Clinical Analysis Of A Novel Splicing Mutation Of SDHB Gene Inducing Paraganglioma Near Inferior Vena Cava

Background and objectivePheochromocytoma/paraganglioma（PPGL）was a rare kind of neuroendocrine tumor that derived from chromaffin tissue and was characterized with paroxysmal or persistent hypertension,headache,palpitations,sweatiness and so on.With the development of molecular detection technology,pathogenic gene screening of PPGL had been carried out generally.At least 17 kinds of pathogenic genes of PPGL had been discovered,including VHL,RET,SDHA,SDHB,SDHC,SDHD,SDHAF2,FH,HIF2A/EPAS1,etc,which was related to 50%PPGL.Malignant PPGL was defined as PPGL metastasis loci in non-chromaffin tissues or organs,the molecular oncology behavior of PPGL was different because of genetic background,and 40%malignant PPGL was associated with SDHB gene mutation.Succinate dehydrogenase was one of the crucial enzymes involved in the tricarboxylic acid cycle and participated in electron transport in the mitochondrial respiratory chain.Succinate dehydrogenase was consisted of four subunits.SDHA and SDHB were activity center of SDH,and SDHC and SDHD anchored the complex on the inner mitochondrial membrane.Astuti et al.found the pathogenic mutation of SDHB gene in an extra-adrenal PPGL family in 2001.SDHB was an iron-sulfur protein containing 280 amino acids and encoded by SDHB gene（（NM-003000））,which located in 1p35-p36.1 and spanned approximately 40 kb.SDHB gene was a kind of tumor suppressor gene and the LOH often occurred in the tumor tissue,which could result in completely deficiency of SDHB.At the same time,dysfunction of mitochondrial respiratory chain led to accumulation of succinate,inhibition of prolyl hydroxylase activity,activation of pseudohypoxia pathway and overexpression of hypoxia-inducible transcription factor,all of that involved in tumorigenesis,invasion and metastasis.In this study,we explored the pathogenic mechanism of a novel splicing mutation of SDHB gene found in a PPGL patient by RT-PCR,evaluated the risk of metastasis of the patient and made a reasonable follow-up plan.Methods1.A 16-year-old adopted and thin girl manifested paroxysmal headache,palpitation and sweatiness for 4 years.Biochemical parameters revealed a significantly increased norepinephrine in urine collected for continuous 24 hours;abdominal CT showed a retroperitoneal mass and ¹³¹I-MIBG imaging was positive.She was diagnosed with PPGL,which was confirmed by pathology result.2.With the approval of the patient and her adoptive parents,we collected the blood sample and tumor tissue sample of the patient and carried out high-throughput sequencing in the target enrichment of 1500 gene exons and adjacent splicing regions of tumor DNA,which was related to endocrine and metabolic diseases.The varitation site was verified by Sanger sequencing in tumor tissue and blood,compared with normal people.3.The RT-PCR technique was applied to explore the change of mRNA,and the effect of the mutation on the structure and function of SDHB was analyzed.4.A reasonable follow-up plan was made based on the risk of metastasis evaluation result.Results1.Genetic sequencing showed a germline heterozygous mutation of SDHB gene:c.423+1G>T in the patient,which had not been reported yet.2.The cDNA sequencing showed that 54 coding bases in the downstream of the fourth exon was cutted,which led to 18-amino-acid deficiency at position 124-141of SDHB and was predicted to be a pathogenic.3.The patient was with more than 50%risk of metastasis according to the evalution system of metastasis potential of pheochromocytoma based on molecular marker.Conclusions1.A novel splicing heterozygous germline mutation of SDHB gene was found in the study:c.423+1G>T,which led to deletion of 54 coding bases in downstream of the 4^th exon in transcription and absence of 18 amino acids from 124-141 in the new peptide chain.It was predicted to be a pathogenic mutation.2.The adolescent PPGL patient with relatively high risk of metastasis because of SDHB gene mutation,large tumor size,locatin in extra-adrenal gland and norepinephrine secretion.