PDGFRB Fusion Research Articles

Recurrent CLTC::SYK fusions and CSF1R mutations in juvenile xanthogranuloma of soft tissue

AbstractJuvenile xanthogranuloma (JXG) is a histiocytic neoplasm that usually presents in the skin. Rarely, extracutaneous localizations occur; the genetic drivers of this clinical variant of JXG remain incompletely characterized. We present detailed clinicopathologic and molecular data of 16 children with extracutaneous JXG and 5 adults with xanthogranulomas confined to the central nervous system (CNS) or soft tissue. Tissue samples were obtained through the Dutch Nationwide Pathology Databank and analyzed with an innovative sequencing technique capable of detecting both small genomic variants and gene rearrangements. Targetable kinase alterations were detected in 16 of 16 children and 1 of 5 adults. Alterations included CLTC::SYK fusions in 6 children and CSF1R mutations in 7 others; all aged <2 years with soft tissue tumors. One child had a CSF1R mutation and MRC1::PDGFRB fusion. Most were treated surgically, although spontaneous regression occurred in 1 of 6 with CLTC::SYK and 2 of 7 with CSF1R mutations, underscoring that treatment is not always necessary. Tumors with CLTC::SYK fusions generally lacked Touton giant cells but exhibited many other histologic features of JXG and concordant methylation profiles. Using multispectral immunofluorescence, phosphorylated–spleen tyrosine kinase expression was localized to CD163+ histiocytes; tumors with CLTC::SYK fusions also demonstrated mTOR activation, cyclin D1 expression, and variable phosphorylated–extracellular signal-regulated kinase expression. BRAFV600E was detected in 1 child and 1 adult with CNS xanthogranulomas; both responded to BRAF inhibition. Finally, a TPM3::NTRK1 fusion or MAP2K1 deletion was detected in 2 children with systemic JXG who experienced spontaneous disease regression. This study advances the molecular understanding of histiocytic neoplasms and may guide diagnostics and clinical management.

Delineation of features, responses, outcomes, and prognostic factors in pediatric PDGFRB-positive acute lymphoblastic leukemia patients: A retrospective multicenter study.

PDGFRB fusions in acute lymphoblastic leukemia (ALL) is rare. The authors identified 28 pediatric PDGFRB-positive ALL. They analyzed the features, outcomes, and prognostic factors of this disease. This multicenter, retrospective study included 6457 pediatric patients with newly diagnosed PDGFRB fusion ALL according to the CCCG-ALL-2015 and CCCG-ALL-2020 protocols from April 2015 to April 2022 in 20 hospitals in China. Of these patients, 3451 were screened for PDGFRB fusions. Pediatric PDGFRB-positive ALL accounted for only 0.8% of the 3451 cases tested for PDGFRB. These patients included 21 males and seven females and 24 B-ALL and 4 T-ALL; the median age was 10 years; and the median leukocyte count was 29.8 × 109/L at baseline. Only one patient had eosinophilia. Three patients had an IKZF1 deletion, three had chromosome 5q31-33 abnormalities, and one suffered from a complex karyotype. The 3-year event-free survival (EFS), overall survival (OS), and cumulative incidence of relapse (CIR) were 33.1%, 65.5%, and 32.1%, respectively, with a median follow-up of 25.5 months. Twenty patients were treated with chemotherapy plus tyrosine-kinase inhibitors (TKIs) and eight were treated without TKI. Complete remission (CR) rates of them were 90.0% and 63.6%, respectively, but no differences in EFS, OS, or CIR. Univariate analyses showed patients with IKZF1 deletion or measurable residual disease (MRD) ≥0.01% after induction had inferior outcomes (p<.05). Pediatric PDGFRB-positive ALL has a poor outcome associated with high-risk features. Chemotherapy plus TKIs can improve the CR rate, providing an opportunity for lower MRD levels and transplantation. MRD ≥0.01% was a powerful adverse prognostic factor, and stratified treatment based on MRD may improve survival for these patients. Pediatric acute lymphoblastic leukemia patients with PDGFRB fusions are associated with high-risk clinical features such as older age, high white blood cell count at diagnosis, high measurable residual disease after induction therapy, and increased risk of leukemia relapse. Chemotherapy plus tyrosine-kinase inhibitors can improve the complete remission rate and provide an opportunity for lower measurable residual disease (MRD) levels and transplantation for pediatric PDGFRB-positive acute lymphoblastic leukemia (ALL) patients. The MRD level was also a powerful prognostic factor for pediatric PDGFRB-positive ALL patients.

The novel TERF2::PDGFRB fusion gene enhances tumorigenesis via PDGFRB/STAT5 signalling pathways and sensitivity to TKI in ph-like ALL.

