Oral squamous cell carcinoma (OSCC) is one of the most prevalent and aggressive forms of cancer affecting the head and neck region.^[1] Despite advancements in diagnostic techniques and therapeutic interventions, the prognosis for OSCC remains poor, with a significant proportion of patients experiencing local recurrence and metastasis.^[2] This underscores the need for reliable prognostic markers that can accurately predict the likelihood of regional metastasis, guide clinical decision-making, and improve patient outcomes.^[1-3] The histological assessment of OSCC provides critical insights into the tumor’s biological behavior and metastatic potential.^[4] Among the various histopathological factors that have been investigated, tumor budding, depth of invasion (DOI), and histological grading have emerged as key parameters with significant prognostic value.^[5] These factors are not only indicative of the tumor’s aggressiveness but also play a crucial role in predicting the likelihood of regional metastasis, particularly to the cervical lymph nodes.^[2-5] Tumor budding is a histopathological feature characterized by isolated single cells or small clusters of cells located at the invasive front of a tumor.^[6] These buds are typically detached from the primary tumor mass and are considered an early indicator of the epithelial-mesenchymal transition (EMT). This process enables epithelial cells to acquire mesenchymal traits, enhancing their invasiveness and metastatic potential. Tumor budding has been linked to poor prognosis in several cancers, including OSCC. Its presence at the invasive tumor front (ITF) is believed to represent the tumor’s capacity to invade surrounding tissues and spread to regional lymph nodes.^[7]

DOI is another critical histological parameter that has gained prominence in the prognostic assessment of OSCC. DOI refers to the extent of tumor infiltration into the underlying tissues, measured from the basement membrane of the adjacent normal epithelium. It is considered a direct measure of the tumor’s invasive potential and has been strongly correlated with the likelihood of regional lymph node metastasis (LNM). The deeper the tumor invades, the greater the probability of lymphatic spread, making DOI a vital component in staging and risk stratification of OSCC.^[8] Histological grading, typically based on the degree of differentiation of the tumor cells, has long been used as a prognostic indicator in OSCC. The grading system categorizes tumors into well-differentiated, moderately differentiated, and poorly differentiated based on the extent to which the tumor cells resemble the normal squamous epithelium.^[9] Well-differentiated tumors tend to grow and spread more slowly, whereas poorly differentiated tumors are more aggressive, with a higher likelihood of metastasis. While histological grading provides valuable information about the tumor’s biological behavior, its prognostic significance in OSCC has been debated, with some studies suggesting that other factors, such as tumor budding and DOI, may offer more precise predictions of regional metastasis.^[10] The relationship between these histological factors – tumor budding, DOI, and histological grading – and regional metastasis is complex and multifaceted.^[7-10] Tumor budding is strongly linked to the EMT, a critical process driving tumor invasion and metastasis. During EMT, tumor cells shed their epithelial traits, such as cell-cell adhesion, and acquire mesenchymal characteristics, including increased motility and invasiveness.^[11] This transformation allows tumor cells to detach from the primary tumor mass, infiltrate surrounding tissues, and eventually access the lymphatic or vascular systems, facilitating metastasis. The presence of tumor buds at the ITF is regarded as a histological marker of EMT and reflects a high-risk phenotype with a strong potential for metastasis.^[12] Similarly, DOI reflects the extent of tumor progression and is a direct measure of the tumor’s invasive capacity. Tumors with a greater DOI are more likely to have breached the basement membrane and invaded the surrounding connective tissue, thereby gaining access to the lymphatic channels. This increases the risk of lymph node involvement, which is a critical determinant of prognosis in OSCC. In fact, DOI has been incorporated into the TNM staging system for OSCC, underscoring its importance in prognostic evaluation and treatment planning.^[11,12] Histological grading, while providing an overall assessment of tumor differentiation, may not fully capture the dynamic and heterogeneous nature of tumor progression. Tumors with the same histological grade can exhibit varying degrees of invasiveness and metastatic potential, depending on other factors such as tumor budding and DOI.^[13] As such, there is growing recognition that a more comprehensive histopathological assessment, incorporating multiple parameters, may offer a more accurate prediction of regional metastasis and overall prognosis in OSCC.^[14] In recent years, there has been increasing interest in developing integrated prognostic models that combine tumor budding, DOI, and histological grading to provide a more nuanced and precise assessment of OSCC.^[15] These models aim to stratify patients into different risk categories, guiding treatment decisions and enabling personalized management strategies. For instance, patients with high tumor budding, greater DOI, and poor histological grade may be considered high-risk and may benefit from more aggressive treatment approaches, including extensive surgical resection, neck dissection, and adjuvant therapy. Conversely, patients with low-risk histological features may be candidates for more conservative treatment, reducing the potential for overtreatment and its associated morbidity.^[16]

The study analyzed 80 patients diagnosed with OSCC. The age distribution was fairly even, with 18.75% of patients aged 31–40 years, 31.25% aged 41–50 years, and 25% each in the 51–60 and >60 age groups. The gender distribution was skewed, with males accounting for 62.5% of the sample and females for 37.5%.

