Establishment Of Risk Model For Recurrence And Metastasis Of Pheochromocytoma/paraganglioma And Preliminary Study Of Metastasis Mechanis

Background:Pheochromocytoma and paraganglioma(PPGL)are neuroendocrine tumors arising from chromaffin cells of the adrenal medulla and extra-adrenal autonomic paraganglia,respectively.Lifelong follow-up is suggested for patients who have undergone surgery.Recurrence after resection occurs in about 3～16%of patients,and metastases occurs in about 10～17%of patients.Till now,few studies focused on the histopathological/clinicopathological scoring systems for predicting the metastatic potential of the disease.All of they were small scale studies,and had some limitations.Objective:To study the characteristics,risk factors and establish prognostic nomograms to improve the prediction of local-regional recurrent and metastatic probability in PPGL patients.Methods:Clinical data were retrospectively reviewed in 127 PPGL patients with local-regional recurrence,228 patients with metastases and 148 patients without recurrence and metastasis.We examined the variants of germline pathogenic genes by Sanger sequencing andnext generation sequencing.Succinate dehydrogenase subunit B(SDHB),alpha-thalassemia/memal retardation syndrome(ATRX),S100 calcium binding protein(S-100),Ki-67 index immunohistochemical staining were assessed.Multivariate COX regression analysis of selected prognostic features was performed,and two nomograms to predict recurrence and metastasis were constructed separately.Discrimination and calibration were employed to evaluate the performance of the nomograms.Results:1.26.3%metastatic patients in our series had synchronous metastases.The median time for metastasis after resection of the primary tumor was about 5.0(3.0,9.0)years after the operation of the primary tumor.The median time for recurrence was 5.0(3.5,9.0)years after the operation of the primary tumor.2.A multivariate COX regression analysis of selected prognostic features was performed for metastasis.SDHB mutation[HR 4.5(2.6,7.9),p<0.001],negative S-100 immunohistochemistry[HR 2.2(1.2,3.9),p=0.009],negative ATRX immunohistochemistry[HR 2.2(1.3,3.8),p=0.003],high Ki-67 index[HR 4.01(1.2,3.9),p<0.001],lager primary tumour size[HR 2.71(1.6,4.6),p<0.001]were independent predictors for metastasis in PPGL patients.All the above factors were included in the nomogram.The nomogram showed good discrimination and calibration.3.A multivariate COX regression analysis of selected prognostic features was performed for recurrence.SDHB mutation[HR4.7(2.3,10.0),p<0.001],and negative ATRX immunohistochemistry[HR 4.9(2.8,8.7),p<0.001],high Ki-67 index[HR 3.1(1.7,5.8),p<0.001],and lager primary tumour size[HR 2.4(1.2,4.7),p=0.011]were independent predictors for recurrence in PPGL patients.All the above factors were included in the nomogram.The nomogram showed good discrimination and calibration.4.Seventy-eight patients carried pathogenic germline SDHB mutations,and 262 patients hadnoSDHB mutations.Compared with patients without SDHB mutation,patients with SDHB mutation had a younger age of onset and diagnosis(29.1±13.8 vs.37.5±13.4;32.2±15.1 vs.40.6±14.5,P<0.001)and a higher incidence of metastasis(93.6%vs.45.8%,P<0.001),a higher proportion of paraganglioma(78.2%vs.43.9%,P<0.001),alarger tumor diameter[6.0cm(5.0,8.0)vs.5.5cm(4.0,8.0),P<0.05],andahigherKi-67 index[5%(2%,10%)vs.1%(<1%,3%),P<0.001].Conclusion:The median time for recurrence and metastasis after resection of the primary tumor was about 5 years after the operation of the primary tumor.Lifelong follow-up is suggested for PPGL patients.SDHB mutation,negative ATRX immunohistochemistry,high Ki-67 index,and lager primary tumour size were independent predictors for recurrence and metastasis in PPGL patients.Negative S-100 immunohistochemistry was independent predictor for metastasis.All the above factors were included in the nomograms,which had good discrimination and calibration to predict the recurrence and metastatic probability.Background:Pheochromocytoma and paraganglioma(PPGL)are neuroendocrine tumorsarising from chromaffin cells of the adrenal medulla and extra-adrenal autonomic paraganglia,respectively.Lifelong follow-up is suggested for patients who have undergone surgery due to all tumors have the potential to recurrence/metastasis.Recurrence after resection occurs in about 3～16%of patients,and metastases occurs in about 10～17%of patients.Succinatedehydrogenase B(SDHB)mutation have the highest risk of recurrence/metastasis.but the mechanism of recurrence/metastasis in these patients is still unclear.Objective:To explore the mechanism of recurrence/metastasis in patients with SDHB gene mutation.Methods:Eight patients with SDHB mutation in our hospital,including 5 patients with recurrence/metastasis and 3 patients without recurrence/metastasis,were included in our study.Total RNA was extracted from frozen tissue.We used DEseq2 to detect differentially expressed mRNA of primary tumour between the two groups and validated somegenes by quantitative real-time PCR(qRT-PCR).Enrichment analysis of differentially expressed genes were performed.The data of 14 patients with SDHB mutation from public databases were compared.including 9 patients with recurrence/metastasis and 5 patients without recurrence/metastasis.Result:1.Inourstudy,1102 genes were differentially expressedbetween the group with recurrence/metastasis and the group without recurrence/metastasis,including 657 up-regulated genes and 445 down-regulatedgenes.Enrichment analysis showed that the differential genes were enriched in the pathways of mitotic spindle checkpoint,mitotic nuclear division,cell cycle,transcriptional misregulation in cancer,regulation of system process,and so on.2.Assembly factor for spindle microtubules(ASPM),spindle and kinetochore associated complex subunit 3(SKA3),structural maintenance of chromosomes condensin Ⅰ complex subunit(NCAPG)were highly expressed in recurrence/metastasis group(27.53±32.85 vs.1.14±0.37;18.47±31,84 vs.1.20±0.87;19.60±32.64 vs.1.32±0.83,P<0.05).Survival analysis suggested that these genes were associated with poor prognosis.3.Glutathione peroxidase 1(GPX1)was lowly expressed in the in recurrence/metastasis group(0.46±0.17 vs.1.06±0.29,P<0.05).Conclusion:ASPM,NCAPG,SKA3,GPX1 were differentially expressed between the group without recurrence/metastasis and the group with recurrence/metastasis,and they may promote the occurrence of recurrence/metastasisin patients with SDHB mutation.