A functional genomic approach to actionable gene fusions for precision oncology.

Fusion genes represent a class of attractive therapeutic targets. Thousands of fusion genes have been identified in patients with cancer, but the functional consequences and therapeutic implications of most of these remain largely unknown. Here, we develop a functional genomic approach that consists of efficient fusion reconstruction and sensitive cell viability and drug response assays. Applying this approach, we characterize ~100 fusion genes detected in patient samples of The Cancer Genome Atlas, revealing a notable fraction of low-frequency fusions with activating effects on tumor growth. Focusing on those in the RTK-RAS pathway, we identify a number of activating fusions that can markedly affect sensitivity to relevant drugs. Last, we propose an integrated, level-of-evidence classification system to prioritize gene fusions systematically. Our study reiterates the urgent clinical need to incorporate similar functional genomic approaches to characterize gene fusions, thereby maximizing the utility of gene fusions for precision oncology.

Pan-TGFß inhibition by SAR439459 relieves immunosuppression and improves antitumor efficacy of PD-1 blockade.

TGFß is a pleiotropic cytokine that may have both tumor inhibiting and tumor promoting properties, depending on tissue and cellular context. Emerging data support a role for TGFß in suppression of antitumor immunity. Here we show that SAR439459, a pan-TGFß neutralizing antibody, inhibits all active isoforms of human and murine TGFß, blocks TGFß-mediated pSMAD signaling, and TGFß-mediated suppression of T cells and NK cells. In vitro, SAR439459 synergized with anti-PD1 to enhance T cell responsiveness. In syngeneic tumor models, SAR439459 treatment impaired tumor growth, while the combination of SAR439459 with anti-PD-1 resulted in complete tumor regression and a prolonged antitumor immunity. Mechanistically, we found that TGFß inhibition with PD-1 blockade augmented intratumoral CD8+ T cell proliferation, reduced exhaustion, evoked proinflammatory cytokines, and promoted tumor-specific CD8+ T cell responses. Together, these data support the hypothesis that TGFß neutralization using SAR439459 synergizes with PD-1 blockade to promote antitumor immunity and formed the basis for the ongoing clinical investigation of SAR439459 in patients with cancer (NCT03192345).

Global impact of somatic structural variation on the DNA methylome of human cancers.

BACKGROUND: Genomic rearrangements exert a heavy influence on the molecular landscape of cancer. New analytical approaches integrating somatic structural variants (SSVs) with altered gene features represent a framework by which we can assign global significance to a core set of genes, analogous to established methods that identify genes non-randomly targeted by somatic mutation or copy number alteration. While recent studies have defined broad patterns of association involving gene transcription and nearby SSV breakpoints, global alterations in DNA methylation in the context of SSVs remain largely unexplored. RESULTS: By data integration of whole genome sequencing, RNA sequencing, and DNA methylation arrays from more than 1400 human cancers, we identify hundreds of genes and associated CpG islands (CGIs) for which the nearby presence of a somatic structural variant (SSV) breakpoint is recurrently associated with altered expression or DNA methylation, respectively, independently of copy number alterations. CGIs with SSV-associated increased methylation are predominantly promoter-associated, while CGIs with SSV-associated decreased methylation are enriched for gene body CGIs. Rearrangement of genomic regions normally having higher or lower methylation is often involved in SSV-associated CGI methylation alterations. Across cancers, the overall structural variation burden is associated with a global decrease in methylation, increased expression in methyltransferase genes and DNA damage response genes, and decreased immune cell infiltration. CONCLUSION: Genomic rearrangement appears to have a major role in shaping the cancer DNA methylome, to be considered alongside commonly accepted mechanisms including histone modifications and disruption of DNA methyltransferases.

A Pan-Cancer Compendium of Genes Deregulated by Somatic Genomic Rearrangement across More Than 1,400 Cases.

A systematic cataloging of genes affected by genomic rearrangement, using multiple patient cohorts and cancer types, can provide insight into cancer-relevant alterations outside of exomes. By integrative analysis of whole-genome sequencing (predominantly low pass) and gene expression data from 1,448 cancers involving 18 histopathological types in The Cancer Genome Atlas, we identified hundreds of genes for which the nearby presence (within 100 kb) of a somatic structural variant (SV) breakpoint is associated with altered expression. While genomic rearrangements are associated with widespread copy-number alteration (CNA) patterns, approximately 1,100 genes-including overexpressed cancer driver genes (e.g., TERT, ERBB2, CDK12, CDK4) and underexpressed tumor suppressors (e.g., TP53, RB1, PTEN, STK11)-show SV-associated deregulation independent of CNA. SVs associated with the disruption of topologically associated domains, enhancer hijacking, or fusion transcripts are implicated in gene upregulation. For cancer-relevant pathways, SVs considerably expand our understanding of how genes are affected beyond point mutation or CNA.

