Analysis of the genomic landscape of primary central nervous system lymphoma using whole-genome sequencing in Chinese patients / 医学前沿

Primary central nervous system lymphoma (PCNSL) is an uncommon non-Hodgkin's lymphoma with poor prognosis. This study aimed to depict the genetic landscape of Chinese PCNSLs. Whole-genome sequencing was performed on 68 newly diagnosed Chinese PCNSL samples, whose genomic characteristics and clinicopathologic features were also analyzed. Structural variations were identified in all patients with a mean of 349, which did not significantly influence prognosis. Copy loss occurred in all samples, while gains were detected in 77.9% of the samples. The high level of copy number variations was significantly associated with poor progression-free survival (PFS) and overall survival (OS). A total of 263 genes mutated in coding regions were identified, including 6 newly discovered genes (ROBO2, KMT2C, CXCR4, MYOM2, BCLAF1, and NRXN3) detected in ⩾ 10% of the cases. CD79B mutation was significantly associated with lower PFS, TMSB4X mutation and high expression of TMSB4X protein was associated with lower OS. A prognostic risk scoring system was also established for PCNSL, which included Karnofsky performance status and six mutated genes (BRD4, EBF1, BTG1, CCND3, STAG2, and TMSB4X). Collectively, this study comprehensively reveals the genomic landscape of newly diagnosed Chinese PCNSLs, thereby enriching the present understanding of the genetic mechanisms of PCNSL.

Clinical Study on the Relationship between Gene Mutation Profile and Prognosis in Pediatric Acute Lymphocyte Leukemia / 中国实验血液学杂志

OBJECTIVE@#To analyze the gene mutation profile in children with acute lymphocyte leukemia (ALL) and to explore its prognostic significance.@*METHODS@#Clinical data of 249 primary pediatric ALL patients diagnosed and treated in the Department of Hematological Oncology of Wuhan Children's Hospital from January 2018 to December 2021 were analyzed retrospectively. Next-generation sequencing (NGS) was used to obtain gene mutation data and analyze the correlation between it and the prognosis of children with ALL.@*RESULTS@#227 (91.2%) were B-ALL, 22 (8.8%) were T-ALL among the 249 cases, and 178 (71.5%) were found to have gene mutations, of which 85 (34.1%) had ≥3 gene mutations. NRAS(23.7%), KRAS (22.9%),FLT3(11.2%), PTPN11(8.8%), CREBBP (7.2%), NOTCH1(6.4%) were the most frequently mutated genes, the mutations of KRAS, FLT3, PTPN11, CREBBP were mainly found in B-ALL, the mutations of NOTCH1 and FBXW7 were mainly found in T-ALL. The gene mutation incidence of T-ALL was significantly higher than that of B-ALL (χ2= 5.573，P＜0.05) and were more likely to have co-mutations (P＜0.05). The predicted 4-year EFS rate (47.9% vs 88.5%, P＜0.001) and OS rate (53.8% vs 94.1%, P＜0.001) in children with tp53 mutations were significantly lower than those of patients without tp53 mutations. Patients with NOTCH1 mutations had higher initial white blood cell count （128.64×109/L vs 8.23×109/L，P＜0.001）, and children with NOTCH1 mutations had a lower 4-year EFS rate than those of without mutations (71.5% vs 87.2%, P＝0.037).@*CONCLUSION@#Genetic mutations are prevalent in childhood ALL and mutations in tp53 and NOTCH1 are strong predictors of adverse outcomes in childhood ALL, with NGS contributing to the discovery of genetic mutations and timely adjustment of treatment regimens.

Analysis of genotypes and phenotypes of three children with Cornelia de Lange syndrome / 中华医学遗传学杂志

OBJECTIVE@#To analyze the clinical phenotype and results of genetic testing in three children with Cornelia de Lange syndrome (CdLS).@*METHODS@#Clinical data of the children and their parents were collected. Peripheral blood samples of the pedigrees were collected for next generation sequencing analysis.@*RESULTS@#The main clinical manifestations of the three children have included growth delay, mental retardation, peculiar facies and other accompanying symptoms. Based on the criteria proposed by the International Diagnostic Consensus, all three children were suspected for CdLS. As revealed by whole exome sequencing, child 1 has harbored NIPBL gene c.5567_5569delGAA insTAT missense variant, child 2 has harbored SMC1A gene c.607A>G missense variant, and child 3 has harbored HDAC8 gene c.628+1G>A splicing variant. All of the variants were de novo in origin.@*CONCLUSION@#All of the children were diagnosed with CdLS due to pathogenic variants of the associated genes, among which the variants of NIPBL and HDAC8 genes were unreported previously. Above finding has enriched the spectrum of pathogenic variants underlying CdLS.

