Adult Acute Myeloid Leukemia with the KMT2A-Mixed Lineage Leukemia T10 Fusion: An Analysis of 10 Cases Showed Common Features and Frequent Mutations in the RAS Signaling Pathway.

Mixed lineage leukemia (MLL) T10 is a relatively rare partner for the KMT2A lysine (K)-specific methyltransferase 2A gene. The common features and coexisting mutations of acute myeloid leukemia (AML) patients with KMT2A-MLLT10 remain unknown. In this study, 10 adult AML patients with KMT2A-MLLT10 fusions were picked up from 496 AML patients by using RT-polymerase chain reaction (PCR) and/or fluorescence in situ hybridization, and then screened for mutations in the 49 genes panel with next-generation sequencing and PCR, followed by direct Sanger sequencing. Of the 10 unique individuals identified, 6 were male and 4 were female (M:F ratio, 1.5:1) with ages ranging from 19 to 52 years (median 39.5 years). Most (90%, 9/10) patients with KMT2A-MLLT10 were accompanied by additional mutations. Twelve mutated genes were detected, averaging 2.1 mutations per patient (range, 0-4). The most frequently mutated gene was NRAS (n = 5). Clinical and laboratory data pointed to common features: French American British-M5 subtype (n = 7), a high rate of relapse, and biomarkers CD33 (n = 10), CD117 (n = 9), CD13 (n = 8), and CD64 (n = 8). Overall, most patients harbored at least one mutation. A high incidence of mutations affecting the RAS signaling pathway or RAS regulating components was found in 50% (5/10) patients. The overall survival is about 12.0 months. Allogeneic-hematopoietic stem cell transplantation trends to improve survival in selected patients.

[Analysis of ASXL1 gene variant in patients with myelodysplastic syndrome].

OBJECTIVE: To detect ASXL1 gene variants among patients with myelodysplastic syndrome (MDS) and explore their correlation with variants of other genes and clinical features of patients. METHODS: For 149 patients with MDS, genomic DNA was amplified by PCR and subject to direct sequencing to identify variants of ASXL1, U2AF1, SF3B1, DNMT3A, TET2, IDH1/2, NPM1, FLT3-ITD and C-KIT genes. RESULTS: ASXL1 variants were found among 37 patients (24.8%). Other commonly mutated genes included U2AF1 (22.8%), TET2 (11.4%), DNMT3A (9.4%), NPM1 (8.1%) and SF3B1 (6.0%). The frequency of concurrent U2AF1 and TET2 variants among patients with ASXL1 variants was slightly higher than that of wild-type patients. No significant difference was found in median age, MDS subtype, karyotype, peripheral leukocytes, hemoglobin, platelet levels, and bone marrow blast counts between the ASXL1-variant and the wild-type groups (P> 0.05). Twenty-nine patients harboring ASXL1 variants were followed up, 37.9% progressed to acute myeloid leukemia (AML). The rate of transformation in ASXL1-variant group was significantly higher than the wild-type group (37.9% vs. 14.1%, P< 0.01). CONCLUSION: ASXL1 showed a high frequency of variant among MDS patients, which was frequently accompanied with U2AF1 and TET2 variants. Compared with the wild type group, patients with ASXL1 variants were more likely to progress to AML.

[Clinical feature and cytogenetic analysis of 28 patients with bone marrow invasion non-Hodgkin's lymphoma].

OBJECTIVE: To study the correlation of hematomorphology, bone marrow cytogenetics and clinical biochemical parameters with the prognosis of non-Hodgkin's lymphoma with bone marrow invasion. METHODS: Morphological analysis of bone marrow cells was performed by routine bone marrow puncture.Chromosome samples were prepared by short-term bone marrow culture. Karyotype analysis was carried out by R-banding in 28 patients. P53 gene was detected by fluorescence in situ hybridization (FISH). Serum lactate dehydrogenase (LDH) of all patients was determined and compared. RESULTS: In all patients, bone marrow morphology showed invasion of lymphoma. Chromosome analysis revealed abnormal karyotypes in 19 cases, which yielded an incidence of 67.85%. The proportion of lymphoma cells in bone marrow among those with an abnormal karyotype was much higher than those with a normal karyotype (60.2% vs. 33.5%, P<0.05). FISH assay showed that 9 (32.14%) patients had P53 gene deletion. And the deletion was much more common among those with an abnormal karyotype (42.11% vs. 11.11%, P<0.05). The serum LDH level in patients with an abnormal karyotype was significantly higher compared with whose with a normal karyotype (1464.37 U/L vs. 294.33 U/L, P<0.05). CONCLUSION: Patients with abnormal karyotypes have a higher rate of P53 gene deletion, and their LDH level is significantly higher than those with a normal karyotype, which predicted a relatively poor prognosis.

