Objective:To investigate the feasibility of segmented total marrow and lymphoid irradiation(TMLI)using Helical TomoTherapy(HT)in adults,and to analyze the impact of positional errors in the treatment on the actually received dose.Methods:Imaging data from four adult patients with acute myeloid leukemia were treated at the First Medical Center of the Chinese PLA General Hospital between November 2022 and December 2024 were retrospectively selected.The prescribed dose was 12 Gy in 3 fractions,and the heights range of patient was from 162 to 178 cm.The planning target volume(PTV)included the entire bone marrow and lymphatic drainage regions.The mid-femur was used as the dividing landmark,and the segmented location of head-in first(upper segmentation)and foot-in first(lower segmentation)were adopted to design plan for each segmentation.The upper segmentation used helical tomotherapy,and the lower segmentation used tomo direct therapy.The dosimetric parameters of planning target volume,and organ at risk(OAR)included eye lens,testicles and lungs were analyzed.The located computed tomography(CT)images of patients were used to simulate the changes of dosimetric parameters of each planning target region and OAR when the positions of patients moved 3,5,10 and 15 mm to outside of rack aperture.The changes of dose distribution of connection region of upper and lower segmentation were further analyzed.Results:The segmented irradiation technique can better achieve the cover(95%)of whole target region of TMLI under the condition of protecting normal organs.Under the condition with different position errors,the cover rate of the most of target region significant decreased.The largest dose of right lens increased 9.272%than primary under the 5 mm position error,and that of left and right lens increased respectively 40.199%and 67.898%under the 10 mm position error,and the max dose and mean dose of testicles increased respectively 5.847%and 5.240%under the 3mm position error.The mean dose of whole lung increased all under the error at different grades,which increased 3.021%under the 10mm position error.The non-uniformity of dose distribution of connection region increased after the position error was introduced,and the largest variation range reached to 4 Gy.Conclusion:The technique of segmented radiotherapy of Helical Tomotherapy has feasibility for TMLI of adult.But the error of the position of patients can reduce the cover of target region,and decrease the control rate for target region,and can lead to the occurrences of excessive doses on eye lens,testicles,lung and other OAR at the same time,which can increase the probability of occurring OAR injury.It should be high pay attention in treatment and implementation.

Objective:To investigate the feasibility of segmented total marrow and lymphoid irradiation(TMLI)using Helical TomoTherapy(HT)in adults,and to analyze the impact of positional errors in the treatment on the actually received dose.Methods:Imaging data from four adult patients with acute myeloid leukemia were treated at the First Medical Center of the Chinese PLA General Hospital between November 2022 and December 2024 were retrospectively selected.The prescribed dose was 12 Gy in 3 fractions,and the heights range of patient was from 162 to 178 cm.The planning target volume(PTV)included the entire bone marrow and lymphatic drainage regions.The mid-femur was used as the dividing landmark,and the segmented location of head-in first(upper segmentation)and foot-in first(lower segmentation)were adopted to design plan for each segmentation.The upper segmentation used helical tomotherapy,and the lower segmentation used tomo direct therapy.The dosimetric parameters of planning target volume,and organ at risk(OAR)included eye lens,testicles and lungs were analyzed.The located computed tomography(CT)images of patients were used to simulate the changes of dosimetric parameters of each planning target region and OAR when the positions of patients moved 3,5,10 and 15 mm to outside of rack aperture.The changes of dose distribution of connection region of upper and lower segmentation were further analyzed.Results:The segmented irradiation technique can better achieve the cover(95%)of whole target region of TMLI under the condition of protecting normal organs.Under the condition with different position errors,the cover rate of the most of target region significant decreased.The largest dose of right lens increased 9.272%than primary under the 5 mm position error,and that of left and right lens increased respectively 40.199%and 67.898%under the 10 mm position error,and the max dose and mean dose of testicles increased respectively 5.847%and 5.240%under the 3mm position error.The mean dose of whole lung increased all under the error at different grades,which increased 3.021%under the 10mm position error.The non-uniformity of dose distribution of connection region increased after the position error was introduced,and the largest variation range reached to 4 Gy.Conclusion:The technique of segmented radiotherapy of Helical Tomotherapy has feasibility for TMLI of adult.But the error of the position of patients can reduce the cover of target region,and decrease the control rate for target region,and can lead to the occurrences of excessive doses on eye lens,testicles,lung and other OAR at the same time,which can increase the probability of occurring OAR injury.It should be high pay attention in treatment and implementation.