Dual MAPK Inhibition Is an Effective Therapeutic Strategy for a Subset of Class II BRAF Mutant Melanomas.

PURPOSE: Dual MAPK pathway inhibition (dMAPKi) with BRAF and MEK inhibitors improves survival in BRAF V600E/K mutant melanoma, but the efficacy of dMAPKi in non-V600 BRAF mutant tumors is poorly understood. We sought to characterize the responsiveness of class II (enhanced kinase activity, dimerization dependent) BRAF mutant melanoma to dMAPKi. EXPERIMENTAL DESIGN: Tumors from patients with BRAF wild-type (WT), V600E (class I), and L597S (class II) metastatic melanoma were used to generate patient-derived xenografts (PDX). We assembled a panel of melanoma cell lines with class IIa (activation segment) or IIb (p-loop) mutations and compared these with WT or V600E/K BRAF mutant cells. Cell lines and PDXs were treated with BRAFi (vemurafenib, dabrafenib, encorafenib, and LY3009120), MEKi (cobimetinib, trametinib, and binimetinib), or the combination. We identified 2 patients with BRAF L597S metastatic melanoma who were treated with dMAPKi. RESULTS: BRAFi impaired MAPK signaling and cell growth in class I and II BRAF mutant cells. dMAPKi was more effective than either single MAPKi at inhibiting cell growth in all class II BRAF mutant cells tested. dMAPKi caused tumor regression in two melanoma PDXs with class II BRAF mutations and prolonged survival of mice with class II BRAF mutant melanoma brain metastases. Two patients with BRAF L597S mutant melanoma clinically responded to dMAPKi. CONCLUSIONS: Class II BRAF mutant melanoma is growth inhibited by dMAPKi. Responses to dMAPKi have been observed in 2 patients with class II BRAF mutant melanoma. These data provide rationale for clinical investigation of dMAPKi in patients with class II BRAF mutant metastatic melanoma.See related commentary by Johnson and Dahlman, p. 6107.

Discordant inheritance of chromosomal and extrachromosomal DNA elements contributes to dynamic disease evolution in glioblastoma.

To understand how genomic heterogeneity of glioblastoma (GBM) contributes to poor therapy response, we performed DNA and RNA sequencing on GBM samples and the neurospheres and orthotopic xenograft models derived from them. We used the resulting dataset to show that somatic driver alterations including single-nucleotide variants, focal DNA alterations and oncogene amplification on extrachromosomal DNA (ecDNA) elements were in majority propagated from tumor to model systems. In several instances, ecDNAs and chromosomal alterations demonstrated divergent inheritance patterns and clonal selection dynamics during cell culture and xenografting. We infer that ecDNA was unevenly inherited by offspring cells, a characteristic that affects the oncogenic potential of cells with more or fewer ecDNAs. Longitudinal patient tumor profiling found that oncogenic ecDNAs are frequently retained throughout the course of disease. Our analysis shows that extrachromosomal elements allow rapid increase of genomic heterogeneity during GBM evolution, independently of chromosomal DNA alterations.

Genomic Profiling on an Unselected Solid Tumor Population Reveals a Highly Mutated Wnt/ß-Catenin Pathway Associated with Oncogenic EGFR Mutations.

