BTK drives neutrophil activation for sterilizing antifungal immunity.

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, BTKi-treated patients, 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 p40phox 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 p47phox 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 susceptible patients.

Primary Large B-cell Lymphomas of Immune-Privileged Sites.

Diffuse large B-cell lymphoma (DLBCL) encompasses a diverse spectrum of aggressive B-cell lymphomas with remarkable genetic heterogeneity and myriad clinical presentations. Multiplatform genomic analyses of DLBCL have identified oncogenic drivers within genetic subtypes that allow for pathologic subclassification of tumors into discrete entities with shared immunophenotypic, genetic, and clinical features. Robust classification of lymphoid tumors establishes a foundation for precision medicine and enables the identification of novel therapeutic vulnerabilities within biologically homogeneous entities. Most cases of DLBCL involving the central nervous system (CNS), vitreous, and testis exhibit immunophenotypic features suggesting an activated B-cell (ABC) origin. Shared molecular features include frequent co-mutations of MYD88 (L265P) and CD79B and frequent genetic alterations promoting immune evasion, which are hallmarks of the MCD/C5/MYD88 genetic subtype of DLBCL. Clinically, these lymphomas primarily arise within anatomic sanctuary sites and have a predilection for remaining confined to extranodal sites and strong CNS tropism. Given the shared clinical and molecular features, the umbrella term primary large B-cell lymphoma of immune-privileged sites (IP-LBCL) was proposed. Other extranodal DLBCL involving breast, adrenal glands, and skin are often ABC DLBCL, but are more heterogeneous in their genomic profile and involve anatomic sites that are not considered immune privileged. In this review, we describe the overlapping clinical, pathologic, and molecular features of IP-LBCL and highlight important considerations for diagnosis, staging and treatment. We also discuss potential therapeutic vulnerabilities of IP-LBCL including sensitivity to inhibitors of Bruton's tyrosine kinase, immunomodulatory agents, and immunotherapy.

Multi-omic profiling of follicular lymphoma reveals changes in tissue architecture and enhanced stromal remodeling in high-risk patients.

Follicular lymphoma (FL) is a generally incurable malignancy that evolves from developmentally blocked germinal center (GC) B cells. To promote survival and immune escape, tumor B cells undergo significant genetic changes and extensively remodel the lymphoid microenvironment. Dynamic interactions between tumor B cells and the tumor microenvironment (TME) are hypothesized to contribute to the broad spectrum of clinical behaviors observed among FL patients. Despite the urgent need, existing clinical tools do not reliably predict disease behavior. Using a multi-modal strategy, we examined cell-intrinsic and -extrinsic factors governing progression and therapeutic outcomes in FL patients enrolled onto a prospective clinical trial. By leveraging the strengths of each platform, we identify several tumor-specific features and microenvironmental patterns enriched in individuals who experience early relapse, the most high-risk FL patients. These features include stromal desmoplasia and changes to the follicular growth pattern present 20 months before first progression and first relapse.

The follicular lymphoma tumor microenvironment at single-cell and spatial resolution.

ABSTRACT: Follicular lymphoma (FL) is a generally incurable malignancy that originates from developmentally blocked germinal center B cells residing, primarily, within lymph nodes (LNs). During the long natural history of FL, malignant B cells often disseminate to multiple LNs and can affect virtually any organ. Nonmalignant LNs are highly organized structures distributed throughout the body, in which they perform functions critical for host defense. In FL, the malignant B cells "re-educate" the lymphoid environment by altering the phenotype, distribution, and abundance of other cells such as T cells, macrophages, and subsets of stromal cells. Consequently, dramatic anatomical changes occur and include alterations in the number, shape, and size of neoplastic follicles with an accompanying attenuation of the T-cell zone. Ongoing and dynamic interactions between FL B cells and the tumor microenvironment (TME) result in significant clinical heterogeneity observed both within and across patients. Over time, FL evolves into pathological variants associated with distinct outcomes, ranging from an indolent disease to more aggressive clinical courses with early death. Given the importance of both cell-intrinsic and -extrinsic factors in shaping disease progression and patient survival, comprehensive examination of FL tumors is critical. Here, we describe the cellular composition and architecture of normal and malignant human LNs and provide a broad overview of emerging technologies for deconstructing the FL TME at single-cell and spatial resolution. We additionally discuss the importance of capturing samples at landmark time points as well as longitudinally for clinical decision-making.

