Wyndham H. Wilson

PD-1 is an inhibitory receptor on T cells that, when bound by PD-L1 or PD-L2, suppresses anti-tumor immunity. Tumors can exploit this pathway through overexpression of PD-L1, often driven by 9p24.1 chromosomal alterations. Within aggressive lymphomas, Chromosome 9p24.1 alterations occur in approximately 60% of mediastinal gray zone lymphoma (MGZL) cases, 14% of non-germinal center B-cell (non-GCB) diffuse large B-cell lymphomas (DLBCL), and over 50% of primary central nervous system (CNS) lymphoma and primary testicular lymphoma (PTL) cases. Outcomes for relapsed or refractory disease remain dismal across these histologies. Given the frequent chromosome 9p24.1 alterations in these lymphomas, PD-1/PD-L1 inhibition represents a rational therapeutic strategy in these malignancies. We conducted a single-institution, open-label, phase 2 study of time-limited pembrolizumab monotherapy 200 mg every 21 days for up to 2 years (35 cycles) in patients with relapsed or refractory MGZL, primary CNS lymphoma, or DLBCL subtypes that share biologic features with primary CNS lymphoma, including PTL and non-GCB DLBCL with extranodal involvement. Eligible patients were aged ≥14 years with adequate organ function and no prior anti-PD-1/PD-L1 therapy. Disease assessment was performed via CT and/or MRI at baseline, after cycles 2, 6, 13, 17, 26, and 35. The primary endpoint was overall response rate (ORR); secondary endpoints included safety, progression free survival (PFS), and overall survival (OS). The study was terminated early due to slow accrual, and 11 patients were evaluable for response. Patients had a median of 2 prior lines of therapy and histologies included 5 primary CNS lymphoma, 2 MGZL, 2 non-GCB DLBCL without CNS involvement, and 2 non-GCB DLBCL with secondary CNS involvement. Responses were observed in two patients: 1 patient with PCNSL with a partial response and 1 patient with MGZL with a CR that was ongoing at 3 years. Overall, the 2-year PFS was 9.1% and the 2-year OS was 64%. No unexpected toxicities were observed. Single-agent pembrolizumab had limited activity in relapsed or refractory extranodal DLBCL including PCNSL. In PCNSL, this may party reflect the immunosuppressive tumor microenvironment, which can impair effective antitumor T-cell responses despite frequent activation of the PD-1/PD-L1 pathway. Clinical factors may also contribute, as even limited CNS progression can cause significant neurologic symptoms and create an urgent need for disease control, potentially narrowing the therapeutic window for agents with delayed response kinetics such as PD-1 inhibitors. However, a durable CR was observed in the one MGZL patient treated, supporting continued investigation of this treatment in select LBCL subtypes.

PURPOSE: Large B-cell lymphomas (LBCLs) are curable, but patients with residual disease after therapy invariably experience progression. Ultrasensitive methods to detect circulating tumor DNA (ctDNA) as minimal residual disease (MRD) may improve the determination of remission. METHODS: We integrated data from five prospective studies of frontline anthracycline-based chemotherapy in patients with LBCL. Tumor-specific phased variants were identified from pretreatment samples and monitored at landmark time points. Serial plasma specimens were blindly analyzed for detectable ctDNA as MRD. MRD status was compared with conventional response criteria for prognosis of progression-free survival (PFS). RESULTS: We studied ctDNA-MRD in 137 patients by monitoring 409 plasma specimens over time. Detectable ctDNA rates decreased during therapy with 55% and 78% of patients achieving undetectable ctDNA after two cycles and at the end of therapy, respectively. After a median follow-up of 37 months, the 2-year PFS for patients with detectable versus undetectable ctDNA after two cycles was 67% versus 96% ( P = .0025; hazard ratio [HR], 6.9) and after therapy was 29% versus 97% ( P < .0001; HR, 28.7), respectively. Ninety-two (94%) patients with undetectable ctDNA at the end of therapy remained alive without progression, while 19 (68%) patients with detectable ctDNA progressed or died. MRD status at the end of therapy had greater prognostic utility than conventional lymphoma response criteria using positron emission tomography (PET) scans (HR, 3.6 for positive PET and 28.3 for detectable ctDNA). CONCLUSION: Ultrasensitive ctDNA detection after frontline LBCL therapy is more prognostic than conventional radiographic response criteria. A refined definition of remission with ctDNA-MRD may improve clinical and psychological outcomes for patients with LBCL.

Supplemental Figure 1. Association between genetic alterations and methylation patterns Supplemental Figure 2. Epitypes characteristics and their relation to other lymphomas. Supplemental Figure 3. Comparison of epitype characteristics in relation to EBV status. Supplemental Figure 4. Validation of epitypes using Nanopore sequencing data. Supplemental Figure 5. Selected characteristics of BL epitypes and relative aSHM frequencies in EBV-positive BL. Supplemental Figure 6. GSEA plots for SignatureDB pathways determined to be significantly different between BL epitypes. Supplemental Figure 7. The Cox proportional hazards models accounting for both BL epitype and TP53 mutation status in adult BLs.