Patients with Philadelphia chromosome-like acute lymphoblastic leukaemia (Ph-like ALL) often face a grim prognosis, with PDGFRB gene fusions being commonly detected in this subgroup. Our study has unveiled a newfound fusion gene, TERF2::PDGFRB, and we have found that patients carrying this fusion gene exhibit sensitivity to dasatinib. Ba/F3 cells harbouring the TERF2::PDGFRB fusion display IL-3-independent cell proliferation through activation of the p-PDGFRB and p-STAT5 signalling pathways. These cells exhibit reduced apoptosis and demonstrate sensitivity to imatinib invitro. When transfused into mice, Ba/F3 cells with the TERF2::PDGFRB fusion gene induce tumorigenesis and a shortened lifespan in cell-derived graft models, but this outcome can be improved with imatinib treatment. In summary, we have identified the novel TERF2::PDGFRB fusion gene, which exhibits oncogenic potential both invitro and invivo, making it a potential therapeutic target for tyrosine kinase inhibitors (TKIs).

Promoter swapping of truncated PDGFRB drives Ph-like acute lymphoblastic leukemia

Philadelphia chromosome (Ph)-like acute lymphoblastic leukemia (ALL) is a subset of ALL that demonstrated a high treatment failure rate. One of the hallmarks of Ph-like ALL is PDGFRB gene fusion, with fusion partner proteins often harboring dimerization domains and enhancing the kinase activity of PDGFRB. We determined a novel oncogenic PDGFRB fusion gene, NRIP1::PDGFRB, from a pediatric patient with ALL, encoding a protein with the carboxy-terminal kinase domain of PDGFRB, without the partner peptide. We confirmed the oncogenic potential of NRIP1::PDGFRB in vitro and the efficacy of all ABL1-specific inhibitor generations, including imatinib, dasatinib, nilotinib, and ponatinib, in suppressing this potential. PDGFRB activation mechanism may include juxtamembrane domain truncation in the predicted peptide. In conclusion, we determined a novel fusion gene pattern in Ph-like ALL.

Feasibility and Outcome of Post-Induction Therapy Incorporating Dasatinib for Patients with Newly Diagnosed ABL-Class Fusion B-Lymphoblastic Leukemia (ABL-class Fusion B-ALL): Children's Oncology Group AALL1131

Background: Approximately 2-3% of B-ALL patients have Philadelphia chromosome like (Ph-like) B-ALL with an ABL-class fusion ( ABL1, ABL2, PDGFRB, or CSF1R fusions), but lacking BCR::ABL1. These patients are predicted to be sensitive to ABL-class tyrosine kinase inhibitors, such as imatinib or dasatinib. Ph-like B-ALL is associated with male sex, older age, higher initial white blood cell (WBC) count, elevated end of induction (EOI) minimal residual disease (MRD), and poor outcome. The AALL1131 Dasatinib arm was designed to evaluate response to therapy following induction of patients with ABL-class fusion B-ALL, when given dasatinib continuously on a modified Berlin-Frankfurt-Münster (MBFM) backbone. Methods: Between February 2012 and March 2019, AALL1131 enrolled patients 1-30 years with newly diagnosed high risk (HR) B-ALL. AALL1131 was amended in August 2016, to include the Dasatinib arm for patients with ABL-class fusions involving ABL1, ABL2, PDGFRB, and CSF1R. Patients with Down Syndrome were not eligible for the Dasatinib arm. ABL-class fusion B-ALL patients with a predicted TKI-sensitive mutation were screened by low density array (LDA) PCR and confirmed by additional molecular testing. Following 4-drug induction, patients with HR B-ALL and ABL-class fusions received MBFM with dasatinib (60 mg/m 2, maximum 140 mg) daily from start of consolidation through end of maintenance. Dasatinib was held only for toxicity. Results: Twenty-two evaluable patients with HR B-ALL were non-randomly assigned to the Dasatinib arm. ABL-class 3' partners included ABL1 (n=4), ABL2 (n=4), PDGFRB (n=12), and CSF1R (n=2). Compared to all patients on AALL1131, patients treated on the Dasatinib arm were older (median age at diagnosis 14 versus 10 years, p=0.008), more often male (77% versus 56%, p=0.046), and had a trend towards an increased initial WBC (median WBC at diagnosis 44,000/µL versus 19,000/µL, p=0.086). Patients with ABL-class fusions had EOI MRD &amp;gt; 0.01% in 20 of 22 patients and available end of consolidation MRD &amp;gt;0.01% in 6 of 10 patients. Only 5 of 22 (22.7%) patients completed prescribed protocol therapy. Reasons for therapy discontinuation included induction failure (n=2), relapse (n=1), alternate therapy (n=1), determined to be in the patient's best interest (n=10), death (n=1), unknown (n=2). Compared to all patients on AALL1131, four-year disease-free survival was 52.5+18.1% versus 86.8+0.7% (p&amp;lt;0.0001) and overall survival 79.4+13.6% versus 89.2+0.4% (p&amp;lt;0.0001). Dasatinib was well tolerated with no unexpected treatment related toxicities. Conclusion: Patients with ABL-class fusions were more likely male, EOI MRD+, and had poorer outcomes. Seventy-seven percent of patients enrolled on the Dasatinib arm did not complete prescribed therapy. While dasatinib was well tolerated, treatment failures occurred early, indicating alternate strategies are needed.

