Dose-dependent inhibition of Akt, ERK1/2 and EGFR.
ERα highNCI-H1975 cells were grown 24 h in fresh medium (Ctrl) or in medium containing 10 nM, 100 nM, 1 μM of the Akt inhibitor MK-2206 (Akti, panel a), 0.1 μM, 1 μM, 10 μM of the ERK1/2 inhibitor U-0126 (ERKi, panel b), 10 nM, 100 nM, 1 μM of the EGFR inhibitor AZD9291/Osimertinib (EGFRi, panel c). As read-out assays, Akt and ERK1/2 activity were measured by ELISA (technical duplicates), EGFR kinase activity was measured by a chemiluminescence-based assay (technical duplicates). Data are means + SD (n=3, biological replicates). *p<0.05, ***p<0.001: treated cells versus Ctrl cells (ANOVA).
ARTICLE ABSTRACT
The response to immune checkpoint inhibitors (ICI) often differs between genders in non–small cell lung cancer (NSCLC), but metanalyses results are controversial, and no clear mechanisms are defined. We aim at clarifying the molecular circuitries explaining the differential gender-related response to anti–PD-1/anti–PD-L1 agents in NSCLC.
We prospectively analyzed a cohort of patients with NSCLC treated with ICI as a first-line approach, and we identified the molecular mechanisms determining the differential efficacy of ICI in 29 NSCLC cell lines of both genders, recapitulating patients’ phenotype. We validated new immunotherapy strategies in mice bearing NSCLC patient-derived xenografts and human reconstituted immune system (“immune-PDXs”).
In patients, we found that estrogen receptor α (ERα) was a predictive factor of response to pembrolizumab, stronger than gender and PD-L1 levels, and was directly correlated with PD-L1 expression, particularly in female patients. ERα transcriptionally upregulated CD274/PD-L1 gene, more in females than in males. This axis was activated by 17-β-estradiol, autocrinely produced by intratumor aromatase, and by the EGFR-downstream effectors Akt and ERK1/2 that activated ERα. The efficacy of pembrolizumab in immune-PDXs was significantly improved by the aromatase inhibitor letrozole, which reduced PD-L1 and increased the percentage of antitumor CD8+T-lymphocytes, NK cells, and Vγ9Vδ2 T-lymphocytes, producing durable control and even tumor regression after continuous administration, with maximal benefit in 17-β-estradiol/ERα highfemale immune-xenografts.
Our work unveils that 17-β-estradiol/ERα status predicts the response to pembrolizumab in patients with NSCLC. Second, we propose aromatase inhibitors as new gender-tailored immune-adjuvants in NSCLC.
