Robust HPV‐16 Detection Workflow for Formalin‐Fixed Cancer Tissue and Its Application for Oral Squamous Cell Carcinoma
ABSTRACT Background Virus‐related cancers are malignancies caused by specific viruses, such as human papillomavirus (HPV), hepatitis B virus, and human T‐cell leukemia virus, contributing significantly to the global cancer burden through persistent infection and oncogenic transformation. The current...
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Wiley
2025-02-01
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| Online Access: | https://doi.org/10.1002/cam4.70544 |
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| author | Shizuka Morodomi Akiyuki Hirosue Akhinur Rahman Kyotaro Nohata Misaki Matsuo Omnia Reda Samiul Alam Rajib Haruki Saito Hiroki Takeda Ryoji Yoshida Masafumi Nakamoto Masatoshi Hirayama Kenta Kawahara Mitsuyoshi Takatori Yorihisa Orita Hideki Nakayama Yorifumi Satou |
| author_facet | Shizuka Morodomi Akiyuki Hirosue Akhinur Rahman Kyotaro Nohata Misaki Matsuo Omnia Reda Samiul Alam Rajib Haruki Saito Hiroki Takeda Ryoji Yoshida Masafumi Nakamoto Masatoshi Hirayama Kenta Kawahara Mitsuyoshi Takatori Yorihisa Orita Hideki Nakayama Yorifumi Satou |
| author_sort | Shizuka Morodomi |
| collection | DOAJ |
| description | ABSTRACT Background Virus‐related cancers are malignancies caused by specific viruses, such as human papillomavirus (HPV), hepatitis B virus, and human T‐cell leukemia virus, contributing significantly to the global cancer burden through persistent infection and oncogenic transformation. The current study aimed to develop a robust HPV‐16 detection method for formalin‐fixed cancer specimens. Materials and Methods To prevent false negatives resulting from DNA fragmentation, a DNA quality check step was added. Additionally, this study used multiplex polymerase chain reaction (PCR) covering the entire HPV‐16 genome to mitigate effects caused by viral sequence variation. To prove this concept, we analyzed genomic DNA extracted from oropharyngeal cancer tissues known as HPV‐16‐positive. Subsequently, the protocol was tested on oral squamous cell carcinoma (OSCC) samples in our cohort. Given the wide variation in HPV‐16 positivity in previous studies, it remains elusive how frequently HPV‐16 is positive in OSCC. Results The results showed faint bands or smears in the multiplex PCR of 7 out of 112 cases. Droplet digital PCR confirmed variable positivity levels of HPV‐16, suggesting two scenarios of HPV‐16 positivity in cancer tissue: cancer cells derived from infected cells or only a portion being HPV‐16‐positive. Finally, we comprehensively analyzed the case and identified the integration of a deleted HPV‐16 genome into the intronic region of the host gene TMEM94 on chromosome 17. To the best of our knowledge, this is the first evidence showing the integration of HPV‐16 in OSCC cells and providing its complete viral sequence. Conclusions The established protocol should be applicable to various cancer tissues for analyzing the association with HPV‐16 infection. |
| format | Article |
| id | doaj-art-cba00170dde247cf8e6a6d98e3ecb3ce |
| institution | DOAJ |
| issn | 2045-7634 |
| language | English |
| publishDate | 2025-02-01 |
| publisher | Wiley |
| record_format | Article |
| series | Cancer Medicine |
| spelling | doaj-art-cba00170dde247cf8e6a6d98e3ecb3ce2025-08-20T03:18:05ZengWileyCancer Medicine2045-76342025-02-01144n/an/a10.1002/cam4.70544Robust HPV‐16 Detection Workflow for Formalin‐Fixed Cancer Tissue and Its Application for Oral Squamous Cell CarcinomaShizuka Morodomi0Akiyuki Hirosue1Akhinur Rahman2Kyotaro Nohata3Misaki Matsuo4Omnia Reda5Samiul Alam Rajib6Haruki Saito7Hiroki Takeda8Ryoji Yoshida9Masafumi Nakamoto10Masatoshi Hirayama11Kenta Kawahara12Mitsuyoshi Takatori13Yorihisa Orita14Hideki Nakayama15Yorifumi Satou16Department of Oral and Maxillofacial Surgery, Faculty of Life Sciences Kumamoto University Kumamoto JapanDepartment of Oral and Maxillofacial Surgery, Faculty of Life Sciences Kumamoto University Kumamoto JapanDivision of Genomics and Transcriptomics, Joint Research Center for Human Retrovirus Infection Kumamoto University Kumamoto JapanDivision of Genomics and Transcriptomics, Joint Research Center for Human Retrovirus Infection Kumamoto University Kumamoto JapanDivision of Genomics and Transcriptomics, Joint Research Center for Human Retrovirus Infection Kumamoto University Kumamoto JapanDivision of Genomics and Transcriptomics, Joint Research Center for Human Retrovirus Infection Kumamoto University Kumamoto JapanDivision of Genomics and Transcriptomics, Joint Research Center