Targeting hyperactive platelet-derived growth factor receptor-β signaling in T-cell acute lymphoblastic leukemia and lymphoma.

T-cell acute lymphoblastic leukemia (T-ALL) and T-cell lymphoblastic lymphoma (T-LBL) are rare aggressive hematologic malignancies. Current treatment consists of intensive chemotherapy leading to 80% overall survival but is associated with severe toxic side effects. Furthermore, 10-20% of patients still die from relapsed or refractory disease providing a strong rationale for more specific, targeted therapeutic strategies with less toxicities. Here, we report a novel MYH9::PDGFRB fusion in a T-LBL patient, and demonstrate that this fusion product is constitutively active and sufficient to drive oncogenic transformation in vitro and in vivo. Expanding our analysis more broadly across T-ALL, we found a T-ALL cell line and multiple patient-derived xenograft models with PDGFRB hyperactivation in the absence of a fusion, with high PDGFRB expression in TLX3 and HOXA T-ALL molecular subtypes. To target this PDGFRB hyperactivation, we evaluated the therapeutic effects of a selective PDGFRB inhibitor, CP-673451, both in vitro and in vivo and demonstrated sensitivity if the receptor is hyperactivated. Altogether, our work reveals that hyperactivation of PDGFRB is an oncogenic driver in T-ALL/T-LBL, and that screening T-ALL/T-LBL patients for phosphorylated PDGFRB levels can serve as a biomarker for PDGFRB inhibition as a novel targeted therapeutic strategy in their treatment regimen.

Myeloid/lymphoid neoplasms with eosinophilia and tyrosine kinase gene fusions: reevaluation of the defining characteristics in a registry-based cohort

In a registry-based analysis of 135 patients with “myeloid/lymphoid neoplasms with eosinophilia and tyrosine kinase gene fusions” (MLN-TK; FIP1L1::PDGFRA, n = 78; PDGFRB, diverse fusions, n = 26; FGFR1, diverse, n = 9; JAK2, diverse, n = 11; ETV6::ABL1, n = 11), we sought to evaluate the disease-defining characteristics. In 81/135 (60%) evaluable patients, hypereosinophilia (>1.5 × 109/l) was observed in 40/44 (91%) FIP1L1::PDGFRA and 7/7 (100%) ETV6::ABL1 positive patients but only in 13/30 (43%) patients with PDGFRB, FGFR1, and JAK2 fusion genes while 9/30 (30%) patients had no eosinophilia. Monocytosis >1 × 109/l was identified in 27/81 (33%) patients, most frequently in association with hypereosinophilia (23/27, 85%). Overall, a blast phase (BP) was diagnosed in 38/135 (28%) patients (myeloid, 61%; lymphoid, 39%), which was at extramedullary sites in 18 (47%) patients. The comparison between patients with PDGFRA/PDGFRB vs. FGFR1, JAK2, and ETV6::ABL1 fusion genes revealed a similar occurrence of primary BP (17/104, 16% vs. 8/31 26%, p = 0.32), a lower frequency (5/87, 6% vs. 8/23, 35%, p = 0.003) of and a later progression (median 87 vs. 19 months, p = 0.053) into secondary BP, and a better overall survival from diagnosis of BP (17.1 vs. 1.7 years, p < 0.0008). We conclude that hypereosinophilia with or without monocytosis and various phenotypes of BP occur at variable frequencies in MLN-TK.

