Acute lymphoblastic leukemia

Hedgehog pathway mutations drive oncogenic transformation in high-risk T-cell acute lymphoblastic leukemia


The role of Hedgehog signaling in normal and malignant T-cell development is controversial. Recently, Hedgehog pathway mutations have been described in T-ALL, but whether mutational activation of Hedgehog signaling drives T-cell transformation is unknown, hindering the rationale for therapeutic intervention. Here, we show that Hedgehog pathway mutations predict chemotherapy resistance in human T-ALL, and drive oncogenic transformation in a zebrafish model of the disease. We found Hedgehog pathway mutations in 16% of 109 childhood T-ALL cases, most commonly affecting its negative regulator PTCH1. Hedgehog mutations were associated with resistance to induction chemotherapy (P = 0.009). Transduction of wild-type PTCH1 into PTCH1-mutant T-ALL cells induced apoptosis (P = 0.005), a phenotype that was reversed by downstream Hedgehog pathway activation (P= 0.007). Transduction of most mutant PTCH1, SUFU, and GLI alleles into mammalian cells induced aberrant regulation of Hedgehog signaling, indicating that these mutations are pathogenic. Using a CRISPR/Cas9 system for lineage-restricted gene disruption in transgenic zebrafish, we found that ptch1 mutations accelerated the onset of notch1-induced T-ALL (P= 0.0001), and pharmacologic Hedgehog pathway inhibition had therapeutic activity. Thus, Hedgehog-activating mutations are driver oncogenic alterations in high-risk T-ALL, providing a molecular rationale for targeted therapy in this disease.

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We thank Julien Ablain for the pCS2-Cas9, pDest-Tol2-CG2-U6-gRNA, pME-MCS-zfCas9-T2A-GFP, and p3E-polyA vectors. We thank Christine Reynolds and Oscar Calzada for technical assistance; and Sofie Peirs, Bjorn Menten, and Pieter Van Vlierberghe for the array CGH analysis. We thank Jessica Blackburn for the rag2-notch1aICD vector; and Jae Cho for the basal cell carcinoma cell line, ASZ001. We are grateful to the patients and families who provided samples for these studies. This work was supported by NIH R01 CA193651 (AG) and the Boston Children’s Hospital Translational Research Program (AG). The Children’s Oncology Group work was supported by U10 CA98543 (COG Chair’s grant), U10 CA98413 (COG Statistical Center), U24 CA114766 (COG Specimen Banking), U10 CA 180886 (COG Operations Center), and U10 CA 180899 (COG Statistics and Data Center). MB is supported by the Children’s Leukemia Research Association, Inc. SPH is the Jeffrey E. Perelman Distinguished Chair in the Department of Pediatrics, Children’s Hospital of Philadelphia. AG is supported by an Investigatorship at Boston Children’s Hospital.

Author contributions:

MAB, ZWL, and AG conceived the project, designed and performed experiments, interpreted data, and wrote the manuscript. ART and MD performed the targeted exon sequencing and analysis. MAB, GPP, and AG analyzed targeted exon sequencing, RNA sequencing and array CGH data. MAB, EAS, and NY performed and interpreted experiments. KES and DSN performed statistical analyses and interpreted data. KES, DSN, SPH, MLL, SSW, KPD, MD, LBS, and SES aided in data collection of primary patient samples, and in analysis and interpretation of data. All authors aided in critical revision and approval of the submitted manuscript.

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Correspondence to Alejandro Gutierrez.

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