Risk stratification for radioactive iodine refractoriness using molecular alterations in distant metastatic differentiated thyroid cancer

Abstract

Objective

Patients with radioactive iodine-refractory differentiated thyroid cancer (RAIR-DTC) are often diagnosed with delay and constrained to limited treatment options. The correlation between RAI refractoriness and the underlying genetic characteristics has not been extensively studied.

Methods

Adult patients with distant metastatic DTC were enrolled and assigned to undergo next-generation sequencing of a customized 26-gene panel (ThyroLead). Patients were classified into RAIR-DTC or non-RAIR groups to determine the differences in clinicopathological and molecular characteristics. Molecular risk stratification (MRS) was constructed based on the association between molecular alterations identified and RAI refractoriness, and the results were classified as high, intermediate or low MRS.

Results

A total of 220 patients with distant metastases were included, 63.2% of whom were identified as RAIR-DTC. Genetic alterations were identified in 90% of all the patients, with BRAF (59.7% vs. 17.3%), TERT promoter (43.9% vs. 7.4%), and TP53 mutations (11.5% vs. 3.7%) being more prevalent in the RAIR-DTC group than in the non-RAIR group, except for RET fusions (15.8% vs. 39.5%), which had the opposite pattern. BRAF and TERT promoter are independent predictors of RAIR-DTC, accounting for 67.6% of patients with RAIR-DTC. MRS was strongly associated with RAI refractoriness (P<0.001), with an odds ratio (OR) of high to low MRS of 7.52 [95% confidence interval (95% CI), 3.96−14.28; P<0.001] and an OR of intermediate to low MRS of 3.20 (95% CI, 1.01−10.14; P=0.041).

Conclusions

Molecular alterations were associated with RAI refractoriness, with BRAF and TERT promoter mutations being the predominant contributors, followed by TP53 and DICER1 mutations. MRS might serve as a valuable tool for both prognosticating clinical outcomes and directing precision-based therapeutic interventions.

Introduction

Although most patients with differentiated thyroid cancer (DTC) achieve an excellent response after radioactive iodine (RAI, ¹³¹I) therapy combined with surgery, a subset of patients is unable to benefit from ¹³¹I therapy, as indicated by either a loss of ability to uptake RAI initially or gradually, completely or partially, or disease progression despite significant RAI uptake, which is referred to as RAI-refractory differentiated thyroid cancer (RAIR-DTC). RAIR-DTC has been proven to be a significant challenge in the management of thyroid cancer. Sampson et al. (1) reported that the 3-year survival rate of patients with distant metastases with high RAI uptake was much better than that of patients with non-RAI-avid metastases (82% vs. 57%). Similar results were reported in subsequent studies (2-5). Once patients develop RAIR-DTC, there is no indication for further RAI treatment alone but active surveillance of disease progression. Recent advances in targeted therapies, including multikinase inhibitors (MKIs) and selective inhibitors, have demonstrated particularly encouraging results. Accordingly, with respect to precision treatment, there is a growing need to distinguish patients with RAI-refractory characteristic as soon as possible.

RAIR-DTC is traditionally defined based on clinical behaviors, including RAI avidity and response to ¹³¹I therapy. However, on the one hand, it is necessary to increase the thyrotropin concentration to enhance the ¹³¹I concentration in tumors before ¹³¹I whole-body scans or treatment, which could also accelerate tumor proliferation. On the other hand, classification by this system proves to be somewhat subjective and time-consuming, leading to inconsistent treatment decisions among different physicians and delaying diagnosis until after treatment has been administered. Furthermore, for patients who are naturally refractory to RAI, the administration of RAI undoubtedly increases their radiation exposure with little benefit. Thus, it is essential to identify an approach to improve the predictability of RAIR-DTC. Over the past several decades, additional studies focused on molecular mechanisms have enriched the understanding of tumor behaviors at a more intrinsic level (6,7). In the case of DTC, BRAF^V600E mutation was initially identified as a contributor to the downregulation of sodium-iodide symporter expression and the reduction of RAI uptake in thyroid and cancer cells, thereby exhibiting a non-RAI-avid pattern (8,9); these results have been broadly reported and verified in subsequent studies (10-12). TERT promoter mutations have been further suggested to play a supporting role in RAI-refractory status (13-15). However, there is still a subset of RAI-refractory disease that cannot be explained by BRAF and/or TERT promoter mutations. Notably, interactions among genes, such as cooccurrence or mutual exclusivity, have been recognized as important causes of tumor evolution and heterogeneity. However, the prevailing understanding of the molecular underpinnings of RAIR-DTC is still predominantly bereft of analyses incorporating a comprehensive genetic profile and clinical manifestations, thereby lacking a holistic view of the onset and progression of advanced DTC, particularly that of RAIR-DTC.