Unusual PDGFRB fusion reveals novel mechanism of kinase activation in Ph-like B-ALL

Unusual PDGFRB fusion reveals novel mechanism of kinase activation in Ph-like B-ALL

Pediatric Acute Lymphoblastic Leukemia with Pdgfrb Fusions: A Multicenter Retrospective Study

Objective: PDGFRB fusions in acute lymphoblastic leukemia (ALL) is rare. PDGFRB fusions was mainly observed in B-ALL, but also in T-ALL. Previous studies were mostly limited to anecdotal cases, suggesting that this subtype of ALL had low rate of remission and poor prognosis. The addition of tyrosine kinase inhibitor (TKI) to intensive chemotherapy may improve the outcome, but lack of long-term survival data. Herein, we retrospectively analyzed the clinical characteristics, survival and prognostic factors for 28 pediatric patients with PDGFRB fusions ALL. Methods: This multicenter, retrospective study included pediatric patients with newly diagnosed PDGFRB fusion ALL after treatments according to the CCCG-ALL-2015 and CCCG-ALL-2020 protocols from April 2015 to April 2022 in 10 hospitals in China. All patients completed the followed up. The baseline characteristics and treatment outcomes of these patients, including complete remission, drug resistance, relapse, treatment-related mortality and minimal residual disease (MRD) at end of induction therapy, as well as hematopoietic stem cell transplantation (HSCT) were collected. The outcome was compared between patients treated with and without TKI. Fisher test and nonparametric rank sum test were used for comparative analysis. 4-year event-free survival (EFS) and 4-year overall survival (OS) were estimated by use of Kaplan-Meier methods, and the 4-year cumulative incidence of relapse was calculated by use of a competing risk model gray test. Univariate and multivariate analyses were performed by use of Cox regression. Results: We identified 28 pediatric patients with newly diagnosed PDGFRB fusion ALL, which accounted for approximately 0.5% of childhood ALL. In all 28 patients, 21 (75%) were male and 7 (25%) were female; the median age was 10 (1-16) years old; median white blood cell (WBC) was 29.8 (1.6-627.7) × 10^9/L; 24 (85.7%) were acute B-lymphoblastic leukemia and 4 (14.3%) were acute T-lymphoblastic leukemia; one case (3.5%) was observed accompanied by the CNS involvement at diagnosis. Twenty of the patients received TKI treatment. Only 6 patients underwent hematopoietic stem cell transplantation. For all 28 patients, the median follow-up was 25.5 (3-77) months, the 4-year EFS was 29.2% (95% CI, 13.6-36.4), the 4-year OS was 42.1% (95% CI, 30.6-61.4), and the 4-year cumulative incidence of relapse was 58% (95% CI, 45·6-70·1). Patients in TKI group had a high rate of complete remission after induction therapy compared with those in no-TKI group (90% vs 63.6%). However, no significant differences were observed in the 4-year EFS and 4-year OS between TKI group and no-TKI group (EFS, 21.1% vs 43.8%, P=0.925; OS, 50.1% vs 36.5%, P=0.807). The cumulative incidences of relapse between these two groups also have no remarkable differences (P >0.05). Among the 6 patients who underwent HSCT, 2 patients who achieved CR1 survived and 4 died. Univariate analysis showed that minimal residual disease (MRD) ≥10-4 after induction therapy for D46 and IKZF1 positive had a poor prognosis (all P <0.05). MRD≥10-4 at the end of induction therapy is an independent risk factor for poor prognosis (hazard ratio of event-free, 9.101 [95% CI 2.03-40.86], P =0·004, compared to those with a MRD of <10-4). Conclusion: Pediatric ALL patients with PDGFRB fusions have a poor outcome. This subtype of ALL is associated with high-risk clinical features such as older age, high WBC count at diagnosis, high MRD after induction therapy, and increased risk of leukemia relapse. The addition of TKI to chemotherapy can improve the early remission rate of treatment, but does not improve the long-term survival. Profitably, the TKI treatment can provide an opportunity for subsequent treatment, including HSCT. Additionally, HSCT has indeed potential to reduce the relapse rate for this subtype patients. HSCT is recommended for this subtype patients who achieve CR1. The MRD≥10-4 or more at the end of induction therapy was independently predictive factor for unfavorable outcome and stratified treatment based on MRD may improve survival of these patients. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

MPN-521 Somatic Mutational Landscape and Clinical Response to Upfront Targeted Tyrosine Kinase Inhibitors Among Patients With Myeloid and Lymphoid Neoplasms With Eosinophilia and Rearrangements of PDGFRA, PDGFRB, FGFR1, JAK2, or FLT3.

