Comparative analysis of post-transplant lymphoproliferative disorders after solid organ and hematopoietic stem cell transplantation reveals differences in the tumor microenvironment

Virchows Archiv https://doi.org/10.1007/s00428-020-02985-4 ORIGINAL ARTICLE Comparative analysis of post-transplant lymphoproliferative disorders after solid organ and hematopoietic stem cell transplantation reveals differences in the tumor microenvironment Mathis Overkamp 1 & Massimo Granai 1,2 & Irina Bonzheim 1 & Julia Steinhilber 1 & Jens Schittenhelm 1 & Wolfgang Bethge 3 & Leticia Quintanilla-Martinez 1 & Falko Fend 1 & Birgit Federmann 1 Received: 19 October 2020 / Revised: 19 October 2020 / Accepted: 1 December 2020 # The Author(s) 2020 Abstract Post-transplant lymphoproliferative disorders (PTLD) occur after solid organ transplantation (SOT) or hematopoietic stem cell transplantation (HCT) and are frequently associated with Epstein-Barr virus (EBV). Because of the complex immune setup in PTLD patients, the tumor microenvironment (TME) is of particular interest to understand PTLD pathogenesis and elucidate predictive factors and possible treatment options. We present a comparative study of clinicopathological features of 48 PTLD after HCT (n = 26) or SOT (n = 22), including non-destructive (n = 6), polymorphic (n = 23), and monomorphic (n = 18) PTLD and classic Hodgkin lymphoma (n = 1). EBV was positive in 35 cases (73%). A detailed examination of the TME with image analysis-based quantification in 22 cases revealed an inflammatory TME despite underlying immunosuppression and significant differences in its density and composition depending on type of transplant, PTLD subtypes, and EBV status. Tumor-associated macrophages (TAMs) expressing CD163 (p = 0.0022) and Mannose (p = 0.0016) were enriched in PTLD after HCT. Double stains also showed differences in macrophage polarization, with more frequent M1 polarization after HCT (p = 0.0321). Higher counts for TAMs (CD163 (p = 0.0008) and cMaf (p = 0.0035)) as well as in the T cell compartment (Granzyme B (p = 0.0028), CD8 (p = 0.01), and for PD-L1 (p = 0.0305)) were observed depending on EBV status. In conclusion, despite the presence of immunosuppression, PTLD predominantly contains an inflammatory TME characterized by mostly M1-polarized macrophages and cytotoxic T cells. Status post HCT, EBV positivity, and polymorphic subtype are associated with an actively inflamed TME, indicating a specific response of the immune system. Further studies need to elucidate prognostic significance and potential therapeutic implications of the TME in PTLD. Keywords Post-transplant lymphoproliferative disease . Solid organ transplantation . Hematopoietic stem cell transplantation . Microenvironment . Macrophages Introduction Post-transplant lymphoproliferative disorders (PTLD) are a heterogeneous group of lymphoid or plasmacytic * Birgit Federmann 1 Institute of Pathology and Neuropathology, University Hospital and Comprehensive Cancer Center Tuebingen, Liebermeisterstraße 8, 72076 Tuebingen, Germany 2 Section of Pathology, Department of Medical Biotechnology, University of Siena, Siena, Italy 3 Department of Internal Medicine Hematology and Oncology, Comprehensive Cancer Center and University Hospital Tuebingen, Tuebingen, Germany proliferations. They develop in patients under immunosuppression after solid organ transplantation (SOT), or less frequently after allogeneic hematopoietic stem cell transplantation (HCT). PTLDs form a spectrum of usually Epstein-Barr virus (EBV) driven polyclonal proliferations to EBV-positive or EBV-negative clonal malignancies resembling lymphomas occurring in immunocompetent patients. According to the current WHO classification, there are four categories of PTLD [1]: Nondestructive PTLDs show preserved architecture and are usually EBV-positive. Polymorphic PTLDs show significant architectural effacement, are usually EBV positive, and comprise the full range of cellular maturation without fulfilling the criteria for malignant lymphoma. At the end of the spectrum are monomorphic PTLDs which fulfill the criteria for the respective B cell or T/NK-cell Virchows Arch lymphomas in immunocompetent patients, and classic Hodgkin lymphoma (CHL). They can be EBV-positive or EBV-negative [1]. PTLD is one of the most serious complications of transplantation with a reported incidence between about 2 and 20% depending on the kind of transplantation and a 3-year survival of about 40 to 55% [2–4]. While the etiology of PTLD is not yet fully understood, the majority of cases, especially early after transplantation, are associated with EBV infection or reactivation, which induces an uncontrolled lymphocyte proliferation [2]. Regarding the etiology of EBV-negative PTLD, hit-and-run EBV infection, the effects of persistent antigen stimulation by the graft, long-term immunosuppression, as well as other infectious agents have been suggested as possible pathogenic mechanisms [2, 5]. Due to advanced conditioning protocols and graft modification, the incidence of EBV-positive PTLD has decreased in recent times resulting in a relative increase of EBV-negative cases [3, 6]. EBV-negative PTLD usually arises late after transplantation and differs in clinicopathological features as well as gene expression profiles from EBV-positive PTLD [3, 5, 7, 8]. This suggests that EBV-negative PTLD might represent a different entity [6, 9] or sporadic lymphoma occurring coincidentally [8]. Adding to its complexity, PTLD can be of donor or host origin. Whereas the vast majority of examined cases of PTLD after HCT is of donor origin [10], PTLD after SOT is usually of host origin [11, 12]. PTLD after HCT is considered to be more aggressive and usually occurs earlier after transplantation [9, 10]. This complex immunologic situation, influenced by the presence of oncogenic EBV, chronic immune stimulation through chronic antigen presentation by the graft, chronic immunosuppression, and interaction of donor-derived immune cells with the host immune cells, makes the tumor microenvironment (TME) of PTLD and interesting focus of research [13], but published data on the TME of PTLD are sparse. The TME represents the specific setting in which a tumor resides and consists of all non-malignant constituents of a neoplasm containing variable numbers of immune cells, mesenchymal cells, blood vessels, and non-cellular components such as extracellular matrix [14]. The composition of the TME has a profound impact on the biological behavior, prognosis, and therapy response in many tumor types including lymphoma, since tumor cells retain a range of dependence on interactions with the non-malignant cells of the TME [13–16]. T cell subsets and tumor-associated macrophages (TAMs) are considered the major immunologically relevant cell types of the TME. TAMs constitute a significant part of the tumor infiltrating microenvironment [14, 17, 18]. They are usually detected using CD163 or CD68 antibodies [19] and further classified corresponding to their functional state as antitumoral M1- and pro-tumoral M2-phenotypes in a simplified view [20–22]. In PTLD, (...truncated)