In this study, patients with distant metastatic DTC (DM-DTC) in China were enrolled to perform next-generation sequencing of a 26-gene panel associated with thyroid cancer. Leveraging this unique dataset, we aim to elucidate the molecular profile of DM-DTC and explore the correlation between genomic alterations and RAI refractoriness, thereby informing future therapeutic strategies.

Materials and methods

Patient cohorts

This was a retrospective clinical study aimed to ascertain the genetic alterations correlated with RAI refractoriness. Patients included in this study met the following key criteria: 1) aged 18 years or older; 2) had undergone thyroid surgery and received a pathological diagnosis of DTC; 3) had distant metastases identified through either pathologic examination of surgical and/or fine needle aspiration specimens or a comprehensive evaluation of biochemical examination and radiologic imaging when pathology samples of metastatic disease were unavailable; 4) attended at Peking Union Medical College Hospital (PUMCH) between 2020 and 2022; and 5) had adequate tissue for next-generation sequencing. The clinical, pathological, and radiographic data were retrieved from existing medical records. Patients were classified as RAIR-DTC or non-RAIR according to comprehensive assessment through ¹³¹I whole-body scans, supplementary imaging modalities, and longitudinal monitoring of serum thyroglobulin and antibody levels. RAIR-DTC was defined in alignment with the criteria set forth in the 2015 American Thyroid Association guidelines for adult patients, namely, which include any of the following: 1) metastatic lesions do not concentrate RAI initially; 2) metastatic lesions gradually lose the ability to concentrate RAI; 3) some lesions concentrate RAI while others do not; and 4) disease progresses despite evident RAI concentration. Patients without enough data to determine the classification of RAIR-DTC or non-RAIR tumors were excluded. This study was approved by the Institutional Review Board of PUMCH (No. JS-2432). All participants provided written informed consent.

DNA isolation

For formalin-fixed paraffin-embedded (FFPE) tissues, tumor-rich areas (>20% of neoplastic cells) were microdissected. This was performed employing one or two unstained histological sections, each 3−5 μm in thickness, with stereomicroscopic guidance provided by an Olympus CX31 microscope. Hematoxylin and eosin-stained slides were used for reference. Genomic DNA was manually extracted from each dissected region using the GeneReadTM DNA FFPE Kit (QIAGEN), in accordance with the manufacturer’s protocols.

Next-generation sequencing

A bespoke 26-gene panel ThyroLead panel (Topgen, Shanghai, China) was engineered to encompass the exonic region for 18 genes (HRAS, KRAS, NRAS, CDC73, CDKN1B, DICER1, IDH1, MEN1, MTOR, PIK3CA, PTEN, TP53, TSHR, CTNNB1, GNAS, PAX8, AKT1, and EIF1AX), a 1,000-bp region in the promoter region of TERT, and both exonic and intronic regions for 7 frequently rearranged genes (BRAF, RET, NTRK1/2/3, ALK and PPARG). A minimum of 30 ng of genomic DNA was extracted from the samples and subsequently fragmented using the Covaris E220 instrument (Covaris, Woburn, USA). Sequence libraries were prepared using the KAPA HyperPlus Library Preparation Kit, initially creating blunt-ended, 5’-phosphorylated fragments; subsequently, dAMP (A-tailing) was appended to the 3’ ends of the dsDNA library fragments. Subsequently, dsDNA featuring with 3’-dTMP was ligated to the A-tailed library fragments. Library fragments possessing appropriate adapter sequences underwent amplification through ligation-mediated precapture polymerase chain reaction (PCR). Library capture was conducted using an Igenetech custom probe system, and the library was biotinylated to allow sequence enrichment by capture using streptavidin-conjugated beads (Thermo Fisher, Waltham, USA). Ultimately, the pooled libraries, comprising captured DNA fragments, were sequenced on the Illumina NextSeq 500 system utilizing the NextSeq 500/550 High Output Kit v2.5 (300 cycles) to yield 150-bp paired-end reads.

