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
1Istituto di Ematologia, Fondazione Policlinico Universitario A. Gemelli IRCSS,2Istituto di Medicina Nucleare, Fondazione Policlinico Universitario A. Gemelli IRCSS,3Istituto di Medicina Nucleare, Fondazione Policlinico A. Gemelli IRCSS- Università Cattolica Sacro Cuore,4Divisione di Radioterapia, Fondazione Policlinico Universitario A. Gemelli IRCSS,5Istituto 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.