Oncogenic epidermal growth factor receptors (EGFRs) can recruit key effectors in diverse cellular processes to propagate oncogenic signals. Targeted and combinational therapeutic strategies have been successfully applied for treating EGFR-driven cancers. However, a main challenge in EGFR therapies is drug resistance due to mutations, oncogenic shift, alternative signaling, and other potential mechanisms. To further understand the genetic alterations associated with oncogenic EGFRs and to provide further insight into optimal and personalized therapeutic strategies, we applied a proprietary comprehensive next-generation sequencing (NGS)-based assay of 435 genes to systematically study the genomic profiles of 1565 unselected solid cancer patient samples. We found that activating EGFR mutations were predominantly detected in lung cancer, particularly in non-small cell lung cancer (NSCLC). The mutational landscape of EGFR-driven tumors covered most key signaling pathways and biological processes. Strikingly, the Wnt/ß-catenin pathway was highly mutated (48 variants detected in 46% of the EGFR-driven tumors), and its variant number topped that in the TP53/apoptosis and PI3K-AKT-mTOR pathways. Furthermore, an analysis of mutation distribution revealed a differential association pattern of gene mutations between EGFR exon 19del and EGFR L858R. Our results confirm the aggressive nature of the oncogenic EGFR-driven tumors and reassure that a combinational strategy should have advantages over an EGFR-targeted monotherapy and holds great promise for overcoming drug resistance.

Comprehensive genomic sequencing detects important genetic differences between right-sided and left-sided colorectal cancer.

OBJECTIVES: Anti-epidermal growth factor receptor (EGFR) therapy has been found to be more effective against left-sided colorectal cancer (LCRC) than right-sided colorectal cancer (RCRC). We hypothesized that RCRC is more likely to harbor genetic alterations associated with resistance to anti-EGFR therapy and tested this using comprehensive genomic sequencing. MATERIALS AND METHODS: A total of 201 patients with either primary RCRC or LCRC were analyzed. We investigated tumors for genetic alterations using a 415-gene panel, which included alterations associated with resistance to anti-EGFR therapy: TK receptors (ERBB2, MET, EGFR, FGFR1, and PDGFRA), RAS pathway (KRAS, NRAS, HRAS, BRAF, and MAPK2K1), and PI3K pathway (PTEN and PIK3CA). Patients whose tumors had no alterations in these 12 genes, theoretically considered to respond to anti-EGFR therapy, were defined as "all wild-type", while remaining patients were defined as "mutant-type". RESULTS: Fifty-six patients (28%) and 145 patients (72%) had RCRC and LCRC, respectively. Regarding genetic alterations associated with anti-EGFR therapy, only 6 of 56 patients (11%) with RCRC were "all wild-type" compared with 41 of 145 patients (28%) with LCRC (P = 0.009). Among the 49 patients who received anti-EGFR therapy, RCRC showed significantly worse progression-free survival (PFS) than LCRC (P = 0.022), and "mutant-type" RCRC showed significantly worse PFS compared with "all wild-type" LCRC (P = 0.004). CONCLUSIONS: RCRC is more likely to harbor genetic alterations associated with resistance to anti-EGFR therapy compared with LCRC. Furthermore, our data shows primary tumor sidedness is a surrogate for the non-random distribution of genetic alterations in CRC.

Clinical application of a cancer genomic profiling assay to guide precision medicine decisions.

AIM: Develop and apply a comprehensive and accurate next-generation sequencing based assay to help clinicians to match oncology patients to therapies. MATERIALS & METHODS: The performance of the CANCERPLEX® assay was assessed using DNA from well-characterized routine clinical formalin-fixed paraffin-embedded (FFPE) specimens and cell lines. RESULTS: The maximum sensitivity of the assay is 99.5% and its accuracy is virtually 100% for detecting somatic alterations with an allele fraction of as low as 10%. Clinically actionable variants were identified in 93% of patients (930 of 1000) who underwent testing. CONCLUSION: The test's capacity to determine all of the critical genetic changes, tumor mutation burden, microsatellite instability status and viral associations has important ramifications on clinical decision support strategies, including identification of patients who are likely to benefit from immune checkpoint blockage therapies.

PRKCI promotes immune suppression in ovarian cancer.

A key feature of high-grade serous ovarian carcinoma (HGSOC) is frequent amplification of the 3q26 locus harboring PRKC-ι (PRKCI). Here, we show that PRKCI is also expressed in early fallopian tube lesions, called serous tubal intraepithelial carcinoma. Transgenic mouse studies establish PRKCI as an ovarian cancer-specific oncogene. Mechanistically, we show that the oncogenic activity of PRKCI relates in part to the up-regulation of TNFα to promote an immune-suppressive tumor microenvironment characterized by an abundance of myeloid-derived suppressor cells and inhibition of cytotoxic T-cell infiltration. Furthermore, system-level and functional analyses identify YAP1 as a downstream effector in tumor progression. In human ovarian cancers, high PRKCI expression also correlates with high expression of TNFα and YAP1 and low infiltration of cytotoxic T cells. The PRKCI-YAP1 regulation of the tumor immunity provides a therapeutic strategy for highly lethal ovarian cancer.

