Lučka Boltežar

Respiratory microbiome alterations, coinfections, and virus intrahost evolution are of great interest in persistently viable SARS-CoV-2 infections in the context of antiviral treatment and immune response. However, samples before, during, and after infection are seldom available to researchers. Therefore, there has been a significant lack of opportunities to comprehensively study microbiota homeostasis, coinfections, and virus intra-host evolution on the consensus and minor variants scale in response to antiviral treatments and patient immune response. A 63-year-old female patient with diffuse large B-cell lymphoma received multiple treatments for SARS-CoV-2 that remained active 169 days. Together, 32 respiratory and 19 serum samples were collected before, during, and after (− 398 to 233 days) COVID-19. Subsets were selected for virus viability testing by culture (20) and subgenomic (sg) RNA (20) measurement, intra-host evolution assessment (18), microbiome composition analysis (28), and coinfection identification (11). IgA/IgG and neutralizing anti-SARS-CoV-2 antibodies were measured 19 times throughout the infection. SARSCoV-2 lineage XBB.1.16.11 persisted and remained viable until 116 days post infection (PI) regardless of treatments. No sgRNA marker tested was suitable for virus viability prediction. IgG/IgA antibodies first appeared after 38 days, but the virus persisted regardless of multiple plasma treatments before neutralizing antibodies appeared (100 days PI) and finally cleared the virus 116 days PI. Consensus-level mutations fluctuated around 102.7 ± 4.0, and minor variants increased from six to 61 with a mutation rate of 4.9 × 10−3 per site per year, with the highest average number of mutations per gene length in S and E (0.013) with surges after every antiviral treatment. The transversion/transition ratio increased from 0.50 (day 0) to 0.57 (day 24) with a steady decrease to 0.48 (day 147). Mutational signature analysis showed dominance of C > T substitutions consistent with APOBEC antiviral enzyme activity. Upper respiratory microbiota showed three distinct profiles with varying α-/β-diversity and an association of Staphylococcus spp. with COVID-19. These findings further elucidate the dynamics of intra-host viral evolution and complexities of virus clearance in individuals with hematological malignancies and highlight the impact of antiviral treatments on the potential of virus variants emergence in longitudinally infectious patients due to delayed immune response.

Background/Objectives: The tumour microenvironment (TME) plays an important role in the development and progression of cancer and it differs among lymphomas, both with respect to the composition and quantity of specific tumour-infiltrating immune cells (TICs), such as FoxP3⁺ regulatory T cells (Tregs). The role of FoxP3⁺ Tregs in the TME of peripheral T-cell lymphomas (PTCLs) is complex, and their impact on overall survival (OS) remains unclear. Consequently, we aim to evaluate and compare the FoxP3⁺ cell quantity in nodal PTCLs and reactive lymph nodes (LNs), with a focus on investigating their impact on OS. Methods: excisional lymph node (LN) biopsies from 105 nodal PTCLs and 17 reactive LNs are immunohistochemically stained for FoxP3. Visual scoring of FoxP3⁺ cells is performed, and different cut-off values are used to evaluate the impact of FoxP3⁺ cell quantity on OS. Results: FoxP3⁺ cells are present in the TME of all cases, except for four cases where FoxP3⁺ is expressed in lymphoma cells. Lower FoxP3⁺ cell quantities are observed in certain nodal PTCL subtypes compared to reactive LNs. Patients with high FoxP3⁺ cell quantities show improved OS. However, the FoxP3⁺ cell quantity is not confirmed as an independent prognostic biomarker. Conclusions: these findings underscore the promise of FoxP3⁺ cell quantities as added value in prognosis and highlight the potential benefits of Treg-stimulating therapies in PTCLs.

