CD38, BCL-2, PD-1, and PD-1L expression in nodal peripheral T-cell lymphoma: Possible biomarkers for novel targeted therapies?
Authors: Francesco Zaja, Valentina Tabanelli, Claudio Agostinelli, Angelica Calleri, Annalisa Chiappella, Marzia Varettoni, Pier Luigi Zinzani, Stefano Volpetti, Elena Sabattini, Renato Fanin, Stefano A. Pileri
Journal: American Journal of Hematology (2016)

Peripheral T-cell lymphomas (PTCL) account for 8–10% of lymphoid tumors and are very heterogeneous as far as clinical presentation, geographic distribution and histotypes. The 2016 revised edition of the WHO-classification of Tumors of the Haematopoietic and Lymphoid Tissues 1 has modified the body of nodal PTCLs, by introducing the new category of angioimmunoblastic (AITL) and other nodal T-cell lymphomas of follicular helper T-cell (TFH) phenotype. With the exception of ALK-positive (ALK+) anaplastic large cell lymphoma (ALCL), nodal PTCLs are generally very aggressive malignancies, with presentation in advanced stage, primary chemo-refractoriness, and 5-year survival of approximately 30%. CHOP ± etoposide and high-dose therapy plus stem cell consolidation in younger patients, represent the standard of care for the majority of patients but, unfortunately, nearly 50% of them relapse or progress during the first year from treatment start and the life expectancy after treatment failure is generally less than 1 year. For patients who fail first line therapy, there is no standard of care. During the last few years, many efforts have been made to find new agents and therapeutic approaches for nodal PTCLs. Unfortunately, with the exception of Brentuximab-Vedotin that showed remarkable activity in ALCLs, the results achieved with other agents such as Romidepsin, Belinostat, and Pralatrexate are less impressive with median progression-free survival of only few months. Quite recently, new targeted agents against CD38, BCL-2, and PD-1/PD-1L have shown impressive activity respectively for the treatment of multiple myeloma, chronic lymphocytic leukemia, and Hodgkin lymphoma, and are now under investigation as single agents or in combination in other hematological malignancies. The expression of these target-molecules has not been extensively investigated in PTCLs, limiting the possible use of targeted therapies also in these lymphoid tumors. The aim of this study was to evaluate the expression of CD38, BCL-2, and PD-1/PD-1L in different nodal PTCL subtypes to establish the rationale for possible future clinical investigations. Formalin-fixed paraffin-embedded tissue blocks of 73 nodal PTCLs/NOS, 48 ALK− ALCLs, 25 ALK+ ALCLs, and 27 AITLs were used for tissue microarrays construction. Tumor samples were collected at disease presentation and diagnosed according to the 2008 WHO criteria. More methodological information can be founded in the Supporting Information. Immunohistochemical results were evaluated by four experienced pathologists (C.A., S.A.P., E.S., V.T.). Cases with an insufficient amount of tumor tissue were considered as "not evaluable." Among the remaining cases, the percentage of positive tumor cells was scored as follows: 4: >75%; 3: 50–75%; 2: 25–49%; 1: 5–24%; 0: <5%, according to Bossard et al. 2. The results of the four biomarkers analyzed on immunohistochemistry are detailed in Table 1. CD38 resulted particularly expressed in AITLs (80%), with variable scores. In particular, 7 of the 20 positive cases (35%) were scored 3 or 4. As in AITLs there is usually a high amount of plasma cells, CD38 positivity of T neoplastic cells was ascertained by concomitant immunostaining with other T-cell markers and anti-CD138 (see above). Fifty-seven percent of PTCLs-NOS were CD38 positive, again with variable numbers of stained lymphomatous elements and a 3–4 score in 50% of them. Conversely, CD38 positivity was rarely recorded in ALCLs (6/36 and 0/10 ALK− and ALK+ cases, respectively). BCL-2 positivity was detected in 88% of AITLs (88%) and 79.5% of PTCLs-NOS, with a 3–4 score in most of them (Table 1). BCL-2 was expressed by 58% ALK− and a minority of ALK+ ALCLs (31%), once again with a prevalent 3–4 score. PD-1 expression was recorded in 70 and 61% of AITLs and PTCLs-NOS respectively with a 3–4 score in nearly half cases. On the contrary, PD-1 positivity was very rarely detected in ALCLs (only in 2 ALK+ cases). Interestingly, the eight AITLs negative for PD-1 expressed at least two other TFH-associated markers, while all the nine PTCLs-NOS scored 4 showed TFH phenotype because of the simultaneous expression of at least another TFH-associated antigen. The latter cases are moved to the category of AITLs and other nodal TCLs of TFH phenotype in the revised WHO Classification. PD-1L was expressed in 72 and 46% of ALK+ and 46% ALK− ALCLs (with a 3–4 score in 11/16 and 5/8 respectively). On the contrary none of neoplastic cells showed PD-1L expression among AITLs and PTCLs-NOS. The results of this analysis are informative regarding the expression of some biomarkers that represent possible therapeutic targets of new biologic agents in the setting of PTCLs. For the first time, we documented that the majority of AITLs and PTCLs-NOS express the CD38 molecule although with variable percentages of positive cells. Several previous studies already indicated possible BCL-2 expression by AITL, PTCL-NOS, and ALCL 3, 4. Our analysis confirms BCL-2 expression in these neoplasms, with higher positivity scores in PTCL-NOS, AITL, and ALK− ALCL. PD-1 has been recently identified as one of the biomarkers associated with the diagnosis of AITL. Our analysis showed similar rates of PD-1 expression in AITLs and PTCLs-NOS (nearly 70 and 60% of cases, respectively) and no expression in ALCLs. These results are in line with previous studies with the exception of PTCLs-NOS, where we had a higher prevalence of positivity (61% vs. 0 to 28%) 5. This finding deserves some comments. First, most series so far tested for PD-1 focused on AITL. Second, nine of our cases classified as PTCL-NOS according to the 2008 edition of the WHO Classification, all scored 4, are moved to the new category of AITL and other nodal T-cell lymphomas of TFH phenotype in the 2016 revision of the classification. Finally, PD-1 positivity does not always imply TFH derivation, as the antigen can also be expressed in other settings because of cell plasticity. On the contrary and similarly to what previously reported in smaller series 6, PD-1L was expressed in 46 and 72% of ALK− and ALK+ ALCLs, respectively, while it was absent in AITLs and PTCLs-NOS. The results of our analysis may represent the background for new clinical trials with Daratumumab and Venetoclax in CD38 and BCL-2 positive AITLs and PTCLs-NOS, respectively. We are aware of the complexity of pathways modulating the expression of checkpoint inhibitors, which may impact on the response to PD-1 and PD-1L inhibitors. With this in mind, the results of our analysis suggest that the frequent expression of PD-1L by ALCL neoplastic cells may herald sensitivity of these tumors to checkpoint inhibitors; on the contrary, no efficacy should be expected in the setting of PTCL-NOS and AITL, which only express PD-1. FZ, VT, SAP: designed the study. FZ and SAP: wrote the manuscript. RF, AC, MV: contributed to manuscript writing and critical review. AC: performed immunostains. VT, CA, ES, PLZ, SAP: rendered the original diagnoses and evaluated marker results. All the authors: contributed to the collection of cases and approved the manuscript. Francesco Zaja1*, Valentina Tabanelli2, Claudio Agostinelli3,4, Angelica Calleri2, Annalisa Chiappella5, Marzia Varettoni6, Pier Luigi Zinzani7, Stefano Volpetti1, Elena Sabattini3, Renato Fanin1 Stefano A. Pileri2,4 1Clinica Ematologica, DISM, Azienda Sanitaria Universitaria Integrata, Udine, Italy; 2Unit of Haematopathology, European Institute of Oncology, Milan, Italyi; 3Unità di Emolinfopatologia, Dipartimento di Ematologia & Oncologia Azienda Ospedaliero – Universitaria, Bologna, Italy; 4Department of Experimental, Diagnostic and Specialty Medicine, Bologna University School of Medicine, Bologna, Italy; 5S.C. Ematologia A.O.U. Città della Salute e della Scienza, Torino, Italy; 6Dipartimento di Ematologia e Oncologia, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy; 7Institute of Hematology "L. e A. Seràgnoli", University of Bologna, Bologna, Italy Additional Supporting Information may be found in the online version of this article. Contract grant sponsor: AIRC 5x1000 n.10007 (to S.A.P). Additional Supporting Information may be found in the online version of this article. Supporting Information Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.