Biallelic mutations in WDR12 are associated with male infertility with tapered-head sperm / 亚洲男科学杂志(英文版)

Teratozoospermia is a rare disease associated with male infertility. Several recurrent genetic mutations have been reported to be associated with abnormal sperm morphology, but the genetic basis of tapered-head sperm is not well understood. In this study, whole-exome sequencing (WES) identified a homozygous WD repeat domain 12 (WDR12; p.Ser162Ala/c.484T>G) variant in an infertile patient with tapered-head spermatozoa from a consanguineous Chinese family. Bioinformatic analysis predicted this mutation to be a pathogenic variant. To verify the effect of this variant, we analyzed WDR12 protein expression in spermatozoa of the patient and a control individual, as well as in the 293T cell line, by Western blot analysis, and found that WDR12 expression was significantly downregulated. To understand the role of normal WDR12, we evaluated its mRNA and protein expression in mice at different ages. We observed that WDR12 expression was increased in pachytene spermatocytes, with intense staining visible in round spermatid nuclei. Based on these results, the data suggest that the rare biallelic pathogenic missense variant (p.Ser162Ala/c.484T>G) in the WDR12 gene is associated with tapered-head spermatozoa. In addition, after intracytoplasmic sperm injection (ICSI), a successful pregnancy was achieved. This finding indicates that infertility associated with this WDR12 homozygous mutation can be overcome by ICSI. The present results may provide novel insights into understanding the molecular mechanisms of male infertility.

Analysis of clinical phenotype and pathogenic variant of a fetus with Cornelia de Lange syndrome type II / 中华医学遗传学杂志

OBJECTIVE@#To explore the prenatal ultrasonographic features and genetic basis for an abortus suspected for type II Cornelia de Lange syndrome (CdLS2).@*METHODS@#A fetus diagnosed with CdLS2 at the Shengjing Hospital Affiliated to China Medical University on September 3, 2019 was selected as the study subject. Clinical data of the fetus and family history was collected. Following induced labor, whole exome sequencing was carried out on the abortus. Candidate variant was verified by Sanger sequencing and bioinformatic analysis.@*RESULTS@#Prenatal ultrasonography (33 weeks of pregnancy) has revealed multiple anomalies in the fetus, which included slightly widened cavity of septum pellucidum, blurred corpus callosum, slightly reduced frontal lobe volume, thin cortex, fusion of lateral ventricles, polyhydramnios, small stomach bubble, and digestive tract atresia. Whole exome sequencing has revealed a heterozygous c.2076delA (p.Lys692Asnfs*27) frameshifting variant in the SMC1A gene, which was found in neither parent and was rated as pathogenic based on the guidelines of American College of Medical Genetics and Genomics (ACMG).@*CONCLUSION@#The CdLS2 in this fetus may be attributed to the c.2076delA variant of the SMC1A gene. Above finding has provided a basis for genetic counseling and assessment of reproductive risk for this family.

miR-218-5p Targeting TPX2 Regulates p53 Pathway and Inhibits Malignant Progression of Lung Adenocarcinoma / 中国肺癌杂志