[Characteristics of a patient with myeloid neoplasm and co-existence of t(7;11)(p15;p15) and t(5;12)(q33;p13) translocations].

OBJECTIVE: To delineate the clinical and molecular characteristics of a patient with myeloid neoplasm and co-existence of t(7;11)(p15;p15) and t(5;12)(q33;p13) translocations. METHODS: Clinical data of the patient was collected. Conventional karyotyping, reverse transcriptase (RT)-PCR and next generation sequencing (NGS) were carried out to delineate its genetic features. RESULTS: The patient has featured recurrent rash, fatigue, loss of appetite and splenomegaly. Laboratory test suggested hyperleukocytosis of FAB-M2-subtype. Neither eosinophilia nor basophilia was presented. NUP98/HOXA9 and ETV6/PDGFRB fusion genes were detected by RT-PCR. NGS and DNA-PCR showed the co-existence of WT1 p.C423Y, KRAS p.G12D and DNMT3A p.R882C mutations. The patient achieved morphological remission after imatinib plus coventional chemotherapy (standard IAC regimen). However, the disease has relapsed shortly after. Treatment was switched to HHT-Ara-C-Acla regimen, no hematological response was observed. The ETV6/PDGFRB fusion gene was undetectable in bone marrow sample, though strong expression of NUP98/HOXA9 was detectable throughout the whole course. CONCLUSION: Acute myeloid leukemia in association with the co-existence of NUP98/HOXA9 and ETV6/PDGFRB fusion genes have unique clinical and genetic features. Imatinib seems to have no impact on the overall survival in such cases.

[Characterization of mutational pattern of patients with core-binding factor acute myeloid leukemia].

OBJECTIVE: To characterize the mutational profile of patients with core-binding factor acute myeloid leukemia (CBF-AML). METHODS: A total of 81 acute myeloid leukemia patients were recruited, which included 36 cases of CBF-AML and 45 cases of cytogenetically normal acute myeloid leukemia (CN-AML) . Mutations of FLT3-ITD, FLT3-TKD, NPM1, c-KIT, NRAS, KRAS, TET2, IDH1/2, RUNX1, DNMT3A, GATA2, ASjXL1, TP53, PTPN11, JAK2V617F, SETBP1 and CEBPA genes were simultaneously detected by DNA-based PCR and Sanger sequencing. RESULTS: Over all, mutations were detected in 68 patients (83.9%), with the most common ones including double CEBPA mutations (n=17), followed by NPM1 (n=15), c-KIT (n=11), NRAS (n=10), TET2 (n=9), FLT3-TKD (n=9), FLT3-ITD (n=8), IDH1 (n=7), RUNX1 (n=7), KRAS (n=7), DNMT3A (n=6), IDH2 (n=4), and GATA2 (n=4) mutations. AML1-ETO and CBFß-MYH11 fusions were present in 21 and 15 patients, respectively. Coexistence of ≥2 mutations was more common in CN-AML comparing with CBF-AML. The mutation rate of NPM1, FLT3-ITD, DNMT3A, IDH1 and CEBPA double mutations were higher in patients with CN-AML. NRAS, c-KIT and KRAS mutations were identified more frequently in patients with CBF-AML (P<0.05). Based on the function, aberration of genes involved in DNA methylation, NPM1 proteins and transcription predominated in CN-AML, while tyrosine kinase receptor signaling and RAS pathways have predominated in CBF-AML. CONCLUSION: The genomic landscape of CBF-AML patients has differed from that of CN-AML patients. Synergy of fusion genes with particular mutations may impact the clinical phenotype and prognosis of patients.

