Prognostic significance and possible therapeutic implications of tumor-infiltrating lymphocytes in de novo canine diffuse large b-cell lymphoma

In human medicine, there is a growing body of evidence that tumor cells are able to escape T-cell mediated anti-tumor immunity [1]. Therefore, immunotherapy represents the new frontier of cancer treatment [2] and clinical trials with customized cancer vaccines are ongoing [3]. Dogs with diffuse large B-cell lymphoma (DLBCL) benefit from the inclusion of immunotherapy in the treatment regimen [4]. However, the immunity status of dogs affected by DLBCL has been scarcely characterized. The aim of this study was to describe the composition of the intra-tumoral non-neoplastic lymphoid population in lymph node aspirates of dogs with DLBCL, and to assess the possible prognostic role of different immune patterns. Twenty-three cases with histopathological diagnosis of DLBCL were retrospectively extracted from the flow cytometric (FC) database of the Department of Veterinary Medicine (University of Milan). All cases were obtained from a single oncological referral center (Centro Oncologico Veterinario), underwent a standardized complete staging workup and received chemo-immunotherapy. The percentage of CD21+, CD5+, CD4+ and CD8+ cells out of small non-neoplastic lymphocytes was extracted from FC data. Hierarchical cluster analysis separated two groups: group 1 (12 dogs) had a lower percentage of small cells and most of them were CD21+; group 2 (11 dogs) had significantly higher percentage of small cells and most of them were CD5+, either CD4+ or CD8+. CD5/CD21 ratio accurately discriminated between the two groups, with a cutoff value of 1.0 having 100% sensitivity and specificity. Breed, sex, age, anemia, thrombocytopenia, LDH levels, disease stage and achievement of complete remission (CR) were equally represented among groups, whereas all symptomatic dogs (n=5) were in group 1. The achievement of CR was the only variable significantly influencing lymphoma-specific survival (LSS). Still, dogs in group 1 had a shorter LSS compared to dogs in group 2 (median 148 days and 623 days, respectively) (p=0.187). Based on our results, about a half of dogs with DLBCL showed a poor number of T-cell infiltrating the tumor. This was associated with uneffective response to immunotherapy, as median LSS was similar to that obtained in dogs treated with chemotherapy and placebo in a previous clinical trial [4]. Although preliminar, our data suggest that a higher component of intraneoplastic T-cells predict a good response to immunotherapy. Further studies are needed to assess the activation status of T-cells in these dogs and the strategies used by the neoplastic cells to escape immune surveillance. A better characterization of these population in DLBCL before treatment will help to stratify dogs and drive therapy, eventually. [1] Curran EK et al. Mechanisms of immune tolerance in leukemia and lymphoma, Trends in immunology, 38(7):513-525, 2017. [2] Ribas A, Wolchok JD. Cancer immunotherapy using checkpoint blockade, Science, 359(6382):1350-1355, 2018. [3] Sahin U, Tureci O. Personalized vaccines for cancer immunotherapy, Science, 359(6382):1355-1360, 2018. [4] Marconato L et al. Randomized, placebo-controlled, double-blinded chemoimmunotherapy clinical trial in a pet dog model of diffuse large B-cell lymphoma, Clinical Cancer Research, 20(3):668-677, 2014. [5] Barber LG, Weishaar KM. Criteria for designation of clinical substage in canine lymphoma: a survey of veterinary oncologists, Veterinary and Comparative Oncology, 14 Suppl 1:32-39, 2016.