BACKGROUND@#Lung adenocarcinoma (LUAD) is a major subtype of lung cancer, and its treatment and diagnosis remain a hot research topic. Targeting protein for Xenopus kinesin-like protein 2 (TPX2) is highly expressed in a variety of cancer cells and may be associated with the progression of LUAD. This study aimed to investigate the effect of TPX2 on the malignant progression of LUAD cells and the regulatory mechanisms.@*METHODS@#The expression of gene TPX2 in LUAD tissues from The Cancer Genome Atlas (TCGA) database was analyzed by bioinformatics analysis techniques. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression levels of TPX2 and miR-218-5p in human lung normal cell lines and human LUAD cell lines. Western blot was used to detect TPX2 protein expression in cell lines and its effect on the expression of key proteins in the p53 signaling pathway. The relationship between TPX2 and miR-218-5p was predicted using bioinformatics and verified by dual luciferase reporter gene assay. Cell counting kit-8 (CCK-8) assay, cell clone formation, cell scratching, Transwell assay, and flow cytometry were used to detect the effects of miR-218-5p and TPX2 on LUAD cell function.@*RESULTS@#TPX2 was significantly overexpressed in LUAD cells, and knockdown of TPX2 inhibited LUAD cell proliferation, migration, and invasion, promoted apoptosis and induced G2/M phase block, and promoted the expression of key proteins in the p53 signaling pathway. miR-218-5p, an upstream regulator of TPX2, could inhibit its expression. Overexpression of miR-218-5p eliminated the malignant development caused by high expression of TPX2, inhibited the malignant processes of LUAD cells such as proliferation and migration as well as promoted the p53 signaling pathway.@*CONCLUSIONS@#miR-218-5p targets and inhibits TPX2 expression and exerts an inhibitory effect on the malignant progression of LUAD cells via p53.

MBOAT1 homozygous missense variant causes nonobstructive azoospermia / 亚洲男科学杂志(英文版)

Nonobstructive azoospermia (NOA) is a common cause of infertility and is defined as the complete absence of sperm in ejaculation due to defective spermatogenesis. The aim of this study was to identify the genetic etiology of NOA in an infertile male from a Chinese consanguineous family. A homozygous missense variant of the membrane-bound O-acyltransferase domain-containing 1 (MBOAT1) gene (c.770C>T, p.Thr257Met) was found by whole-exome sequencing (WES). Bioinformatic analysis also showed that this variant was a pathogenic variant and that the amino acid residue in MBOAT1 was highly conserved in mammals. Quantitative polymerase chain reaction (Q-PCR) analysis showed that the mRNA level of MBOAT1 in the patient was 22.0% lower than that in his father. Furthermore, we screened variants of MBOAT1 in a broader population and found an additional homozygous variant of the MBOAT1 gene in 123 infertile men. Our data identified homozygous variants of the MBOAT1 gene associated with male infertility. This study will provide new insights for researchers to understand the molecular mechanisms of male infertility and will help clinicians make accurate diagnoses.

Diagnosis of a child with mitochondrial myopathy and cerebellar atrophy with ataxia due to compound heterozygous variants of MSTO1 gene / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic basis for a child with myopathy and cerebellar atrophy with ataxia.@*METHODS@#Clinical examinations and laboratory testing were carried out for the patient. The proband and the parents' genomic DNA was extracted from peripheral blood samples and subjected to trio whole-exome sequencing. Candidate variant was validated by Sanger sequencing.@*RESULTS@#The 1-year-and-8-month-old boy manifested motor developmental delay, ataxia, hypomyotonia, increased serum creatine kinase. Cranial MRI showed cerebellar atrophy with progressive aggravation. Genetic testing revealed that the patient has harbored compound heterozygous variants of the MSTO1 gene, namely c.13delG (p.Ala5ProfsTer68) and c.971C>T (p.Thr324Ile), which were respectively inherited from his mother and father. The former was unreported previously and was predicted to be likely pathogenic, whilst the latter has been reported previously and was predicted to be of uncertain significance.@*CONCLUSION@#The compound heterozygous c.13delG (p.Ala5ProfsTer68) and c.971C>T (p.Thr324Ile) variants probably underlay the disease in the proband. Above finding has enriched the spectrum of MSTO1 gene variants underlying mitochondrial myopathy and cerebellar atrophy with ataxia.

Bioinformatics-based identification of key genes CDC5L and related pathways in osteosarcoma and Ewing's sarcoma / 中国骨伤

OBJECTIVE@#Osteosarcoma(OS) and Ewing's sarcoma (EWS) are the two most common primary malignant bone tumors in children. The aim of the study was to identify key genes in OS and EWS and investigate their potential pathways.@*METHODS@#Expression profiling (GSE16088 and GSE45544) were obtained from GEO DataSets. Differentially expressed genes were identified using GEO2R and key genes involved in the occurrence of both OS and EWS were selected using venn diagram. Gene ontology and pathway enrichment analyses were performed for the ensembl. Protein-protein interaction (PPI) networks were established by STRING. Further, UCSC was used to predict the transcription factors of the cell division cycke 5-like(CDC5L) gene, and GEPIA was used to analyze the correlation between the transcription factors and the CDC5L gene.@*RESULTS@#The results showed that CDC5L gene was the key gene involved in the pathogenesis of OS and EWS. The gene is mainly involved in mitosis, and is related to RNA metabolism, processing of capped intron-containing pre-mRNA, mRNA and pre-mRNA splicing.@*CONCLUSION@#CDC5L, as a key gene, plays a role in development of OS and EWS, which may be reliable targets for diagnosis and treatment of these primary malignant tumors.