[Characterizing the molecular cytogenetics in acute monocytic leukemia].

OBJECTIVE: To characterize the molecular genetics of 81 patients with acute monocytic leukemia (AML). METHODS: Fluorescence in situ hybridization (FISH) was employed to detect MLL gene rearrangements. Combined mutations of 17 genes were detected by DNA-based PCR and Sanger sequencing. RESULTS: Sixty seven patients were found to harbor at least one mutation. The most commonly mutated gene was NPM1 (n=18), which was followed by FLT3-ITD (n=16), NRAS (n=16), DNMT3A (n=15), TET2 (n=12), RUNX1 (n=11) and KRAS (n=9). Based on the functions of mutated genes, the most frequently involved genes were those involved in DNA methylation (38.27%), tyrosine kinase receptor signaling (32.1%), transcription regulation (28.4%), and RAS pathway (24.7%). Single gene mutation predominated in patient with cytogenetic abnormalities, while coexistence of 2 mutations have predominated in patient with normal cytogenetic findings. Stratified by cytogenetic findings, patients with single gene mutations (intermediate-risk group) had significantly higher complete remission (CR) rates than those with ≥2 gene mutations (unfavorable-risk group) (91.7% vs. 57.6% , 87.5% vs. 25.0%, P =0.0319, 0.0117, respectively). CONCLUSION: Over 80% of AML patients were found to harbor at least one mutation. Their clinical phenotype and prognosis may be impacted by the synergy of MLL gene rearrangement and multiple mutations. For patients under the same risk stratification, the number of mutations is reversely correlated with the CR rate.

[Mutation analysis of 77 patients with normal-karyotype myelodysplastic syndrome].

OBJECTIVE: To carry out mutation analysis for patients with myelodysplastic syndromes (MDS) and a normal karyotype. METHODS: Targeted capture and next-generation sequencing (NGS) was carried out using a customized 49-gene panel. FLT3 internal tandem duplication (FLT3-ITD), CALR, NPM1 and CEBPA mutations were detected by PCR and Sanger sequencing. RESULTS: Sixty-two patients (80.5%) were found to harbor at least one mutation. Each patient has carried 2.21 mutations in average. Coexistence of ≥ 3 mutations was common (43.7%). The most commonly mutated genes were RUNX1 (23.4%, 18/77), ASXL1 (18.2%, 14/77), NPM1 (15.6%, 12/77), U2AF1 (15.6%, 12/77), DNMT3A (11.7%, 9/77). Patients with SF3B1 mutations were significantly older than those with ASXL1 mutations (P=0.023). Mutations of the DNMT3A gene were significantly associated with the blood platelet level compared with BCOR mutations (P=0.02). No significant difference was found in the number and rate of mutations between those under or above 60-year-old. Among 67 patients with clinical follow-up, 20 (29.8%) has transformed to acute myeloid leukemia, and the time of transformation has ranged from 1 to 44 months, with a average of 5.3 months. RUNX1, U2AF1 and FLT3 mutations are associated with leukemic transformation. CONCLUSION: Coexistence of ≥ 3 mutations are frequent among patients with normal-karyotype MDS. Certain mutations are associated with age and leukemic transformation.

[Clinical and genetic features of a patient with myeloid neoplasm in association with PDGFRA and EVI1 gene rearrangements].

OBJECTIVE: Todelineate the clinical and genetic features of a patient with myeloproliferative neoplasm (MPN) in association with PDGFRA and EVI1 genes rearrangements. METHODS: Clinical data of the patient was collected. Conventional cytogenetics, fluorescence in situ hybridization (FISH) and nested PCR were carried out for the patient. RESULTS: The patient has featured recurrent rash, joint pain, and intermittent fever. Laboratory tests showed hyperleukocytosis and marked eosinophilia. Physical examination revealed splenomegaly. His karyotype was 46,XY,t(3;5)(q26;q15)[6]/46,XY[10]. FISH assay showed that both PDGFRA and EVI1 genes were rearranged. Molecular studies of the mRNA suggested that there was a in-frame fusion between exon 12 of the PDGFRA gene and exon 9 of the FIP1L1 gene. Imatinib was initiated at a dosage of 200 mg, and after 10 months, the signal of the FIP1L1-PDGFRA fusion gene was undetectable in bone marrow sample. However, the expression of EVI1 mRNA was stable, with no significant difference found between the patient and 10 healthy controls. CONCLUSION: MPN in association with PDGFRA and EVI1 genes rearrangements have unique clinical and genetic features. Genetic testing is helpful for early diagnosis. Imatinib may be effective for the treatment.