Modeling Genomic Instability and Selection Pressure in a Mouse Model of Melanoma.

Tumor evolution is an iterative process of selection for pro-oncogenic aberrations. This process can be accelerated by genomic instability, but how it interacts with different selection bottlenecks to shape the evolving genomic landscape remains understudied. Here, we assessed tumor initiation and therapy resistance bottlenecks in mouse models of melanoma, with or without genomic instability. At the initiation bottleneck, whole-exome sequencing revealed that drug-naive tumors were genomically silent, and this was surprisingly unaffected when genomic instability was introduced via telomerase inactivation. We hypothesize that the strong engineered alleles created low selection pressure. At the therapy resistance bottleneck, strong selective pressure was applied using a BRAF inhibitor. In the absence of genomic instability, tumors acquired a non-genomic drug resistance mechanism. By contrast, telomerase-deficient, drug-resistant melanomas acquired highly recurrent copy number gains. These proof-of-principle experiments demonstrate how different selection pressures can interact with genomic instability to impact tumor evolution.

A Pan-Cancer Proteogenomic Atlas of PI3K/AKT/mTOR Pathway Alterations.

Molecular alterations involving the PI3K/AKT/mTOR pathway (including mutation, copy number, protein, or RNA) were examined across 11,219 human cancers representing 32 major types. Within specific mutated genes, frequency, mutation hotspot residues, in silico predictions, and functional assays were all informative in distinguishing the subset of genetic variants more likely to have functional relevance. Multiple oncogenic pathways including PI3K/AKT/mTOR converged on similar sets of downstream transcriptional targets. In addition to mutation, structural variations and partial copy losses involving PTEN and STK11 showed evidence for having functional relevance. A substantial fraction of cancers showed high mTOR pathway activity without an associated canonical genetic or genomic alteration, including cancers harboring IDH1 or VHL mutations, suggesting multiple mechanisms for pathway activation.

Systematic Epigenomic Analysis Reveals Chromatin States Associated with Melanoma Progression.

The extent and nature of epigenomic changes associated with melanoma progression is poorly understood. Through systematic epigenomic profiling of 35 epigenetic modifications and transcriptomic analysis, we define chromatin state changes associated with melanomagenesis by using a cell phenotypic model of non-tumorigenic and tumorigenic states. Computation of specific chromatin state transitions showed loss of histone acetylations and H3K4me2/3 on regulatory regions proximal to specific cancer-regulatory genes in important melanoma-driving cell signaling pathways. Importantly, such acetylation changes were also observed between benign nevi and malignant melanoma human tissues. Intriguingly, only a small fraction of chromatin state transitions correlated with expected changes in gene expression patterns. Restoration of acetylation levels on deacetylated loci by histone deacetylase (HDAC) inhibitors selectively blocked excessive proliferation in tumorigenic cells and human melanoma cells, suggesting functional roles of observed chromatin state transitions in driving hyperproliferative phenotype. Through these results, we define functionally relevant chromatin states associated with melanoma progression.

Utility of comprehensive genomic sequencing for detecting HER2-positive colorectal cancer.

HER2-targeted therapy is considered effective for KRAS codon 12/13 wild-type, HER2-positive metastatic colorectal cancer (CRC). In general, HER2 status is determined by the use of immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). Comprehensive genomic sequencing (CGS) enables the detection of gene mutations and copy number alterations including KRAS mutation and HER2 amplification; however, little is known about the utility of CGS for detecting HER2-positive CRC. To assess its utility, we retrospectively investigated 201 patients with stage I-IV CRC. The HER2 status of the primary site was assessed using IHC and FISH, and HER2 amplification of the primary site was also assessed using CGS, and the findings of these approaches were compared in each patient. CGS successfully detected alterations in 415 genes including KRAS codon 12/13 mutation and HER2 amplification. Fifty-nine (29%) patients had a KRAS codon 12/13 mutation. Ten (5%) patients were diagnosed as HER2 positive because of HER2 IHC 3+, and the same 10 (5%) patients had HER2 amplification evaluated using CGS. The results of HER2 status and HER2 amplification were completely identical in all 201 patients (P < .001). Nine of the 10 HER2-positive patients were KRAS 12/13 wild-type and were considered possible candidates for HER2-targeted therapy. CGS has the same utility as IHC and FISH for detecting HER2-positive patients who are candidates for HER2-targeted therapy, and facilitates precision medicine and tailor-made treatment.

