Understanding Immune Dysregulation in Tylosis with Oesophageal Cancer

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Tylosis is a rare focal non-epidermolytic palmoplantar keratoderma, first described in 18801.Subsequently, tylosis has been stratified into two separate groups; type A which typicallypresents with hyperkeratosis by the ages of 7 or 8, and type B which presents withhyperkeratosis during the first year of life2. Importantly, type B is considered benign3, whereastype A is associated with a high lifetime risk of developing oesophageal squamous cellcarcinoma (OSCC) and is separately classified as tylosis with oesophageal cancer (TOC[OMIM 148500])4.TOC is inherited as an autosomal dominant trait with complete penetrance and followingmapping to a 42.5 kb region on chromosome 17q255, Blaydon et al. identified missensemutations within the RHBDF2 gene to be associated with TOC in 3 separate families4.Subsequently, other TOC families have been identified with novel RHBDF2 mutations thatcluster only a few amino acids apart and are described in Tab.1 alongside pedigrees with yetundefined mutational status.HypothesisThe gain-of-function TOC RHBDF2 mutation leads to immune dysregulation and subsequentepithelial dysplasia and keratinocyte hyperproliferation.Primary AimTo characterise the immune phenotype of TOC in the periphery and within the tissue, with aparticular focus on the oesophagus and skin as clinically relevant sites of disease.Secondary AimTo elucidate the mechanisms that underpin immune dysregulation in TOC and unravelwhether this is a direct or indirect consequence of the TOC RHBDF2 mutation.Results and Future WorkThe peripheral blood of TOC patients has been well characterised and an expansion of CD8+TEMRA cells has been observed. However, the functional relevance and mechanistic basis forthis phenotype is yet to be explored.In the immediate term, this work will be extended in two RHBDF2 mouse models; knock-outand TOC gain-of-function knock in. Using flow cytometry, the lymphoid organs (thymus,spleen, lymph nodes) of these mice will be fully immunophenotyped to observe the effects ofiRhom2 on immune development and the peripheral immune compartment, and whether theCD8+ TEMRA expansion observed in human samples is replicated in mice.Secondly, computational analyses have identified elevated numbers of mast cells and M2macrophages in TOC dysplastic oesophageal biopsies, which is indicative of a type 2 immuneresponse21. It's prudent to note that there were some patient by patient discrepancies betweenthe CIBERSORT and xCell outputs; therefore, these findings require validation.Predominantly, immunofluorescence staining for mast cells, M2 macrophages and other type2-related immune cells such as Th2 cells is planned in a bank of TOC skin and oesophagealtissue sections. As the clinically relevant sites of TOC, it will be interesting to observe whetherthere are immunophenotypic differences between the oesophagus and skin. Additionally, bothflow cytometry and immunofluorescence staining will be performed on both skin andoesophageal sections from both the RHBDF2 knock-out and TOC knock-in mouse models.