for cell survival. In contrast to the finding that pediatric cancers harbor fewer somatic mutations, we found a similar complexity of genetic dependencies in pediatric cancer cell lines compared to that in adult models. Findings from the pediatric cancer dependency map provide preclinical support for ongoing precision medicine clinical trials. The vulnerabilities observed in pediatric cancers were often distinct from those in adult cancer, indicating that repurposing adult oncology drugs will be insufficient to address childhood cancers.

A Combination CDK4/6 and IGF1R Inhibitor Strategy for Ewing Sarcoma.

PURPOSE: Novel targeted therapeutics have transformed the care of subsets of patients with cancer. In pediatric malignancies, however, with simple tumor genomes and infrequent targetable mutations, there have been few new FDA-approved targeted drugs. The cyclin-dependent kinase (CDK)4/6 pathway recently emerged as a dependency in Ewing sarcoma. Given the heightened efficacy of this class with targeted drug combinations in other cancers, as well as the propensity of resistance to emerge with single agents, we aimed to identify genes mediating resistance to CDK4/6 inhibitors and biologically relevant combinations for use with CDK4/6 inhibitors in Ewing. EXPERIMENTAL DESIGN: We performed a genome-scale open reading frame (ORF) screen in 2 Ewing cell lines sensitive to CDK4/6 inhibitors to identify genes conferring resistance. Concurrently, we established resistance to a CDK4/6 inhibitor in a Ewing cell line. RESULTS: The ORF screen revealed IGF1R as a gene whose overexpression promoted drug escape. We also found elevated levels of phospho-IGF1R in our resistant Ewing cell line, supporting the relevance of IGF1R signaling to acquired resistance. In a small-molecule screen, an IGF1R inhibitor scored as synergistic with CDK4/6 inhibitor treatment. The combination of CDK4/6 inhibitors and IGF1R inhibitors was synergistic in vitro and active in mouse models. Mechanistically, this combination more profoundly repressed cell cycle and PI3K/mTOR signaling than either single drug perturbation. CONCLUSIONS: Taken together, these results suggest that IGF1R inhibitors activation is an escape mechanism to CDK4/6 inhibitors in Ewing sarcoma and that dual targeting of CDK4/6 inhibitors and IGF1R inhibitors provides a candidate synergistic combination for clinical application in this disease.

Evaluation of Gentamicin Exposure in the Neonatal Intensive Care Unit and Hearing Function at Discharge.

OBJECTIVE: To characterize the association between gentamicin dosing, duration of treatment, and ototoxicity in hospitalized infants. STUDY DESIGN: This retrospective cohort study conducted at 330 neonatal intensive care units (2002-2014) included inborn infants exposed to gentamicin with available hearing screen results, and excluded infants with incomplete dosing data and major congenital anomalies. Our primary outcome was the final hearing screen result performed during hospitalization: abnormal (failed or referred for further testing in one or both ears) or normal (bilateral passed). The 4 measures of gentamicin exposure were highest daily dose, average daily dose, cumulative dose, and cumulative duration of exposure. We fitted separate multivariable logistic regression models adjusted for demographics, comorbidities, and other clinical events. RESULTS: A total of 84 808 infants met inclusion/exclusion criteria; median (25th, 75th percentile) gestational age and birth weight were 35 weeks (33, 38) and 2480 g (1890, 3184), respectively. Failed hearing screens occurred in 3238 (3.8%) infants; failed screens were more likely in infants of lower gestational age and birth weight, who had longer hospital lengths of stay, higher rates of morbidities, and were small for gestational age. Median highest daily dose, average daily dose, and cumulative dose were 4.0 mg/kg/day (3.0, 4.0), 3.8 mg/kg/day (3.0, 4.0), and 12.1 mg/kg (9.1, 20.5), respectively. Median cumulative duration of exposure was 3 days (3, 6). In adjusted analysis, gentamicin dose and duration of therapy were not associated with hearing screen failure. CONCLUSIONS: Gentamicin dosing and duration of treatment were not associated with increased odds of failed hearing screen at the time of discharge from initial neonatal intensive care unit stay.