Understanding Immune Dysregulation in Tylosis with Oesophageal Cancer

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 typically
presents with hyperkeratosis by the ages of 7 or 8, and type B which presents with
hyperkeratosis during the first year of life2. Importantly, type B is considered benign3, whereas
type A is associated with a high lifetime risk of developing oesophageal squamous cell
carcinoma (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 following
mapping to a 42.5 kb region on chromosome 17q255, Blaydon et al. identified missense
mutations within the RHBDF2 gene to be associated with TOC in 3 separate families4.
Subsequently, other TOC families have been identified with novel RHBDF2 mutations that
cluster only a few amino acids apart and are described in Tab.1 alongside pedigrees with yet
undefined mutational status.

Hypothesis
The gain-of-function TOC RHBDF2 mutation leads to immune dysregulation and subsequent
epithelial dysplasia and keratinocyte hyperproliferation.
Primary Aim
To characterise the immune phenotype of TOC in the periphery and within the tissue, with a
particular focus on the oesophagus and skin as clinically relevant sites of disease.
Secondary Aim
To elucidate the mechanisms that underpin immune dysregulation in TOC and unravel
whether this is a direct or indirect consequence of the TOC RHBDF2 mutation.

Results and Future Work
The 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 for
this phenotype is yet to be explored.
In the immediate term, this work will be extended in two RHBDF2 mouse models; knock-out
and 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 of
iRhom2 on immune development and the peripheral immune compartment, and whether the
CD8+ TEMRA expansion observed in human samples is replicated in mice.
Secondly, computational analyses have identified elevated numbers of mast cells and M2
macrophages in TOC dysplastic oesophageal biopsies, which is indicative of a type 2 immune
response21. It's prudent to note that there were some patient by patient discrepancies between
the CIBERSORT and xCell outputs; therefore, these findings require validation.
Predominantly, immunofluorescence staining for mast cells, M2 macrophages and other type
2-related immune cells such as Th2 cells is planned in a bank of TOC skin and oesophageal
tissue sections. As the clinically relevant sites of TOC, it will be interesting to observe whether
there are immunophenotypic differences between the oesophagus and skin. Additionally, both
flow cytometry and immunofluorescence staining will be performed on both skin and
oesophageal sections from both the RHBDF2 knock-out and TOC knock-in mouse models.