Diffuse large B-cell lymphoma involving the central nervous system: biologic rationale for targeted therapy.

Diffuse large B-cell lymphoma (DLBCL) is an aggressive B-cell lymphoma curable even in advanced stages. DLBCL involving the central nervous system (CNS) is more difficult to cure and fewer treatment options exist. Primary CNS lymphoma (PCNSL) refers to aggressive lymphomas confined to the CNS, and are almost always DLBCL. Standard approaches for PCNSL use high-dose methotrexate-based combinations as induction therapy and younger patients often receive dose-intensive consolidation. However, dose-intensive therapies are not suitable for all patients, and older patients have fewer effective treatment options. Patients with relapsed or chemotherapy-refractory disease have a very poor prognosis. Secondary CNS lymphoma (SCNSL) describes aggressive lymphomas involving the CNS at initial presentation or relapses within the CNS after treatment for systemic DLBCL. Isolated CNS relapse is often managed as PCNSL, but patients with synchronous involvement of DLBCL in both the periphery and the CNS pose a unique clinical challenge. Insights into the molecular circuitry of DLBCL have identified distinct genetic subtypes including cases with a predilection for CNS invasion. PCNSL and subsets of SCNSL are characterized by chronically activated B-cell receptor and NFκB signaling along with genetic evidence of immune evasion which may be exploited therapeutically. Improved mechanistic understanding of targetable pathways underpinning CNS lymphomas has led to numerous clinical trials testing targeted agent combinations and immunotherapy approaches with promising early results. Biologically rational strategies may further improve the cure rate of CNS lymphomas, either by overcoming intrinsic or acquired treatment resistance and/or by being broadly applicable to patients of all ages.

Clinical applications of circulating tumor DNA in indolent B-cell lymphomas.

Indolent B-cell lymphomas are generally incurable with standard therapy and most patients have a prolonged disease course that includes multiple treatments and periods of time in which they do not require therapy. Currently available tools to monitor disease burden and define response to treatment rely heavily on imaging scans that lack tumor specificity are unable to detect disease at the molecular level. Circulating tumor DNA (ctDNA) is a versatile and promising biomarker being developed across multiple lymphoma subtypes. Advantages of ctDNA include high tumor specificity and limits of detection that are significantly lower than imaging scans. Potential clinical applications of ctDNA in indolent B-cell lymphomas include baseline prognostication, early signs of treatment resistance, measurements of minimal residual disease, and a noninvasive method to directly monitor disease burden and clonal evolution after therapy. Clinical applications of ctDNA have not yet proven clinical utility but are increasingly used as translational endpoints in clinical trials testing novel approaches and the analytic techniques used for ctDNA continue to evolve. Advances in therapy for indolent B-cell lymphomas include novel targeted agents and combinations that achieve very high rates complete response which amplifies the need to improve our current methods to monitor disease.

Phase I Study of Acalabrutinib Plus Danvatirsen (AZD9150) in Relapsed/Refractory Diffuse Large B-Cell Lymphoma Including Circulating Tumor DNA Biomarker Assessment.