Molecular Mechanism of Aurora Kinase A Regulating the Meiosis of Oocyte / 中国医学科学院学报

Aurora kinase A (AURKA),a family member of aurora kinases,is involved in mitotic entry,maturation and separation of centrosome,assembly and stabilization of bipolar spindle,and condensation and separation of chromosome.Studies have demonstrated that AURKA plays a similar role in meiosis,while the specific mechanism and the similarities and differences in its role between meiosis and mitosis remain unclear.Therefore,we reviewed the studies about the localization and activation of AURKA in oocyte meiosis,and compared the role of AURKA in regulating spindle formation,activating spindle assembly checkpoint,and correcting the kinetochore-microtubule attachment between the meiosis of oocytes and the mitosis of somatic cells.This review will lay a theoretical foundation for revealing the mechanism of AURKA in the regulation of cell division and for the clinical research related to cancer and reproduction.

A case of neonatal Cornelia de Lange syndrome caused by a novel variant of SMC1A gene / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic etiology of a neonate with suggestive features of Cornelia de Lange Syndrome (CdLS).@*METHODS@#Chromosome karyotyping, copy number variation sequencing (CNV-seq) and whole exome sequencing (WES) were carried out for the child. Meanwhile, peripheral venous blood samples were taken from his parents for verifying the suspected pathogenic variants detected in the child.@*RESULTS@#The child has exhibited developmental delay, microcephaly, ptosis, micrognathia, and low ear setting, and was suspected as CdLS. No abnormality was found by karyotyping and CNV-seq analysis. WES has detected 5 heterogeneous variants and 1 hemizygous variant on the X chromosome. Combining the genetic pattern and result of family verification, a hemizygous C.3500T>C (p.ile1167thr) of the SMC1A gene was predicted to underlay the clinical manifestations of the patient. This variant was not recorded in the dbSNP and gnomAD database. PolyPhen2, Provean, SIFT all predicted the variant to be harmful, and PhastCons conservative prediction is was a conservative mutation. ACMG variant classification standard evidence supports are PM2, PP2, and PP3.@*CONCLUSION@#The novel c.3500T>C (p.Ile1167Thr) missense mutation of the SMC1A gene probably underlay the genetic etiology of CdLS in this child. Above results has enriched the mutation spectrum of CdLS type II, and facilitated clinical counseling for this family.

Effect of CDK1 Interferes with the Regulation of PLK1, Aurora B and TRF1 on the Proliferation of Leukemia Cells / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of CDK1 interference regulation of PLK1, Aurora B and TRF1 on the proliferation of leukemia cells.@*METHODS@#The human myelogenous leukemia cell line HL-60 was selected as the research object, and the effect of TRF1 expression and its changes on cell proliferation and cycle was investigated by regulating intracellular CDK1 expression. The objects were divided into 5 groups, including control group, shRNA-NC group, CDK1-shRNA group, pcDNA group and pcDNA-CDK1 group. RT-PCR was used to detect the CDK1 expression of cells in each group; colony formation was used to detect the proliferation of the cells. Western blot was used to detect the expression of CDK1, PLK1, Aurora B, TRF1, and cyclin p53, p27, cyclinA.@*RESULTS@#The phosphorylation level of PLK1, Aurora B and the expression of TRF1 in the CDK1-shRNA group were significantly down-regulated as compared with those in the control group (P<0.05). Compared with the control group, the cells in CDK1-shRNA group showed lower clone formation rate, the increasing of cycle-associated proteins p53 and p27 and the decreasing of cyclinA expression (P<0.05). It was shown that interfered CDK1 expression could inhibit the proliferation of HL-60 cells and prolong the time that they enter mitosis, thereby extending the cell cycle. Compared with the control group, the overexpressed CDK1 in the pcDNA-CDK1 group made the phosphorylation level of PLK1, Aurora B, and TRF1 expression increase significantly (P<0.05), also the colony formation rate (P<0.05). The cycle-related proteins p53 and p27 was down-regulated, while cyclinA expression was up-regulate significantly (P<0.05). The results indicted that overexpressed CDK1 could stimulate adverse reactions, thereby promoting the proliferation of HL-60 cells and shortening the cell cycle.@*CONCLUSION@#Knocking out CDK1 can inhibit the phosphorylation of PLK1 and Aurora B and negatively regulate TRF1, thereby inhibiting the proliferation of leukemia cells.