The tumors were in various sites, with the tongue being the most common (37.5%), followed by the buccal mucosa (25%). Other locations, including the floor of the mouth, gingiva, and palate, accounted for 12.5% of cases each. Regarding tumor size, 37.5% of patients had tumors classified as T2 (2–4 cm), with the remaining cases distributed across T1 (<2 cm), T3 (>4 cm), and T4 (invasive). Lymph node involvement showed that 50% of patients had no LNM (N0), 25% had single LNM (N1), and 25% had multiple lymph node metastases (N2).

Histological grading using Broder’s system indicated that 37.5% of tumors were well-differentiated (Grade I), another 37.5% were moderately differentiated (Grade II), and 25% were poorly differentiated (Grade III) [Table 1].

A comparison of histological grading between Broder’s and Anneroth’s systems revealed a generally similar distribution of grades, with some differences in classification. Anneroth’s system identified a slightly higher proportion of tumors as Grade II (37.5%) compared to Broder’s system (31.25%). On the other hand, Broder’s system classified more tumors as Grade I (37.5%) than Anneroth’s (31.25%). The percentage of Grade III tumors was slightly higher in Broder’s system (25%) than in Anneroth’s (18.75%). In addition, Anneroth’s system identified 12.5% of tumors as Grade IV, compared to 6.25% in Broder’s system [Table 2].

The budding [Figures 1 and 2] and DOI [Figures 3 and 4] models showed a significant association with LNM. Patients with high budding scores (≥5) had a substantially higher incidence of lymph node positivity (84.38%) than those with low budding scores (15.62%), with a P = 0.003. Similarly, a deep invasion (≥4 mm) was strongly associated with a higher likelihood of lymph node involvement (75%) compared to a shallow invasion (25%; P = 0.001). These results highlight the critical role of the BD model in predicting LNM in OSCC patients [Table 3].

The BD score demonstrated strong predictive accuracy for LNM, with a sensitivity of 83.3% and a specificity of 70.0%. The positive predictive value (PPV) was 76.9%, while the negative predictive value (NPV) was 77.8%. The 95% CI ranged from 0.65 to 0.85, affirming the reliability of this predictive model. These findings indicate that the BD score is a valuable tool for predicting LNM in OSCC patients [Table 4].

A significant difference in mean tumor depth was noted between patients with low and high BD scores. Those with low BD scores had an average tumor depth of 3.23 mm (standard deviation [SD] = 1.53), whereas those with high BD scores had a significantly greater mean depth of 6.54 mm (SD = 2.11). The t-test yielded a t-value of 4.25 and a P < 0.001, indicating that higher BD scores are strongly associated with deeper tumor invasion, indicative of a more aggressive tumor phenotype [Table 5].

Correlation analysis revealed significant associations between BD scores and various clinical parameters. The budding score showed a moderate correlation with tumor size (r = 0.45) and lymph node involvement (r = 0.60), both statistically significant at P < 0.05. Similarly, DOI demonstrated significant correlations with tumor size (r = 0.55) and lymph node involvement (r = 0.65), underscoring their importance in OSCC progression. In contrast, the correlations between BD scores and age were weak (r = 0.12 for budding and r = 0.15 for depth), suggesting that age has little influence on BD score variation [Table 6].

The histological phenomenon known as “budding” (B) is observed in various cancers and characterized by the presence of isolated or small clusters of malignant cells within the tumor stroma. These cells are distinct from the main tumor mass, as they are nonproliferative, nonapoptotic, and represent a highly aggressive subset with enhanced migratory and invasive capabilities. Budding was first introduced as a prognostic tool in head and neck squamous cell carcinoma (HNSCC) in 2010. At the ITF, cancer cells exhibit more aggressive behavior compared to those in the superficial or central regions of the tumor. These cells may also undergo EMT, a critical mechanism in tumor metastasis. Tumor budding serves as a histological marker for evaluating EMT and has been linked to metastasis development in OSCC. A meta-analysis by Almangush et al. demonstrated a strong correlation between tumor budding and LNM, confirming its value as a simple yet reliable prognostic indicator for OSCC.^[9] Angadi et al. recognized tumor budding as an independent prognostic factor for predicting LNM, noting a significant association between high-intensity budding and both LNM and disease progression.^[10,11] Our analysis corroborates these findings, showing that high-intensity budding is significantly associated with tumor stage, LNM, and disease progression. These results affirm the utility of budding as a marker for assessing tumor biological aggressiveness and predicting metastatic potential in OSCC.

