Objective:To explore the characteristics and patterns of gene mutations in tyrosine kinase inhibitor (TKI)-resistant chronic myeloid leukemia (CML) patients and their relationship with TKI-resistant CML.Methods:A retrospective case series study was performed. Clinical data and next-generation sequencing results from TKI-resistant CML patients in Nanfang Hospital of Southern Medical University and Yuebei People's Hospital of Shantou University Medical College from August 2018 to November 2022 were retrospectively analyzed, and the gene mutations of the patients in general and at different disease stages were analyzed.Results:Sixty patients were enrolled, with the age [ M ( Q1, Q3)] of 41.5 years old (32 years old, 53 years old); 38 cases (63.33%) were male and 22 cases (36.67%) were female; 43 cases were in the chronic stage, and 17 cases were in the progression stage (3 cases were in the accelerated stage and 14 cases were in the blast stage). non-ABL1 mutations were detected in 30 patients (50.00%) including 45 times of 15 non-ABL1 genes. The number of non-ABL1 mutation gene was 1 (0, 2) in 60 patients. Of the 60 patients, 21 (35.00%) had ASXL1 mutations, 5 (8.33%) had DNMT3A mutations, 5 (8.33%) had RUNX1 mutations, and 3 (5.00%) had SETBP1 mutations; the proportions of patients with 1 and ≥2 non-ABL1 mutations were 33.33% (20/60) and 16.67% (10/60), respectively. The total detection rates of non-ABL1 mutations were 52.94% (9/17) and 48.84% (21/43), and the detection rates of ≥2 non-ABL1 mutations were 23.53% (4/17) and 13.95% (6/43) in patients with progression and patients with chronic disease, and the differences were not statistically significant ( χ2 = 0.08, P = 0.774; χ2 = 0.80, P = 0.370). Seventeen of 60 patients (28.33%) had mutations in the ABL1 kinase region, of which 14 (82.35%) had non-ABL1 mutations; of these 17 cases, 6 patients with progressive disease all had non-ABL1 mutations, in 11 patients with chronic disease, 8 patients had non-ABL1 mutations, and the difference was not statistically significant ( P = 0.452). Conclusions:Patients with TKI-resistant CML have high frequencies of non-ABL1 mutations, and there is a trend for higher mutation rates in patients with progressive disease than in patients with chronic disease, and these may be related to the abnormal activation of ABL1 kinase by BCR-ABL1 fusion gene in patients with drug-resistant CML, which leads to the genome-level and epigenome-level mutations, and driving disease progression from chronic phase to accelerated or blast phase.

Objective:To investigate the clinical characteristics and next generation sequencing (NGS) results of acute myeloid leukemia (AML) patients dying early.Methods:A retrospective case series study was performed. The clinical data of 49 AML patients dying early in the Affiliated Yuebei People's Hospital of Shantou University Medical College from January 2016 to May 2021 were retrospectively analyzed. All patients were divided into 10 cases in the very early death group (death occurred within 3 d after diagnosis) and 38 cases in the non-very early death group (death occurred within 4-30 d after diagnosis). NGS was used to detect 196 mutant genes related to hematological malignancies.Results:The white blood cell count, creatinine level, lactate dehydrogenase level, bone marrow original cells proportion in the very early death group were higher than those in the non-very early death group, and the differences were statistically significant (all P < 0.05). Among 48 AML patients dying early, 34 cases had NGS results, among which the very early death occurred in 5 cases and the non-very early death occurred in 29 cases. Gene mutations were detected in 34 patients; finally 32 mutant genes were detected and 33 cases (97.06%) harbored more than 2 gene mutations, and the median number [ M ( Q1, Q3)] of gene mutations was 3 (2, 4). Conclusions:AML patients dying early harbor more than 2 gene mutations involving multiple signaling pathways. The clinical characteristics of AML patients in the very early death group are different from those of patients in the non-very early death group.