Analysis of ALMS1 gene variants in seven patients with Alström syndrome / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic basis for 7 patients with Alström syndrome.@*METHODS@#DNA was extracted from peripheral blood samples of the patients and their parents. Whole exome sequencing was carried out for the patients. Suspected variant was verified by Sanger sequencing and bioinformatic analysis.@*RESULTS@#Genetic testing revealed 12 variants of the ALMS1 gene among the 7 patients, including 7 nonsense and 5 frameshift variants, which included c.5418delC (p.Tyr1807Thrfs*23), c.10549C>T (p.Gln3517*), c.9145dupC (p.Thr3049Asnfs*12), c.10819C>T (p.Arg3607*), c.5701_5704delGAGA (p.Glu1901Argfs*18), c.9154_9155delCT (p.Cys3053Serfs*9), c.9460delG (p.Val3154*), c.9379C>T (p.Gln3127*), c.12115C>T (p.Gln4039*), c.1468dupA (p.Thr490Asnfs*15), c.10825C>T (p.Arg3609*) and c.3902C>A (p.Ser1301*). Among these, c.9154_ 9155delCT, c.9460delG, c.9379C>T, and c.1468dupA were unreported previously. Based on the standards and guidelines of American College of Medical Genetics and Genomics, the c.9379C>T and c.12115C>T variants of the ALMS1 gene were predicted to be likely pathogenic (PVS1+PM2), whilst the other 10 variants were predicted to be pathogenic (PVS1+ PM2+ PP3+PP4).@*CONCLUSION@#ALMS1 variants probably underlay the Alström syndrome in the 7 patients, and genetic testing can provide a basis for the clinical diagnosis of this syndrome. The discovery of four novel variants has expanded the mutational spectrum of Alström syndrome.

Phenotypic and mutation analysis of a fetus with Cornelia de Lange syndrome Ⅰ / 中华医学遗传学杂志

OBJECTIVE@#To carry out genetic testing for an abortus suspected with Cornelia de Lange syndrome (CdLS).@*METHODS@#History of gestation and the family was taken. Combined with prenatal ultrasonography and the phenotype of the abortus, a diagnosis was made for the proband. Fetal tissue and peripheral blood samples of its parents were collected for the extraction of genomic DNA. Whole exome sequencing was carried out to detect mutations related to the phenotype. Suspected mutations were verified in the parents through Sanger sequencing.@*RESULTS@#Prenatal ultrasound found that the forearms and hands of the fetus were anomalous, in addition with poorly formed vermis cerebellum, slight micrognathia, and increased echo of bilateral renal parenchyma. Examination of the abortus has noted upper limb and facial malformations. Whole exome sequencing revealed that the fetus carried a heterozygous c.2118delG (p.Lys706fs) frameshift mutation of the NIPBL gene. The same mutation was not found in either parent.@*CONCLUSION@#The heterozygous c.2118delG (p.Lys706fs) frameshift mutation of the NIPBL gene probably underlies the CdLS in the fetus. Above finding has provided a basis for the genetic counseling for the family.

