Abstract
Background Cutaneous squamous cell carcinomas (cSCC) are the primary cause of premature deaths in patients suffering from the rare skin-fragility disorder recessive dystrophic epidermolysis bullosa, which is in marked contrast to the rarely metastasizing nature of these carcinomas in the general population. This remarkable difference is attributed to the frequent development of chronic wounds caused by an impaired skin integrity. However, the specific molecular and cellular changes to malignancy, and whether there are common players in different types of aggressive cSCCs, remain relatively undefined.
Methods MiRNA expression profiling was performed across various cell types isolated from skin and cSCCs. Microarray results were confirmed by qPCR and by an optimized in situ hybridization protocol. Functional impact of overexpression of a dysregulated miRNA was assessed in migration and 3D spheroid assays. Sample-matched transcriptome data was generated to support the identification of disease relevant miRNA targets.
Results Several miRNAs were identified as dysregulated in cSCCs as compared to controls. These included the metastasis-linked miR-10b, which was significantly upregulated in primary cell cultures and in archival biopsies. At the functional level, overexpression of miR-10b conferred the stem cell-characteristic of 3D-spheroid formation capacity to keratinocytes, and impaired their mobility. Analysis of miR-10b downstream effects identified a novel putative target of miR-10b, the actin- and tubulin cytoskeleton-associated protein DIAPH2.
Conclusion The discovery that miR-10b confers an aspect of cancer stemness – that of enhanced tumor cell adhesion, known to facilitate metastatic colonization - provides an important avenue for future development of novel therapies targeting this metastasis-linked miRNA.
Footnotes
↵* MW and RZ equally contributed to the work.
mo.wimmer{at}salk.at
rolan.zauner{at}salk.at
m.ablinger{at}salk.at
j.d.pinon{at}salk.at
c.gruber{at}salk.at
m.reisenberger{at}salk.at
t.lettner{at}salk.at
norbert.niklas{at}o.roteskreuz.at
johannes.proell{at}jku.at
mila.sajinovic{at}unsw.edu.au
p.desouza{at}westernsydney.edu.au
s.hainzl{at}salk.at
t.kocher{at}salk.at
e.murauer{at}salk.at
joh.bauer{at}salk.at
dirk.strunk{at}pmu.ac.at
julia.reichelt{at}newcastle.ac.uk
a.mellick{at}unsw.edu.au
v.wally{at}salk.at
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE130767
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE130925
List of abbreviations
- cSCC
- cutaneous squamous cell carcinomas
- CSCs
- cancer stem cells
- CTCs
- circulating tumor cells
- DIAPH2
- Diaphanous Related Formin 2
- ECM
- extracellular matrix
- EMT
- epithelial to mesenchymal transition
- FDR
- false discovery rate
- FFPE
- formalin-fixed paraffin embedded
- GAPDH
- Glycerinaldehyd-3-phosphat-dehydrogenase
- H&E
- Hematoxilin & eosin
- HC
- healthy control
- HNSCC
- head and neck squamous cell carcinoma
- HOXD
- homeobox D
- IHC
- immunohistochemistry
- KC
- keratinocytes
- LNA
- locked-nucleid acid
- MET
- mesenchymal to epithelial transition
- miRNAs
- micro-RNAs
- onco-miRs
- oncogenic miRNAs
- PCA
- principal component analysis
- RDEB
- recessive-dystrophic epidermolysis bullosa
- LM
- lymphnode metastasis
- RNA-seq
- RNA sequencing
- SCC
- squamous cell carcinomas
- SCR
- scrambled
- SEM
- standard error of mean
- SSC
- saline-sodium citrate
- TCGA
- The Cancer Genome Atlas
- TUBA1
- tubulin alpha 1