Genetic profile of advanced thyroid carcinomas using targeted next generation sequencing and its relation to clinical prognosis

서론: 갑상선암 중 일부 진행성 갑상선 암은 예후가 매우 불량하고 치료가 어려운데, 원격 전이가 동반되거나 침습적 변이형 분화 갑상선암 (DTC), 저분화 갑상선 암(PDTC) 또는 역형성 갑상선 암 (ATC)과 같은 드물지만 공격적인 아형이 이에 해당한다. 이러한 진행성 갑상선암의 체계적인 유전자 프로파일과 임상 결과와의 연관성에 관한 자료는 매우 제한적이다. 재료 및 방법: 진행성 갑상선 (원격 전이 또는 공격적 변이가 있는 DTC, PDTC, ATC) 환자 100명의 원발 갑상선 종양 및 원격 전이 조직에서 50개의 유전자를 사용한 표적 차세대 염기서열 분석이 시행되었다. 50개 유전자 중 functional group (FG)을 구성하는 유전자는 histone methyltransferases (HMTs), SWI/SNF subunit, 그리고 the PI3K/AKT/mTOR 신호전달체계 세가지로 분류하여 분석하였다. 하위 그룹 분석 (subgroup analyses)을 시행하여 BRAFV600E 돌연변이가 있는 PTC 환자 50명에서 FG 돌연변이가 사망률에 미치는 영향을 평가하였다. 결과: 100명의 환자로부터 얻은 118개의 조직 중 99개 조직은 원발성 종양, 19개는 원격 전이 조직이었다. 원발성 종양에서 관찰된 가장 흔한 돌연변이는 BRAFV600E 돌연변이 (58%)였고, 그 다음이 TERT 프로모터 (41%) 및 RAS (16%) 돌연변이였다. TP3는 6%에서 변이가 발견되었다. FG를 구성하는 HMTs, SWI/SNF subunit 및 PI3K/AKT/mTOR 신호전달체계와 관련 유전자의 돌연변이는 각각 30.3%, 10.1% 및 13.1%에서 확인되었다. 갑상선 원발 종양과 이에 상응하는 원격 전이 조직 15쌍과 서로 다른 부위의 원격전이 조직 1쌍에서 돌연변이 상태를 분석한 결과 일치도가 높았다. TERT 프로모터 (56% vs. 58%) 및 BRAFV600E (41% vs. 42%)뿐만 아니라 HMTs (42% vs. 37%), SWI/SNF subunit (12% vs. 10%) 및 PI3K/AKT/mTOR 신호전달체계 (22% vs. 26%)에서 유사한 돌연변이 빈도가 관찰되었다. 동일한 주요 초기 및 후기 돌연변이가 원발 종양-원격 전이 조직 쌍에 항상 존재함을 확인할 수 있었다. BRAFV600E 돌연변이가 있는 갑상선 유두암 (PTC) 환자의 하위 그룹 분석에서 15명의 환자는 BRAFV600E 돌연변이만 있었고 (그룹 1), 22명은 BRAFV600E 와 FG 이외의 돌연변이가 있었고 (그룹 2), 13명은 BRAFV600E 와 FG 돌연변이(그룹 3)가 있었다. 전체 생존 (overall survival, OS)은 FG 돌연변이가 있는 환자에서 그렇지 않은 환자보다 유의하게 낮았고(p=0.001), 그룹 3의 환자는 생존율이 가장 낮았다(p=0.004). OS는 FG 돌연변이 부위 개수 (없거나 1개 또는 2개) 에 따라 유의하게 달랐는데, FG 돌연변이가 2개 있는 군이 나머지 군에 비하여 OS가 낮았다 (p=0.005). FG 돌연변이의 존재는 사망률 증가와 독립적으로 연관됨이 확인되었다 (위험비 11.65, 95% 신뢰 구간 1.39– 97.58, p=0.024). 결론: 원격 전이가 있는 진행된 갑상선암에서 BRAFV600E, TERT 프로모터 및 RAS 돌연변이가 빈번하게 발견되었다. 원발 갑상선 종양과 상응하는 원격 전이 조직의 유전적 프로파일은 높은 일치도를 보였다. 또한, BRAFV600E의 돌연변이와 FG를 코딩하는 유전자의 공존은 높은 사망률과 관련이 있습니다. BRAFV600E 돌연변이를 동반한 PTC에서 FG 돌연변이 여부의 식별은 이 군의 위험도를 분류하는 데 유용한 지표가 될 것으로 생각한다.Docto

Mutation in Genes Encoding Key Functional Groups Additively Increase Mortality in Patients with BRAF(V600E)-Mutant Advanced Papillary Thyroid Carcinoma