Data analysis

The sequences obtained from Illumina were first filtered to remove adapter sequences and low-quality reads (i.e., reads with >40% of bases failed Q25; reads shorter than 70 bp; and reads with low complexity) with Fastp. The remaining sequences were aligned to the hg19 human genome reference using the maximal exact matches of the Burrows-Wheelers-Alignment tool (BWA-MEM), and duplicate sequences were marked using Picard MarkDuplicates. Indel realignment and variant calling were then conducted using the commercial software Sentieon TNseq. The identified variants were subsequently filtered to retain high-confidence variants only. Variants were ultimately annotated through a variety of databases, such as the catalogue of somatic mutations in cancer (COSMIC), 1000 Genomes Project (1000G), Exome Sequencing Project (ESP), Exome Aggregation Consortium (ExAC), Genome Aggregation Database (gnomAD) and Clinical Relevant Sequence Variants (ClinVar), with Variant Effect Predictor (VEP) and Annotate Variation (ANNOVAR). Each variant was graded based on its pathogenicity according to American College of Medical Genetics and Genomics and the Association for Molecular Pathology (ACMG-AMP) guidelines. Variants graded as benign or likely benign were excluded. To ensure the reliability of variant calling and pathogenicity grading, manual inspection was performed on individual variants. Quality control (QC) was applied at the sample level to examine the uniformity of coverage, average deduplicated depth across targeted regions and insert size distribution. Samples with uneven depth of coverage, low deduplicated depth or aberrant insert sizes were removed from further analysis. Only samples that passed QC were included in further analyses.

Molecular risk stratification (MRS)

In light of the observed correlation between molecular alterations and RAI refractoriness, we divided the detected genetic alterations into 3 MRSs: high, intermediate, and low. High MRS included genetic alterations, which were identified as positively independent predictors of RAIR-DTC. The genetic alterations included in the intermediate MRS were positively correlated with RAIR-DTC but cannot serve as independent indicators. Finally, the low MRS included genetic alterations that did not fit within either the high or intermediate MRS categories.

Statistical analysis

Analyses were performed using R software (Version 4.1.1; R Foundation for Statistical Computing, Vienna, Austria) and SPSS Statistics (Version 25.0; IBM Corp., NewYork, USA). The Kruskal-Wallis test was used to compare the differences in age and mutational load between the RAIR-DTC and non-RAIR groups. The Chi-square test or Fisher’s exact test was performed to test the associations between categorical covariates and RAI refractoriness, and between gene interactions. A two-sided P value less than 0.05 was considered to indicate statistical significance. The Bonferroni method was applied for correcting multiple comparisons of gene interactions. Univariate and multivariate analyses were performed using logistic regression to identify independent predictors of RAI refractoriness. Several clinical variables and genetic alterations were included in the univariate analysis, and variables with P values less than 0.05 and variables with potential clinical relationships were included in the subsequent multivariate analysis.

Results

Clinicopathological characteristics

Of all 220 patients, 139 (63.2%) presented with RAIR-DTC, while 81 (36.8%) were non-RAIR. The clinicopathological characteristics are summarized in Table 1. The median age at diagnosis was greater in the RAIR-DTC group than in the non-RAIR group (34.15 vs. 46.72 years, P<0.001). No significant differences were observed in terms of gender, pathological type, or T (tumor) stage and N (lymph node) stage according to the American Joint Committee on Cancer. Compared to those in the non-RAIR group, those in the RAIR-DTC group had a greater propensity for developing distant metastases beyond the lung and bone (1.2% vs. 17.3%, P=0.004). Additionally, the RAIR-DTC group exhibited a greater mutational load than the non-RAIR group.

Table 1. Clinicopathologic features of 220 patients with distant metastatic differentiated thyroid cancer.

Clinicopathologic features	n (%)		P
	Non-RAIR (N=81)	RAIR-DTC (N=139)
IQR, interquartile range; PTC, papillary thyroid cancer; FTC, follicular thyroid cancer; PDTC, poorly differentiated thyroid cancer; T, tumor; N, lymph node; RAIR, radioactive iodine-refractory; RAIR-DTC, radioactive iodine-refractory differentiated thyroid cancer; *, Part of data are not available; a, Brain, liver, pancreas, pleura, muscle, or distant metastatic lymph nodes, with or without lung and bone metastases.
Age at diagnosis (year) [median (IQR)]	34.15 (28.80, 41.72)	46.72 (35.27, 56.13)	<0.001
Gender: female	51 (63.0)	78 (56.1)	0.320
Pathology			0.441
PTC	74 (91.4)	118 (84.9)
FTC	5 (6.2)	12 (8.6)
PTC and FTC	1 (1.2)	2 (1.4)
PDTC	1 (1.2)	7 (5.0)
T stage*			0.064
1−3a	47 (63.5)	58 (46.8)
3b−4b	27 (36.5)	66 (53.2)
N stage*			0.145
0	11 (14.1)	11 (8.9)
1a	5 (6.4)	10 (8.1)
1b	62 (79.5)	102 (82.9)
Distant metastatic organs			0.004
Lung	69 (85.2)	96 (69.1)
Bone	3 (3.7)	5 (3.6)
Lung and bone	8 (9.9)	14 (10.1)
Othersa	1 (1.2)	24 (17.3)
Mutational load [median (IQR)]	1 (1, 2)	2 (1, 2)	<0.001

