Spatial Mapping and Immunomodulatory Role of the OX40/OX40L Pathway in Human Non-Small Cell Lung Cancer

Purpose: To evaluate the tissue distribution and clinical significance of OX40 and OX40L in human non-small cell lung cancer (NSCLC).

Experimental design: Using multiplexed quantitative immunofluorescence, we conducted simultaneous and localized measurements of OX40 and OX40L proteins, major T-cell subsets, and conventional type 1 dendritic cells (cDC1) in 614 primary NSCLCs from three independent cohorts represented in tissue microarrays. We also measured OX40L protein in samples from a phase I clinical trial of intratumor administration of a lipid nanoparticle encapsulated mRNA encoding OX40L (mRNA-2416) in human solid tumors. Finally, we studied the OX40 pathway in 212 uterine/ovarian serous carcinomas.

Results: OX40 protein was expressed in approximately 90% of NSCLCs, and OX40L was detected in approximately 10% of cases. Increased expression of OX40 was associated with higher CD4⁺ and CD8⁺ T lymphocytes, as well as cDC1s. Elevated expression of OX40L was consistently associated with increased CD4⁺ tumor-infiltrating lymphocytes and longer overall survival. No association was found between OX40 or OX40L levels and oncogenic driver mutations in EGFR and KRAS in lung adenocarcinomas. Delivering OX40L mRNA using intratumor mRNA-2416 injection mediated increased local OX40L protein levels that was most prominent in a patient with ovarian serous carcinoma. Detectable OX40L protein levels were observed in 15% of primary uterine/ovarian serous malignancies and associated with longer survival.

Conclusions: The OX40 pathway is expressed in a fraction of NSCLCs and is associated with a favorable immune contexture. Although OX40L is uncommonly expressed in NSCLC and serous malignancies, it is associated with better prognosis and can be introduced using exogenous mRNA.