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01.12.2017 | Case report | Ausgabe 1/2017 Open Access

BMC Cancer 1/2017

Stepwise addition of genetic changes correlated with histological change from “well-differentiated” to “sarcomatoid” phenotypes: a case report

Zeitschrift:
BMC Cancer > Ausgabe 1/2017
Autoren:
Taichiro Goto, Yosuke Hirotsu, Hitoshi Mochizuki, Takahiro Nakagomi, Toshio Oyama, Kenji Amemiya, Masao Omata
Wichtige Hinweise

Electronic supplementary material

The online version of this article (doi:10.​1186/​s12885-017-3059-1) contains supplementary material, which is available to authorized users.
Abbreviations
EGFR
Epidermal growth factor receptor
KRAS
Kirsten rat sarcoma viral oncogene homolog
NRAS
Neuroblastoma ras oncogene
PIK3CA
Phosphatidylinositol 3-kinase catalytic, alpha polypeptide
STK11
Serine/threonine kinase 11
TP53
Tumour protein p53

Background

Sarcomatoid carcers are rare malignant biphasic tumors composed of carcinomatous and sarcomatous components. However, it is still unclear whether the carcinomatous or sarcomatous component develops first, or whether they develop simultaneously from adjacent areas. We recently encountered a patient who underwent surgery for sarcomatoid cancer that was composed of three different histological areas. To examine whether the different histological components resulted from genetic divergence, targeted sequencing was performed in each histological component. Herein, we report the case findings and the genomic analysis of sarcomatoid differentiation.

Case presentation

The patient was a 54-year-old man. He complained of hemosputum, and chest X-ray showed a mass shadow in the right upper lung field (Fig.  1a). He had a history of smoking 20 cigarettes per day for 30 years. Chest computed tomography revealed an irregularly-shaped mass in segment 1 of the right lung (Fig.  1b). Bronchoscopic biopsy led to a diagnosis of adenocarcinoma.
Under a diagnosis of lung cancer (cT2aN0M0), he underwent right upper lobectomy and lymph node dissection. Bilateral legs had been swelling preoperatively, which subsided spontaneously after the surgery. Pleural lavage cytology led to a diagnosis of disseminated adenocarcinoma (pT2aN0M1a, stage IV); histopathological analysis showed that tumor was composed of three different histological areas; portion A: well-differentiated adenocarcinoma, portion B: poorly differentiated adenocarcinoma, portion C: sarcomatoid cancer, and portion B was a major component (Fig.  1c-g). As the sarcomatoid cancer component accounted for less than 10% of the entire tumor area, the cancer did not meet the diagnostic criteria for pleomorphic carcinoma. To examine whether different histological components resulted from genetic divergence, tumor cells of each different histological area were collected from formalin-fixed paraffin-embedded tissues by using laser capture microdissection (Fig.  1d-g). A panel targeting the exon of 53 lung cancer-associated genes was established in-house, as we previously reported [ 14]. Subsequently targeted sequencing of those 53 lung cancer-related genes was performed. The sequencing data were processed using standard Ion Torrent Suite software running on a Torrent Server (Thermo Fisher Scientific). Variants calling were performed using an Ion Reporter Server System (Thermo Fisher Scientific), and peripheral blood DNA was used as a control to detect somatic mutations in tumours using filtration of “confident somatic variants” in a Tumour-Normal pair pipeline [ 4, 5]. This study was approved by the institutional review board, and the patient provided written informed consent. In total, there were 26 mutations found in the cancer (Fig.  2a and Additional file 1: Table S1), and mutations with an allele fraction ≥30% were determined as significant mutations (Table  1). Common missense mutations in TP53 and KRAS genes were detected in the portion A, B and C (Table  1, Fig.  2a). These mutations were validated by Sanger sequencing (Fig.  2b). The positions and patterns of the amino acid changes and base-pair substitutions in those co-mutations were consistent (Table  1).
Table 1
Mutation analysis in different histological areas
Histological area
Gene
Mutation
Position
Ref
Variant in tumor
AF
Portion A: well diff. Adeno
TP53
p.V118F
chr17:7578461
C
A
47%
 
KRAS
p.G12V
chr12:25398284
C
A
30%
Portion B: poorly diff. Adeno
KRAS
p.G12V
chr12:25398284
C
A
97%
 
TP53
p.V118F
chr17:7578461
C
A
86%
Portion C: sarcomatoid
TP53
p.V118F
chr17:7578461
C
A
70%
 