[Correlation of cytogenetic changes with VEGF and TRacp-5b levels among 60 elderly patients with multiple myeloma].

OBJECTIVE: To assess the correlation of cytogenetic changes with serum vascular endothelial growth factor (VEGF) and serum tartrate resistant acid phosphatase (TRacp-5b) levels among elderly patients with multiple myeloma (MM). METHODS: Chromosomal changes were analyzed with a modified culturing method in the presence of IL-6. Serum levels of VEGF and TRacp-5b were determined with enzyme-linked immunosorbent assays (ELISA). RESULTS: Among the 60 MM patients, chromosomal abnormalities were found in 27 cases, including 22 with numerical abnormalities and 15 with structural abnormalities. Many patients had both numerical and structural abnormalities. For 33 patients with a normal karyotype, the levels of VEGF and TRacp-5b were 117.35 ± 55.26 pg/mL and 4.15 ± 2.15 U/L, respectively, while for 27 patients with an abnormal karyotype, the levels of VEGF and TRacp-5b were 190.26 ± 85.74 pg/ml and 5.96 ± 2.24 U/L, respectively. The difference between the two groups was significant (P<0.05). CONCLUSION: Compared with MM patients with a normal karyotype, the levels of VEGF and TRacp-5b are higher in those with cytogenetic abnormalities.

[8p11 myeloproliferative syndrome with CEP110-FGFR1 fusion in a patient].

OBJECTIVE To explore the clinical and laboratory features of a patient with 8p11 myeloproliferative syndrome (EMS) and CEP110-FGFR1 fusion. METHODS Combined bone marrow cytology, fluorescence in situ hybridization, fusion gene detection was used to analyze the patient. RESULTS Clinically, the patient had many features similar to those with chronic myelomonocytic leukemia, which included hyperleukocytosis, marked eosinophilia, monocytosis, myeloid hyperplasia and hyperplasia. Fluorescence in situ hybridization analysis for FGFR1 gene rearrangement was positive. Further study of the mRNA also confirmed an in-frame fusion between exon 38 of the CEP110 gene and exon 9 of FGFR1 gene. CONCLUSION EMS with CEP110-FGFR1 fusion is a very rare and distinct myeloproliferative neoplasm. FISH and molecular studies may improve its diagnosis.

[Analysis of CSF3R Gene Mutations and Clinical Characteristics in Patients with t(8;21) Acute Myeloid Leukemia].

OBJECTIVE: To investigate the occurrence of CSF3R mutation in patients with t(8;21) acute myeloid leukemia (AML) and its correlation with some clinical parameters. METHODS: The clinical and laboratory data of 167 newly diagnosed AML patients with t(8;21) translocation were analyzed retrospectively. High-throughput DNA sequencing technology combined with Sanger sequencing method was used to detect 112 gene mutations. The occurrence of CSF3R gene mutation and its influence on the remission rate after chemotherapy were analyzed. RESULTS: Among 167 patients with t(8;21) AML, 15 patients (9.0%) carried CSF3R mutations, including 6 cases of membrane proximal region mutations and 9 cases of truncation mutations in the cytoplasmic tail. The most common coexisting mutations of CSF3R were KIT (40.0%), TET2 (33.3%), DNMT3A (26.7%), FLT3 (20.0%), CBL (20.0%), IDH1 (13.3%), etc. Compared with the wild type, the CSF3R mutant group had a higher mutation rate of DNA methylation-related genes（P <0.001）. The median peripheral white blood cell (WBC) count of patients with CSF3R gene mutation was 5.80 (3.20-8.56)×109/L at initial diagnosis, which was significantly lower than 8.80 (5.26-19.92)×109/L of the CSF3R wild-type patients (P =0.017). There was no significant difference between the two groups in sex, median age, FAB classification, hemoglobin level, platelet count, etc. (P >0.05). The CR rate of the CSF3R gene mutation group (100%) was significantly higher than that of the wild-type group (86.8%), but the difference was not statistically significant (P >0.05). The CSF3R gene mutation group had a significantly higher CD19 positive rate and a higher -X rate than the wild group (86.7% vs 47.4%, P =0.004; 33.3% vs 13.2%, P =0.037). CONCLUSION: There is a high incidence of CSF3R mutation in t (8;21) AML patients. The clinical characteristics and coexisting mutation genes of CSF3R mutation-positive patients are different from those of wild-type patients.