for Human Retrovirus Infection Kumamoto University Kumamoto JapanDepartment of Otolaryngology‐Head and Neck Surgery Kumamoto University Graduate School of Medicine Kumamoto JapanDepartment of Otolaryngology‐Head and Neck Surgery Kumamoto University Graduate School of Medicine Kumamoto JapanDepartment of Oral and Maxillofacial Surgery, Faculty of Life Sciences Kumamoto University Kumamoto JapanDepartment of Oral and Maxillofacial Surgery, Faculty of Life Sciences Kumamoto University Kumamoto JapanDepartment of Oral and Maxillofacial Surgery, Faculty of Life Sciences Kumamoto University Kumamoto JapanDepartment of Oral and Maxillofacial Surgery, Faculty of Life Sciences Kumamoto University Kumamoto JapanDivision of Genomics and Transcriptomics, Joint Research Center for Human Retrovirus Infection Kumamoto University Kumamoto JapanDepartment of Otolaryngology‐Head and Neck Surgery Kumamoto University Graduate School of Medicine Kumamoto JapanDepartment of Oral and Maxillofacial Surgery, Faculty of Life Sciences Kumamoto University Kumamoto JapanDivision of Genomics and Transcriptomics, Joint Research Center for Human Retrovirus Infection Kumamoto University Kumamoto JapanABSTRACT Background Virus‐related cancers are malignancies caused by specific viruses, such as human papillomavirus (HPV), hepatitis B virus, and human T‐cell leukemia virus, contributing significantly to the global cancer burden through persistent infection and oncogenic transformation. The current study aimed to develop a robust HPV‐16 detection method for formalin‐fixed cancer specimens. Materials and Methods To prevent false negatives resulting from DNA fragmentation, a DNA quality check step was added. Additionally, this study used multiplex polymerase chain reaction (PCR) covering the entire HPV‐16 genome to mitigate effects caused by viral sequence variation. To prove this concept, we analyzed genomic DNA extracted from oropharyngeal cancer tissues known as HPV‐16‐positive. Subsequently, the protocol was tested on oral squamous cell carcinoma (OSCC) samples in our cohort. Given the wide variation in HPV‐16 positivity in previous studies, it remains elusive how frequently HPV‐16 is positive in OSCC. Results The results showed faint bands or smears in the multiplex PCR of 7 out of 112 cases. Droplet digital PCR confirmed variable positivity levels of HPV‐16, suggesting two scenarios of HPV‐16 positivity in cancer tissue: cancer cells derived from infected cells or only a portion being HPV‐16‐positive. Finally, we comprehensively analyzed the case and identified the integration of a deleted HPV‐16 genome into the intronic region of the host gene TMEM94 on chromosome 17. To the best of our knowledge, this is the first evidence showing the integration of HPV‐16 in OSCC cells and providing its complete viral sequence. Conclusions The established protocol should be applicable to various cancer tissues for analyzing the association with HPV‐16 infection.https://doi.org/10.1002/cam4.70544genetic diagnosisgenomic analysishead and neck cancerhuman papillomavirus (HPV)multiplex PCR |
| spellingShingle | Shizuka Morodomi Akiyuki Hirosue Akhinur Rahman Kyotaro Nohata Misaki Matsuo Omnia Reda Samiul Alam Rajib Haruki Saito Hiroki Takeda Ryoji Yoshida Masafumi Nakamoto Masatoshi Hirayama Kenta Kawahara Mitsuyoshi Takatori Yorihisa Orita Hideki Nakayama Yorifumi Satou Robust HPV‐16 Detection Workflow for Formalin‐Fixed Cancer Tissue and Its Application for Oral Squamous Cell Carcinoma Cancer Medicine genetic diagnosis genomic analysis head and neck cancer human papillomavirus (HPV) multiplex PCR |
| title | Robust HPV‐16 Detection Workflow for Formalin‐Fixed Cancer Tissue and Its Application for Oral Squamous Cell Carcinoma |
| title_full | Robust HPV‐16 Detection Workflow for Formalin‐Fixed Cancer Tissue and Its Application for Oral Squamous Cell Carcinoma |
| title_fullStr | Robust HPV‐16 Detection Workflow for Formalin‐Fixed Cancer Tissue and Its Application for Oral Squamous Cell Carcinoma |
| title_full_unstemmed | Robust HPV‐16 Detection Workflow for Formalin‐Fixed Cancer Tissue and Its Application for Oral Squamous Cell Carcinoma |
| title_short | Robust HPV‐16 Detection Workflow for Formalin‐Fixed Cancer Tissue and Its Application for Oral Squamous Cell Carcinoma |
| title_sort | robust hpv 16 detection workflow for formalin fixed cancer tissue and its application for oral squamous cell carcinoma |
| topic | genetic diagnosis genomic analysis head and neck cancer human papillomavirus (HPV) multiplex PCR |
| url | https://doi.org/10.1002/cam4.70544 |
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