Carbon dating cancer: defining the chronology of metastatic progression in colorectal cancer

Abstract

Background: Patients often ask oncologists how long a cancer has been present before causing symptoms or spreading to other organs. The evolutionary trajectory of cancers can be defined using phylogenetic approaches but lack of chronological references makes dating the exact onset of tumours very challenging.

Patients and methods: Here, we describe the case of a colorectal cancer (CRC) patient presenting with synchronous lung metastasis and metachronous thyroid, chest wall and urinary tract metastases over the course of 5 years. The chest wall metastasis was caused by needle tract seeding, implying a known time of onset. Using whole genome sequencing data from primary and metastatic sites we inferred the complete chronology of the cancer by exploiting the time of needle tract seeding as an in vivo 'stopwatch'. This approach allowed us to follow the progression of the disease back in time, dating each ancestral node of the phylogenetic tree in the past history of the tumour. We used a Bayesian phylogenomic approach, which accounts for possible dynamic changes in mutational rate, to reconstruct the phylogenetic tree and effectively 'carbon date' the malignant progression.

Results: The primary colon cancer emerged between 5 and 8 years before the clinical diagnosis. The primary tumour metastasized to the lung and the thyroid within a year from its onset. The thyroid lesion presented as a tumour-to-tumour deposit within a benign Hurthle adenoma. Despite rapid metastatic progression from the primary tumour, the patient showed an indolent disease course. Primary cancer and metastases were microsatellite stable and displayed low chromosomal instability. Neo-antigen analysis suggested minimal immunogenicity.

Conclusion: Our data provide the first in vivo experimental evidence documenting the timing of metastatic progression in CRC and suggest that genomic instability might be more important than the metastatic potential of the primary cancer in dictating CRC fate.

Keywords: cancer evolution; metastatic colorectal carcinoma; mutational analysis; phylogenetic tree; synchronous metastases; whole genome sequencing.

Figure 1.

Overview of clinical, radiological and pathological progression of a CRC patient. The patient underwent anterior resection for a _pT_4pN₂ sigmoid cancer in November 2008. A lung nodule of unknown significance at the time of diagnosis was kept under surveillance and was subsequently proved to be metastatic in nature (biopsy in November 2011) and was resected in January 2012. An FDG-avid thyroid nodule was resected in August 2012 and showed a Hurthle adenoma containing foci of metastatic CRC. Two years after the lung biopsy, a chest wall mass on the site of previous lung biopsy was histologically confirmed and treated with chemo-radiotherapy. Six months after treatment the chest wall metastasis showed disease progression and a new site of metastasis along the left ureter was histologically confirmed during the insertion of a urinary stent. The timeline shows the dates that metastatic sites became apparent on imaging. Haematoxylin and Eosin (H&E) staining as well as FDG-PET-CT images of primary and metastatic sites are shown.

Figure 2.

Histopathology and molecular profiling of the primary CRC and metastatic sites. (A) Microscopy of the primary and metastatic sites including H&E staining, proliferation (nuclear Ki-67) and apoptosis (Caspase-3). (B) Microscopy of the CRC lung metastasis that led to the needle tract seeding to the chest wall. Cells stained negative for TTF1 and positive for CDK20, CDX2 and CEA. (C) Gross pathology and microscopy of the Hurthle adenoma containing a CRC metastasis. Macroscopic examination showed the Hurthle adenoma containing a haemorrhagic or cystic area. Different magnifications show the Hurthle adenomatous cells (background large eosinophilic cells) with an invasive front of CRC metastasis. The dotted line indicates the border between the normal thyroid and the Hurthle adenoma. The black rectangle indicates the area of invasive CRC within the adenoma. (D) Microscopy of the urothelial metastasis. H&E and CDK20 staining confirmed the colonic origin of the metastasis. (E) Drivers of CRC detected by WGS and validated by targeted exome sequencing. (F) Copy number changes in primary and metastatic sites.

Figure 3.

Chronology of the patient’s CRC evolution. (A) Whole-genome sequencing of multiple lesions from the patient’s malignancy allowed phylogenetic reconstruction of the tumour tree. In the phylogenetic tree, dates within brackets indicate the time of clinical diagnosis whereas dates in italic highlight the estimated times of the different lesions. (B) Illustrative cartoon of the patient disease progression and samples taken. At time $t_c$ the first colorectal cancer cell arose, giving rise to the primary tumour. At time $t_l$ the first metastatic clone emerged, giving rise to the lung metastasis, quickly followed by a second metastasis to the thyroid emerging at time $t_t$. At time $t_c^R$ the sample from the primary tumour was collected (resection) and analysed. During the lung biopsy, the needle tract seeding event spread cancer cells in the chest wall at time $t_{cw}$. A few weeks later, at time $t_l^R$ the lung metastasis was resected and profiled. Finally at time $t_{cw}^R$ the chest wall metastasis was also sampled.