In 2016, WHO put together a category of myeloid and lymphoid neoplasms with eosinophilia (MLN-Eo) with rearrangement of PDGFRA, PDGFRB, FGFR1, or PCM1-JAK2. Gene fusions involving FLT3 and JAK2 are considered potential provisional entities. Here, we describe the mutational landscape of MLN-Eo and clinical outcomes of upfront targeted TK inhibitor (TKI) therapy. We retrospectively reviewed clinical and molecular data on patients with MLN-Eos. Next-generation sequencing was performed using a customized or commercially available panel. Twenty-three patients had PDGFRA fusion or activating mutations; 4 PDGFRB fusion; 8 FGFR1 fusion; 2 JAK2 fusion; and 4 FLT3 fusion. Twelve (92%) had at least one other coexisting mutation. At least 2 patients harbored TET2, RUNX1, TP53, and PTPN11 mutations. Pathogenic PTPN11 mutation was present in all activating PDGFRA mutations but absent in other cases. Among 23 patients with the chronic-phase disease, 14 (61%) received upfront TKI. None transformed into blastic phase. Six patients with disease in the de novo blastic phase received upfront TKI and achieved complete remission. Upfront TKI use was associated with longer overall survival (OS) in univariate analysis (HR 0.065, 95% CI, 0.008-0.50. P =.008). The improved OS remained significant in a multivariate analysis (HR 0.043, 95% CI, 0.003-0.63, p = 0.021) when adjusted for sex, age, hypereosinophilia, PDGFRA/B fusion, complex cytogenetics, the blastic- phase, and transplant status. Patients with MLN-Eo have high frequency of secondary somatic mutations like other myeloid malignancies. TKI therapies are associated with excellent outcomes and should be used in the upfront setting.

Myeloid neoplasm with ETV6::ACSl6 fusion: landscape of molecular and clinical features

ABSTRACT Objectives Since the publication of the third edition, the WHO classification of tumors of hematopoietic and lymphoid disorders has introduced the disease entity of ‘myeloid/lymphoid neoplasms with eosinophilia and PDGFRB rearrangement’, in which the most common chromosomal abnormality is t(5;12) (q32;p13.2), and this abnormality generates the ETV6::PDGFRB fusion gene. However, there have been patients with hematologic features and chromosomal abnormalities that are extremely similar to those carrying ETV6::PDGFRB fusion. These rare disorders harbor ETV6::ACSL6 fusion, and only sporadic cases have been reported at present. Methods We report a patient with chronic eosinophilic leukemia (CEL) carrying chromosome translocation t(5;12)(q32;p13.2), and we present the clinical features. In addition, we conducted a literature review to collect all reported cases and summarized the genetic and clinical profiling as well as the treatments and outcomes. Result In addition to our patient, a total of 19 cases have been previously reported, including 6 variants of ETV6::ACSL6 and 3 reciprocals. We identified a novel variant of the ETV6::ACSL6 transcript in our patient, and the breakpoint was flanked by exon 2 of ETV6 and exon 2 of ACSL6. The cellular morphology features consisted of myeloproliferative neoplasm (MPN); myelodysplastic/myeloproliferative neoplasm (MDS/MPN), specifically CEL; and acute myelocytic leukemia (AML). The treatments and outcomes varied greatly depending on the type of disease, although tyrosine kinase inhibitors (TKIs) were not effective. Conclusion In contrast to neoplasms with ETV6::PDGFRB fusion, myeloid neoplasms with ETV6::ACSL6 fusion have unique characteristics.

Histopathologic and Molecular Characterization of Uterine Leiomyoma-like Inflammatory Myofibroblastic Tumor: Comparison to Molecular Subtypes of Uterine Leiomyoma.

Uterine leiomyoma (UL) is a common benign neoplasm which can sometimes be difficult to differentiate from the uterine inflammatory myofibroblastic tumor (IMT) based on morphology alone. IMT is a myofibroblastic/fibroblastic neoplasm which has typically been considered to be rare in the uterus. Its clinical behavior is usually indolent although aggressive variants exist. The majority of IMTs harbor genomic rearrangement of anaplastic lymphoma kinase ( ALK ), while ALK fusion has not been thus far detected in ULs. We analyzed 2263 ULs of which 9 (0.4%) had tyrosine-kinase activation. Seven of the samples were ALK immunopositive: 6 had an ALK fusion gene and 1 overexpressed an ALK transcript skipping exons 2 to 3, Moreover, 1 sample had a RET , and 1 a PDGFRB fusion gene. While no recurrent somatic mutations were found, 1 patient had an ALK germline mutation. Seven tumors showed leiomyoma-like morphology, 1 tumor had slightly loose, and 1 fibrous growth pattern. Six tumors had mild to moderate lymphocyte infiltration, while no immune cell infiltration was detected in 3 cases. None of the tumors showed aggressive behavior. Except for strong ALK positivity (7/9 tumors) the protein expression profile of the tumors was identical to ULs and distinct from other mesenchymal uterine tumors. In gene expression level, these tumors and the known UL subclasses did not separate perfectly. However, vitamin C metabolism and epithelial-mesenchymal transition pathways were uniquely enriched in these lesions. The overall similarity of the analyzed tumors to UL raises the question whether an UL diagnosis would be more proper for a subset of uterine IMTs.

