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Thiopurines are effective immunosuppressants and anticancer agents, but intracellular accumulation of their active metabolites (6-thioguanine nucleotides, 6-TGN) causes dose-limiting hematopoietic toxicity. Thiopurine S-methyltransferase deficiency is known to exacerbate thiopurine toxicity. However, many patients are highly sensitive to thiopurines for unknown reasons. We show that multidrug-resistance protein 4 (Mrp4) is abundant in myeloid progenitors and tested the role of the Mrp4, an ATP transporter of monophosphorylated nucleosides, in this unexplained thiopurine sensitivity. Mrp4-deficient mice experienced Mrp4 gene dosage-dependent toxicity caused by accumulation of 6-TGNs in their myelopoietic cells. Therefore, Mrp4 protects against thiopurine-induced hematopoietic toxicity by actively exporting thiopurine nucleotides. We then identified a single-nucleotide polymorphism (SNP) in human MRP4 (rs3765534) that dramatically reduces MRP4 function by impairing its cell membrane localization. This SNP is common (>18%) in the Japanese population and indicates that the increased sensitivity of some Japanese patients to thiopurines may reflect the greater frequency of this MRP4 SNP. [Cancer Res 2008;68(13):4983–9]

Supplementary Figure 1 from Transporter-Mediated Protection against Thiopurine-Induced Hematopoietic Toxicity

Supplementary Table 1 from Transporter-Mediated Protection against Thiopurine-Induced Hematopoietic Toxicity

Glucocorticoids (GCs) are a mainstay of contemporary, multi-drug chemotherapy in the treatment of childhood acute lymphoblastic leukemia (ALL), and resistance to GCs remains a major clinical concern. Resistance to GCs is predictive of ALL relapse and poor clinical outcome, and therefore represents a major hurdle limiting further improvements in survival rates. While advances have been made in identifying genes implicated in GC resistance, there remains an insufficient understanding of the impact of cis-regulatory disruptions in resistance. To address this, we mapped the gene regulatory response to GCs in two ALL cells using functional genomics and high-throughput reporter assays and identified thousands of GC-responsive changes to chromatin state, including the formation of over 250 GC-responsive super-enhancers and a depletion of AP-1 bound cis-regulatory elements implicated in cell proliferation and anti-apoptotic processes. By integrating our GC response maps with genetic and epigenetic datasets in primary ALL cells from patients, we further uncovered cis-regulatory disruptions at GC-responsive genes that impact GC resistance in childhood ALL. Overall, these data indicate that GCs initiate pervasive effects on the leukemia epigenome, and that alterations to the GC gene regulatory network contribute to GC resistance.

Inherited non-coding genetic variants confer significant disease susceptibility in many cancers. However, the molecular processes of by which germline variants contribute to somatic lesions are poorly understood. We performed targeted sequencing in 5,008 patients and identified a key regulatory germline variant in GATA3 strongly associated with Philadelphia chromosome-like acute lymphoblastic leukemia (Ph-like ALL). By creating an isogenic cellular model with CRISPR-Cas9 system, we showed that this variant activated a strong enhancer that significantly upregulated GATA3 transcription, which in turn reshaped the global chromatin accessibility and 3D genome organization. Remarkably, this genotype switch induced a chromatin loop between the CRLF2 oncogene and a distal enhancer, similar to the somatically acquired super-enhancer hijacking event in patients. GATA3 genotype-related alterations in transcriptional control and 3D chromatin organization were further validated in Ph-like ALL patients. Finally, we showed that GATA3 directly regulates CRLF2 and potentiates the oncogenic effects of JAK-STAT signaling in leukemogenesis. Altogether, our results provide evidence for a novel mechanism by which a germline non-coding variant contributes to oncogene activation epigenetic regulation and 3D genome reprogramming. Citation Format: Hongbo Yang, Hui Zhang, Yu Luan, Tingting Liu, Kathryn Roberts, Mao-xiang Qian, Bo Zhang, Wenjian Yang, Virginia Perez-Andreu, Jie Xu, Sriranga lyyanki, Da Kuang, Shalini Reshmi, Julie Gastier-Foster, Colton Smith, Ching-Hon Pui, William Evans, Stephen Hunger, Stephen Hunger, Leonidas Platanias, Mary Relling, Charles Mullighan, Mignon Loh, Feng Yue, Jun Yang. Non-coding germline GATA3 variants alter chromatin topology and contribute to pathogenesis of acute lymphoblastic leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2118.