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Molecular Characterization of Oral Epithelial Dysplasia and Oral Squamous Cell Carcinoma Using EGFR, CDKN2A, and HRAS Alterations
by
, , , , and
Satoshi Okubo
1,†, 
Satoru Miyabe
1,*,†,
Masahiro Fukumura
1,
Jun Sasaki
1,
Hitoshi Fujii
1,
Fumitaka Terasawa
1,
Satoshi Watanabe
1,
Soma Okada
1,
Megumi Miyabe
2,
Katsuyuki Miyabe
3,
Yoshihiko Sugita
4,
Hatsuhiko Maeda
4,
Sanako Nakaya
1,
Kaori Sakane
1,
Seiji Yamada
5,
Nitin Bhola
6,
Saman Warnakulasuriya
7,
Toru Nagao
1 and
Mitsuo Goto
1 1
Department of Oral and Maxillofacial Surgery, School of Dentistry, Aichi Gakuin University, Nagoya 464-8651, Japan
2
Department of Internal Medicine, School of Dentistry, Aichi Gakuin University, Nagoya 464-8650, Japan
3
Department of Gastroenterology, Japanese Red Cross Aichi Medical Center Nagoya Daini Hospital, Nagoya 464-8650, Japan
4
Department of Oral Pathology/Forensic Odontology, School of Dentistry, Aichi Gakuin University, Nagoya 464-8650, Japan
5
Division of Analytical Pathology, Oncology Innovation Center, Research Promotion Headquarters, Fujita Health University School of Medicine, Toyoake 470-1192, Japan
6
Department of Oral & Maxillofacial Surgery, Sharad Pawar Dental College, Datta Meghe Institute of Medical Sciences, Wardha 442001, India
7
Faculty of Dentistry, Oral and Craniofacial Sciences, King’s College London, WHO Collaborating Centre for Oral Cancer, London SE1 9RT, UK
*
Author to whom correspondence should be addressed.
†
These authors contributed equally to this work.
Cancers 2025, 17(24), 3949; https://doi.org/10.3390/cancers17243949 (registering DOI)
Submission received: 5 November 2025
/
Revised: 4 December 2025
/
Accepted: 9 December 2025
/
Published: 10 December 2025
(This article belongs to the Special Issue Molecular Mechanisms in Head and Neck Cancer)
Simple Summary
We tested molecular markers to improve early diagnosis and risk assessment of patients diagnosed with oral cancer. Using routine biopsy tissue, the application of FISH technique for EGFR amplification and CDKN2A deletion distinguished oral squamous cell carcinoma from non-cancerous dysplasia with high specificity, improved diagnostic performance, and identified patients with poorer survival. In cancers without EGFR amplification, high HRAS expression indicated aggressive disease and unfavorable prognosis. This two-step biomarker panel (EGFR/CDKN2A by FISH, then HRAS IHC if EGFR-negative) may help clinicians decide which oral lesions require aggressive treatment and closer follow-up.
Abstract
Background/Objectives: Oral squamous cell carcinoma (OSCC) often presents at an advanced stage; therefore, the early detection of precursor lesions is crucial. However, the risk assessment of precursor lesions such as oral epithelial dysplasia (OED) remains challenging because of the subjectivity of histopathological grading. We aimed to identify molecular markers that enhance the diagnostic accuracy and prognostic stratification of OSCC and explore the differences in the molecular characterization of OED and OSCC using a few selected markers. Methods: A two-step diagnostic workflow was applied: (1) FISH evaluation of EGFR amplification and CDKN2A deletion to distinguish OED from OSCC and identify EGFR-dependent tumors, and (2) HRAS immunohistochemistry performed exclusively in EGFR-negative OSCCs to stratify EGFR-independent cases. Fluorescence in situ hybridization (FISH) was used to assess seven EGFR/cell cycle-related genes (CCND1, CDKN2A, EGFR, PIK3CA, PTEN, TP53, and 1p36 locus) in 117 formalin-fixed paraffin-embedded samples (66 OED and 51 OSCC) and 10 normal mucosa samples. HRAS expression was evaluated using immunohistochemistry (IHC) in 36 EGFR amplification-negative OSCCs samples. Results: EGFR amplification was frequent in OSCC, whereas CDKN2A deletion was common in OED. The EGFR-amplified/ CDKN2A-intact profile showed high specificity for OSCC and improved diagnostic performance (area under the curve = 0.77) when combined with the Ki-67 labeling index. It also predicted poor disease-free survival (hazard ratio [HR] = 5.08, p = 0.016) and overall survival (HR = 6.10, p = 0.047). Among EGFR-negative OSCCs, HRAS overexpression was associated with advanced-stage disease and a poor prognosis (HR = 6.15, p = 0.043). Conclusions: EGFR amplification was frequent in OSCC, and CDKN2A deletion was prevalent in OED, supporting their use as molecular markers for differential diagnoses. FISH for EGFR/CDKN2A and HRAS IHC can stratify OSCC by diagnosis and prognosis, enabling practical molecular subclassification, including EGFR-negative cases.