Ruxolitinib or Dasatinib in Combination with Chemotherapy for Patients with Relapsed/Refractory Philadelphia (Ph)-like Acute Lymphoblastic Leukemia: A Phase I-II Trial

Ruxolitinib or Dasatinib in Combination with Chemotherapy for Patients with Relapsed/Refractory Philadelphia (Ph)-like Acute Lymphoblastic Leukemia: A Phase I-II Trial

Uterine mesenchymal tumours: recent advances.

Almost all uterine mesenchymal tumours have been historically classified as either smooth muscle or endometrial stromal neoplasms. Recent application of molecular techniques has identified numerous lesions with distinctive genetic abnormalities and clinicopathological characteristics. Newly discovered uterine sarcoma subtypes include high-grade endometrial stromal sarcomas with BCOR genetic abnormalities, fibrosarcoma-like uterine sarcomas with NTRK rearrangements and COL1A-PDGFRB fusions, as well as undifferentiated uterine sarcomas with SMARCA4 mutations. Novel PLAG1 and PGR fusions have been identified in subsets of myxoid and epithelioid leiomyosarcomas. Some uterine tumours resembling ovarian sex-cord tumour harbour GREB1 and ESR1 rearrangements. Histological and immunophenotypical features as well as underlying genetic abnormalities defining these lesions are discussed.

A Fast and Accurate Diagnostic Method for Ph-like ALL Using the Ncounter System

Ph-like acute lymphoblastic leukemia (also known as BCR-ABL1-like ALL) is a new disease entity of B-cell ALL (B-ALL) that exhibits a mRNA expression profile similar to that of Philadelphia chromosome-positive ALL (Ph+ ALL). Ph-like signature is presumably driven by kinase-activating gene alterations. Thus, both gene expression pattern and DNA mutational status should be assessed to make a definitive diagnosis for Ph-like ALL. A variety of approaches combining multiple methods, including RNA sequencing (RNA-seq), Taqman low-density array (LDA), fluorescence in situ hybridization (FISH) and targeted DNA sequencing, are being tested; however, such multi-omics approaches are available only in limited institutions. Since Ph-like ALL patients generally exhibit poor response to standard chemotherapy, and tyrosine kinase inhibitors (TKIs) may benefit them when used in a timely manner, a fast, accurate and generalizable diagnostic method is critically needed. In the present study, we have developed a nCounter-based diagnostic method for Ph-like ALL and validated it using a cohort of Japanese adult B-ALL cases. To identify genes that are uniquely expressed (or not expressed) in Ph+ B-ALL, we first obtained publicly-available gene expression datasets comprising 1146 B-ALL cases and identified 82 differentially-expressed genes in Ph+ ALL cases. We then assessed expression levels of those genes in an independent cohort using the nCounter, which enables fast, sensitive and accurate RNA detection. We also tested whether nCounter-based methods can detect fusion transcripts relevant for Ph-like ALL pathogenesis using probes targeting ABL1, ABL2, CSF1R, PDGFRB, and JAK2. We analyzed 123 samples (Ph+ = 42, Ph- = 81, age 16 to 67) obtained from newly-diagnosed adult B-ALL patients enrolled in two clinical trials conducted by the Fukuoka Blood and Marrow Transplantation Group (FBMTG) (Nagafuji et al. Eur J Haematol 2019). Unsupervised hierarchical clustering successfully stratified 123 cases into two disease clusters: Ph+ and Ph- subgroups. As expected, Ph+ subgroup included almost all Ph+ ALL cases (40 out of 42 cases), while 18 out of 81 Ph- ALL cases (22%) were categorized into the Ph+ subgroup. We defined these cases as Ph-like ALL. To validate the nCounter-based Ph-like ALL classification, we performed RNA-seq and target-capture DNA sequencing of all Ph- ALL cases. As expected, we detected kinase-activating fusions/rearrangements, including CRLF2 rearrangements (7 cases), PDGFRB fusions (3 cases), JAK2 fusions (2 cases), EPOR rearrangements (2 cases), ABL1 fusion (1 case), and FLT3 internal tandem duplication (1 case) in 16 Ph-like ALL cases, while no genetic alterations were detected in 2 cases. Fusion genes involving PDGFRB were consistently detected by nCounter (3/3); however, detection of those involving JAK2 (1/2) and ABL1 (0/1) were inconsistent. JAK2 and/or RAS mutations were detected in 5 of 7 Ph-like ALL cases harboring CRLF2 rearrangements. Of note, CRLF2 protein expression was detected by FACS in all CRLF2-rearranged cases. We next assessed significance of the Ph-like signature on clinical outcomes using a cohort of 40 Ph- ALL cases, in which minimal/measurable residual disease (MRD) status, assessed by IgH and/or TCR rearrangements, as well as clinical data were available (Nagafuji et al. Eur J Haematol 2019). Ph-like ALL cases exclusively exhibited MRD positivity after induction therapy as compared to non-Ph-like cases (p=0.04), indicative of the chemo-resistant nature of Ph-like ALL as previously reported (Roberts et al. N Engl J Med, 2014 and Roberts et al. J Clin Oncol, 2017). As expected, Ph-like ALL cases exhibited significantly poor disease-free survival compared with non-Ph-like ALL cases (p=0.04); however, no significant difference was evident in overall survival (p=0.62) presumably due to the fact that all MRD-positive cases were subjected to allo-HSCT after induction therapy. These data indicate that MRD-based therapy stratification could overcome chemo-resistant nature of Ph-like ALL. Our data suggest that nCounter-based diagnostic method is fast and accurate to identify Ph-like ALL. Since Ph-like signature generally dictates poor clinical outcomes, and upfront TKI therapy may improve them, our method could facilitate precision medicine in the treatment of Ph- B-ALL. Disclosures Akashi: Sumitomo Dainippon, Kyowa Kirin: Consultancy; Celgene, Kyowa Kirin, Astellas, Shionogi, Asahi Kasei, Chugai, Bristol-Myers Squibb: Research Funding.

