Objective: To explore the characteristics of genomic variation patterns in Chinese patients with laryngeal and pharyngeal squamous cell carcinoma (SCC) and their correlation with differentiation and clinical significance. Methods: We analyzed genomic variations in 45 patients. Mutation patterns were evaluated using the 688 panel. We evaluated the correlation among degree of differentiation, patient prognosis, and mutation status and also analyzed 564 HNSCC samples from the UALCAN database. Results: Significant differences were observed in overall survival (OS) and progression-free survival (PFS) among patients with different degrees of differentiation. Based on the DriverML model, we found that the genes with the highest mutation rates were neurogenic locus notch homolog protein 1 (NOTCH1), tumor protein 53 (TP53), FAT atypical cadherin 1 (FAT1), and mitogen-activated protein kinase kinase kinase 4 (MAP3 K4) (over 30%). We are the first to our knowledge to propose that MAP3 K4 (33%) may be a driving gene for Chinese SCC patients. Moreover, NOTCH1 and CUB and sushi multiple domains 3 (CSMD3) were mutually exclusive (p < 0.05). CSMD3 mutations were primarily found in poorly differentiated patients (83%, 5/6). Furthermore, NOTCH1(wild) and MAP3 K4(wild) were mainly present in poorly differentiated patients (p = 0.011) as well. We also validated the differential expression of NOTCH1 and MAP3 K4 and their association (p < 0.05) with tumor differentiation using 564 HNSCC samples from the UALCAN database. Conclusion: We identified a potential new driving gene, MAP3 K4, in Chinese SCC patients and confirmed that the interaction between NOTCH1-MAP3 K4 may affect the differentiation of laryngeal and pharyngeal SCC. However, further exploration and large-scale sample validation are needed.