Objective:To evaluate the safety, efficacy and performance of disposable cholangiopancreatoscope in the diagnosis and treatment of bile duct diseases.Methods:A total of 20 subjects were selected and 16 subjects were enrolled in the prospective and exploratory clinical study which were performed in the Digestive Endoscope Center of Hangzhou First People's Hospital Affiliated to Zhejiang University School of Medicine from July 2021 to August 2021. The disposable cholangiopancreatoscope was used to diagnose bile duct diseases in routine endoscopic retrograde cholangiopancreatography. Biopsies were performed in cases where malignancy was considered. The use related indexes and performance indexes of the instruments were analyzed.Results:Among the 16 patients, there were 6 cases of biliary space-occupying lesions, 6 cases of biliary calculi, and 4 cases of benign biliary stricture before operation. The success rate of the disposable insertion was 100.00% (16/16), and the success rate of observation was 100.00% (16/16). Three biliary space-occupying lesions diagnosed as malignant tumor under cholangiopancreatoscope were confirmed by pathology after operation. Diagnosis of other patients was consistent with preoperative diagnose, so no biopsy was conducted. The rate of positive feedback from operators in directional control was 81.25% (13/16), the image failure rate was 18.75% (3/16), and the rate of positive feedback for image clarity was 93.75% (15/16). In terms of clinical performance, the imaging quality of excellence was 93.75% (15/16), the flexible degree of excellence was 81.25% (13/16), and other indexes were all 100.00% excellence. During the period, there were no instrument defects, pancreatitis, perforation, bleeding or other instrument-related adverse events.Conclusion:The effectiveness, safety and performance indexes of domestic disposable cholangiopancreatoscope have reached the standards of clinical application with high pixel, integration, and portability. It's worthy of clinical recommendation.

Objective:To explore the driver mutations in patients with classical myeloproliferative neoplasms (MPN) and their relationships with clinical characteristics.Methods:The clinical data of 186 patients with classical MPN in the Affiliated Yuebei People's Hospital of Shantou University Medical College from January 2013 to October 2019 who met the World Health Organization 2016 MPN diagnostic criteria were retrospectively analyzed. The mutations of diver genes JAK2, CALR and MPL and clinical characteristics, such as white blood cell count, hemoglobin, and platelet count in patients with essential thrombocythemia (ET), polycythemia vera (PV) and primary myelofibrosis (PMF) were analyzed.Results:Among the 186 MPN patients, 100 were male and 86 were female, with a median age of onset of 62.0 years old (24.0-93.0 years old). There were 125 patients (67.20%) with ET, 44 patients (23.66%) with PV, and 17 patients (9.14%) with PMF. The JAK2V617F mutation was found in 133 patients (71.51%, 133/186). The JAK2V617F mutation rates in patients with ET, PV and PMF were 66.40% (83/125), 88.64% (39/44) and 64.71% (11/17), respectively. Two patients (1.08%, 2/186) with PV were found with JAK2 exon 12 mutation. The CALR exon 9 mutation was found in 32 patients (17.20%, 32/186), with the CALR mutation rates of 24.00% (30/125), 0 and 11.76% (2/17) in patients with ET, PV and PMF, respectively. The MPL exon 10 mutation was found in one ET patient (0.54%, 1/186). CALR mutated ET patients showed higher platelet count [(1 155±537)×10 9/L vs. (997±330)×10 9/L, t = -2.095, P = 0.038], and lower leukocyte count ( t = 2.434, P = 0.017) and hemoglobin ( t = 3.087, P = 0.003) than JAK2V617F mutated ET patients. Two cases of MPN had rare concurrent driver mutations, of which one ET patient with JAK2V617F and CALR mutations and one PMF patient with JAK2V617F and MPLW515L mutations. Conclusions:The detection result of driver mutations is an important basis for precision health care for MPN. Different types of MPN have different detection rates of driver mutations, which are of great significances for judging the clinical characteristics of patients.

