Durable recurrence-free survival after pneumonectomy for late lung metastasis from rectal cancer: case report with genetic and epigenetic analyses

In 1990, a 40-year-old male underwent anterior resection of rectal adenocarcinoma (Dukes B2; preoperative carcinoembryonic antigen (CEA) serum level: 35 ng/mL), that was diagnosed synchronously with a 9-cm metastasis in liver segments V-VIII. After postoperative 5-fluorouracil based chemotherapy, the CEA serum level was slightly decreased (22 ng/mL); liver metastasis was markedly downsized and was radically resected. Follow-up revealed no evidence of residual disease until 2002; follow-up was then intentionally interrupted because the patient was considered to be free of disease. On March 2007, the patient developed dyspnea and cough, and a chest computed tomography (CT) scan revealed a 5-cm right hilar mass bulging into the main bronchus and infiltrating the upper lobe (Figs. 1 and 2a). Bronchoscopy demonstrated a neoplastic growth that was obstructing the right main bronchus 2 cm distal to the carina; biopsy revealed moderately differentiated adenocarcinoma, compatible with rectal cancer recurrence. Colonoscopy and CT of the abdomen and pelvis showed no other evidence of neoplastic disease, and the CEA serum level was normal (1 ng/mL). On April 2007, the patient was aged 57 years and was fit for major thoracic surgery. He was treated by means of right pneumonectomy and regional lymphadenectomy to obtain complete resection (Fig. 2b). Histology of the excised lung metastasis (Fig. 2c) revealed an adenocarcinomatous proliferation with morphological features overlapping those of primary rectal cancer and of previously resected hepatic metastasis. The primary cancer and all of the metastatic sites were characterized by extensive complex tubular gland formation with sparse centroglandular necrotic foci. Nuclei were moderately or highly atypical, elongated and stratified. The mitotic index ranged from 17 (in rectal cancer) to 24 × 10 high power fields (in metastatic sites). A mild peritumoral lymphocytic infiltrate (cluster of differentiation 3 positive) was present. Budding was absent and there was no lymphovascular invasion. The immunophenotype included caudal-type homeobox protein 2 and cytokeratin (CK) 20 (CK 20) positivity (Fig. 2c, inset), whereas thyroid transcription factor-1 and CK 7 were negative. The main bronchus resection margin was tumor-free. Of the 18 lymph nodes excised, 4 (all peribronchial) were metastatic. The early postoperative course was complicated by empyema that was successfully treated with antibiotics and drainage. Follow-up with CT scans of the chest and abdomen at 6 months after pneumonectomy revealed enlarged left supraclavicular lymph nodes (Fig. 2d). Biopsy of the latter revealed a metastatic adenocarcinomatous growth with a high mitotic index (14 × 10 high-power fields) and a morphological and immunohistochemical profile identical to that previously described (Fig. 2e). There were no other signs of relapse. Systemic chemotherapy (leucovorin, 5-fluorouracil and oxaliplatin regimen) was then initiated and continued until July 2008, resulting in complete clinical remission. Subsequent follow-up with semi-annual assessments of CEA level and total body CT and/or a positron emission tomography scans revealed no evidence of recurrence. At the last examination, in February 2015, the patient was relapse-free and enjoyed a good quality of life, at 7 years after pneumonectomy for lung metastasis and at 24 years after rectal cancer resection. Genetic and epigenetic profiles of the primary tumor and the three matched metastases (pulmonary, hepatic and lymph nodal) were assessed to verify the clonal origin of the four tumor samples and to characterize key biological traits during the tumor evolution. Tables 1 and 2 summarize the molecular results. Methylation specific-multiplex ligation dependent probe amplification (MS-MLPA) was performed by using the ME001 MS-MLPA tumor suppressor-1 kit and the ME002 MS-MLPA tumor suppressor-2 Kit (MRC-Holland, Amsterdam, The Netherlands) to simultaneously analyze the methylation status of 33 tumor suppressor genes and copy number alterations (CNA) of 53 genes [11]. The primary rectal carcinoma showed a high frequency of CNAs at multiple chromosome regions suggesting an unstable karyotype in this tumor. Clonal CNAs including chromosome gains at 7q, 9pq, 11pq, 13q, 19p and 20q were recurrent in the primary tumor and in the matched metastases, demonstrating a common origin of the four tumor samples (Table 1). Similarly, clonal gene specific hypermethylation was observed at adenomatous polyposis coli (APC) and cadherin 13 (CDH13) loci, while additional hypermethylated genes, namely WT1, DAPK1, CHFR, PAX5 and GATA5, were found in the lung and in supraclavicular lymph node metastases (Table 2). Microsatellite instability (MSI) analysis, carried out using a pentaplex panel of monomorphic mononucleotide repeats (BAT25, BAT26, NR-21, NR-22 and NR-24) as previously reported [12], demonstrated the absence of MSI in all tumor samples. The Maldi-TOF mass spectrometry platform and the myriapod colon status kit (Diatech Pharmacogenetics, Jesi, Italy) were used to profile the patient’s four tumor samples, analyzing the 216 common hot-spot cancer mutations in the four major oncogenes involved in CRC pathogenesis (KRAS, BRAF, PIK3CA and NRAS). This analysis demonstrated no gene mutation in the primary rectal adenocarcinoma, and interestingly this genetic profile remained relatively well preserved in the three metastatic sites.

Written informed consent was obtained from the patient for publication of this case report and any accompanying images. A copy of the written consent is available for review by the Editor of this journal.