Molecular profile of DM-DTC and its relationship with RAI refractoriness

Genetic alterations were identified in 197 of 220 (90%) patients using the ThyroLead panel, while no alterations were detected in 23 (10%) patients. Sixty-one (28%) patients exhibited gene fusions; specifically, 54 had RET fusions, 6 had NTRK1 fusions and one had a BRAF fusion. One hundred and sixty patients (73%) were found to harbor point mutations, 97 of whom had BRAF mutations, 67 of whom had TERT promoter mutations, and 26 of whom had RAS mutations (including HRAS, NRAS and KRAS mutations). An overview of these gene alterations is illustrated in Figure 1.

Figure 1.

Genetic landscape and clinicopathological characteristics of 220 adult patients with distant metastatic differentiated thyroid cancer. RAIR, radioactive iodine-refractory; T, tumor; N, lymph node; SNV, single nucleotide variation.

When comparing the genetic alterations between the non-RAIR and RAIR-DTC groups, mutations in BRAF (17.3% vs. 59.7%, P<0.001), TERT promoter (7.4% vs. 43.9%, P<0.001), and TP53 (3.7% vs. 11.5%, P=0.047) were more frequently detected in the RAIR-DTC group, while RET fusions were more prevalent in the non-RAIR group (39.5% vs. 15.8%, P<0.001). Concerning RAS and other genes, no difference was identified between the two groups. After removing variants of uncertain significance, the mutational alterations were largely similar between the two groups, except DICER1 mutations, which were identified only in the RAIR-DTC group (0 vs. 6.5%, P=0.019).

In the RAIR-DTC subgroup, the most common alterations were BRAF mutations (59.7%), followed by TERT promoter mutations (43.9%), RET fusions (15.8%), TP53 mutations (11.5%), RAS mutations (10.1%), DICER1 mutations (7.9%), and IDH1 mutations (5.0%). Overall, 33.1% of cases harbored isolated genetic alterations, while 60.4% had multiple alterations. Importantly, patients with BRAF or TERT promoter mutations, with or without other alterations, accounted for 67.6% of patients. Additionally, TP53 or DICER1 mutations could explain the molecular setting of 7.2% of RAIR-DTC cases. In addition, 17.3% of patients harbored RAS mutations or RET fusions, with or without other mutations, such as PIK3CA, PTEN, and EIF1AX. One patient had an isolated MTOR mutation, and the other had both an NTRK1 fusion and a TSHR mutation. Patients with currently known genetic backgrounds accounted for 93.5% of the population. Notably, actionable alterations or targetable alterations, which refer to gene alterations with corresponding targeted drugs (including BRAF^V600E, RET, NTRK fusions, ALK and MTOR) were identified in 77.7% of patients with RAIR-DTC.

Gene interaction patterns

Gene interaction analysis revealed 4 mutually exclusive gene pairs and 2 cooccurring gene pairs (all P<0.05; Figure 2). The mutually exclusive gene pairs were BRAF and RAS mutations, BRAF mutations and RET fusions, RAS mutations and RET fusions, and RET fusions and TERT promoter mutations. BRAF and TERT promoter mutations were found to be the most common cooccurring gene pairs in the whole DM-DTC population (50/220, 22.7%), as well the most common in the RAIR-DTC group (50/139, 36.0%). Notably, this gene pair was exclusively present in the RAIR-DTC group (P<0.001). RAS and EIF1AX mutations also significantly cooccurred, albeit less frequently, at a rate of 1.4% (3/220); one instance was in the RAIR-DTC subgroup and two were in the non-RAIR subgroup. Notably, even though the difference was not statistically significant, the cooccurring gene pair of TP53 and DICER1 was only identified in the RAIR-DTC group, for which the frequency was 2.9% (4/139).

Figure 2.