PURPOSE: Novel targeted and immunotherapies have improved outcomes in relapsed/refractory (R/R) diffuse large B-cell lymphoma (DLBCL), but toxicities limit widespread use. The selective Bruton tyrosine kinase (BTK) inhibitor acalabrutinib has activity in patients with R/R DLBCL but durable responses are uncommon. STAT3 inhibition has demonstrated clinical activity in DLBCL. PATIENTS AND METHODS: Final results of the phase I study of acalabrutinib plus STAT3 inhibitor (danvatirsen; AZD9150) in patients with R/R DLBCL are reported. Danvatirsen 200 mg intravenous infusion [Days 1, 3, 5 (Cycle 1); weekly infusions starting Day 8, Cycle 1] was administered in combination with oral acalabrutinib 100 mg twice daily until progressive disease (PD) or unacceptable toxicity. Primary endpoints were safety and tolerability. Secondary endpoints included efficacy, pharmacokinetics, and immunogenicity. RESULTS: Seventeen patients received combination treatment. One dose-limiting toxicity (Grade 3 liver transaminase) occurred in 1 patient. The most common reason for treatment discontinuation was PD (65%). In evaluable patients (n = 17), objective response rate was 24%; median duration of response was 1.9 months. All responders with available DLBCL cell-of-origin data were either activated B-cell or nongerminal center B-cell like subtype. Genetic subtype did not correlate with response. Baseline and longitudinal plasma cell-free DNA (cfDNA) concentrations were mostly higher in nonresponding patients. cfDNA changes were generally concordant with imaging. Pretreatment circulating B-cell levels were higher in responders versus nonresponders. CONCLUSIONS: Targeting both STAT3 and BTK in combination is safe and tolerable but efficacy is limited in R/R DLBCL. Results support evaluation of circulating tumor DNA as a biomarker for clinical response.

Interferon alfa-2b in patients with low-grade lymphomatoid granulomatosis and chemotherapy with DA-EPOCH-R in patients with high-grade lymphomatoid granulomatosis: an open-label, single-centre, phase 2 trial.

BACKGROUND: Lymphomatoid granulomatosis is a rare Epstein-Barr virus-associated B-cell lymphoproliferative disorder with a median overall survival of less than 2 years. In this study, we hypothesised that low-grade lymphomatoid granulomatosis is immune-dependent and high-grade lymphomatoid granulomatosis is immune-independent. On the basis of this hypothesis, we investigated the activity and safety of new treatment with immunotherapy in patients with low-grade disease and standard chemotherapy in patients with high-grade disease. METHODS: In this open-label, single-centre, phase 2 trial, we enrolled patients aged 12 years or older with untreated, or relapsed or refractory lymphomatoid granulomatosis at the National Cancer Institute (National Institutes of Health, Bethesda, MD, USA). Patients with low-grade disease received dose-escalated interferon alfa-2b, starting at 7·5 million international units subcutaneously three times per week for up to 1 year past best response, and patients with high-grade disease received six cycles every 3 weeks of intravenous, dose-adjusted etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin, and rituximab (DA-EPOCH-R). Starting doses were 50 mg/m2 per day as a continuous intravenous infusion from day 1 to day 4 (96 h) for etoposide; 60 mg/m2 twice daily by mouth from day 1 to day 5 for prednisone; 0·4 mg/m2 per day as a continuous intravenous infusion from day 1 to day 4 (96 h) for vincristine; 750 mg/m2 intravenous on day 5 for cyclophosphamide; 10 mg/m2 per day as a continuous intravenous infusion from day 1 to day 4 (96 h) for doxorubicin; and 375 mg/m2 intravenous on day 1 for rituximab. The doses of doxorubicin, etoposide, and cyclophosphamide were adjusted up or down on the basis of neutrophil and platelet nadirs. Patients with residual or progressive disease after initial therapy crossed over to alternative therapy. The primary endpoint was the proportion of patients who had an overall response and the 5-year progression-free survival after initial or cross-over treatment. Analysis of response included all participants who underwent restaging imaging; safety analysis included all patients who received any dose of study drugs. The trial is open for enrolment and is registered at ClinicalTrials.gov, NCT00001379. FINDINGS: 67 patients were enrolled between Jan 10, 1991, and Sept 5, 2019 (42 [63%] were male). 45 patients received initial treatment with interferon alfa-2b (16 of whom crossed over to DA-EPOCH-R) and 18 received initial treatment with DA-EPOCH-R (eight of whom crossed over to interferon alfa-2b); four underwent surveillance only. After initial treatment with interferon alfa-2b, the overall response was 64% (28 of 44 evaluable patients) with 61% (27 of 44) having a complete response, whereas, after cross-over treatment with interferon alfa-2b, the overall response was 63% (five of eight evaluable patients) with 50% (four of eight) having a complete response. After initial treatment with DA-EPOCH-R, the overall response was 76% (13 of 17 evaluable patients) with 47% (eight of 17) having a complete response, whereas, after cross-over treatment with DA-EPOCH-R, the overall response was 67% (ten of 15 evaluable patients) with 47% (seven of 15) having a complete response. 5-year progression-free survival was 48·5% (95% CI 33·2-62·1) after initial treatment with interferon alfa-2b, 50·0% (15·2-77·5) after cross-over treatment with interferon alfa-2b, 25·4% (8·2-47·2) after initial treatment with DA-EPOCH-R, and 62·5% (34·9-81·1) after cross-over treatment with DA-EPOCH-R. The most common grade 3 or worse adverse events in patients treated with interferon alfa-2b included neutropenia (27 [53%] of 51 patients), lymphopenia (24 [47%]), and leukopenia (24 [47%]). The four most common grade 3 or worse adverse events in patients treated with DA-EPOCH-R included neutropenia (29 [88%] of 33 patients), leukopenia (28 [85%]), infection (18 [55%]), and lymphopenia (17 [52%]). Serious adverse events occurred in 13 (25%) of 51 patients receiving treatment with interferon alfa-2b and 21 (64%) of 33 patients receiving DA-EPOCH-R, with five treatment-related deaths: one thromboembolic, one infection, and one haemophagocytic syndrome with interferon alfa-2b, and one infection and one haemophagocytic syndrome with DA-EPOCH-R. INTERPRETATION: Interferon alfa-2b is efficacious for treating low-grade lymphomatoid granulomatosis and hence reducing progression to high-grade disease, whereas patients with high-grade lymphomatoid granulomatosis showed expected responses to chemotherapy. Uncontrolled immune regulation of Epstein-Barr virus is hypothesised to result in the emergence of low-grade disease after chemotherapy, for which treatment with interferon alfa-2b is efficacious. FUNDING: Intramural Research Programs of the National Cancer Institute and National Institute of Allergy and Infectious Diseases, National Institutes of Health.