[Molecular Genetic Characteristics of Acute Myeloid Leukemia Patients with CBFß-MYH11 Positive].

OBJECTIVE: To explore mutational characteristics of acute myeloid leukemia (AML) patients with CBFß-MYH11+ and analyze the correlation between the mutations and partial clinical characteristics. METHODS: A total of 62 AML patients with CBFß-MYH11+ were included and 51 candidate genes were screened for their mutations using targeted next-generation sequencing (NGS). The exon 12 of NPM1 , FLT3-ITD , and TAD, bZIP domains of CEBPA were detected by genomic DNA-PCR combined with sanger sequencing. RESULTS: Compared with RUNX1-RUNX1T1 + group, the patients with CBFß-MYH11+ showed higher age, peripheral WBC level, initial induced complete remission (CR) rate, more commonly carried chromosomal abnormalities such as +22, and lower deletion ratio of sex chromosome (-X or -Y) (P<0.05). In AML patients with CBFß-MYH11+, the most common mutation was NRAS , followed by KIT, KRAS , and FLT3-TKD . Compared with RUNX1-RUNX1T1+ group, NRAS and FLT3-TKD were more frequently mutated in patients with CBFß-MYH11+ (51.6% vs 18.7%, 17.7% vs 3.8%) (P<0.05). CONCLUSION: The genomic landscape and clinical characteristics of AML patients with CBFß-MYH11+ are different from patients with RUNX1-RUNX1T1 +.

Molecular genetic and clinical characterization of acute myeloid leukemia with trisomy 8 as the sole chromosome abnormality.

INTRODUCTION: The aim of the study was to determine molecular genetic and clinical characterization of acute myeloid leukemia (AML) with trisomy 8 as the sole chromosome abnormality, a recurrent but rare chromosomal abnormality in AML. METHODS: Interphase fluorescence in situ hybridization, reverse transcriptase-quantitative polymerase chain reaction for gene rearrangement and next-generation sequencing (NGS) were performed on sole trisomy 8 AML patients. RESULTS: A total of 35 AML patients with trisomy 8 as the sole chromosome abnormality were screened. The most frequently mutated genes were DNMT3A(37.1%), RUNX1(28.6%), FLT3-ITD(28.6%), IDH2(22.9%), NPM1(17.1%), and ASXL1 (14.3%). The sole +8 AML patients exhibited more mutations in RUNX1 (28.6% vs. 4.8%, P = 0.001) and ASXL1 (14.3% vs. 4.8%, P = 0.039) by comparing with normal karyotype AML (NK AML) patients(n = 63). The sole +8 AML patients(n = 35) with RUNX1 or IDH2 mutations showed significantly lower WBC counts, while FLT3-ITD showed higher white blood cell (WBC) counts as compared to the corresponding wild-type groups. Total of 45.7% patients achieved complete remission (CR) after the first induction therapy. The CR rate of patients with FLT3-ITD or IDH1 mutation was significantly lower than that in the corresponding wild-type cases (P = 0.047, 0.005, respectively). The median overall survival (OS) and disease-free survival (PFS) were 18.0 (95% CI: 10.8-25.2) and 10 (95% CI: 6.7-13.3) months, respectively. FLT3-ITD mutations and allogeneic hematopoietic stem cell transplantation (allo-HSCT) were independent prognostic markers for OS in multivariable analysis. CONCLUSION: The results suggest a possible association between trisomy 8 and additional mutations that may influence clinical feature and prognosis.

PTPN11 mutations in adult acute myeloid leukaemia: Prevalence and clinical implications in the context of NPM1 mutation.