Gene interaction relationships in 220 patients with distant metastatic differentiated thyroid cancer. FDR, false detection rate.

Risk factors for RAI refractoriness

In the univariate logistic analysis, we enrolled genes with mutation frequencies greater than 10% in this cohort, an additional two genes of interest (TP53 and DICER1) and clinical characteristics listed in Table 1. The outcomes of this analysis are depicted in Figure 3A. RAIR-DTC was positively associated with age, T stage, extrapulmonary metastasis, BRAF mutations, TERT promoter mutations and mutational load but negatively related to RET fusions. After adjustment with the multivariate logistic regression analysis of these 7 parameters, it was found that only BRAF and TERT promoter mutations remained as independent predictors of RAI refractoriness (P<0.001 and P=0.030, respectively), while RET fusions and mutational load lost significance (P=0.697 and P=0.689). Clinical variables such as the presence of extrapulmonary metastases, patient age and high T stage did not sustain their statistical significance in predicting RAIR-DTC (Figure 3B). The risk of RAIR-DTC was 5.25 times greater in patients with BRAF mutations than in those without these mutations and 3.54 times greater in patients with TERT promoter mutations than in those without these alterations.

Figure 3.

Forest map of univariate analysis (A) and multivariate analysis (B). PDTC, poorly differentiated thyroid cancer; OR, odds ratio; 95% CI, 95% confidence interval.

MRS for RAI refractoriness

Based on the above findings, mutations in the BRAF and/or TERT promoters were categorized as high MRS, while the TP53 and DICER1 mutations were allocated to intermediate MRS; all the other genetic alterations had a low MRS. A statistical association was observed between this MRS and RAI refractoriness (Figure 4A, P<0.001). RAIR-DTC cases accounted for 82.5%, 66.7% and 38.5% of patients with high, intermediate and low MRS, respectively (Figure 4B). The odds ratio (OR) of high relative to low MRS was 7.52 [95% confidence interval (95% CI), 3.96−14.28; P<0.001], and the OR of intermediate relative to low MRS was 3.20 (95% CI, 1.01−10.14; P=0.041).

Figure 4.

Percentage of each molecular risk stratification (high, intermediate, low) in patients with RAIR-DTC and non-RAIR, respectively (A), and percentages of RAIR and non-RAIR in each molecular risk stratification (B). RAIR-DTC, radioactive iodine-refractory differentiated thyroid cancer.

Discussion

To our knowledge, this study represents the most extensive cohort study of adult patients with DM-DTC, with a specific focus on elucidating the genetic correlates of RAI refractoriness. After systematically investigating the genetic alterations related to RAI avidity, the BRAF and TERT promoter were unsurprisingly found to be the most powerful genetic duet for predicting RAI refractoriness, similar to reports in patients with recurrent and persistent diseases (14,16-18). BRAF mutations were demonstrated to be the most common genetic alterations in patients with RAIR-DTC (59.7%), followed by TERT promoter mutations, for which the frequency was 43.9%. Individuals harboring either BRAF or TERT promoter mutations constituted 67.6% of the patients with RAIR-DTC in this cohort. Of note, either of them could also appear in patients with RAI-avid disease, while the coexistence of BRAF and TERT promoter mutations was detected only in those with RAIR-DTC, suggesting a synergistic effect in mediating the process of RAI refractoriness. Previous studies have shown that BRAF^V600E indirectly activates the mutant TERT promoter via the mitogen-activated protein kinase (MAPK)/familial olfactory seduction (FOS)/GA binding protein (GABP) signaling pathway to selectively recruit activated extracellular signal-regulated kinase (ERK), which can phosphorylate specificity protein 1 (Sp1), thereby resulting in histone deacetylase 1 (HDAC1) dissociation and sustaining an active chromatin state of the TERT promoter (19,20). In turn, TERT promoter mutations enhance MAPK signaling output by providing de novo consensus motifs for the E-twenty-six (ETS) family of transcription factors triggered by MAPK signaling. Moreover, TERT can promote the dedifferentiation of BRAF-mutant thyroid cancer cells by regulating ribosomal RNA expression and protein synthesis (21,22). Furthermore, Landa et al. found that telomerase-upregulated tumors displayed anaplastic-like features in BRAF^V600E-mutant mice, which also involved nontelomeric effects (23). Gene interaction analysis further revealed the statistical significance of the coexistence of the BRAF and TERT promoter mutations. This dual gene pair was also the most common cooccurring gene mutation pair in patients with RAIR-DTC, with a frequency of 36.0% in the present cohort. Moreover, multivariate analysis further proved the independent value of both BRAF and TERT promoter for predicting RAIR-DTC, revealing their predominant involvement in impairing the efficacy of RAI therapy. This evidence allows practitioners to safely decrease empirical RAI therapy.