High-grade B-cell lymphomas with MYC and BCL2 and/or BCL6 rearrangements are known for their aggressive clinical course and so are the ones with MYC and BCL2 protein overexpression. The optimal therapy for these lymphomas remains to be elucidated. A retrospective analysis of all diffuse large B-cell lymphomas and high-grade B-cell lymphomas with MYC and BCL2 and/or BCL6 rearrangements diagnosed between 2017 and 2021 at the Institute of Oncology Ljubljana, Slovenia, has been performed. Only patients with double-expressor lymphoma (DEL), double-hit lymphoma (DHL), or triple-hit lymphoma (THL) were included. Demographic and clinical parameters were assessed, as well as progression-free survival (PFS) and overall survival (OS). In total, 161 cases out of 309 (161/309; 52,1%) were classified as DEL. Sixteen patients had DHL, MYC/BCL2 rearrangement was observed in eleven patients, and MYC/BCL6 rearrangement was observed in five patients. Five patients were diagnosed with THL. Out of 154 patients (according to inclusion/exclusion criteria) included in further evaluation, one-hundred and thirty-five patients had double-expressor lymphoma (DEL), sixteen patients had DHL, and three patients had THL. In total, 169 patients were treated with R-CHOP, 10 with R-CHOP and intermediate-dose methotrexate, 19 with R-DA-EPOCH, and 16 with other regimens. The median follow-up was 22 months. The 5-year OS for the whole DEL group was 57.1% (95% CI 45.9–68.3%) and the 5-year PFS was 76.5% (95% CI 72.6–80.4%). The log-rank test disclosed no differences in survival between treatment groups (p = 0.712) while the high-risk international prognostic index (IPI) carried a significantly higher risk of death (HR 7.68, 95% CI 2.32–25.49, p = 0.001). The 5-year OS for DHL patients was 32.4% (95% CI 16.6–48.2%) while all three TH patients were deceased or lost to follow-up. Our analyses of real-life data disclose that the R-CHOP protocol with CNS prophylaxis is a successful and curative treatment for a substantial proportion of DEL patients.

Background: t(14;18)(q32;q21) translocation is an important genetic feature of follicular lymphoma resulting in antiapoptotic B-cell lymphoma 2 (BCL2) protein overexpression. On chromosome 18 breakpoint-site variation is high but does not affect BCL2. Breakpoint most commonly occurs at major breakpoint region (MBR) but may happen at minor cluster region (mcr) and between MBR and mcr at 3′MBR and 5′mcr. The aim of this study was to analyze the correlation of t(14;18)(q32;q21) breakpoint site with clinical characteristics in follicular lymphoma. Patients and methods: We included patients diagnosed with follicular lymphoma who received at least 1 cycle of systemic treatment and had the t(14;18)(q32;q21) translocation detected by polymerase chain reaction (PCR) at MBR, mcr or 3′MBR prior to first treatment. Among patients with different breakpoints, sex, age, disease grade, stage, B-symptoms, follicular lymphoma international prognostic index (FLIPI), presence of bulky disease, progression free survival and overall survival were compared. Results: Of 84 patients, 63 had breakpoint at MBR, 17 at mcr and 4 at 3′MBR. At diagnosis, the MBR group had a significantly lower disease stage than the mcr group. Although not significant, in the MBR group we found a higher progression-free survival (PFS) and overall survival (OS), lower grade, age, FLIPI, and less B-symptoms. Conclusions: Compared to patients with mcr breakpoint, those with MBR breakpoint seem to be characterised by more favourable clinical characteristics. However, a larger study would be required to support our observation.

Not many treatment options exist for patients with relapsed or refractory (R/R) B-cell non-Hodgkin lymphoma (NHL) in whom first- and second-line therapies were unsuccessful. This is especially true for patients with aggressive lymphomas. The innovative agent pixantrone has shown some promising results in terms of disease-free and overall survival, both in monotherapy as well as in combinations. However, recent trials (Phase III and real-world studies) reported unsatisfactory results, thereby raising the question about the role of pixantrone in the current treatment of R/R aggressive lymphomas. Nonetheless, there might still be a potential position for this drug in combinations, for use as first-line treatment of patients with cardiac dysfunction. This article summarizes the definition, structure, mechanism of action and reduced cardiotoxicity of pixantrone as well as efficacy and toxicity both in monotherapy and in combinations, as treatment for aggressive and indolent non-Hodgkin lymphomas.