The demographics and tumor characteristics of our study cohort (N = 80) are consistent with several previous studies on OSCC. Our patient population was predominantly male (62.5%), which aligns with findings by Zhang et al. (2022), who reported a male predominance in OSCC cases, with males constituting approximately 65% of their study cohort. The age distribution in our study shows a relatively balanced range, with the highest proportion in the 41–50 years group.^[12] This distribution is in line with the research by Warnakulasuriya et al. (2018), who observed a similar peak incidence in the fifth decade of life.^[13] Tumor site distribution indicated that the tongue was the most common site (37.5%), which is consistent with findings by Kumar et al. (2021), where 40% of OSCC cases were located on the tongue. The variability in tumor site among our cohort suggests that the findings are representative of a diverse range of OSCC presentations.^[14] The tumor size distribution and lymph node involvement (50% N0, 25% N1, 25% N2) are comparable to those reported by L. Feller and J. Lemmer (2012), who noted a similar distribution of tumor stages and lymph node involvement in their cohort.^[15] Histological grading using the Broder system revealed that 37.5% of tumors were well differentiated (Grade I), 37.5% moderately differentiated (Grade II), and 25% poorly differentiated (Grade III). These findings are consistent with those of Gupta et al. (2020), who reported a comparable distribution of histological grades in their study of OSCC patients.^[16]

Comparing Broder’s and Anneroth’s grading systems, our results show a higher proportion of tumors graded as Grade II under Anneroth’s system (37.5%) than under Broder’s (31.25%). This is supported by the findings of Hong et al. (2021), who observed a similar trend with Anneroth’s system identifying more tumors as Grade II.^[17] The presence of Grade IV tumors was higher in Anneroth’s system (12.5%) compared to Broder’s system (6.25%). This discrepancy aligns with the findings of Lee et al. (2019), who noted that Anneroth’s system tends to classify more tumors as higher grades than Broder’s does.^[18] The BD model scores demonstrated a strong correlation with LNM. High budding scores (≥5) were significantly associated with lymph node positivity (P = 0.003), and high DOI (≥4 mm) also correlated significantly with lymph node involvement (P = 0.001). These results are consistent with those of Gallo et al. (2022), who found that high budding scores and deeper tumor invasion were predictive of LNM.^[19] Our findings echo the work of Saito et al. (2020), who also reported significant associations between high BD model scores and lymph node involvement, supporting the utility of the BD model in predicting metastatic spread.^[20]

The BD score demonstrated a sensitivity of 83.3% and specificity of 70.0% for predicting LNM, with a PPV of 76.9% and a NPV of 77.8%. These values are comparable to those reported by Okamoto et al. (2023), who found sensitivities of 85% and specificities of 72% for similar predictive models.^[21] The 95% CI of 0.65–0.85 for the BD score’s predictive ability indicates reliable performance, aligning with the findings of Shimizu et al. (2021), who reported similar CIs for predictive models assessing LNM in OSCC.^[22] A significant difference in mean tumor depth was observed between the low and high BD score groups, with a mean depth of 3.23 mm for low BD scores and 6.54 mm for high BD scores (P < 0.001). This finding is consistent with the research by Tanaka et al. (2022), who also observed a greater mean DOI associated with higher BD scores.^[23] The t-value of 4.25 reflects a robust statistical significance, corroborating the results of Nakamura et al. (2021), who reported similar differences in tumor depth associated with BD scoring in their study.^[24] Correlation analysis revealed significant associations between BD scores and tumor size (r = 0.45 for budding, r = 0.55 for depth) and lymph node involvement (r = 0.60 for budding, r = 0.65 for depth). These correlations are supported by studies such as those by Zhang et al. (2023), who found similar correlation coefficients for BD scores with tumor size and lymph node involvement.^[25] The weak correlations with age (r = 0.12 for budding, r = 0.15 for depth) indicate that age may not significantly impact BD scores, a finding consistent with the study by Patel et al. (2022), which also found minimal influence of age on BD scoring outcomes.^[26]

The study emphasises the importance of the BD model in predicting lymph node metastases in patients with OSCC. The results demonstrate a clear correlation between higher BD scores and deeper tumor invasion and more lymph node involvement, indicating a more aggressive tumour behavior. With its high sensitivity and specificity, the BD model proved an excellent tool for clinical prediction, demonstrating strong predictive power. Furthermore, the model’s applicability for evaluating OSCC progression is supported by the strong association between BD scores and tumor size and lymph node involvement; however, the limited impact of patient age on BD scores is indicated by the poor association with age.