Novel GTF2I\u2013PDGFRB and IKZF1\u2013TYW1 fusions in pediatric leukemia with normal karyotype

BackgroundMany cases of acute lymphoblastic leukemia (ALL) carry visible acquired chromosomal changes of pathogenetic, diagnostic, and prognostic importance. Nevertheless, from one-fourth to half of newly diagnosed ALL patients have no visible chromosomal changes detectable by G-banding analysis at diagnosis. The introduction of powerful molecular methodologies has shown that many karyotypically normal ALLs carry clinically important submicroscopic aberrations.Case presentationWe used fluorescence in situ hybridization (FISH), array comparative genomic hybridization (aCGH), RNA sequencing, reverse transcription (RT) and genomic polymerase chain reaction (PCR), as well as Sanger sequencing to investigate a case of pediatric ALL with a normal karyotype. FISH with a commercial PDGFRB breakapart probe showed loss of the distal part of the probe suggesting a breakpoint within the PDGFRB locus. aCGH revealed submicroscopic deletions in chromosome bands 5q32q35.3 (about 30 Mb long, starting within PDGFRB and finishing in the CANX locus), 7q34 (within TCRB), 9p13 (PAX5), 10q26.13 (DMBT1), 14q11.2 (TRAC), and 14q32.33 (within the IGH locus). RNA sequencing detected an in-frame GTF2I–PDGFRB and an out-of-frame IKZF1–TYW1 fusion transcript. Both fusion transcripts were verified by RT-PCR together with Sanger sequencing and interphase FISH. The GTF2I–PDGFRB fusion was also verified by genomic PCR and FISH. The corresponding GTF2I–PDGFRB fusion protein would consist of almost the entire GTF2I and that part of PDGFRB which harbors the catalytic domain of the tyrosine kinase. It would therefore seem to lead to abnormal tyrosine kinase activity in a manner similar to what has been seen for other PDGFRB fusion proteins.ConclusionsThe examined pediatric leukemia is a Ph-like ALL which carries novel GTF2I–PDGFRB and IKZF1–TYW1 fusion genes together with additional submicroscopic deletions. Because hematologic neoplasms with PDGFRB-fusion genes can be treated with tyrosine kinase inhibitors, the detection of such novel fusions may be clinically important. Since the GTF2I–PDGFRB could be detected only after molecular studies of the leukemic cells, further investigations of ALL-cases, perhaps especially but not exclusively with a normal karyotype, are needed in order to determine the frequency of GTF2I–PDGFRB in leukemia, and also to find out which clinical impact the fusion may have.

Sustained Response to Imatinib in a Pediatric Patient with Concurrent Myeloproliferative Disease and Lymphoblastic Lymphoma Associated with a CCDC88C-PDGFRB Fusion Gene

Background: The WHO defined myeloid and lymphoid neoplasms (MLN) with eosinophilia associated with PDGFRB, PDGFRA, FGFR1 rearrangements as a new entity in 2016. PDGFRB-rearranged MLN sensitive to imatinib were described in adult patients. We report the first pediatric patient with PDGFRB-rearranged myeloproliferative disorder associated with T-lymphoblastic lymphoma bearing the t(5; 14)(q33;q32) translocation who was successfully treated with imatinib only. Methods/Aims: Analysis of bone marrow and peripheral blood cells by fluorescent in situ hybridization identified the PDGFRB partner as CCDC88C. Whole genome sequencing of the patient’s DNA identified the exact junction site, confirmed by PCR amplification and Sanger sequencing. A real-time quantitative PCR assay was designed to quantify the fused CCDC88C-PDGFRB product. Results: A 2.5-year-old boy was diagnosed with myeloproliferative disorder and eosinophilia associated with lymphoblastic lymphoma both bearing the CCDC88C-PDGFRB fusion. Imatinib therapy resulted in rapid clinical, hematological, and cytogenetic response. Molecular response to treatment was monitored by a real-time PCR assay specific for the CCDC88C- PDGFRB fusion. Conclusion: This is the first description of MLN with eosinophilia in the pediatric age group. Response to treatment with imatinib only was monitored by specific quantitative PCR assay with sustained remission lasting 5.5 years from diagnosis.

