## 1 Introduction  
Oral cancer, dominated up to 90% by oral squamous cell carcinoma (OSCC), is a highly prevalent cancer across the world. In general, OSCC has a poor prognosis because of its aggressive and metastatic nature, in which the recurrence arises in about 30% of cases, leading to a low 5-year survival and high mortality rates [1, 2]. The use of anticancer drugs is imperative for the treatment of advanced cancers and metastases [3]. The arsenal of cancer pharmacologic treatment can be simply divided into three classes: cytotoxic chemotherapy for halting the rapid and uncontrolled proliferation of cancer cells, targeted therapy for directing the molecular targets involved in the tumorigenic processes, and immunotherapy for targeting the immune checkpoints implicated in the immune surveillance [4, 5]. Among the panel of approved cytotoxic chemotherapeutic agents, 5-Fluorouracil (5-FU),an antimetabolite drug, is commonly used to treat many cancers. The cytotoxicity of 5-FU is ascribed to the inhibition of essential biosynthetic processes and the incorporation into macromolecules (such as DNAs and RNAs) to suppress their normal function [6]. A past study demonstrated the anticancer effects of 5-FU in the cultured oral cancer cells, with apoptosis as the major mechanism that determined the therapeutic efficacy [7]. However, a phase III trial in patients with recurrent head and neck cancer reported that the overall response rate to 5-FU treatment was as low as 13% [8]. In addition to the low response rate, high drug resistance also presents a critical obstacle to the clinical use of 5-FU [9]. Therefore, identification of new therapeutic agents for their potential applicability combined with 5-FU in OSCC treatment may help reduce adverse effects and improve patient survival. The mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) pathway is crucial for cancer cell growth, survival, and differentiation [10]. By examining the biopsy specimens of OSCC, it was found that ERK1/2 was overexpressed and related to the proliferation in OSCC [11]. The extracellular signal-regulated kinase (MEK) is the downstream of BRAF in the RAS-RAF-MEK-ERK pathway, which is important for signal transduction to ERK [10]. The MEK inhibition provides an encouraging strategy for targeted therapy due to the high selectivity for its target pathway RAS-RAF-MEK-ERK, thereby halting cell proliferation and promoting apoptosis [12; 13]. Furthermore, the combination of MEK inhibitors with other therapies, including chemotherapy, targeted therapy, and immunotherapy, is considered a hopeful prospect for clinical use [13]. Refametinb is a potent, highly selective, and allosteric inhibitor of MEK1/2. Its excellent pharmacokinetic profile and potency have led to its clinical development. [12; 13]. In a subset of cancer patients, Refametinb has demonstrated lower toxicities compared to first-generation MEK inhibitors [14; 15]. This suggests that it could be a promising new option for the treatment of ERK-associated malignancies. In several clinical trials, Refametinb has shown positive results as a monotherapy or in combination therapy, effectively inhibiting tumor growth in various solid tumors through MEK inhibition [16; 17; 18]. From this point of view, in the present study, we investigated the chemosensitivity of a human OSCC cell line (SCC4) to 5-FU, either alone or in combination with Refametinb, to determine whether this orally bioavailable selective MEK1/2 inhibitor may increase the effectiveness of 5-FU for OSCC treatment. Improving our understanding of the mechanisms behind drug combinations would help us enhance the therapeutic efficacy of cancer treatments for patients. One crucial factor that allows cancer cells to multiply, spread, and metastasize is the absence of antitumor immunity [4]. The programmed death-ligand 1 (PD-L1) is a vital immune checkpoint that functions as an immunological brake in the modulation of T-cell activation, thereby compromising the immune surveillance.