S1. DMRs EBVpos vs CB S2. DMRs EBVneg vs CB S3. Shared DMRs EBV S4. DMRs EBV status S5. Methylation gene corrs EB S6. GO pathways enrichment S7. LPS selected probes S8. Epitype assignment S9. DMRs eptiype S10. DMR gene corrs epitype S11. JASPAR TF results epitype S12. JASPAR TF results EBV S13. aSHM epitype S14. DEG epitpye S15. SignatureDB pathways enrichment

The genetic subtypes of Burkitt lymphoma have been defined, but the role of epigenetics remains to be comprehensively characterized. We searched genomic DNA from 218 patients across four continents for recurrent DNA methylation patterns and their associations with clinical and molecular features. We identified DNA methylation patterns that were not fully explained by the Epstein–Barr virus status or mutation status, leading to two epitypes described here as HypoBL and HyperBL. Each is characterized by distinct genomic and clinical features including global methylation, mutation burden, aberrant somatic hypermutation, and survival outcomes. Methylation, gene expression, and mutational differences between the epitypes support a model in which each arises from a distinct cell of origin. These results, pending validation in external cohorts, point to a refined risk assessment for patients with Burkitt lymphoma who may experience inferior outcomes. Significance: Burkitt lymphoma can be divided into two epigenetic subtypes (epitypes), each carrying distinct biological, transcriptomic, genomic, and clinical features. Epitype is more strongly associated with clinical and mutational features than the Epstein–Barr virus status or genetic subtype, highlighting an important additional layer of Burkitt lymphoma pathogenesis.

The identification of oncogenic mutations in diffuse large B-cell lymphoma (DLBCL) has led to the development of drugs that target essential survival pathways, but whether targeting multiple survival pathways may be curative in DLBCL is unknown. We performed a single-center, phase 1b-2 study of a regimen of venetoclax, ibrutinib, prednisone, obinutuzumab, and lenalidomide (ViPOR) in relapsed or refractory DLBCL. In phase 1b, which included patients with DLBCL and indolent lymphomas, four dose levels of venetoclax were evaluated to identify the recommended phase 2 dose, with fixed doses of the other four drugs. A phase 2 expansion in patients with germinal-center B-cell (GCB) and non-GCB DLBCL was performed. ViPOR was administered every 21 days for six cycles. In phase 1b of the study, involving 20 patients (10 with DLBCL), a single dose-limiting toxic effect of grade 3 intracranial hemorrhage occurred, a result that established venetoclax at a dose of 800 mg as the recommended phase 2 dose. Phase 2 included 40 patients with DLBCL. Toxic effects that were observed among all the patients included grade 3 or 4 neutropenia (in 24% of the cycles), thrombocytopenia (in 23%), anemia (in 7%), and febrile neutropenia (in 1%). Objective responses occurred in 54% of 48 evaluable patients with DLBCL, and complete responses occurred in 38%; complete responses were exclusively in patients with non-GCB DLBCL and high-grade B-cell lymphoma with rearrangements of MYC and BCL2 or BCL6 (or both). Circulating tumor DNA was undetectable in 33% of the patients at the end of ViPOR therapy. With a median follow-up of 40 months, 2-year progression-free survival and overall survival were 34% (95% confidence interval [CI], 21 to 47) and 36% (95% CI, 23 to 49), respectively. Treatment with ViPOR was associated with durable remissions in patients with specific molecular DLBCL subtypes and was associated with mainly reversible adverse events. (Funded by the Intramural Research Program of the National Cancer Institute and the National Center for Advancing Translational Sciences of the National Institutes of Health and others; ClinicalTrials.gov number, NCT03223610.).

We describe a previously unappreciated role for Bruton’s tyrosine kinase (BTK) in fungal immune surveillance against aspergillosis, an unforeseen complication of BTK inhibitors (BTKi) used for treating B cell lymphoid malignancies. We studied BTK-dependent fungal responses in neutrophils from diverse populations, including healthy donors, patients who were treated with BTKi, and X-linked agammaglobulinemia patients. Upon fungal exposure, BTK was activated in human neutrophils in a TLR2-, Dectin-1-, and FcγR-dependent manner, triggering the oxidative burst. BTK inhibition selectively impeded neutrophil-mediated damage to Aspergillus hyphae, primary granule release, and the fungus-induced oxidative burst by abrogating NADPH oxidase subunit p40 ^phox and GTPase RAC2 activation. Moreover, neutrophil-specific Btk deletion in mice enhanced aspergillosis susceptibility by impairing neutrophil function, not recruitment or lifespan. Conversely, GM-CSF partially mitigated these deficits by enhancing p47 ^phox activation. Our findings underline the crucial role of BTK signaling in neutrophils for antifungal immunity and provide a rationale for GM-CSF use to offset these deficits in patients who are susceptible.