Role of Artificial Intelligence in PET/CT Imaging for Management of Lymphoma.

Our review shows that AI-based analysis of lymphoma whole-body FDG-PET/CT can inform all phases of clinical management including staging, prognostication, treatment planning, and treatment response evaluation. We highlight advancements in the role of neural networks for performing automated image segmentation to calculate PET-based imaging biomarkers such as the total metabolic tumor volume (TMTV). AI-based image segmentation methods are at levels where they can be semi-automatically implemented with minimal human inputs and nearing the level of a second-opinion radiologist. Advances in automated segmentation methods are particularly apparent in the discrimination of lymphomatous vs non-lymphomatous FDG-avid regions, which carries through to automated staging. Automated TMTV calculators, in addition to automated calculation of measures such as Dmax are informing robust models of progression-free survival which can then feed into improved treatment planning.

Doxorubicin pharmacokinetics and toxicity in patients with aggressive lymphoma and hepatic impairment.

Aggressive lymphomas are curable with doxorubicin-based chemotherapy. In patients presenting with elevated serum bilirubin, doxorubicin is commonly dose reduced or delayed based on limited pharmacokinetic data. We evaluated plasma pharmacokinetics of doxorubicin and its metabolite doxorubicinol as well as toxicity in 59 patients with normal bilirubin levels and 10 patients with elevated bilirubin levels. Patients received full-dose EPOCH +/-rituximab. Median (range) age was 51 (18-75) years. Patients with elevated bilirubin levels had higher international prognostic index and poorer performance status. Although median doxorubicin clearance was lower and median plasma doxorubicin and doxorubicinol concentrations were higher in patients with elevated bilirubin levels, values were within the concentration range observed in patients with normal levels. Rates of febrile neutropenia were similar between groups, but there was greater grade 4 neutropenia and thrombocytopenia during the first but not subsequent treatment cycles in patients with elevated bilirubin. More grade 3/4 gastrointestinal and neurotoxicity occurred in patients with elevated bilirubin during the first but not subsequent cycles. Although toxicity was greater on cycle 1, the adverse effects were managed safely. These results show that empiric dose reductions of continuous infusion doxorubicin may not be necessary in patients with elevated bilirubin levels. This trial was registered at www.clinicaltrials.gov as #NCT00001337, #NCT00069238, and #NCT00005780.

