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Updated: Nov 9, 2025
Generation and Expansion of Primary, Malignant Pleural Mesothelioma Tumor Lines
Published on: April 21, 2022
P14/ARF-Positive Malignant Pleural Mesothelioma: A Phenotype With Distinct Immune Microenvironment
Federica Pezzuto1, Francesca Lunardi1, Luca Vedovelli1
1Department of Cardiac, Thoracic, Vascular Sciences and Public Health, University of Padova, Padova, Italy.
Introduction:
The CDKN2A gene plays a central role in the pathogenesis of malignant pleural mesothelioma (MPM). The gene encodes for two tumor suppressor proteins, p16/INK4A and p14/ARF, frequently lost in MPM tumors. The exact role of p14/ARF in MPM and overall its correlation with the immune microenvironment is unknown. We aimed to determine whether there is a relationship between p14/ARF expression, tumor morphological features, and the inflammatory tumor microenvironment.
Methods:
Diagnostic biopsies from 76 chemo-naive MPMs were evaluated. Pathological assessments of histotype, necrosis, inflammation, grading, and mitosis were performed. We evaluated p14/ARF, PD-L1 (tumor proportion score, TPS), and Ki-67 (percentage) by immunohistochemistry. Inflammatory cell components (CD3+, CD4+, CD8+ T lymphocytes; CD20+ B-lymphocytes; CD68+ and CD163+ macrophages) were quantified as percentages of positive cells, distinguishing between intratumoral and peritumoral areas. The expression of p14/ARF was associated with several clinical and pathological characteristics. A random forest-based machine-learning algorithm (Boruta) was implemented to identify which variables were associated with p14/ARF expression.
Results:
p14/ARF was evaluated in 68 patients who had a sufficient number of tumor cells. Strong positivity was detected in 14 patients (21%) (11 epithelioid and 3 biphasic MPMs). At univariate analysis, p14/ARF-positive epithelioid mesotheliomas showed higher nuclear grade (G3) (p = 0.023) and higher PD-L1 expression (≥50%) (p = 0.042). The percentages of CD4 and CD163 in peritumoral areas were respectively higher and lower in p14/ARF positive tumors but did not reach statistical significance with our sample size (both p = 0.066). The Boruta algorithm confirmed the predictive value of PD-L1 percentage for p14/ARF expression in all histotypes.
Conclusions:
p14/ARF-positive epithelioid mesotheliomas may mark a more aggressive pathological phenotype (higher nuclear grade and PD-L1 expression). Considering the results regarding the tumor immune microenvironment, p14/ARF-negative tumors seem to have an immune microenvironment less sensitive to immune checkpoint inhibitors, being associated with low PD-L1 and CD4 expression, and high CD163 percentage. The association between p14/ARF-positive MPMs and PD-L1 expression suggests a possible interaction of the two pathways. Confirmation of our preliminary results could be important for patient selection and recruitment in future clinical trials with anticancer immunotherapy.
Insights
The tumor suppressor p14/ARF expression in malignant pleural mesothelioma (MPM) correlates with aggressive features and PD-L1 levels. p14/ARF-negative MPMs may show less response to immunotherapy, impacting patient selection for clinical trials.
Area of Science:
- Oncology
- Immunology
- Pathology
Background:
- The CDKN2A gene, encoding tumor suppressors p16/INK4A and p14/ARF, is frequently altered in malignant pleural mesothelioma (MPM).
- The specific role of p14/ARF in MPM pathogenesis and its relationship with the tumor immune microenvironment remain unclear.
- This study investigates the association between p14/ARF expression, MPM morphology, and the inflammatory tumor microenvironment.
Purpose of the Study:
- To determine the relationship between p14/ARF expression and clinicopathological features in MPM.
- To investigate the correlation of p14/ARF expression with the immune cell infiltrate in the tumor microenvironment.
- To explore the potential implications of p14/ARF expression for immunotherapy response in MPM.
Main Methods:
- Pathological assessment of 76 chemo-naive MPM biopsies, including histotype, grade, and mitosis.
- Immunohistochemical evaluation of p14/ARF, PD-L1 (TPS), and Ki-67.
- Quantification of immune cell populations (CD3+, CD4+, CD8+ T cells, CD20+ B cells, CD68+, CD163+ macrophages) in intratumoral and peritumoral areas.
- Application of a random forest machine-learning algorithm (Boruta) to identify predictors of p14/ARF expression.
Main Results:
- p14/ARF expression was evaluated in 68 MPM patients; 21% showed strong positivity.
- p14/ARF-positive epithelioid MPMs exhibited higher nuclear grade (G3) and increased PD-L1 expression (≥50%).
- Machine learning confirmed PD-L1 as a significant predictor of p14/ARF expression across all histotypes.
Conclusions:
- p14/ARF-positive epithelioid MPMs may represent a more aggressive phenotype associated with higher nuclear grade and PD-L1 expression.
- p14/ARF-negative MPMs might have an immune microenvironment less responsive to immune checkpoint inhibitors, indicated by low PD-L1 and CD4, and high CD163.
- The link between p14/ARF and PD-L1 suggests potential pathway interactions, warranting further investigation for patient selection in immunotherapy trials.
