This study is to investigate transcription factors involved in cisplatin resistance in ovarian cancer cells. The transcriptome of cisplatin resistant and sensitive A2780 epithelial ovarian cancer cells was obtained from GSE15372. Ovarian transcriptome data GSE62944 was downloaded from TCGA and applied for transcription regulatory network analysis. The analysis results were confirmed using quantitative polymerase chain reaction. The roles of SREBP2 in cisplatin-resistant cells were investigated by RNA inference and cell viability analysis. Transcription regulatory network analysis found that 12 transcription factors and their targets were involved in cisplatin resistant in A2780 cells. Among these factors, the targets of EZH2 and SREBP2 revealed by Transcriptional Regulatory Relationships Unraveled by Sentence-based Text mining were also enriched in differentially expressed genes between cisplatin resistant and cisplatin sensitive cells. Their targets were enriched mainly in cell cycle and cholesterol metabolic process, respectively. Bioinformatic analysis illustrated three known targets of SREBP2, namely LDLR, FDFT1, and HMGCR were increased in A2780-resistant cell lines. Additionally, the three genes and SREBP2 were also elevated in live cells after cisplatin treatment via quantitative polymerase chain reaction. Importantly, RNA inference of SREBP2 in A2780 cell line resulted in a decrease of cell viability after cisplatin treatment. SREBP2 played important roles in cisplatin resistance and cholesterol metabolic process might be a novel target for cancer therapy. Impact statement Transcriptome of cisplatin resistant and sensitive A2780 epithelial ovarian cancer cells was obtained from GSE15372 and TCGA. Twelve transcription factors and their targets were involved in cisplatin resistant. Among these factors, the targets of EZH2 and SREBP2 revealed by Transcriptional Regulatory Relationships Unraveled by Sentence-based Text mining were also enriched in differentially expressed genes. Their targets were enriched mainly in cell cycle and cholesterol metabolic process. Three targets of SREBP2, namely LDLR, FDFT1, and HMGCR were increased in A2780-resistant cell lines and were found elevated in live cells after cisplatin treatment via qPCR. RNAi of SREBP2 in A2780 cell line resulted in a decrease of cell viability after cisplatin treatment. SREBP2 played important roles in cisplatin resistance and might be a novel target for cancer therapy.

Neurodevelopmental delay accompanied unexplained dyspnea is a highly lethal disease in clinic. This study is to investigate the performance characteristics of trio whole exome sequencing (Trio-WES) in a pediatric setting by presenting our patient cohort and displaying the diagnostic yield. A total of 31 pediatric patients showing neurodevelopmental delay accompanied unexplained dyspnea were admitted to our hospital and referred for molecular genetic testing using Trio-WES. Eight genes namely MMACHC, G6PC, G6PT, ETFDH, OTC, NDUFAF5, SLC22A5, and MAGEL2 were suspected to be responsible for the onset of the clinical symptoms and 6 variants were novel. Standard interpretation according to ACMG guideline showed that the variants were pathogenic. Finally, diagnosis of methylmalonic aciduria and homocystinuria, glycogen storage disease, ornithine transcarbamylase deficiency, glutaric acidemia II, mitochondrial complex 1 deficiency, carnitine deficiency, and Schaaf-Yang syndrome was made in 12 out of the 31 patients. Trio-WES is an effective means for molecular diagnosis of infantile neurodevelopmental delay accompanied unexplained dyspnea. As for molecular etiology identification, when routine potential monogenetic inheritance patterns including de novo, autosomal recessive, autosomal dominant, and X-linked recessive inheritance analysis is negative, physicians should take into account imprinted genes.

This retrospective study aims to investigate the diagnostic yields of multiple strategies of next-generation sequencing (NGS) for children with rare neurological disorders (NDs). A total of 220 pediatric patients with NDs who visited our hospital between Jan 2017 and Dec 2018 and had undergone NGS were included. Most patients were 5 years old or younger, and the number of patients visiting the hospital decreased with age. Seizures were the most common symptom in this cohort. The positive rates for targeted NGS panels (Panel), whole-exome sequencing (WES), and copy number variation sequencing (CNVseq) were 26.5% (9/34), 36.6% (63/172), and 16.7% (22/132), respectively. The positive rate for patients undergoing a combination of WES and CNVseq (WES + CNVseq) was 47.8% (54/113), which was significantly better than the positive rate for patients who underwent WES alone (32.7%, 37/113). A total of 83 variants were found in 42 genes, and SCN1A was the most frequently mutanted gene. Twenty-four CNVs were identified in 22 patients: two CNVs were inherited from the mother; 12 CNVs were de novo; and the CNV origins could not be determined in 10 patients. WES + CNVseq may potentially be the mostly effective NGS approach for diagnosis of rare NDs in pediatric patients. K E Y W O R D Scopy number variation sequencing, diagnostic yield, neurological disorders, targeted nextgeneration sequencing panels, whole-exome sequencing

