Background Response to modern treatment strategies, which combine cytotoxic compounds with immune stimulatory agents and targeted treatment is highly variable among MCL patients. Thus, providing prognostic and predictive markers for risk adapted therapy is warranted and molecular information that can help in patient stratification is a necessity. In relapsed MCL, biopsies are rarely available and molecular information from tumor tissue is often lacking. Today, the main tool to access risk is the MCL international prognostic index (MIPI), which does not include detailed biological information of relevance for different treatment options. To enable continuous monitoring of patients, non-invasive companion diagnostic tools are needed which can further reduce cost and patient distress and enable efficient measurements of biological markers. Methods We have assessed if serum-based protein profiling can identify immune related proteins that stratify relapsed MCL patients based on risk. Overall, 371 scFv targeting 158 proteins were assessed using an antibody microarray platform. We profiled patients (n = 44) who had been treated within the MCL6-Philemon trial combining targeted and immune-modulatory treatment. Results The downstream processing led to the identification of the relapsed immune signature (RIS) consisting of 11 proteins with potential to stratify patients with long and short overall survival (OS). Moreover, in this population, MIPI alone failed to separate high, intermediate and low risk patients, but a combined index based on MIPI together with RIS, MIPIris, showed improved performance and significantly stratified all three risk groups based on OS. Conclusions Our results show that addition of biological parameters to previous prognostic indices improves patient stratification among patients treated with BTK inhibitor triplet combination, particularly, in the identification of an extreme high risk group.

The outcome for patients with mantle cell lymphoma (MCL) has drastically improved with new treatments directed towards the tumor immune microenvironment, where macrophages play an important role. In MCL, the presence of M2 macrophages defined by CD163 expression in diagnostic biopsies has been associated with a worse prognosis. An alternative way to assess the abundance of M2 macrophages is by measuring the level of soluble CD163 in serum (sCD163). We aimed to investigate the prognostic value of sCD163 in 131 MCL patients. We found that high sCD163 at diagnosis was associated with shorter progression-free survival (PFS) and shorter overall survival (OS) (p=0.002 and p<0.001) in 81 newly diagnosed chemoimmunotherapy-treated patients. The same was seen in a cohort of 50 relapsed MCL patients mainly treated within the phase II Philemon-trial with rituximab, ibrutinib and lenalidomide (PFS p=0.016 and OS p=0.035). In newly diagnosed patients with low levels of sCD163, five-year survival was 97%. There was a moderate correlation between sCD163 and tissue CD163 (r=0.64, p=0.014). The association with a poor prognosis was independent of MIPI, Ki67, p53 status and blastoid morphology, as assessed in a multivariable Cox proportional hazards model. Here, high sCD163 was associated with both shorter PFS (HR 3.48 95% CI: 1.42-8.54) and shorter OS (HR 4.33 95% CI: 1.32-14.2), showing that high levels of the M2 macrophage marker sCD163 is an independent negative prognostic factor in MCL, both in the chemoimmunotherapy and ibrutinib/lenalidomide era. In addition, low sCD163 levels identify MCL patients with a very good prognosis.

The use of many essential drugs is restricted due to their deleterious effects on the liver. Molecules that can prevent or protect the liver from drug-induced liver injury (DILI) would be invaluable in such situations. We used a transgenic line in zebrafish with a hepatocyte-specific expression of bacterial nitroreductase to cause temporally controlled liver damage. A whole organism-based chemical screen using the transgenic line identified BML-257, a potent small molecule AKT inhibitor, that protected the liver against metronidazole-induced liver injury. BML-257 also showed potent prophylactic and pro-regenerative activity in this liver damage model. BML-257 was tested in two independent toxicological models of liver injury caused by acetaminophen and isoniazid and was found to be protective against damage. This suggests that BML-257 has the potential to protect against multiple kinds of DILI.

