Differentially expressed full-length, fusion and novel isoforms transcripts-based signature of well-differentiated keratinized oral squamous cell carcinoma

Abstract

Highly keratinized oral squamous cell carcinoma (OSCC) exhibits an improved response to treatment and prognosis compared with weakly keratinized OSCC. Therefore, we aimed to develop gene transcript signature and to identify novel full-length isoforms, fusion transcript and non-coding RNA to differentiate welldifferentiated (WD) with Moderately Differentiated (MD)/Poorly Differentiated (PD)/WD-lymphadenopathy OSCC through, HTA, Isoform sequencing, and NanoString. Additionally, specific copy number gain and loss were also identify in WD keratinized OSCC through Oncoscan array and validated through Realtime PCR in histopathologically characterized FFPE-WD keratinized OSCC. Threehundred-thirty-eight (338) differentially expressed full-length (FL) transcript isoforms (317 upregulated and 21 down-regulated in OSCC) were identified through Isoform Sequencing using the PacBio platform. Thirty-four (34) highly upregulated differentially expressed transcripts from IsoSeq data were also correlated with HTA2.0 and validated in 42 OSCC samples. We were able to identify 18 differentially expressed transcripts, 12 fusion transcripts, and two long noncoding RNAs. These transcripts were involved in increased cell proliferation, dysregulated metabolic reprogramming, oxidative stress, and immune system markers with enhanced immune Research Paper Oncotarget 3228 www.oncotarget.com rearrangements, suggesting a cancerous nature. However, an increase in proteasomal activity and hemidesmosome proteins suggested an improved prognosis and tumor cell stability in keratinized OSCC and helped to characterize WD with MD/PD/WD with lymphadenopathy OSCC. Additionally, novel isoforms of IL37, NAA10, UCHL3, SPAG7, and RAB24 were identified while in silico functionally validated SPAG7 represented the premalignant phenotype of keratinized (K4) OSCC. Most importantly we found copy number gain and overexpression of EGFR suggest that TKIs may also be used as therapeutics in WD-OSCCs.

Single‐cell RNA‐seq reveals actinic keratosis‐specific keratinocyte subgroups and their crosstalk with secretory‐papillary fibroblasts

“…The RBPJ, SPAG7, and ESRRA TFs were all highly expressed in AK compared with NS as a whole (Figure S4F), which was in accordance with the role they played in the development of SCC. [23][24][25] Moreover, they were more highly expressed in clusters 9 and 11 (Figure S4E). These results suggested that specific regulons were active in AK-specific keratinocytes whose activity differed between AK clusters, reinforcing the heterogeneity of AK, which might be important in AK biology.…”

Investigating Cutaneous Squamous Cell Carcinoma in vitro and in vivo: Novel 3D Tools and Animal Models

“…Overall, epidermis and mucosa undergo to tissue-specific mechanical stress and exposure to environmental stimuli, including contacts with secretes (such as mucus) and enzymes characteristic of the tissue, as well as pathogens. Furthermore, with regard to keratin expression in neoplastic versus healthy tissues, both proteomic and transcriptomic profiles reveal the presence of a tumor dependent signatures, with a defined keratin pattern associated with proliferation and differentiation status of the cells ( 34 – 36 ). For example, while Keratin 13 is absent in cancers from the gingivo-buccal complex ( 35 ), it becomes indicative of a neoplastic and malignant conversion in skin cancers ( 37 – 39 ).…”

<p>Comprehensive Characterization of Stage IIIA Non-Small Cell Lung Carcinoma</p>

“…We performed microarray assay on 10 NSCLC stage IIIA and 5 control samples to establish gene-expression profiles using GeneChip® Human Transcriptome Array 2.0 (HTA 2.0, Affymetrix, Santa Clara). 10 We measured the RNA concentrations with the Qubit 3.0 Fluorometer (Invitrogen, USA). We processed 500ng RNA samples with the WT PLUS Reagent kit, followed by hybridization on the HTA 2.0 microarray chips.…”

“…NanoString geneexpression analysis was performed as previously done. 10 In brief, hybridization of 100 ng of total RNA with customized reporter code set (for gene-transcript expression or gene fusion-transcript) and capture probe set was performed in a NanoString Prep Station (NanoString Technologies), and the mRNA molecules counted with the NanoString nCounter (NanoString Technologies). The nSolver™ Analysis Software 3.0 (NanoString Technologies) was used to perform data handling, including automated background subtraction, spike-in-control normalization, and reference gene normalization.…”