KRAS
p.G12V
chr12:25398284
C
A
52%
The table listed somatic mutations with an allele fraction ≥30%
chr chromosome, Ref reference sequence, AF allele fraction
For further inference of phylogenies and estimation of evolutionary distances, the Neighbor-Joining method was implemented to cluster the nonsilent mutations and the phylogenetic tree was constructed (Fig.  3a and b) [ 6]. The ‘ape’ and ‘phangon’R (3.2.3 on linux) packages were utilized for these analyses. As a result, sarcomatoid component clusters together with poorly differentiated adenocarcinoma and that cluster segregates away from well-differentiated adenocarcinoma (Fig.  3a). From the phylogenetic tree, it can be inferred, under the assumption of normal cells without any mutations being roots, that well-differentiated adenocarcinoma first branched off from the roots, and that another branch evolved through poorly differentiated adenocarcinoma into sarcomatoid carcinoma (Fig.  3b). According to the branch length, which indicates the evolutionary distance, the distance between the poorly differentiated adenocarcinoma and the sarcomatoid carcinoma is relatively short.
Postoperative adjuvant chemotherapy was performed, but the patient developed lung and brain metastases and died of cancer 5 months postoperatively.

Discussion

Sarcomatoid carcinomas are carcinomas with sarcoma or sarcoma-like features. They are rare entities and are characterized by poor prognosis and resistance to conventional chemotherapy or radiotherapy. On evaluating the mutational profile of sarcomatoid cancer, Fallet et al. reported that the most frequently detected mutations were KRAS, EGFR, TP53, STK11, NOTCH1, NRAS, and PIK3CA [ 7].
In our case, comparative genetic analysis of different histological areas revealed intratumoral homogeneity for some mutations. These concordant mutations across different sites reinforced the hypothesis that a single cell clone evolved and progressed later by addition of other mutations into different histological phenotypes, involving epithelial-to-mesenchymal transition. In portions B and C of the tumor, TP53 and KRAS mutations were found at high allelic fractions over 70%, which strongly indicates the incidence of allelic losses or amplifications of these genes.
Some hypotheses have been proposed regarding the development of sarcomatoid carcinoma, such as: (i) simultaneous malignant transformation of epithelial elements and stroma, (ii) malignant transformation of cancer-derived stroma, (iii) sarcomatous change of carcinoma, (iv) carcinomatous change of sarcoma. With regard to the development of sarcomatoid carcinoma in the present case, the mutation analysis results indicated the occurrence of a sarcomatous change.
From the phylogenetic analyses, it can be concluded that stepwise addition of genetic changes caused the sarcomatous change in this cancer. Sarcomatous cancer is reported to be resistant to conventional chemotherapy or radiation therapy and to show poor prognosis. Reflecting on these observations, the mutations responsible for those phenotypic changes may be relate to worse clinical outcomes. Further understanding of the molecular pathology of sarcomatoid cancer may help in selection of sarcomatoid cancer patients for newly developed molecular-targeted therapies.

Conclusions

The different histological components in lung cancer result from genetic divergence. Sarcomatous change of carcinoma occurs in the case of sarcomatoid or pleomorphic cancer. Phenotypic changes to sarcomatoid cancer are associated with the addition of mutation patterns.

Acknowledgements

We thank Hidetoshi Shigetomo, Yumi Kubota, Ritsuko Yokouchi for their help.

Funding

This study was supported by a Grant-in-Aid for Genome Research Project from Yamanashi Prefecture (Y.H. and M.O.) and the grant from The YASUDA Medical Foundation (Y.H.).

Availability of data and materials

The datasets generated during the current study are not publicly available because of patient privacy, but are available from the corresponding author on reasonable request.

Authors’ contributions

TG wrote the manuscript. TG and TN performed surgery. TO carried out the pathological examination. TG, YH, HM and KA participated in the design and carried out genomic analyses, and help with data analysis. MO supervised the experimental design and laboratory processes. MO and YH were involved in the final editing. All authors have read and approved the final manuscript.

Competing interests

The authors declare that they have no competing interests.

Consent for publication

Written informed consent was obtained from the patient’s relatives for publication of this case report and the accompanying images.

Ethics approval and consent to participate

The study was reviewed and approved by the Yamanashi Prefectural Central Hospital.
Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License ( http://​creativecommons.​org/​licenses/​by/​4.​0/​), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver ( http://​creativecommons.​org/​publicdomain/​zero/​1.​0/​) applies to the data made available in this article, unless otherwise stated.
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