Background: The possibility to monitor patient's serum proteome during treatment can provide deepened understanding of the biology associated with response to specific drugs. Non-invasive serum sampling provides an opportunity for sustainable repetitive sampling of patients, which allows for more frequent evaluation of the biological response and enhanced flexibility in treatment selection in contrast to tissue biopsies. Aim: To pin-point biologically relevant changes in pre-and on-treatment serum proteome samples in relapsed mantle cell lymphoma (MCL) patients, leading to insight into mechanisms behind response to treatment in sub-groups of patients. Methods: Pre-and on-treatment serum samples from relapsed MCL patients treated with a triple combination therapy of rituximab, ibrutinib and lenalidomide were available for the study, together with detailed clinicopathological information. A microarray technology targeting 158 serum proteins using 371 antibody-fragments was used to compare the serum proteome at the two time-points. Results: Proteins modulated by the treatment were shown to be associated to a MCL sub-group with ATM/TP53 alterations, which emphasizes the importance of treatment stratification. Absolute values of serum protein levels in on-treatment samples were highly variable and showed no correlation to outcome. To circumvent the challenge of variability in absolute serum protein levels, the velocity of change of individual serum proteins was used to identify proteins associated with clinical response. Increased values of TGF-β1, CD40 and complement component 4 comparing pre-and on-treatment samples were associated with remaining minimal residual disease (MRD) and increased BTK was associated with short progression-free survival (PFS). Conclusion: We show that the genetic sub-type of MCL affects the biological response to treatment in serum and that the change in defined serum proteins reveals the biology associated with clinical response.

Introduction: Mantle cell lymphoma (MCL) is an incurable disease and we have previously shown that presence of CD163+ cells in the tumor microenvironment in MCL is associated with an adverse outcome. We therefore profiled CD163+ cells to better understand their impact on this tumor and identify possible targetable markers. We hypothesize that, not only the presence of CD163+ cells, a marker of M2 macrophages, in the tumor tissue, but also the spatial context of CD163+ cells might be relevant for MCL development and treatment response. Material and Methods: Sixty-nine protein targets were measured by the Nanostring GeoMx Digital Spatial Profiler. This method allowed for preserved spatial context, in tissue microarrays from a population-based cohort of de novo MCL patients while investigating different markers. A total of 235 cores from 131 different patients were sampled. Regions of interest for CD20+, CD3+ and CD163+ cells were selected, with more than 600 areas of illumination (AOI) collected. From those, 85 AOI, belonging to 61 patients, were from CD163+ cells. These were subsequently classified according to their spatial profile: CD163+ cells within the tumor area and CD163+ cells excluded from the tumor area. Differentially expressed proteins between different tissue types were also explored. Results: Among the highest expressed proteins in the CD163 AOI were CD45, HLA-DR and CD68. The higher expression of both HLA-DR and CD68 suggests that macrophages in MCL microenvironment may display mixed phenotypes between pro- and anti-inflammatory stages, the called M1 and M2 polarization types. Interestingly, p53 and Ki67 were highly correlated among the CD163 AOI collected, which might suggest higher aggressiveness of CD163+ cells. We explored the differently expressed proteins between CD163 AOI sampled located within the tumor and excluded from the tumor/tumor barrier. We identified significant differences in the expressed proteins between the two groups, such as p53, VISTA, ARG1, LAG3 and HLA-DR. p53 was higher in the CD163 AOI within the tumor area, whereas VISTA was highest expressed in CD163 AOI excluded from the tumor cells/tumor barrier. Among the 85 CD163+ AOIs, 47 were sampled from lymph node biopsies, 14 from bone marrow, 10 from tonsil samples, four from the gastrointestinal tract, six from other tissue types and four from unknown origin. Principal component analysis and k-means clustering showed that most of bone marrow and gastrointestinal tract biopsies clustered apart from the remaining samples, with a few differentially expressed proteins, such as NF1 and CD66b. Conclusions: Our study demonstrates that spatial localization within the MCL tumor affects the expression of CD163+ macrophages, adding to the premise that macrophages in tumor are not easily characterized by the concept of M1 and M2 types. Our results shed light on targetable features of the MCL tumor microenvironment. Citation Format: Joana Matos Rodrigues, Lavanya Lokhande, Anna Gerdtsson, Anna Nikkarinen, Peter Hollander, Anna Porwit, Ingrid Glimelius, Mats Jerkeman, Sara Ek. Spatially resolved multiplexed analysisreveals how macrophages adapt to the mantle cell lymphoma microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2026.