Burkitt lymphoma.

Burkitt lymphoma (BL) is an aggressive form of B cell lymphoma that can affect children and adults. The study of BL led to the identification of the first recurrent chromosomal aberration in lymphoma, t(8;14)(q24;q32), and subsequent discovery of the central role of MYC and Epstein-Barr virus (EBV) in tumorigenesis. Most patients with BL are cured with chemotherapy but those with relapsed or refractory disease usually die of lymphoma. Historically, endemic BL, non-endemic sporadic BL and the immunodeficiency-associated BL have been recognized, but differentiation of these epidemiological variants is confounded by the frequency of EBV positivity. Subtyping into EBV+ and EBV- BL might better describe the biological heterogeneity of the disease. Phenotypically resembling germinal centre B cells, all types of BL are characterized by dysregulation of MYC due to enhancer activation via juxtaposition with one of the three immunoglobulin loci. Additional molecular changes commonly affect B cell receptor and sphingosine-1-phosphate signalling, proliferation, survival and SWI-SNF chromatin remodelling. BL is diagnosed on the basis of morphology and high expression of MYC. BL can be effectively treated in children and adolescents with short durations of high dose-intensity multiagent chemotherapy regimens. Adults are more susceptible to toxic effects but are effectively treated with chemotherapy, including modified versions of paediatric regimens. The outcomes in patients with BL are good in high-income countries with low mortality and few late effects, but in low-income and middle-income countries, BL is diagnosed late and is usually treated with less-effective regimens affecting the overall good outcomes in patients with this lymphoma.

SOHO State of the Art Updates and Next Questions: Prophylaxis and Management of Secondary CNS Lymphoma.

Secondary CNS lymphoma (SCNSL) is a rare but frequently fatal complication of systemic lymphoma. There is no standard treatment for SCNSL, and patients who develop SCNSL at diagnosis or after frontline therapy often receive highly intensive chemotherapy regimens that are inactive against primary chemorefractory disease and too toxic for older, frail patients to tolerate. Because the prognosis of SCNSL is so poor, management has historically emphasized prevention, but the current methods of CNS prophylaxis are not universally effective. To improve both the prevention and management of SCNSL, better characterization of the molecular determinants of CNS invasion is needed. Novel treatments that are currently being studied in SCNSL include targeted pathway inhibitors and cellular therapy, but SCNSL patients are often excluded from clinical trials of promising new therapies.

Circulating tumor DNA predicts therapeutic outcome in mantle cell lymphoma.

Mantle cell lymphoma (MCL) is biologically and clinically heterogeneous and would benefit from prognostic biomarkers to guide management. Circulating tumor DNA (ctDNA) is a novel prognostic biomarker in diffuse large B-cell lymphoma that may have applicability in MCL. We analyzed ctDNA dynamics in previously untreated patients with MCL who received induction therapy with bortezomib and DA-EPOCH-R for 6 cycles followed by random assignment to observation or bortezomib maintenance in responding patients in a prospective phase 2 study. Most patients also underwent initial treatment window of bortezomib alone prior to induction. Serum was collected pretreatment, after the window, after cycles 1 and 2, at the end of induction, and at each follow-up visit along with restaging computed tomography scans. Next-generation sequencing was used to identify and quantify ctDNA encoding the immunoglobulin receptor sequences in serum as markers of minimal residual disease. Fifty-three patients were enrolled, with a median follow-up of 12.7 years. Patients without detectable ctDNA after 2 cycles of induction had longer progression-free survival (PFS) and overall survival (OS) compared with those with detectable ctDNA (median PFS, 2.7 vs 1.8 years; overall P = .005; median OS, 13.8 vs 7.4 years; overall P = .03). Notably, in vivo assessment of ctDNA dynamics during the bortezomib window was not prognostic, and there was no difference in PFS or OS with bortezomib maintenance. ctDNA monitoring after induction showed that molecular relapse preceded clinical relapse in some cases. In conclusion, interim ctDNA negativity strongly correlates with improved survival and supports the investigation of response-adapted strategies. This trial was registered at www.clinicaltrials.gov as #NCT00114738.