Abstract
We report changes in the genomic landscape in the development of head and neck squamous cell carcinomas HNSCC from potentially premalignant lesions (PPOLS) to malignancy and lymph node metastases. Likely pathological mutations predominantly involved a relatively small set of genes reported previously (TP53, KMT2D, CDKN2A, PIK3CA, NOTCH1 and FAT1) but also other predicted cancer drivers (MGA, PABPC3, NR4A2, NCOR1 and MACF1). Notably, all these mutations arise early and are present in PPOLs. The most frequent genetic changes, which follow acquisition of immortality and loss of senescence, are of consistent somatic copy number alterations (SCNAs) involving chromosomal regions enriched for genes in known and previously unreported cancer-related pathways. We mapped the evolution of SCNAs in HNSCC progression. One of the earliest SCNAs involved deletions of CSMD1 (8p23.2). CSMD1 deletions or promoter hypermethylation were present in all of the immortal PPOLs and occurred at high frequency in the immortal HNSCC cell lines. Modulation of CSMD1 in cell lines revealed significant suppression of proliferation and invasion by forced expression, and significant stimulation of invasion by knockdown of expression. Known cancer drivers NOTCH1, PPP6C, RAC1, EIF4G1, PIK3CA showed significant increase in frequency of SCNA in transition from PPOLs to HNSCC that correlated with their expression. In the later stages of progression, HNSCC with and without nodal metastases showed some clear differences including high copy number gains of CCND1, hsa-miR-548k and TP63 in the metastases group.
Highlights
Head and neck carcinomas account for over 550,000 new cases per annum with a mortality of approximately 275,000 cases per year[1]
Exceptions to this are the study by Bhattacharya and colleagues[9] and Wood and colleagues[19], which reported a comprehensive analysis of copy number variation in primary PPOLs and HNSCC in the later study, the PPOL analyses was confined to metachronous lesions
We extend their findings by a combination of exome/targeted sequencing, methylation and SNP/CGH array analyses, using our unique panel of fully characterised mortal cultures and immortal cell lines derived from both PPOLS and HNSCC, to show that these genetic alterations are mostly associated with cellular immortalisation and increase with the stage of tumour progression in this class of squamous cell carcinoma (SCC) keratinocyte
Summary
Head and neck carcinomas account for over 550,000 new cases per annum with a mortality of approximately 275,000 cases per year[1]. They lack inactivation of TP53 and CDKNA, but our limited previous investigations show that they are genetically stable They often have extended replicative lifespans[16,17], possess neoplastic phenotypes, such as resistance to suspension-induced terminal differentiation[16] and have expression signatures which are distinct from both immortal cells and normal cells[12]. A very recent study reported the status of SCNAs and common driver mutations in several keratinocyte cultures derived from metachronous normal and dysplastic lesions characterised for replicative lifespan[20] We extend their findings by a combination of exome/targeted sequencing, methylation and SNP/CGH array analyses, using our unique panel of fully characterised mortal cultures and immortal cell lines derived from both PPOLS and HNSCC, to show that these genetic alterations are mostly associated with cellular immortalisation and increase with the stage of tumour progression in this class of SCC keratinocyte. Alterations of CSMD1 were common in PPOL cultures and may be early events in SCC development
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