Prognostic significance of the mad1l1 1673 g:a polymorphism in ovarian adenocarcinomas

Abstract Background: Ovarian cancer is the most lethal gynecologic cancer. Although most patients respond adequately to the first-line therapy, up to 85% experience a recurrence of disease, which carries a poor prognosis. Mitotic arrest deficiency 1 is a protein that helps in the assembly of the mitotic spindle assembly checkpoint by preventing anaphase until all chromatids are properly aligned. A single-nucleotide polymorphism in the MAD1L1 gene is prevalent in patients with advanced epithelial ovarian cancer and alters the way in which it responds to chemotherapy. Objective: The objective of the study was to study the relationship between the rs1801368 polymorphism of MAD1L1 and prognosis of ovarian adenocarcinoma. Methods: A total of 118 patients in whom the MAD1L1 gene was sequenced were analyzed using descriptive and comparative statistics. Results: Patients carrying the wild-type genotype had a higher distribution of early-stage disease. Having a MAD1L1 polymorphic allele increased the risk of being non-sensitive to chemotherapy. The median disease-free survival for patients with the wild-type MAD1L1 was 46.93 months, compared to 10.4 months for patients with at least one polymorphic allele. Conclusions: The rs1801368 polymorphism of MAD1L1 gene worsens prognosis in patients with ovarian adenocarcinoma. Traditional therapy for ovarian cancer might not be optimal in patients carrying this polymorphism.

REC8 suppresses tumor angiogenesis by inhibition of NF-κB-mediated vascular endothelial growth factor expression in gastric cancer cells

BACKGROUND: Tumor angiogenesis is an essential event for tumor growth and metastasis. It has been showed that REC8, a component of the meiotic cohesion complex, played a vital role in Epithelial-Mesenchymal Transition (EMT) in gastric cancer. However, the role of REC8 in gastric cancer angiogenesis remains to be identified. RESULTS: Inhibition of REC8 expression in gastric cancer cells contributed to tumor angiogenesis in the gastric cancer microenvironment. The clinical analysis demonstrated that the loss of REC8 in gastric cancer with enrichment of MVD. Depletion of REC8 expression in gastric cancer cells significantly increased tube formation of human umbilical vein endothelial cells (HUVECs), which is attributed to enhancement of vascular endothelial growth factor (VEGF) secretion caused by REC8 slicing. While addition of neutralizing antibody targeted VEGF into supernatant drastically reversed the effect of REC8 loss in gastric cancer cells on tube formation. Mechanistic analyses indicated that ablation of REC8 promotes nuclear factor-κB (NF-κB) p65 activity and its downstream gene VEGF expression, leading to tube formation. CONCLUSIONS: These results demonstrated a novel REC8 function that suppressed tumor angiogenesis and progression by attenuation of VEGF in gastric cancer microenvironment.

Mining Polo-Box domain of Polo-like kinase 1 as a new therapeutic target for cancer / 生物工程学报

Polo-like kinase 1 (Plk1) is widely regarded as one of the most promising targets for cancer therapy due to its essential role in cell division and tumor cell survival. At present, most Plk1 inhibitors have been developed based on kinase domain, some of which are in clinical trial. However, inhibitors targeting kinase domain face off-target effect and drug resistance owing to the conserved nature and the frequent mutations in the ATP-binding pocket. In addition to a highly conserved kinase domain, Plk1 also contains a unique Polo-Box domain (PBD), which is essential for Plk1's subcellular localization and mitotic functions. Inhibitors targeting Plk1 PBD show stronger selectivity and less drug resistance for cancer therapy. Therefore, Plk1 PBD is an attractive target for the development of anti-cancer agents. In this review, we will summarize the up-to date drug discovery for targeting Plk1 PBD, including the molecular structure and cellular functions of Plk1 PBD. Small-molecule inhibitors targeting Plk1 PBD not only provide an opportunity to specifically inhibit Plk1 activity for cancer treatment, but also unveil novel biological basis regarding the molecular recognition of Plk1 and its substrates.

Carfilzomib inhibits the growth of lung adenocarcinoma via upregulation of Gadd45a expression / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Proteasome inhibitors have shown remarkable success in the treatment of hematologic neoplasm. There has been a lot of attention to applying these drugs for solid tumor treatment. Recent preclinical study has signified the effectiveness on cell proliferation inhibition in lung adenocarcinoma treated by carfilzomib (CFZ), a second generation proteasome inhibitor. However, no insight has been gained regarding the mechanism. In this study, we have systematically investigated the CFZ functions in cell proliferation and growth, cell cycle arrest, and apoptosis in lung adenocarcinoma cells. Flow cytometry experiments showed that CFZ significantly induced G2/M cell cycle arrest and apoptosis in lung adenocarcinoma. MTS and colony formation assays revealed that CFZ substantially inhibited survival of lung adenocarcinoma cells. All results were consistently correlated to the upregulation expression of Gadd45a, which is an important gene in modulating cell cycle arrest and apoptosis in response to physiologic and environmental stresses. Here, upregulation of Gadd45a expression was observed after CFZ treatment. Knocking down Gadd45a expression suppressed G2/M arrest and apoptosis in CFZ-treated cells, and reduced cytotoxicity of this drug. The protein expression analysis has further identified that the AKT/FOXO3a pathway is involved in Gadd45a upregulation after CFZ treatment. These findings unveil a novel mechanism of proteasome inhibitor in anti-solid tumor activity, and shed light on novel preferable therapeutic strategy for lung adenocarcinoma. We believe that Gadd45a expression can be a highly promising candidate predictor in evaluating the efficacy of proteasome inhibitors in solid tumor therapy.

