A STUDY ON CANCER ASSOCIATED FIBROBLAST USING ALPHA SMOOTH MUSCLE ACTIN IMMUNOHISTOCHEMISTRY IN ORAL SQUAMOUS CELL CARCINOMA

Abstract

Background Oral-squamous cell carcinoma (O-SCC) represents a significant global health burden with complex pathophysiology involving TME interactions. The tumour stroma, particularly CAFs, plays a crucial role in tumour advancement, invasion, as well as metastasis. CAFs, identified by alpha- SMA expression, influence tumour behavior through ECM remodelling and pro-tumorigenic signalling. Despite emerging evidence of their prognostic significance, the relationship between CAF expression patterns and clinicopathological parameters in O-SCC remains inadequately characterized. Objectives This study aimed to detect CAFs using α-SMA IHC in O-SCC and evaluate their association with LNM and pTNM staging, potentially identifying new prognostic markers for clinical management. Methodology This laboratory-based analytical study was conducted at Sri Devaraj Urs Medical College between September 2022 and December 2023. Histopathologically confirmed O-SCC cases (n=88) treated by composite resection and cervical lymph node dissection were included, excluding recurrent cases, patients who received neoadjuvant chemotherapy, and second primary cancers. Haematoxylin and eosin slides were reviewed for LNM and pTNM staging. Immunohistochemical staining for α-SMA was performed on 4μm formalinfixed paraffin-embedded tissue sections using heat-induced antigen retrieval, followed by incubation with primary antibody and HRP-conjugated secondary antibody. CAF expression was quantified using Kellermann's scoring system (Score 1is less than 1%, Score 2 is between 1 and 50%, Score 3 is more than 19 50% stained cells) and categorized by distribution pattern (focal, network, or spindle). Additional parameters assessed included TSR, WPOI, T-B, and TILs. Results The study population comprised 88 patients (69.3% female, 30.7% male) with an average age of 56.97 years. The buccal mucosa represented the most frequent site (50%), with well-differentiated tumours predominating (80.7%). Pathological staging revealed stage IVA as most prevalent (33%), followed by stage III (31.8%). Regarding CAF expression, score 3 (abundant CAFs) was observed in 55.7% of cases, Score 2 in 40.9%, and Score 1 in only 3.4%. Network pattern CAF distribution predominated (38.6%), with equal representation of focal and spindle configurations (30.7% each). Statistical analysis revealed no significant association between CAF scores and LNM (p=0.758); however, CAF distribution patterns demonstrated a statistically significant association with pTNM staging (χ²=26.716, p=0.001), with advanced stages showing distinct pattern shifts toward network and spindle arrangements. Notably, significant correlations were observed between TSR and CAF score (p<0.0001), T-B and CAF score (p=0.021), and T-B and LNM (p=0.047). More aggressive invasion patterns demonstrated higher CAF scores and increased TB intensity. Conclusion While CAF scores alone did not predict LNM, CAF architectural patterns demonstrated significant associations with pathological staging in O-SCC. The correlations between CAF expression, T-B, and invasion patterns suggest that CAF distribution, rather than mere presence, may serve as a valuable prognostic indicator. These findings highlight the potential of CAF architectural evaluation as an adjunctive histopathological parameter for risk stratification in O-SCC patients