Intratumor heterogeneity and clonal evolution in an aggressive papillary thyroid cancer and matched metastases

Abstract

The contribution of intratumor heterogeneity to thyroid metastatic cancers is still unknown. The clonal relationships between the primary thyroid tumors and lymph nodes (LN) or distant metastases are also poorly understood. The objective of this study was to determine the phylogenetic relationships between matched primary thyroid tumors and metastases. We searched for non-synonymous single-nucleotide variants (nsSNVs), gene fusions, alternative transcripts, and loss of heterozygosity (LOH) by paired-end massively parallel sequencing of cDNA (RNA-Seq) in a patient diagnosed with an aggressive papillary thyroid cancer (PTC). Seven tumor samples from a stage IVc PTC patient were analyzed by RNA-Seq: two areas from the primary tumor, four areas from two LN metastases, and one area from a pleural metastasis (PLM). A large panel of other thyroid tumors was used for Sanger sequencing screening. We identified seven new nsSNVs. Some of these were early events clonally present in both the primary PTC and the three matched metastases. Other nsSNVs were private to the primary tumor, the LN metastases and/or the PLM. Three new gene fusions were identified. A novel cancer-specific KAZN alternative transcript was detected in this aggressive PTC and in dozens of additional thyroid tumors. The PLM harbored an exclusive whole-chromosome 19 LOH. We have presented the first, to our knowledge, deep sequencing study comparing the mutational spectra in a PTC and both LN and distant metastases. This study has yielded novel findings concerning intra-tumor heterogeneity, clonal evolution and metastases dissemination in thyroid cancer.

Heterogeneity of Thyroid Cancer

“…However, only 33–40% of multifocal PTC cases showed a discordant BRAF status between foci [57]. To determine the phylogenetic relationships between matched primary thyroid tumours and metastases, Le Pennec et al [58] performed paired-end massively parallel sequencing of 2 areas of the primary tumour, including 1 pleural metastasis (PLM) and 2 LNMs from a patient with aggressive PTC. Although all of the examined tissue samples were composed of several cellular subclones, the analysed areas of the primary tumour and one of the LNMs displayed similar genetic profiles, whereas the other LNM presented more divergent mutations and fusions.…”

“…These data indicate that genetic alterations accumulate after clonal PLM separation from the primary clone. Although there is currently no clear answer as to whether cancer cells simultaneously spread from the primary thyroid tumour to the regional LNs and distant locations or first spread to the lymph node and then to further locations, Le Pennec et al [58] suggested that the PLM originated from a subclone of the LNM.…”

MicroRNA‑mRNA integrated analysis based on a case of well‑differentiated thyroid cancer with both metastasis and metastatic recurrence

“…Despite increasing WDTC globally, few studies have investigated metastatic sites of WDTC from a genomic perspective. RNA-Seq analysis of metastatic sites, including two LNM and one pleural metastasis from a single patient, revealed intratumor heterogeneity and clonal evolution (32). Another miRNA study analyzed miRNA transcriptomes of PTC with four LNM and noted that downregulation of miRNAs was a common feature in PTC tumorigenesis (33).…”

Intratumoral Genetic Heterogeneity in Papillary Thyroid Cancer: Occurrence and Clinical Significance

“…By the analysis of single nucleotide variants, gene fusions, and loss of heterozygosity, the authors showed that some of the genetic alterations were ubiquitously detected in all the tumor samples from the patient and others were detected only in some tumor areas, indicating the presence of several subclones in the neoplastic tissues. Interestingly, the two selected areas of the primary tumor and the two selected areas of on regional metastasis presented similar genetic profiles, whereas the two selected areas of another regional metastasis had more divergent mutations and fusions [55]. Striking heterogeneity was also observed between paired primary tumors and metastases in studies done by means of NGS and WES studies [58,60,63,64].…”

“…Accordingly, in PTC the allelic frequency must be normalized for the percentage of tumor cells in the sample, a measurement not always easy to perform [22,29]. Indeed, the number of the studies included in the present review which report data on the genetic characterization and/or ITH in TC, not evaluating tumor purity [27, are significantly more numerous than those which normalized the data for the percentage of tumor cells [7,20,22,29,[53][54][55]. Thus, data on ITH obtained without considering the purity of the tumor must be considered with caution.…”