Objective:To investigate the occurrence of stroke and its associated risk factors in patients with Philadelphia chromosome (Ph)-negative myeloproliferative neoplasms (MPN).Methods:The data of patients diagnosed as Ph-negative MPN in the Affiliated Yuebei People's Hospital of Shantou University Medical College from January 2016 to December 2019 were retrospectively analyzed. The occurrence of stroke in these patients and the clinical characteristics were summarized. Logistic regression was used to analyze the risk factors of stroke in MPN patients.Results:A total of 193 Ph-negative MPN patients were collected, including 103 males and 90 females. The median age of onset was 62 years old (24-93 years old). There were 129 patients (66.84%) with essential thrombocythemia, 46 patients (23.83%) with polycythemia vera, and 18 patients (9.33%) with primary myelofibrosis. In 193 patients with MPN, there were 31 patients (16.06%) with stroke, including 30 cases (15.54%) of ischemic stroke and 1 case (0.52%) of hemorrhagic stroke, and the incidence of ischemic stroke was higher than that of hemorrhagic stroke, the difference was statistically significant (χ 2 = 54.258, P < 0.01). Among the patients with stroke, JAK2V617F mutation was observed to be the most common driver mutation (80.65%, 25/31). The small-artery occlusive cerebral infarction was the most common in ischemic stroke (63.33%, 19/30). Compared with MPN patients without stroke, those with stroke displayed higher hemoglobin level [(156±35) g/L vs. (138±40) g/L] and concurrent JAK2V617F and CALR mutations rate [3.23% (1/31) vs. 0.62% (1/162)], and lower CALR mutation rate [3.23% (1/31) vs. 19.14% (31/162)], the differences were statistically significant (all P < 0.05). Logistic regression analysis revealed that hemoglobin ≥ median level (140 g/L) was a risk factor for stroke in MPN patients ( OR = 2.903, 95% CI 1.163-7.244, P = 0.022), and CALR mutation acted as a protective factor for stroke ( OR = 0.090, 95% CI 0.009-0.932, P = 0.044). Conclusions:Ischemic stroke is more common than hemorrhagic stroke in Ph-negative MPN patients, and the small-artery occlusive cerebral infarction is also more frequently found in these patients. Hemoglobin ≥140 g/L is a risk factor for stroke in MPN patients, and CALR mutation is a protective factor.

Objective:To analyze the genetic mutations and clinical features of the subtypes of classical BCR-ABL-negative myeloproliferative neoplasm (MPN) .Methods:Mutations of 108 newly diagnosed BCR-ABL-negative MPN patients [including 55 patients with essential thrombocytopenia (ET) , 24 with polycythemia vera (PV) , and 29 with primary myelofibrosis (PMF) ] were identified using next-generation sequencing with 127-gene panel, and the relationship between gene mutations and clinical features were analyzed.Results:Total 211 mutations in 32 genes were detected in 100 MPN patients (92.59% ) , per capita carried (1.96±1.32) mutations. 85.19% (92/108) patients carried the driver gene (JAK2, CALR, MPL) mutations, 69.56% (64/92) of these patients carried at least 1 additional gene mutation. In descending order of mutation frequency, the highest frequency was for activation signaling pathway genes (42.2% , 89/211) , methylation genes (17.6% , 36/211) , and chromatin-modified genes (16.1% , 34/211) . There was a significant difference in the number of mutations in the activation signaling pathway genes, epigenetic regulatory genes, spliceosomes, and RNA metabolism genes among the three MPN subgroups. The average number of additional mutations in PMF patients was higher than that in ET and PV patients (1.69±1.39, 0.67±0.70, 0.87±1.22, χ2=13.445, P=0.001) . MPN-SAF-TSS (MPN 10 score) ( P=0.006) and myelofibrosis level ( P=0.015) in patients with ≥ 3 mutant genes were higher and the HGB level ( P=0.002) was lower than in those with<3 mutations. Twenty-six patients (24.1% ) carried high-risk mutation (HMR) , and patients with HMR had lower PLT ( P=0.017) , HGB levels ( P<0.001) , and higher myelofibrosis level ( P=0.010) and MPN10 score ( P<0.001) . The frequency of ASXL1 mutations was higher in PMF than in PV patients (34.5% vs. 4.2% , P=0.005) . PMF patients with ASXL1 had lower levels of PLT and HGB ( P=0.029 and 0.019) . Conclusion:69.56% of MPN patients carry at least one additional mutation, and 24.1% patients had HMR. Each subgroup had different mutation patterns. PMF patients had a higher average number of additional gene mutations, especially a higher frequency of ASXL1 mutation; PLT and HGB levels were lower in ASXL1 mutation PMF patients.