More than 90% of NPM1-mutated acute myeloid leukaemia (NPM1mut AML) patients have been determined to harbour other known concurrent mutations. However, there is limited data on the frequency of PTPN11 and its clinical impact in NPM1mut AML. Next-generation sequencing(NGS) was performed retrospectively on 112 genes in 254 patients with NPM1mut AML. Overall, 33 PTPN11 mutations were detected in 29 of the 254 patients (11.42%) screened. PTPN11 alterations were exclusively missense mutations affecting residues located within the N-SH2 (n = 25) and PTP (n = 8) domains and clustered overwhelmingly in exon 3 (n = 25). NPM1mut AML patients with mutant PTPN11 had significant associations with mutations in epigenetic regulators(TET2, IDH1/2, and DNMT3A) (72.41% vs. 46.67%, P = 0.009) and cohesins (RAD21,SMC1A, and SMC3)(10.34% vs. 1.33%, P = 0.02) compared to patients with wild-type PTPN11. Among the patients treated with intensive chemotherapy, the differences in disease-free survival (DFS) and overall survival (OS) between those with mutant and wild-type PTPN11 were not significant (P = 0.4, 0.83, respectively). In conclusion, PTPN11 mutations were frequently identified in NPM1mut AML patients and clustered in exon 3. PTPN11 mutations more frequently co-occurred with mutations involving epigenetic regulators but had no impact on prognosis.

Companion gene mutations and their clinical significance in AML with double or single mutant CEBPA.

INTRODUCTION: We report the co-mutations in AML with CEBPAsm or CEBPAdm and their clinical features in a large cohort (n = 302) of CEBPAmut AML patients. MATERIALS AND METHODS: We retrospectively sequenced 112 genes in 302 patients with CEBPAmut using NGS, and studied the spectrum and clinical impact of co-mutations in CEBPAdm and CEBPAsm. RESULTS: ① The average number of mutations in CEBPAsm and CEBPAdm AML was comparable, but not significant (P = 0.17). ② CEBPAdm patients exhibited more mutations in CSF3R (P = 0.037), GATA2 (P = 0.022), and WT1 (P = 0.046). In contrast, CEBPAsm patients more frequently harbored mutations in NPM1 (P = 0.000), FLT3-ITD (P = 0.025) and NOTCH2 (P = 0.043), as well as mutations in signaling pathways and spliceosomes (P = 0.064, P = 0.027, respectively). ③ Patients with CEBPAsm/TET2mut or CEBPAsm /GATA2mut had higher platelet counts (both P = 0.011), while patients with CEBPAdm /TET2mut had significantly higher hemoglobin levels (P = 0.009). The CR rate of patients with FLT3-ITD mutations was significantly lower in the CEBPAsm group than the CEBPAdm group (P = 0.028). CONCLUSIONS: CEBPAsm and CEBPAdm AML are each associated with their own complex co-mutation cluster. Some co-mutations influence the clinical features and CR rate differently in patients with different CEBPA mutational status.

[Analysis of Coexisting Gene with NRAS in Acute Myeloid Leukemia].