Genomic landscape of colorectal cancer in Japan: clinical implications of comprehensive genomic sequencing for precision medicine.

BACKGROUND: Comprehensive genomic sequencing (CGS) has the potential to revolutionize precision medicine for cancer patients across the globe. However, to date large-scale genomic sequencing of cancer patients has been limited to Western populations. In order to understand possible ethnic and geographic differences and to explore the broader application of CGS to other populations, we sequenced a panel of 415 important cancer genes to characterize clinically actionable genomic driver events in 201 Japanese patients with colorectal cancer (CRC). METHODS: Using next-generation sequencing methods, we examined all exons of 415 known cancer genes in Japanese CRC patients (n = 201) and evaluated for concordance among independent data obtained from US patients with CRC (n = 108) and from The Cancer Genome Atlas-CRC whole exome sequencing (WES) database (n = 224). Mutation data from non-hypermutated Japanese CRC patients were extracted and clustered by gene mutation patterns. Two different sets of genes from the 415-gene panel were used for clustering: 61 genes with frequent alteration in CRC and 26 genes that are clinically actionable in CRC. RESULTS: The 415-gene panel is able to identify all of the critical mutations in tumor samples as well as WES, including identifying hypermutated tumors. Although the overall mutation spectrum of the Japanese patients is similar to that of the Western population, we found significant differences in the frequencies of mutations in ERBB2 and BRAF. We show that the 415-gene panel identifies a number of clinically actionable mutations in KRAS, NRAS, and BRAF that are not detected by hot-spot testing. We also discovered that 26% of cases have mutations in genes involved in DNA double-strand break repair pathway. Unsupervised clustering revealed that a panel of 26 genes can be used to classify the patients into eight different categories, each of which can optimally be treated with a particular combination therapy. CONCLUSIONS: Use of a panel of 415 genes can reliably identify all of the critical mutations in CRC patients and this information of CGS can be used to determine the most optimal treatment for patients of all ethnicities.

The SMARCA2/4 ATPase Domain Surpasses the Bromodomain as a Drug Target in SWI/SNF-Mutant Cancers: Insights from cDNA Rescue and PFI-3 Inhibitor Studies.

The SWI/SNF multisubunit complex modulates chromatin structure through the activity of two mutually exclusive catalytic subunits, SMARCA2 and SMARCA4, which both contain a bromodomain and an ATPase domain. Using RNAi, cancer-specific vulnerabilities have been identified in SWI/SNF-mutant tumors, including SMARCA4-deficient lung cancer; however, the contribution of conserved, druggable protein domains to this anticancer phenotype is unknown. Here, we functionally deconstruct the SMARCA2/4 paralog dependence of cancer cells using bioinformatics, genetic, and pharmacologic tools. We evaluate a selective SMARCA2/4 bromodomain inhibitor (PFI-3) and characterize its activity in chromatin-binding and cell-functional assays focusing on cells with altered SWI/SNF complex (e.g., lung, synovial sarcoma, leukemia, and rhabdoid tumors). We demonstrate that PFI-3 is a potent, cell-permeable probe capable of displacing ectopically expressed, GFP-tagged SMARCA2-bromodomain from chromatin, yet contrary to target knockdown, the inhibitor fails to display an antiproliferative phenotype. Mechanistically, the lack of pharmacologic efficacy is reconciled by the failure of bromodomain inhibition to displace endogenous, full-length SMARCA2 from chromatin as determined by in situ cell extraction, chromatin immunoprecipitation, and target gene expression studies. Furthermore, using inducible RNAi and cDNA complementation (bromodomain- and ATPase-dead constructs), we unequivocally identify the ATPase domain, and not the bromodomain of SMARCA2, as the relevant therapeutic target with the catalytic activity suppressing defined transcriptional programs. Taken together, our complementary genetic and pharmacologic studies exemplify a general strategy for multidomain protein drug-target validation and in case of SMARCA2/4 highlight the potential for drugging the more challenging helicase/ATPase domain to deliver on the promise of synthetic-lethality therapy.

