European Journal of Cardiovascular Medicine

Oral squamous cell carcinoma (OSCC) is one of the most common malignancies of the oral cavity, representing approximately 3% of all cancers worldwide and more than 90% of all oral cancers.¹ It remains a major public health challenge due to its high incidence, morbidity, and mortality. The prognosis of OSCC is closely linked with the stage at diagnosis. When detected at an early stage, the five-year survival rate is nearly 85%. Unfortunately, the majority of cases present in advanced stages, where survival decreases drastically to 30–50%.² Despite advances in surgical techniques, radiotherapy, chemotherapy, and targeted therapy, the overall global survival rate of OSCC has not improved significantly over recent decades.Biomarkers are measurable biological molecules that indicate the presence or progression of disease. In the era of personalized medicine, biomarkers hold tremendous promise for improving cancer outcomes³. In OSCC, immunohistochemical detection of proteins, mRNA expression profiling, and microRNA (miRNA) analysis have been widely studied as potential diagnostic tools. Several molecular signatures have been proposed for predicting prognosis and guiding therapy. For example, overexpression of proteins such as cyclin D1, p16, and Ki-67 has been linked with tumor progression, while specific miRNAs like miR-21 and miR-31 have been associated with tumor aggressiveness and poor prognosis⁴. These molecular insights pave the way for developing highly sensitive and specific diagnostic tests and even targeted therapies tailored to individual patients.Among the different diagnostic fluids available, saliva has emerged as a particularly attractive medium for biomarker discovery in OSCC. Saliva is often referred to as a “mirror of the body” because it contains a diverse array of locally and systemically derived biomolecules, including proteins, peptides, nucleic acids, metabolites, and microbiome-derived products. Its direct contact with oral lesions makes it an especially relevant diagnostic tool in OSCC.⁵ Collection of saliva is non-invasive, simple, cost-effective, and well-accepted by patients, offering a practical advantage over serum or tissue biopsies. Studies have demonstrated that salivary biomarkers can reliably reflect the molecular changes occurring in oral epithelial cells during carcinogenesis.Salivary biomarkers can be broadly categorized into genetic, proteomic, and metabolomic markers.^5,6 These genes are involved in inflammation, apoptosis, and cell proliferation, and their aberrant expression has been consistently detected in OSCC saliva samples. Proteomic biomarkers include cytokines like interleukin-6 (IL-6), interleukin-8 (IL-8), tumor necrosis factor-alpha (TNF-α), and proteins such as alpha-amylase, lactoferrin, mucins, histatins, and cystatins. Elevated levels of these proteins reflect tumor-induced inflammation, tissue remodeling, and immune evasion. Metabolomic biomarkers, on the other hand, provide insights into altered metabolic pathways in cancer cells, including amino acid metabolism, polyamine synthesis, and energy production³.This hierarchical cascade underscores the potential of using saliva for multi-omics biomarker discovery.⁷The clinical utility of salivary biomarkers extends beyond early diagnosis. They also hold promise for risk prediction, monitoring of disease recurrence, and evaluation of treatment response. The exploration of salivary biomarkers offers a non-invasive, reliable, and cost-effective approach for early detection, prognosis, and therapeutic monitoring. With advances in genomics, proteomics, metabolomics, and bioinformatics, the integration of salivary biomarker panels into routine clinical practice appears increasingly feasible^7,8. Future research should focus on validating these biomarkers in large, diverse populations and developing standardized, point-of-care diagnostic platforms that can transform OSCC management and improve patient outcomes.

AIM

To assess the Salivary biomarkers in oral squamous cell carcinoma at a tertiary care centre

T This was a case–control study conducted in the department of dentistry, Govt medical college, Barmer. A total of 58 participants were enrolled, consisting of 29 histopathologically confirmed cases of oral squamous cell carcinoma (OSCC) and 29 age-sexmatched healthy controls. Patients with newly diagnosed and untreated OSCC were included in the study. Healthy volunteers without any oral lesions or systemic illness were recruited as controls. Patients with recurrent or previously treated OSCC, other malignancies, systemic inflammatory or autoimmune diseases, and metabolic disorders were excluded. Individuals who did not provide informed consent were also excluded. This design ensured homogeneity of the study groups and minimized confounding factors

Table 1:Age Group Distribution

The majority of study participants in both groups were in the 50–59 years age group, followed by 40–49 years. Fewer participants were aged ≥70 years in both cases and controls.

Table 2: Gender distribution

In this study, males predominated in both groups, with 21 cases and 19 controls. Females were fewer in number, comprising 8 cases and 10 controls.

Table 3:Lifestyle Risk Factors (Smoking and Alcohol Consumption) in Cases and Controls

A significantly higher proportion of cases were smokers (79.5%) and alcohol consumers (68.4%) compared to controls, where only 20.5% were smokers and 31.6% consumed alcohol. This highlights the strong association of tobacco and alcohol use with OSCC.

Table 4: Lymph node involvement and tumor staging

Among the OSCC cases, lymph node metastasis was present in 16 patients while 13 had no nodal involvement. Tumor staging revealed 11 patients in stage T I–II and 18 patients in stage T III–IV, whereas controls showed no such findings.

Table 5: biomarkers range in cases and controls

The mean levels of IL-8, IL-6, TNF-α, MMP-9, CYFRA 21-1, and LDH were significantly elevated in OSCC cases compared to controls, whereas AZGP1 levels were markedly reduced. These differences highlight the diagnostic potential of salivary biomarkers in distinguishing OSCC patients from healthy individuals.

Table 6:Salivary Biomarkers According to Tumor Stage

When compared across tumor stages, salivary biomarker levels showed no major differences between early (I–II) and late (III–IV) OSCC, though LDH and CYFRA 21-1 tended to be slightly higher in advanced stages. Overall, biomarker alterations were consistent irrespective of tumor stage.

