Immunotherapy – Biomechanisms
A. Cuccaro1, I. Zangrilli1, F. Corrente1, E. Cupelli1, F. Fatone1, E. Galli1, E. Maiolo1, S. Annunziata2, V. Rufini3, M. Balducci4, F. D‘Alò1, S. Bellesi1, V. De Stefano5, S. Hohaus5
1 Istituto di Ematologia, Fondazione Policlinico Universitario A. Gemelli IRCSS, 2 Istituto di Medicina Nucleare, Fondazione Policlinico Universitario A. Gemelli IRCSS, 3 Istituto di Medicina Nucleare, Fondazione Policlinico A. Gemelli IRCSS- Università Cattolica Sacro Cuore, 4 Divisione di Radioterapia, Fondazione Policlinico Universitario A. Gemelli IRCSS, 5 Istituto di Ematologia, Fondazione Policlinico A. Gemelli IRCSS- Università Cattolica Sacro Cuore
The neutrophil-lymphocyte ratio (N/L ratio) has been reported as predictor of progression-free survival (PFS) in cancer patients including classical Hodgkin lymphoma (cHL). We reasoned that the immune checkpoint axis between the Programmed cell death protein 1 (PD-1) and its ligand PD-L1 could be involved in the neutrophil-lymphocyte interaction in cHL. We evaluated N/L ratio and PD-L1/PD-1 axis in the peripheral blood at diagnosis of HL using flow cytometry and RT-PCR and analyze for associations with clinical characteristics and outcome. We analyzed 408 patients diagnosed with cHL between 1999 and 2017 for the prognostic impact of N/L ratio. We studied expression of PD-L1 and PD-1 using RT-PCR in buffy-coats of 82 patients, and in cell fractions from peripheral blood of normal donors (n = 4) and patients (n = 24) separated using magnetic beads. PD1 (CD279, BD Biosciences) and PD-L1 (CD274, Beckman Coulter) was also prospectively analyzed by flow cytometry in a group of 14 patients and 5 normal volunteers. PFS was evaluated with logrank and Cox regression models. We first confirmed the prognostic significance of the N/L ratio in our case series of 408 patients. A high N/L ratio (≥6) was associated with poor PFS (p = 0.003) in univariate analysis and retained its significance in a multivariate analysis including interim PET, as independent prognosticator (HR 2.4; 95% C.I. 1.3–4.2, p = 0.004). We then analyzed PD-L1 and PD-1 expression using flow cytometry and RT-PCR. Both methodologies showed that PD-L1 expression was highest in neutrophils and PD-1 expression was highest in the T cell fraction. We found a correlation between expression of PD-L1 on neutrophils and PD-1 on T cells (r = 0.6 p = 0.008). Both methodologies also showed a significantly increased expression of PD-L1 on neutrophils from patients when compared to controls, with particular high PD-L1 expression on neutrophils in patients with Stage IV. PD-L1 expression on neutrophils did not correlate with neutrophil count but with the N/L ratio (p = 0.004). PD-L1 expression on neutrophils was associated with a poor PFS (p < 0.001), also in a multivariate analysis including N/L ratio (p = 0.03). Our study identifies an additional checkpoint axis of prognostic importance that could be a target during therapy with monoclonal antibodies interferering with the axis.