Simple SummaryThe BRAF(V600E) point mutation is the most common driver mutation in papillary thyroid carcinoma (PTC) and is known to be associated with aggressive clinical features. However, the negative prognostic impact of BRAF(V600E) on PTC mostly depends on tumor characteristics, not on itself. Moreover, the prognosis of BRAF(V600E)-mutant PTCs varies widely implying the genetic diversity of this subtype. Additional genetic alterations other than BRAF(V600E) may be responsible for the aggressiveness of this group but to date, no mutations other than TERT promoter mutation have been identified. This study aimed to investigate the effect of additional genetic alterations, focusing on the mutations in genes encoding functional groups on survival in BRAF(V600E)-mutant PTCs. We observed that coexistence of mutations in BRAF(V600E) and the three functional groups had the worst survival in patients with PTCs compared with mutations in BRAF(V600E) and genes other than those associated with functional groups or mutations in only BRAF(V600E).The prognosis of BRAF(V600E)-mutant papillary thyroid carcinoma (PTC) ranges from indolent to highly aggressive courses. To better define the genetic diversity of this subtype, we evaluated the survival according to the presence of an additional mutation in genes encoding functional groups (FGs) in BRAF(V600E)-mutant advanced PTC patients. Targeted next-generation sequencing was performed in primary tumors of 50 BRAF(V600E)-mutant PTCs with distant metastasis or aggressive variants. The mutation in genes encoding FGs included alterations in histone methyltransferases, SWI/SNF subunit, and the PI3K/AKT/mTOR pathway. The primary outcome was overall survival (OS). Fifteen patients only had the BRAF(V600E)-mutation (group 1), 22 had BRAF(V600E) and mutation other than FGs (group 2), and 13 had BRAF(V600E) and FG mutation (group 3). OS was significantly lower in patients with FG mutations (p = 0.001) than those without, and group 3 patients had the worst survival (p = 0.004). OS significantly varied among none, one, or two FG mutation sites (p = 0.005). Presence of FG mutation was independently associated with increased mortality (hazard ratio 11.65, 95% confidence interval 1.39-97.58, p = 0.024). Coexistence of mutations in BRAF(V600E) and genes encoding FGs was associated with high mortality. Identification of FG mutation in BRAF(V600E)-mutant PTCs may be valuable in risk stratifying this subtype

Immune Profiling of Advanced Thyroid Cancers Using Fluorescent Multiplex Immunohistochemistry

Background: Advanced thyroid cancers, including differentiated thyroid carcinoma (DTC) with distant metastasis, and anaplastic thyroid carcinoma (ATC), are associated with poor clinical outcomes and limited treatment options. This study aimed to determine the immune profiles of advanced thyroid cancers using fluorescent multiplex immunohistochemistry (F-MIHC) and multispectral imaging (MSI). Methods: Twenty-eight tissue samples were collected from 12 patients who had DTC with distant metastasis and from 16 with ATC. The samples were assessed using F-MIHC and MSI with antibodies against the cell surface molecules, cluster of differentiation (CD)4, CD8, programmed cell death-1 (PD-1), PD ligand 1 (PD-L1), forkhead box protein 3, and cytokeratin (CK). The expression of PD-L1 was evaluated using tumor proportion score (TPS) and combined positive score (CPS). Results: Significantly, more PD-L1-positive tumor cells (CK+PD-L1+) per mm2 were found in ATC samples than in DTC samples (183.5 vs. 0.03, p < 0.001). Lymphocyte infiltration was significantly increased in ATC compared with DTC, with significantly more PD-L1- or PD-1-positive lymphocytes in ATC samples than in DTC samples. The TPS and CPS for PD-L1 expression were negative in all DTC samples but positive in 81% and 94% of ATC samples, respectively. Conclusions: Immune profiling revealed significant differences between advanced DTC and ATC, particularly in terms of PD-L1 expression and lymphocyte infiltration. Therefore, immune profiling using F-MIHC and MSI can provide invaluable information regarding tumor microenvironments, which could help select candidates for immunotherapy

Genetic profiles of aggressive variants of Papillary thyroid carcinomas

Aggressive variants of papillary thyroid carcinoma (PTC) have been described with in-creasing frequency and are associated with unfavorable clinical outcomes. However, limited data exist on the comprehensive genetic profile of these variants. We performed targeted next-generation sequencing in 36 patients with aggressive variants of PTC and compared it to PTC from The Cancer Genome Atlas (TCGA) project and poorly differentiated thyroid cancers (PDTCs)/anaplastic thyroid cancers (ATCs) from the Memorial Sloan Kettering Cancer Center (MSKCC). BRAF mutation was the most prevalent (89%) in aggressive variants of PTC compared to that in other thyroid can-cers. RAS mutation was identified in one patient (3%), which was less frequent than in others. TERT promoter mutation (17%) ranged between that of PTCs (9%) and PDTCs (40%). Tumor suppressor genes, ZFHX3, TP53, and CHEK2, were mutated in 14%, 3%, and 6% of aggressive variants of PTC, respectively. The mutation rate of TP53 (3%) was significantly higher than that of PTCs (0.7%) and lower than that of ATCs (73%). Mutations in three functional groups, histone methyl transferases, SWI/SNF chromatin remodeling complex, and the PI3K/AKT/mTOR pathway, were present in 11%, 14%, and 11% of samples, respectively. In conclusion, aggressive variants of PTC had higher BRAF and lower NRAS mutation prevalence than other thyroid cancers. The prevalence of mutations in the TERT promoter, TP53, and genes encoding three functional groups ranged between that of PTCs and PDTCs/ATCs