In the present study, the age distribution of participants showed that the highest proportion belonged to the 50–59 years group, comprising 11 cases and 13 controls.The next common group was 40–49 years with 9 cases and 7 controls.Participants in the 60–69 years group included 7 cases and 6 controls.The least represented group was ≥70 years, with only 2 cases and 3 controls.This indicates that the majority of oral squamous cell carcinoma (OSCC) patients as well as controls were middle-aged to early elderly.Thus, the disease burden was observed predominantly in the 5th and 6th decades of life.

The sex-wise distribution of participants revealed a male predominance in both cases and controls.Among the cases, 21 were males while only 8 were females.Similarly, in the control group, 19 were males and 10 were females.This reflects a higher representation of males compared to females in the study population.The findings are consistent with the known higher prevalence of oral squamous cell carcinoma (OSCC) in males, often attributed to greater exposure to risk factors such as tobacco and alcohol.Overall, the male-to-female ratio was higher in cases, indicating gender-related differences in disease occurrence.

Among the cases, 79.5% were smokers, whereas only 20.5% of the controls reported smoking.Similarly, alcohol consumption was seen in 68.4% of the cases compared to 31.6% of the controls.This clearly indicates a higher prevalence of these risk factors among OSCC patients.The findings reinforce the role of tobacco and alcohol as significant contributors to the development of oral cancer.

In the present study, lymph node involvement was observed in more than half of the OSCC cases.In a study by Kumar, Deepak;⁹Patadiya et al the first table provides a summary of the participants’ clinical and demographic characteristics. Seventy percent of the OSCC group’s members were male, and their mean age was 56.3 ± 8.5 years. The control group consisted of 66% men and had a mean age of 54.7 ± 9.2 years. Compared to the control group, where 24% of participants reported smoking and 20% reported drinking, a larger proportion of individuals in the OSCC group reported smoking (80%) and alcohol consumption (64%) .

Out of 29 patients, 16 had lymph node metastasis while 13 showed no nodal spread.

This indicates a considerable proportion of advanced disease presentation at the time of diagnosis.Tumor staging further revealed that 11 patients were in the early stages (T I–II).However, the majority, 18 patients, were categorized in advanced stages (T III–IV).No such findings were noted among the control group, highlighting the disease-specific nature of these parameters.In a study by Jain, A., Kotimoole, C.N., Ghoshal, S. et al.¹⁰  the total recruited cases, 48% (n = 24) had no regional lymph node metastasis (N0) while 52% (n = 26) had regional lymph node metastasis (N+). 36% (n = 18) had a primary tumour of stage T1/T2 and 64% (n = 32) had a primary tumour of stage T3/T4. Post-treatment disease status was recorded for all cases.

In this study, salivary biomarker analysis showed distinct differences between OSCC cases and controls.The mean levels of pro-inflammatory cytokines IL-8, IL-6, and TNF-α were markedly elevated in cases compared to controls.Similarly, matrix metalloproteinase MMP-9 and tumor marker CYFRA 21-1 showed significantly higher values in OSCC patients.LDH, an indicator of tissue damage and tumor metabolism, was also substantially raised in the case group.In contrast, the salivary level of AZGP1 was found to be significantly reduced in OSCC cases.These findings underline the potential role of salivary biomarkers in early detection and diagnosis of oral cancer.Sahibzada HA, Khurshid Z, et al ¹¹reviews the expression of levels of specific cytokines i.e., IL-8, IL-6 and TNF-α, their signaling pathways in the development of oral cancer, and how they are essential for the diagnosis of OSCC and updates related to it. Riccardi, G.; Bellizzi, M.G.;Fatuzzo et al¹² reviewed literature of biomarker proteins in saliva that could also be evaluated as probabletargets for non-invasive screening of oral neoplasm such as cytokines, matrix metalloproteinases, andacute-phase response proteins. In Bastías, D.; Maturana, A.;Marín et al ¹³study TNF-α, IL-1β, IL-6 IL-8, LDH, and MMP-9 were the most studied, with almost all studies reporting high sensitivity and specificity values. TNF-α, IL-1β, IL-6 IL-8, LDH, and MMP-9 are the most promising salivary biomarkers.

The stage-wise analysis of salivary biomarkers in OSCC revealed largely comparable values between early (I–II) and late (III–IV) stages.IL-8, IL-6, and TNF-α levels remained consistently elevated across both groups with minimal variation.MMP-9 showed a marginally higher mean in early stages compared to late stages.CYFRA 21-1 values were slightly increased in advanced tumors, suggesting a trend toward higher expression with progression.AZGP1 levels remained low in both groups without significant stage-wise differences.LDH levels, however, were notably higher in late-stage patients, indicating its potential association with tumor burden and disease advancement.

The present study highlights the significant role of salivary biomarkers in the diagnosis of oral squamous cell carcinoma (OSCC). Demographic analysis showed that OSCC was most prevalent in middle-aged to elderly individuals, with a marked male predominance, reflecting higher exposure to risk factors. Lifestyle assessment confirmed the strong association of tobacco smoking and alcohol consumption with OSCC development. More than half of the patients presented with lymph node metastasis and advanced-stage tumors, indicating late diagnosis in many cases. Biomarker profiling demonstrated significantly elevated levels of IL-8, IL-6, TNF-α, MMP-9, CYFRA 21-1, and LDH in OSCC cases, while AZGP1 was consistently reduced, reinforcing their diagnostic value. Although most biomarkers did not show major stage-wise variation, LDH and CYFRA 21-1 tended to be higher in advanced stages, suggesting their potential link with disease progression. Overall, salivary biomarkers emerge as a promising, non-invasive diagnostic tool for the early detection and monitoring of OSCC.

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