However, there remains confusion regarding RAIR-DTC with neither BRAF mutations nor TERT promoter mutations, which means that BRAF and TERT promoter mutations cannot explain the whole situation of the undesirable response to RAI treatment. Within this subset of patients in our cohort, 22.2% were identified as carrying mutations in either TP53 or DICER1. Furthermore, the concomitant presence of TP53 and DICER1 mutations was exclusively observed in the RAIR-DTC group, indicating a potential synergistic role in the pathogenesis of RAIR-DTC. TP53 mutations have been recognized as late molecular events involved in tumor dedifferentiation since they are prevalent in anaplastic thyroid carcinoma and some PDTCs but absent in well-differentiated cancer (24-27). Jung et al. (15) demonstrated that at least one mutation in either the TP53, PLEKHS1 promoter or TERT promoter is a significant predictor of RAI-refractory disease. DICER1 mutations are related to miRNA dysfunction, which induces the loss of function of tumor suppressor genes, thereby leading to the occurrence of various malignant tumors (28). Previous literature has reported the prevalence of DICER1 mutations at 10% in pediatric multinodular goiter and 20% in pediatric DTC, respectively (29). Related reports in adults are scarce, and Chong et al. (30) reported that DICER1 mutations were detected in only 1.4% of patients with thyroid nodules. In the present study involving an adult metastatic DTC cohort, DICER1 mutations seemed more frequent (6.4%), which suggested the involvement of DICER1 in aggressive DTC. A high cooccurrence frequency of TP53 and DICER1 was reported in pleuropulmonary blastoma (31). Additionally, possible crosstalk between miRNA biogenesis and TP53 was suggested in anaplastic thyroid cancer (32). Our findings suggest that the coexistence of TP53 and DICER1 mutations may significantly increase the likelihood of RAI refractoriness.

At present, genetic testing has become a ubiquitous practice in medical oncology. For patients with DM-DTC, there are two primary objectives for conducting gene sequencing. One objective is to prognosticate patient outcomes with high precision, necessitating that the genetic testing panel incorporates molecular markers with proven prognostic value. The second objective is to pinpoint actionable genetic alterations that can inform subsequent therapeutic strategies. Given the substantial patient population with DTC and the heterogeneity in economic conditions, MRS serves as a useful tool for optimizing patient outcomes (Figure 5). In our study, to better identify the probability of RAI refractoriness, we further allocated the detected molecular alterations into a three-level MRS system according to the significance associated with RAIR-DTC. Compared to those with low MRS, those with high MRS (containing BRAF and/or TERT promoter) and intermediate MRS (containing TP53 and/or DICER1) have a greater probability of developing RAI-refractory disease with ORs of 7.52 and 3.20, respectively. In addition, actionable alterations were detected in 77.7% of patients with RAIR-DTC in this cohort. Elucidating the molecular profile of patients with DM-DTC holds the promise of tailoring individualized diagnostic modality and targeted therapies, especially for those who have symptomatic or progressive disease and have been refractory to prior treatment regimens (33,34).

Figure 5.

Implication of genetic testing for differentiated thyroid cancer. RAIR-DTC, radioactive iodine-refractory differentiated thyroid cancer.

The current study is not without limitations. Primarily, the genetic sequencing data were predominantly sourced from primary tumors, as the collection of metastatic lesions was constrained by both logistical and ethical concerns. Second, the scope of the current investigation is confined to DNA-level alterations; as such, it cannot discriminate between expressed and unexpressed gene modifications. Additionally, the study lacked coverage of copy-number variations, which could potentially influence the findings. In addition, although the ThyroLead panel could reveal genetic contributors to 93.5% of RAIR-DTC cases, a more detailed approach, such as genome-wide genetic and epigenetic profiling, may be needed to determine the genetic background for the remaining cases.

Conclusions

This study delineates the molecular profile of an extensive cohort of patients with DM-DTC. BRAF and TERT promoter mutations are predominantly responsible for RAI refractoriness. TP53 and DICER1 mutations were associated with RAI refractoriness. Our findings suggest that MRS might serve as a valuable tool for both prognosticating clinical outcomes and directing precision-based therapeutic interventions.