Silver–Russell syndrome (SRS) is a rare, but well-recognized disease characterized by growth disorder. To date, there are two reports arguing IGF2 mutation for the onset of SRS. Herein, we present another sporadic case harboring IGF2 mutation. The male proband was the first and only child of a non-consanguineous Chinese couple. He was small for gestational age, with relative macrocephaly at birth. Severe feeding difficulties, low feeding, and growth retardation were revealed during neonatal period. At 4.5 years old, obvious body asymmetry was noted. Whole exome sequencing identified a novel de novo c.101G > A (p.Gly34Asp, NM_000612) variant in IGF2 and Sanger sequencing validated the variant. Amplification refractory mutation system polymerase chain reaction demonstrated that the IGF2 variant was on the paternal allele. Alignment shows the variant is evolutionarily conserved. Structural modeling argues that the variant site might be important for the binding of IGF2 to its receptor. Our study provides further evidence that IGF2 mutation may be another mechanism of SRS, and we consider that IGF2 should be included in a disease specific gene panel in case it is designed for SRS routine diagnostics.

Objective This study is to investigate the diagnostic yield of the combination of trio whole exome sequencing (Trio-WES) and copy number variation sequencing (CNVseq) for rare neurodevelopmental disorders (NDDs). Methods Clinical data from consecutive pediatric patients who were diagnosed with rare NDDs that were suspected to be monogenic disorders, who were admitted to our hospital from April 2017 to March 2019, and who underwent next generation sequencing (NGS) were extracted from the medical records. Patients for whom Trio-WES and CNVseq data were available were enrolled in this study. Sanger sequencing was applied for the validation of the variants identified by Trio-WES. Sequence alignment and structural modeling were conducted for analyzing the possibility of the variants in the onset of the NDDs. Results In total, 54 patients were enrolled in this study, with the median age of 15 (8–26) months. A total of 242 phenotypic abnormalities belonging to 20 different systems were identified in the cohort. Twenty-four patients were diagnosed by Trio-WES, eight patients were diagnosed by CNVseq, and one case was identified by both WES and CNVseq. Compared with Trio-WES, the diagnosis rate of Trio-WES accompanied by CNVseq was significantly higher ( P = 0.016). Trio-WES identified 36 variants in 26 different genes, among which 27 variants were novel. CNVseq detected four duplications and eight deletions, ranging from 310 kb to 23.27 Mb. Our case examples demonstrated the high heterogeneity of NDDs and showed the challenges of rare NDDs for physicians. Conclusion The significantly higher diagnosis rate of Trio-WES accompanied by CNVseq makes this strategy a potential alternative to the most widely used approaches for pediatric children with rare and undiagnosed NDDs.

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Multiple mitochondrial dysfunctions syndrome (MMDS) is an autosomal recessive disorder of systemic energy metabolism. This study is to present the diagnosis of two MMDS Chinese sufferers. Physical and auxiliary examination was performed. Next generation sequencing (NGS) was conducted to identify candidate causal genes and Sanger sequencing was adopted to validate the variants detected. Fluorescence quantitative polymerase chain reaction (FQ-PCR) amplification was carried out to testify allelic loss existence. Structural investigation was performed to study the possibility of the candidate variants for disease onset. Physical examination showed that the children were with neurological impairment. Auxiliary examination demonstrated energy metabolism disturbance and abnormal brain signals. NGS found that the probands had homozygous mutation of c.545 + 5G > A and compound heterozygous variants of exon 4 deletion and c.721G > T in NFU1, respectively. NFU1 was considered as candidate molecular etiology and indicating that the kids were with MMDS. Sanger sequencing confirmed the variants. FQ-PCR amplification characterized that patient 1 had a de novo allele mutation while patient 2 inherited from his parents. Structural investigation demonstrated that the variants were possible for MMDS occurrence. This is the first report of patients diagnosed as MMDS with novel mutation types from the Asia-Pacific region.