HNSCC is the 4th highest incidence of cancer and 6th of cancer death of the males in Taiwan. Because the patients were mainly middle-aged male, the disease eventually resulted in a huge loss of labor force, productivity and a huge burden of family supports and medicinal costs. Currently, the primary treatments of HNSCC are mainly surgery, radiotherapy, chemotherapy or targeted therapy or concurrent chemoradiotherapy. Compared to oral cavity cancer, patients with pharyngeal cancer would possibly harbor HPV infections and have better treatment outcomes, prognosis and survival with clinically significance; however, the investigator's reports showed quite the opposite prognostic value in oral cavity cancer. The inconsistent data urges us to investigate further. Fortunately, in recent years, The investigator have developed a new method for isolation and detection of CTCs in HNSCC patients.The investigator's data found that high level of CTCs in patients with HNSCC and might be associated with disease prognosis, response to treatment and distant metastasis. This novel tool enhances the studies addressing on metastases or recurrence process in HNSCC patients. However, the investigator did not focus whether if the dynamic change of CTCs and specific surface markers on CTCs, such as P16+ CTCs are clinically meaningful.

Therefore, in the first year, the investigator will utilize the investigator's developing device and protocol to isolate high-purity CTCs to further identify P16+ on CTCs. In the following 2 years of the project, the investigator will enroll 150 freshly diagnosed patients with oral cavity, oropharyngeal, hypopharyngeal and laryngeal cancer at all stages (75 P16+ and 75 P16- patients) and 30 healthy donors for cell line tests, and then analyze CTCs, background white blood cells signals, and their initial biopsied tissue for P16 positivity test. Further statistical tests with clinical conditions (disease status, treatment effects, progression or distant metastasis and death) will be performed to elucidate their clinical significance.Hopefully, the investigator will clarify the clinical significance of circulating P16 expression status on CTCs by this study and provide a new biomarker for clinical cancer care.

1. Establish a protocol and a practical platform to longitudinally isolate, enumerate CTCs in patients with HPV+-head and neck cancer during the treatment course.
2. Elucidate the clinical significance of P16 INK4A -expressing CTCs to cancer status and treatment response.
3. Verify that dynamically monitoring P16 status and changes during anti-cancer treatment is feasible and clinically meaningful to survival or treatment responses.
4. Analyze different habitual conditions, i.e. alcohol, cigarette and betel nut use to P16-expressing CTCs.
5. Establish a good model to follow-up a specific surface marker on CTCs, which could be possibly utilized in other cancers with their particular markers (i.e. CEA on gastric cancer, Her2 on breast cancer)

Inclusion Criteria:

• histologically or cytopathologically proven head and neck squamous cell carcinoma Age ≥ 20 years. Measurable or evaluable disease according to Response Evaluation Criteria In Solid Tumors(RECIST) criteria.

Ability to sign informed consent.

Exclusion Criteria:

• Prior cancers within 5 years, except for non-melanoma skin cancers, and in situ cervical cancers.

Inability to completely with study and/or follow-up procedures.

The study will be conducted in three branch hospitals of Chang Gung Memorial Hospital, Taiwan including Keelung, Linkou, Taipei. Locally advanced or recurrent/metastatic head and neck cancer