Objective To exploring the correlation between the expression level of serum carbohydrate antigen 211(CA211)and the number of natural killer(NK)cells and prognosis in non-small cell lung cancer(NSCLC)pa-tients.Methods 132 NSCLC patients admitted to the Affiliated Hospital of Hangzhou Normal University from June 2019 to July 2022 were selected as the test group,and 132 patients with benign lung lesions during the same period were selected as the control group.Data were collected from the laboratory to analyze the relationship between serum CA211 expression and NK cell count in NSCLC with clinical prognosis,as well as the related factors affecting the prognosis of NSCLC patients.Results The neutrophil to lymphocyte ratio(NLR)and serum CA211 expression in the test group were higher than those in the control group(P＜0.05),while the count of NK cells was less than that in the control group(P＜0.05);The expression level of serum CA211 in the test group was negatively cor-related with the count of NK cells(r=-0.405,P＜0.001);There were significant differences in lymph node metastasis and TNM staging among patients with different levels of serum CA211 expression and NK cell count(P＜0.05);The one-year survival rate of patients with low expression of CA211 was significantly higher than that of patients with high expression of CA211(P＜0.05),and the one-year survival rate of patients with high count of NK cells was higher than that of patients with low count of NK cells(P＜0.05);Lymph node metasta-sis,TNM staging and CA211 level were all risk factors affecting the prognosis of NSCLC patients(P＜0.05),while counting of NK cells was protective factor for the prognosis of NSCLC patients(P＜0.05).Conclusions The level of serum CA211 and the number of NK cells in NSCLC patients are closely related to pathological characteristics and clinical prognosis.

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

OBJECTIVETo investigate the effects of microRNA(miRNA)-29b on the proliferation and migration of breast cancer cells and its molecular mechanism.

METHODSThe recombinant lentiviral expression vector (lenti-miRNA-29b) was constructed and transfected into 293T cells to obtain lentivirus particles that were used to infect breast cancer MCF-7 cells. Transfection efficiency of lenti-miRNA-29b in MCF-7 cells was identified by the expression of green fluorescent protein (GFP). The expression of miRNA-29b was detected by real-time PCR. The cell proliferation and migration were detected by CCK8 assay and Transwell assay, respectively. The bioinformatics softwares were used to predict and screen the downstream target genes regulated by miRNA-29b, which were verified by double luciferase reporter gene assay, RT-PCR and Western blot. The effects of screened target gene RTKN on the growth and migration of MCF-7 cells were verified by RTKN siRNA.

RESULTSRecombinant lentiviral expression vector of miRNA-29b were successfully constructed. About 90% and 60% of the breast cancer cells showed green fluorescence in lenti-miRNA-29b and lenti-miRNA-NC groups, respectively. The expression of miRNA-29b in lenti-miRNA-29b group increased significantly compared with the lenti-miRNA-NC group and blank control group (all<0.05); the proliferation and migration ability of MCF-7 cells significantly reduced compared with the control group (all<0.05). The screening with bioinformatics softwares found that the 3'UTR coding region RTKN had the binding site to miRNA-29b; the dual luciferase reporter gene assay showed that the luciferase activity decreased significantly after the MCF-7 cells were co-transfected with wild type RTKN-WT-3'UTR and miRNA-29b mimics report gene vector (<0.05). The RTKN proteins in MCF-7 cells were significantly decreased after transfection with siRNA-RTKN, and the proliferation and migration ability of MCF-7 cells were significantly reduced (all<0.05).

CONCLUSIONSMiRNA-29b can inhibit the proliferation, invasion and metastasis of breast cancer cells by inhibiting the expression of RTKN.

