Histopathological Spectrum of Primary Non-Small Cell Lung Carcinomas and Metastatic Carcinomas to Lung in Bronchoscopic Biopsies: Immunohistochemical and Molecular Profiling Study

Lung cancer remains the most common cause of cancer-related deaths globally and represents a major public health challenge [1,2]. Non-small cell lung carcinoma (NSCLC) comprises approximately 85% of lung cancers and includes histologically and molecularly distinct subtypes, predominantly adenocarcinoma and squamous cell carcinoma, with differing therapeutic implications [3].

 

With the advent of targeted therapy and immunotherapy, accurate histological subtyping and molecular characterization have become mandatory for optimal patient management [4,5]. The WHO 2021 classification of thoracic tumors underscores a morphology-first diagnostic approach, followed by selective use of immunohistochemistry (IHC) and molecular testing, particularly in small biopsy specimens [1,6].

 

Bronchoscopic biopsies often yield limited tissue, increasing diagnostic uncertainty and resulting in categories such as NSCLC-NOS or poorly differentiated carcinoma when morphology alone is used [7,8]. Immunohistochemical markers such as TTF-1, Napsin-A, p40, and CK5/6 have been validated as reliable tools for distinguishing adenocarcinoma from squamous cell carcinoma in small biopsies [9–11]. Additionally, molecular testing for actionable driver mutations—including EGFR, ALK, KRAS, BRAF, and MET exon 14 alterations—is now standard of care in lung adenocarcinoma, especially in Asian and Indian populations where EGFR mutation prevalence is high [12–15].

 

This study analyses the histopathological spectrum of primary NSCLC and metastatic carcinomas to the lung diagnosed on bronchoscopic biopsies, evaluates the diagnostic contribution of immunohistochemistry, and assesses the molecular profile of lung adenocarcinomas.   

Study Design & Causes

A retrospective observational study was conducted on 210 bronchoscopic lung biopsy specimens.

 

Inclusion & Exclusion criteria:

Only treatment-naïve patients with histologically confirmed NSCLC or metastatic carcinomas to the lung were included. Cases with inconclusive biopsy reports, recurrent tumors, or prior treatment were excluded.

 

Clinical Data

Clinical parameters included age, sex, and smoking status. Never-smokers were defined as individuals who had smoked fewer than 100 cigarettes in their lifetime.

 

Histopathological Evaluation

All biopsy specimens were formalin-fixed and paraffin-embedded. Initial diagnosis was rendered on H&E-stained sections.

 

Immunohistochemistry

IHC was applied in NSCLC-NOS cases, poorly differentiated carcinomas, and tumors suspected to be metastatic, using lineage-specific and organ-specific markers as appropriate [9,10].

 

Molecular Profiling

Molecular testing was performed in lung adenocarcinoma cases using RT-PCR and IHC. Driver mutations evaluated included EGFR, KRAS, ALK, BRAF, and MET exon 14 alterations, following CAP–IASLC–AMP recommendations [16,17].

Clinicodemographic Profile

The study included 210 patients, of whom 168 (80%) were males and 42 (20%) were females. The median age was 62 years (range: 21–89 years). Ninety patients (42.9%) had a history of smoking (Table 1).

 

Table 1. Clinicodemographic Characteristics of the Study Population (n = 210)

 

Histopathological Diagnosis and IHC Correlation

Initial H&E diagnosis showed squamous cell carcinoma in 106 cases, adenocarcinoma in 44 cases, NSCLC-NOS in 30 cases, poorly differentiated carcinoma in 20 cases, adenosquamous carcinoma in 2 cases, and metastatic tumors in 3 cases (Table 2).

Table 2. Initial Histopathological Diagnosis on H&E and Immunohistochemical (IHC) Correlation

 

 

Immunohistochemistry enabled significant diagnostic refinement. Among 30 NSCLC-NOS cases, 20 were reclassified as adenocarcinoma, 6 as squamous cell carcinoma, and 4 retained a null phenotype. All poorly differentiated carcinomas were lineage-assigned (12 squamous cell carcinoma, 8 adenocarcinoma). Metastatic tumors were accurately characterized using organ-specific markers.

 

The final histological classification after IHC integration is shown in Table 3. Squamous cell carcinoma constituted 59% of cases, while adenocarcinoma accounted for 34.3%. The diagnostic utility of IHC in ambiguous tumors is summarized in Table 4.