OBJECTIVE: To investigate the coexisting mutations and clinical significance of Homo sapiens neuroblastoma RAS viral oncogene homolog (NRAS) gene in acute myeloid leukemia (AML) patients. METHODS: High-throughput DNA sequencing and Sanger sequencing were used to detect 51 gene mutations. The occurrence, clinical characteristics and treatment efficacy of coexisting genes with NRAS were investigated. RESULTS: A total of 57 NRAS mutations (17.5%) were detected in 326 patients with AML. Compared with the patients in NRAS non-mutation group, patients in the mutant group were younger (P=0.018) and showed lower platelet count (P=0.033), but there was no significant difference in peripheral leukocyte count, hemoglobin, and sex. For FAB classification, NRAS mutation and M2 subtype showed mutually exclusive (P=0.038). Among 57 patients carried with NRAS mutation, 51 (89.5%) patients carried with other gene mutations, 25 (43.9%) carried with double gene mutations, 10 (17.5%) carried with 3 gene mutations, and 16 (28.1%) corried with ≥ 4 gene mutations. The most common coexisting gene mutation was KRAS (24.6%, 14/57), followed by FLT3-ITD (14.0%, 8/57), RUNX1 (12.3%, 7/57), NPM1 (10.5%, 6/57), PTPN11 (10.5%, 6/57), DNMT3A (10.5%, 6/57) and so on. The age (P=0.013, P=0.005) and peripheral platelet count (P=0.007, P=0.021) of patients with NPM1 or DNMT3A mutations were higher than those of the patients with wild type, but there was no significant difference in peripheral leukocyte count and hemoglobin. Also, there was no significant difference in age, peripheral leukocyte count, hemoglobin, and peripheral platelet count between the patients in KRAS, FLT3-ITD, RUNX1 or PTPN11 mutant group and the wild group. Patients with FLT3-ITD mutations showed a lower complete remission (CR) rate (P=0.044). However, there was no significant difference in CR rate between the patients with KRAS, NPM1, RUNX1, PTPN11 or DNMT3A mutations and the wild group. The CR rate of the patents with single gene mutation, double gene mutations, 3 gene mutations, and≥ 4 gene mutations were decreased gradually, and there was no significant difference in CR rate between pairwise comparisons. CONCLUSION: The mutation rate of NRAS mutation is 17.5%, 89.5% of AML patients with NRAS mutation coexist with additional gene mutations. The type of coexisting mutations has a certain impact on clinical characteristics and CR rate of patients with AML.

Molecular genetic characterization of acute myeloid leukemia with isolated trisomy of chromosomes 4, 11, and 21.

BACKGROUND: Autosomal trisomy is a relatively rare abnormality observed in AML, occurring singly or as a secondary event in association with other karyotypic changes, and associated with prognosis. The molecular genetic and clinical characterizations of acute myeloid leukemia (AML) with isolated trisomy 4, 11, or 21 have been poorly investigated. MATERIALS AND METHODS: Interphase fluorescence in situ hybridization, reverse transcriptase-quantitative polymerase chain reaction for 41 chromosomal gene translocations/fusion genes, and next-generation sequencing (NGS) were performed on 29 AML patients with trisomy 4, 11, or 21 as the sole chromosomal anomaly. RESULTS: Of the 29 patients, one or more mutations were detected in 93.1% of patients. CEBPA had the highest mutation frequency, followed by TET2, NPM1, DNMT3A, and FLT3-ITD. The sole +11 AML patients exhibited more mutations in FLT3-ITD (P = .031) than the sole +21 AML patients, while CEBPA mutation was more frequently found in the sole +21 AML patients than that in the sole +11 AML patients(P = .07). The median overall survival (OS) and disease-free survival (DFS) for patients with +11 were shorter than those with +4(P = .015, 0.046) or +21 (0.057, 0.064), but no difference was found between +4 patients and +21 patients. In the whole cohort, only the FLT3-ITD mutation was significantly associated with inferior OS (18 vs. 35 months, P = .023) and DFS (12 months vs. NR, P = .046). There were no significant differences in OS and DFS according to the gene mutation status of CEBPA, TET2, NPM1, DNMT3A, and IDH1/2. CONCLUSION: There was a significantly different mutation profile among the sole +4, +11, +21 AML patients. Our research provided new insight into the molecular characteristics of AML with isolated trisomy.

Mutational landscape of chronic myelomonocytic leukemia and its potential clinical significance.

We evaluated the mutational landscape of chronic myelomonocytic leukemia (CMML) and its potential clinical significance. We analyzed 47 samples with a panel of 112 genes using next-generation sequencing. Forty-five of the 47 patients (95.74%) had at least one mutation identified, with an average of 3.7 (range 0-9) per patient. The most common mutation was NRAS, followed by ASXL1, TET2, SRSF2, RUNX1, KRAS, and SETBP1. Patients 60 years and older more frequently had mutations in TET2 (56% vs. 9.09%, P = 0.001) and ASXL1 (48% vs. 18.18%, P = 0.031) than patients younger than 60 years. Median overall survival (OS) in patients with CMML was 22.0 months (95% CI 19.7-24.3 months). ASXL1 (18 vs. 22 months, P = 0.012), RUNX1 (17 vs. 22 months, P = 0.001), and SETBP1 (20 vs. 27 months, P = 0.032) mutations predicted inferior OS. However, only RUNX1 mutation was significantly associated with inferior acute myeloid leukemia (AML)-free survival. Our data showed that mutation profile differed significantly between CMML patients aged 60 years and older versus those younger than 60 years, and some of these mutations impact the progression and prognosis of the disease to a certain extent.