Utilizing murine inducible telomerase alleles in the studies of tissue degeneration/regeneration and cancer.

Telomere dysfunction-induced loss of genome integrity and its associated DNA damage signaling and checkpoint responses are well-established drivers that cause tissue degeneration during ageing. Cancer, with incidence rates greatly increasing with age, is characterized by short telomere lengths and high telomerase activity. To study the roles of telomere dysfunction and telomerase reactivation in ageing and cancer, the protocol shows how to generate two murine inducible telomerase knock-in alleles 4-Hydroxytamoxifen (4-OHT)-inducible TERT-Estrogen Receptor (mTERT-ER) and Lox-Stopper-LoxTERT (LSL-mTERT). The protocol describes the procedures to induce telomere dysfunction and reactivate telomerase activity in mTERT-ER and LSL-mTERT mice in vivo. The representative data show that reactivation of telomerase activity can ameliorate the tissue degenerative phenotypes induced by telomere dysfunction. In order to determine the impact of telomerase reactivation on tumorigenesis, we generated prostate tumor model G4 PB-Cre4 Pten(L/L) p53(L/L) LSL-mTERT(L/L) and thymic T-cell lymphoma model G4 Atm(-/-) mTERT(ER/ER). The representative data show that telomerase reactivation in the backdrop of genomic instability induced by telomere dysfunction can greatly enhance tumorigenesis. The protocol also describes the procedures used to isolate neural stem cells (NSCs) from mTERT-ER and LSL-mTERT mice and reactivate telomerase activity in NSCs in vitro. The representative data show that reactivation of telomerase can enhance the self-renewal capability and neurogenesis in vitro. Finally, the protocol describes the procedures for performing telomere FISH (Fluorescence In Situ Hybridization) on both mouse FFPE (Formalin Fixed and Paraffin Embedded) brain tissues and metaphase chromosomes of cultured cells.

Co-clinical assessment identifies patterns of BRAF inhibitor resistance in melanoma.

Multiple mechanisms have been described that confer BRAF inhibitor resistance to melanomas, yet the basis of this resistance remains undefined in a sizable portion of patient samples. Here, we characterized samples from a set of patients with melanoma that included individuals at baseline diagnosis, on BRAF inhibitor treatment, and with resistant tumors at both the protein and RNA levels. Using RNA and DNA sequencing, we identified known resistance-conferring mutations in 50% (6 of 12) of the resistant samples. In parallel, targeted proteomic analysis by protein array categorized the resistant samples into 3 stable groups, 2 of which were characterized by reactivation of MAPK signaling to different levels and 1 that was MAPK independent. The molecular relevance of these classifications identified in patients was supported by both mutation data and the similarity of resistance patterns that emerged during a co-clinical trial in a genetically engineered mouse (GEM) model of melanoma that recapitulates the development of BRAF inhibitor resistance. Additionally, we defined candidate biomarkers in pre- and early-treatment patient samples that have potential for predicting clinical responses. On the basis of these observations, we suggest that BRAF inhibitor-resistant melanomas can be actionably classified using protein expression patterns, even without identification of the underlying genetic alteration.

Identification of a novel fusion gene, IRF2BP2-RARA, in acute promyelocytic leukemia.

Acute promyelocytic leukemia (APL) is characterized by the fusion of retinoic acid receptor alpha (RARA) with promyelocytic leukemia (PML) or, rarely, other gene partners. This report presents a patient with APL with a novel fusion between RARA and the interferon regulatory factor 2 binding protein 2 (IRF2BP2) genes. A bone marrow examination in a 19-year-old woman who presented with ecchymoses and epistaxis showed morphologic and immunophenotypic features consistent with APL. PML oncogenic domain antibody was positive. Results of fluorescence in situ hybridization, conventional cytogenetics, reverse transcription-polymerase chain reaction (RT-PCR), and oligonucleotide microarray for PML-RARA and common APL variant translocations were negative. Next-generation RNA-sequencing analysis followed by RT-PCR and direct sequencing revealed distinct breakpoints within IRF2BP2 exon 2 and RARA intron 2. The patient received all-trans retinoic acid, arsenic, and gemtuzumab ozogamicin, and achieved complete remission. However, the disease relapsed 10 months later, 2 months after consolidation therapy. This is the first report showing involvement of IRF2BP2 in APL, and it expands the list of novel RARA partners identified in APL.