 

Table 3. Final Histological Classification After IHC Integration

  

Table 4. Diagnostic Utility of Immunohistochemistry in Ambiguous Tumors

 

Histological Patterns of Adenocarcinoma

Among 72 adenocarcinoma cases, acinar (37%) and solid (27%) patterns were most frequent, followed by micropapillary, papillary, lepidic, and enteric patterns (Table 5).

Table 5. Histological Patterns of Lung Adenocarcinoma (n = 72)

 

Molecular Profiling

Driver mutations were detected in 24 adenocarcinoma cases. EGFR mutations were most common (41.7%), followed by KRAS (29.2%), ALK (16.7%), BRAF (8.3%), and MET exon 14 alterations (4.1%). Molecular alterations are detailed in Table 6.

 

Table 6. Molecular Alterations Detected in Lung Adenocarcinoma (n = 24)

 

Accurate classification of lung cancer on bronchoscopic biopsy specimens remains challenging due to limited tissue and tumor heterogeneity. In the present study, nearly one-fourth of cases were initially categorised as NSCLC-NOS or poorly differentiated carcinoma on morphology alone, consistent with previous reports [7,8].

 

The application of a focused immunohistochemical panel enabled definitive lineage assignment in 86.7% of NSCLC-NOS cases and all poorly differentiated carcinomas (Table 4). This finding aligns with earlier studies demonstrating the effectiveness of selective IHC in small biopsy specimens while preserving tissue for molecular testing [7,18].

 

Squamous cell carcinoma was the predominant subtype in our cohort (Table 3), reflecting smoking-associated lung cancer patterns reported in Indian studies [14,19]. Morphological features of well-differentiated squamous cell carcinoma were evident on H&E sections (Figure 1), while poorly differentiated variants required immunohistochemical confirmation (Figure 2).

Figure 1: Well-differentiated squamous cell carcinoma of lung showing nests of malignant squamous cells with keratin pearl formation (H&E 40X).

Figure 2: Poorly differentiated squamous cell carcinoma with sheets of pleomorphic tumor cells (A) H&E 40X), showing CK 5/6 Cytoplasmic positivity (B) and nuclear p40 positivity (C).

Adenocarcinoma constituted approximately one-third of cases, with acinar and solid patterns being most frequent (Table 5). Micropapillary and lepidic patterns, which carry prognostic significance, were identifiable even in small biopsy samples (Figures 3 and 4), highlighting the diagnostic adequacy of bronchoscopic biopsies when optimally processed.

 

Figure 3: Micropapillary adenocarcinoma of lung showing small clusters of tumor cells lacking fibrovascular cores (H&E 40X).

Figure 4: Adenocarcinoma Lung lepidic type(A) H&E 40X and TTF 1 positivity(B).

Metastatic tumors to lung, though infrequent, were accurately identified using organ-specific immunomarkers (Figures 5–7). This emphasizes the importance of clinical correlation and targeted IHC to avoid misclassification of metastases as primary lung tumors [20,21].

 

Molecular profiling revealed actionable driver mutations in one-third of adenocarcinoma cases (Table 6). The high frequency of EGFR mutations (41.7%) is consistent with Indian and Asian studies, underscoring the importance of routine EGFR testing in lung adenocarcinoma [12–15]. Detection of KRAS, ALK, BRAF, and MET exon 14 alterations further highlights the molecular heterogeneity of lung adenocarcinoma and the therapeutic relevance of comprehensive molecular testing [22–25].

 

Overall, our findings support the WHO 2021 and CAP–IASLC–AMP guidelines advocating an integrated diagnostic approach combining morphology, immunohistochemistry, and molecular profiling in small biopsy specimens [1,16,17].

 

Limitations

The retrospective design and lack of survival or treatment outcome data are acknowledged limitations. Expanded next-generation sequencing and uniform PD-L1 testing were not available in all cases, reflecting real-world constraints in resource-limited settings [17,26].

Immunohistochemistry is indispensable for refining lung cancer diagnosis on bronchoscopic biopsies, particularly in NSCLC-NOS and poorly differentiated tumors. Molecular profiling of lung adenocarcinomas reveals actionable driver mutations in a substantial proportion of cases. A morphology-first, tissue-sparing diagnostic algorithm ensures accurate classification and facilitates personalized therapy.

 

 

Figure 7: Metastatic paraganglioma involving lung showing (A) Zell Ballen pattern (H&E, 40X) with sustentacular cells highlighted by S-100 (B), Chromogranin positivity (C) and synaptophysin positivity (D).

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