Frequency and clinical impact of WT1 mutations in the context of CEBPA-mutated acute myeloid leukemia.

INTRODUCTION: Several studies have confirmed that mutations in the Wilms tumor 1 (WT1) gene occur in adult acute myeloid leukemia (AML). However, few data are available regarding the incidence of WT1 mutations in CEBPAmut AML and their impact. METHODS: We retrospectively analyzed the frequency and clinical impact of WT1 mutations in 220 newly diagnosed AML patients with CEBPA mutations(CEBPAmut). Chromosome karyotype analysis was performed by R or G banding method and further confirmed either by fluorescence in situ hybridization (FISH) and/or by multiple reverse transcription polymerase chain reaction (multiple RT-PCR). Mutations were detected with a panel of 112mutational genes using next-generation sequencing (NGS). RESULTS: Overall, 30 WT1 mutations were detected in 29 of the 220 CEBPAmut AML patients (13.18%) screened. These mutations clustered overwhelmingly in exon 7 (n=16). WT1 mutations were found to be significantly more frequent in AML patients with double-mutated CEBPA (CEBPAdm) than in AML patients with single-mutated CEBPA (17.36%vs. 8.08%, P = 0.043). Among WT1-mutated patients, the most common co-mutation was FLT3-ITD (n = 7, 24.14%), followed by NRAS (n = 5, 17.24%), CSF3R (n = 4, 13.79%), GATA2 (n = 4, 13.79%), and KIT (n = 4, 13.79%). The most frequent functional pathway was signaling pathways inas many as 62.07% of cases. Notably,the concomitant mutations in epigenetic regulatorswere inversely correlated with WT1 mutations(P = 0.003). CEBPAdm AML patients with WT1 mutations had inferior relapse-free survival, event-free survival and overall survival compared with patients CEBPAdm AML without WT1 mutations (P = 0.002, 0.004, and 0.010, respectively). CONCLUSION: Our data showed that WT1 mutations are frequently identified in CEBPAmut AML, especially in CEBPAdm AML. CEBPAmut AML patients with WT1 mutations show distinct spectrum of comutations. In the context of CEBPAdm AML, WT1 mutations predict a poor prognosis.

Clinical and molecular features of sacrum chordoma in Chinese patients.

Background: Chordoma is a rare malignant bone tumor with high recurrence and metastasis rates. Little is known about the mutational process of this incurable disease. The aim of our research was to explore the potential driver genes and signal pathways in the pathogenesis of chordoma and provide a new idea for the study of molecular biological therapy of chordoma. Methods: We performed whole-exome-sequencing (WES) on 8 sacrum chordoma tissue samples (matched to peripheral blood samples that had been drawn from patients before surgery) to identify genetic alterations in Chinese patients. We analyzed the sequencing data from known driver genes, pathway enrichment analysis and significantly mutated genes (SMGs) after quality control of sequencing, comparison of reference genomes, analysis of mutations and identification of somatic mutations. Immunohistochemistry staining, Sanger sequencing and GeneChip were used to verify the related genes obtained from the analysis of sequencing data. Results: The driver genes Phosphatidylinositol-4,5-Bisphosphate 3-Kinase Catalytic Subunit Alpha (PIK3CA), Phosphoinositide-3-Kinase Regulatory Subunit 1 (PIK3R1), and Phosphatase And Tensin Homolog (PTEN) were enriched in the Phosphatidylinositol 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) signaling pathway and could be potential therapeutic targets for the treatment of sacrum chordoma. The significantly mutated gene Claudin 9 (CLDN9) may play a critical role in the development and progression of sacrum chordoma. Conclusions: Collectively, our results identified the genetic signature of sacrum chordoma and could be used to develop a potential promising therapeutic strategy for the treatment of sacrum chordoma in Chinese patients.