PDGFRB mutation and tyrosine kinase inhibitor resistance in Ph-like acute lymphoblastic leukemia

AbstractPhiladelphia chromosome (Ph)-like acute lymphoblastic leukemia (ALL) comprises ∼10% to 15% of childhood ALL cases, many of which respond exquisitely to tyrosine kinase inhibitors (TKIs), for example, imatinib in PDGFRB-rearranged ALL. However, some cases developed drug resistance to TKIs and the mechanisms are poorly understood. In this study, we identified a novel PDGFRB fusion gene, namely AGGF1-PDGFRB, and functionally characterized its oncogenic potential in vitro. Further genomic profiling of longitudinally collected samples during treatment revealed the emergence of a mutation, PDGFRBC843G, which directly conferred resistance to all generations of ABL TKIs, including imatinib, dasatinib, nilotinib, and ponatinib. PDGFRB-mutant leukemia cells are highly sensitive to multitarget kinase inhibitor CHZ868, suggesting potential therapeutic options for some patients resistant to ABL TKIs. In summary, we describe a complex clonal evolution pattern in Ph-like ALL and identified a novel PDGFRB point mutation that drives leukemia relapse after ABL TKI treatment.

Novel PDGFRB fusions in childhood B- and T-acute lymphoblastic leukemia.

Acute lymphoblastic leukemia (ALL) is among the most curable childhood cancers with a 90% overall survival in the good-risk category.1 Though the remission and cure rates among newly diagnosed patients have improved, the prognosis for those with relapsed disease remains poor. Better risk stratification integrating cytogenetics and genomics has enabled improvements in therapy and prediction of relapse. Recently the clinical utility of comprehensive genomic profiling (CGP) has been recognized, in particular, for risk stratification of about 25–30% of childhood ALL patients who do not appear to harbor any of the established chromosomal abnormalities (Supplementary Information).2 By way of illustration, a novel subgroup of BCR-ABL1-like ALL has been described, in which patients lack the BCR-ABL1 fusion but exhibit a gene expression profile similar to that of BCR-ABL1-positive ALL.3 Furthermore, some of these patients have been demonstrated to harbor tyrosine kinase-activating fusion genes, notably including PDGFRB and JAK2 fusions. The EBF1-PDGFRB gene fusion has been well studied and found to be responsive to the ABL1 tyrosine kinase inhibitor (TKI), imatinib mesylate.4, 5, 6 Here, we have identified three novel PDGFRB fusions (PDGFRB fused to each of SATB1, AGGF1 and DOCK2) in children with either precursor B-cell (B-ALL) or T-cell ALL (T-ALL). We report the clinical utility of CGP to characterize these previously undescribed activating fusion genes, in conjunction with childhood B- or T-ALL, and describe responses to TKIs.

Platelet-derived growth factor receptors (PDGFRs) fusion genes involvement in hematological malignancies.

To investigate oncogenic platelet-derived growth factor receptor(PDGFR) fusion genes involvement in hematological malignancies, the advances in the PDGFR fusion genes diagnosis and development of PDGFR fusions inhibitors. Literature search was done using terms "PDGFR and Fusion" or "PDGFR and Myeloid neoplasm" or 'PDGFR and Lymphoid neoplasm' or "PDGFR Fusion Diagnosis" or "PDGFR Fusion Targets" in databases including PubMed, ASCO.org, and Medscape. Out of the 36 fusions detected, ETV6(TEL)-PDGFRB and FIP1L1-PDGFRA fusions were frequently detected, 33 are as a result of chromosomal translocation, FIP1L1-PDGFRA and EBF1-PDGFRB are the result of chromosomal deletion and CDK5RAP2- PDGFRΑ is the result of chromosomal insertion. Seven of the 34 rare fusions have detectable reciprocals. RNA aptamers are promising therapeutic target of PDGFRs and diagnostic tools of PDGFRs fusion genes. Also, PDGFRs have variable prospective therapeutic strategies including small molecules, RNA aptamers, and interference therapeutics as well as development of adaptor protein Lnk mimetic drugs.