OBJECTIVE: To analyze the clinical features of chronic myeloid leukemia (CML) with T315 I mutation (CML-T315I) and compare the effectiveness of different treatments. METHODS: We retrospectively analyzed the clinical data and outcomes of 19 patients with CML-T315I receiving different treatments. The T315 I mutations in these patients were detected by examination of BCR-ABL kinase domain (KD) mutation by RTQ-PCR and Sanger sequencing. The relapse following the treatments, defined as hematological, cytogenetic and molecular biological recurrences, were analyzed in these patients. RESULTS: Of the 19 patients with CML-T315I, 14 (73.7%) were in CML-CP stage at the initial diagnosis, and 13 (81.2%) were high-risk patients based on the Sokal scores. All the 19 patients were treated with TKI after the initial diagnosis, and during the treatment, 15 (78.9%) patients were found to have additional chromosomal aberrations, and 10 (52.6%) had multiple mutations; 13 (68.4%) of the patients experienced disease progression (accelerated phase/blast crisis) before the detection of T315I mutation, with a median time of 40 months (5-120 months) from the initial diagnosis to the mutation detection. After detection of the mutation, 12 patients were treated with ponatinib and 7 were managed with the conventional chemotherapy regimen, and their overall survival rates at 3 years were 83.3% and 14.2%, respectively ( < 0.001). CONCLUSIONS CML patients resistant to TKI are more likely to have T315I mutations, whose detection rate is significantly higher in the progressive phase than in the chronic phase. These patients often have additional chromosomal aberrations and multiple gene mutations with poor prognoses and a high recurrence rate even after hematopoietic stem cell transplantation. Long-term maintenance therapy with ponatinib may improve the prognosis and prolong the survival time of the patients.
Drug Resistance, Neoplasm ; Fusion Proteins, bcr-abl ; Humans ; Imidazoles ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; Mutation ; Pyridazines ; Retrospective Studies

Objective: To analyze the association of cytogenetic abnormalities with the prognosis of chronic myeloid leukemia (CML) patients in tyrosine kinase inhibitors (TKI) era. Methods: Karyotype analysis of chromosome G-banding was carried out in 387 newly diagnosed CML patients by short-term culture of bone marrow cells. The correlation of cytogenetic abnormalities and CML progression was explored in combination with ABL tyrosine point mutations. Result: Of 387 patients with positive BCR-ABL fusion gene assayed by fluorescence in situ hybridization (FISH) technique, 94.1% (364/387) patients were Ph positive and 5.9% (23/387) Ph negative; 320 patients (87.9%) had a translocation t (9;22) (q34;q11) and 5 (1.4%) a variant translocation t (v;22) . Additional cytogenetic aberrations (ACA) at diagnosis were found in 10.7% (39/387) Ph⁺ patients, major route ACA in 22 (56.4%) cases and minor route ACA in 15 (38.5%) cases and 2 patients (5.1%) lacked the Y chromosome (−Y) ; 23.4% (71/303) patients occurred ACA during TKI treatment and the most frequent abnormalities were abnormal chromosome numbersd, which were likely associated with high proportion of disease progression (χ²=168.21, P<0.001) and ABL tyrosine point mutations (χ²=29.04, P<0.001) . Newly diagnosed CML-CP patients with t (9;22) (q34;q11) had a longer event-free survival (EFS) and disease-free survival (DFS) rates than that of patients with ACA (P=0.037; P=0.003) , while the overall survival (OS) had no significant differences (P=0.209) . As for CML-CP patients that occurred ACA during TKI therapy would have a marked low OS, EFS and DFS (all P<0.001) compared with no ACA occurred patients. Survival of advanced patients that occurred ACA were dramatically reduced. Conclusion ACA often emerged during the disease progress in CML patients, regular and timely detection of chromosomes karyotype and ABL tyrosine point mutations during TKI treatment was important for therapeutic evaluation, progress and prognosis of CML.