Objective: To explore the method of extracting chaperone antigen peptide complexes from gastric cancer stem cells and its immune function. Methods: Gastric cancer stem cells and gastric cancer cells were screened by low temperature ultrasonic lysis. After salting out and dialysis, the lysate supernatant was processed with SDS-PAGE to analyze the expression of chaperone antigen peptide complexes, and then was separated and purified with CNBr-activated SepharoseTM 4B. Reverse high pressure liquid chromatography (HPLC), SDS-PAGE and Western blotting were used to analyze the purity and nature of the acquired albumen. Lymphocyte proliferation assay and lymphocytotoxicity assay were used to ditermine the immunological activity of the chaperone-antigen peptide complexes. Results: The chaperone antigen peptide complexes of gastric cancer stem cells were prepared and identified successfully, of which the main components were the antigen peptides of HSP60, HSP70, HSP90 and HSP110. 0.75 μg and 1.00 μg HSP70-antigen peptide and 1.00 μg HSP90-antigen peptide activated lymphocytes significantly. Their A₄₉₀ values were 0.26±0.03, 0.45±0.05 and 0.32±0.04, respectively, while the corresponding doses of HSP60-antigen peptide and HSP110-antigen peptide did not activate lymphocytes. The killing rates of 1.00 μg HSP70-antigen peptide and 1.00 μg HSP70 were (45.0±2.0)% and (16.0±2.0)%, respectively, showing a significant difference (P=0.012). Similarly, the killing rates of 1.00 μg HSP90-antigen peptide and 1.00 μg HSP90 were (36.0±5.0)% and (13.0±4.0)%, respectively, also showing a significant difference (P=0.048). Conclusions The amount of chaperone antigen peptide complexes in gastric cancer cells is extremely low, but it is obviously increased in gastric cancer stem cells. After purification, the chaperone antigen peptide complexes with high purity can be prepared. The extracted chaperone antigen peptide complexes have stronger immunogenicity, and can be used to make tumor vaccine in vitro, which may have a good application value in the targeted therapy of gastric cancer.

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.

Purpose The diagnosis, treatment and prognosis of benign and malignant biliary stricture are significantly different. This study aims to evaluate the quantitative analysis of biliary structures using magnetic resonance cholangiopancreatography (MRCP) combined with dynamic contrast enhanced CT (DCE-CT).Materials and Methods The quantitative parameters of MRCP and DCE-CT imaging data from 27 patients with benign biliary stricture (benign group) and 30 patients with malignant biliary stricture (malignant group) were retrospectively analyzed. The wall thickness, stricture length and diameter, proximal ductal dilatation and degree of enhancement in two groups were compared, and its correlation was analyzed to evaluate the accuracy of MRCP and DCE-CT.Results There were significant differences in wall thickness [(3.2±2.0) mmvs (2.1±0.6) mm], stricture length [(15.8±8.1) mmvs (9.5±6.5) mm] and diameter [0 mmvs (2.0±0.9) mm], proximal ductal dilatation and the degree of enhancement [(12.7±3.6) mmvs (9.3±2.7) mm] between the two groups (t=2.825, 3.270, 4.025,P<0.001;Z=-3.909,P<0.001). Multivariable stepwise regression analysis showed that the wall thickness and diameter, and the CT HU in portal venous and equilibrium phases combined with CT plain scanning were significant predictors of malignant biliary strictures (t=-6.424-2.309,P<0.05). The sensitivity, specificity, inter-modality agreement and Youden index of MRCP and DCE-CT in diagnosing 57 patients with biliary stricture were 96.67%, 100.00%, 98.25% and 0.97, respectively; with statistical significance in predicting benign and malignant biliary stricture (AUC=0.994,P<0.001).Conclusion Using MRCP and DCE-CT, the wall thickness and diameter of the stricture, and the difference in CT HU in portal venous and equilibrium phases combined with CT plain scanning are valuable in differential diagnosis of benign and malignant biliary stricture.