The role of tyrosine phosphatase Shp2 in spermatogonial differentiation and spermatocyte meiosis / 亚洲男科学杂志(英文版)

The transition from spermatogonia to spermatocytes and the initiation of meiosis are key steps in spermatogenesis and are precisely regulated by a plethora of proteins. However, the underlying molecular mechanism remains largely unknown. Here, we report that Src homology domain tyrosine phosphatase 2 (Shp2; encoded by the protein tyrosine phosphatase, nonreceptor type 11 [Ptpn11] gene) is abundant in spermatogonia but markedly decreases in meiotic spermatocytes. Conditional knockout of Shp2 in spermatogonia in mice using stimulated by retinoic acid gene 8 (Stra8)-cre enhanced spermatogonial differentiation and disturbed the meiotic process. Depletion of Shp2 in spermatogonia caused many meiotic spermatocytes to die; moreover, the surviving spermatocytes reached the leptotene stage early at postnatal day 9 (PN9) and the pachytene stage at PN11-13. In preleptotene spermatocytes, Shp2 deletion disrupted the expression of meiotic genes, such as disrupted meiotic cDNA 1 (Dmc1), DNA repair recombinase rad51 (Rad51), and structural maintenance of chromosome 3 (Smc3), and these deficiencies interrupted spermatocyte meiosis. In GC-1 cells cultured in vitro, Shp2 knockdown suppressed the retinoic acid (RA)-induced phosphorylation of extracellular-regulated protein kinase (Erk) and protein kinase B (Akt/PKB) and the expression of target genes such as synaptonemal complex protein 3 (Sycp3) and Dmc1. Together, these data suggest that Shp2 plays a crucial role in spermatogenesis by governing the transition from spermatogonia to spermatocytes and by mediating meiotic progression through regulating gene transcription, thus providing a potential treatment target for male infertility.

The association of stromal antigen 3 (STAG3) sequence variations with spermatogenic impairment in the male Korean population / 亚洲男科学杂志(英文版)

The stromal antigen 3 (STAG3) gene, encoding a meiosis-specific cohesin component, is a strong candidate for causing male infertility, but little is known about this gene so far. We identified STAG3 in patients with nonobstructive azoospermia (NOA) and normozoospermia in the Korean population. The coding regions and their intron boundaries of STAG3 were identified in 120 Korean men with spermatogenic impairments and 245 normal controls by using direct sequencing and haplotype analysis. A total of 30 sequence variations were identified in this study. Of the total, seven were exonic variants, 18 were intronic variants, one was in the 5'-UTR, and four were in the 3'-UTR. Pathogenic variations that directly caused NOA were not identified. However, two variants, c.3669+35C>G (rs1727130) and +198A>T (rs1052482), showed significant differences in the frequency between the patient and control groups (P = 0.021, odds ratio [OR]: 1.79, 95% confidence interval [CI]: 1.098-2.918) and were tightly linked in the linkage disequilibrium (LD) block. When pmir-rs1052482A was cotransfected with miR-3162-5p, there was a substantial decrease in luciferase activity, compared with pmir-rs1052482T. This result suggests that rs1052482 was located within a binding site of miR-3162-5p in the STAG3 3'-UTR, and the minor allele, the rs1052482T polymorphism, might offset inhibition by miR-3162-5p. We are the first to identify a total of 30 single-nucleotide variations (SNVs) of STAG3 gene in the Korean population. We found that two SNVs (rs1727130 and rs1052482) located in the 3'-UTR region may be associated with the NOA phenotype. Our findings contribute to understanding male infertility with spermatogenic impairment.