Objective: To investigate the clinical implications of p16 gene deletion in adult Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph⁺ ALL) . Methods: Retrospective analysis of clinical, immunophenotypic, cytogenetics, molecular characteristics and prognosis of 80 newly diagnosed Ph⁺ ALL patients with p16 deletion. Results: Of 80 adult Ph⁺ ALL, the prevalence of p16 gene deletion was 31.3%. p16 gene deletion carriers frequently accompanied with high WBC counts (WBC≥30×10⁹/L) and CD20 expression. The incidence of complex chromosome abnormality in p16 gene deletion group was higher than that in non-deletion group, with alternations in chromosome 7, 8, 19 and der (22) more frequently observed. There was no difference occurred between patients with or without p16 gene deletion in complete remission (CR) rate following induction chemotherapy combined with tyrosine kinase inhibitors (TKIs) . However, after three cycles of chemotherapy, the MMR and CMR rate in the p16 gene deletion group was lower than patients with wild-type p16 gene (P=0.034, P=0.036) . The p16 gene deletion patients showed no significant differences in MMR, CMR and relapse rate between Imatinib or Dasatinib plus chemotherapy (P>0.05) . Deletion of p16 gene was significantly associated with poor outcomes including worse overall survival (OS) (37.1% vs 54.1%, P=0.037) , lower disease free-survival (DFS) (12.4% vs 45.9%, P=0.026) , and increased cumulative incidence of relapse (P=0.033) . Among the 25 patients with p16 deletion, 14 underwent allo-HSCT and the median survival was 21 months, better than that of patients received chemotherapy alone (12 months) (P=0.030) . Conclusion This study indicated that deletion of p16 was associated with poor prognosis in adult Ph⁺ ALL, and the utility of second-generation TKI (Dasatinib) does not necessarily have an edge on efficacy over Imatinib, but allo-HSCT has the potential of elongating life expectancy. It is an important significance to define the status of p16 in Ph⁺ ALL for predicting prognosis and guiding therapy decision-making.

Objective To establish a transfer model for excess relative risk (ERR) for radiation-related leukemia from Japanese population to Chinese population.Methods Combined ERR of several subtypes of leukemia published in 1994, with the corresponding leukemia baseline incidence rates obtained from Cancer Incidence in Five Continents Vol.Ⅸ (CI5-Ⅸ) for Japanese population and Chinese population, a weighted risk transfer model was employed between an additive model and a multiplicative model, to execute ERR transfer.Results A range of weighing factors was proposed for risk transfer models:weighing factor was 0.4 for male and 0.3 for female, acute lymphoblastic leukemia, acute myeloid leukemia and chronic myeloid leukemia.The uncertainty for ERR transfer was characterized by lognormal distribution.Conclusions Based on the difference of baseline incidence rate for subtypes of leukemia between Japanese population and Chinese population, the transfer model and these weighing factors discussed in the present study could be applicable to transfer ERR for radiation-related leukemia from Japanese population to Chinese population.