Despite major progress in mantle cell lymphoma (MCL) therapeutics, MCL remains a deadly disease with a median survival not exceeding four years. No single driver genetic lesion has been described to ...solely give rise to MCL. The hallmark translocation t(11;14)(q13;q32) requires additional genetic alterations for the malignant transformation. A short list of recurrently mutated genes including ATM, CCND1, UBR5, TP53, BIRC3, NOTCH1, NOTCH2, and TRAF2 recently emerged as contributors to the pathogenesis of MCL. Notably, NOTCH1 and NOTCH2 were found to be mutated in multiple B cell lymphomas, including 5-10% of MCL, with most of these mutations occurring within the PEST domain of the protein. The NOTCH genes play a critical role in the early and late phases of normal B cell differentiation. In MCL, mutations in the PEST domain stabilize NOTCH proteins, rendering them resistant to degradation, which subsequently results in the upregulation of genes involved in angiogenesis, cell cycle progression, and cell migration and adhesion. At the clinical level, mutated NOTCH genes are associated with aggressive features in MCL, such as the blastoid and pleomorphic variants, a shorter response to treatment, and inferior survival. In this article, we explore in detail the role of NOTCH signaling in MCL biology and the ongoing efforts toward targeted therapeutic interventions.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Plasma SARS-CoV-2 RNA may represent a viable diagnostic alternative to respiratory RNA levels, which rapidly decline after infection. Quantitative PCR with reverse transcription (RT-qPCR) reference ...assays exhibit poor performance with plasma, probably reflecting the dilution and degradation of viral RNA released into the circulation, but these issues could be addressed by analysing viral RNA packaged into extracellular vesicles. Here we describe an assay approach in which extracellular vesicles directly captured from plasma are fused with reagent-loaded liposomes to sensitively amplify and detect a SARS-CoV-2 gene target. This approach accurately identified patients with COVID-19, including challenging cases missed by RT-qPCR. SARS-CoV-2-positive extracellular vesicles were detected at day 1 post-infection, and plateaued from day 6 to the day 28 endpoint in a non-human primate model, while signal durations for 20-60 days were observed in young children. This nanotechnology approach uses a non-infectious sample and extends virus detection windows, offering a tool to support COVID-19 diagnosis in patients without SARS-CoV-2 RNA detectable in the respiratory tract.
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GEOZS, IJS, IMTLJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBMB, UL, UM, UPUK, ZAGLJ
The safety and efficacy of ibrutinib (420 mg) in chronic lymphocytic leukemia (CLL) were evaluated in a phase 2 study; 51 patients had TP53 aberration (TP53 cohort) and 35 were enrolled because of ...age 65 years or older (elderly cohort). Both cohorts included patients with treatment-naive (TN) and relapsed/refractory (RR) CLL. With the median follow-up of 4.8 years, 49 (57.0%) of 86 patients remain on study. Treatment was discontinued for progressive disease in 20 (23.3%) patients and for adverse events in 5 (5.8%). Atrial fibrillation occurred in 18 (20.9%) patients for a rate of 6.4 per 100 patient-years. No serious bleeding occurred. The overall response rate at 6 months, the primary study endpoint, was 95.8% for the TP53 cohort (95% confidence interval, 85.7%-99.5%) and 93.9% for the elderly cohort (95% confidence interval, 79.8%-99.3%). Depth of response improved with time: at best response, 14 (29.2%) of 48 patients in the TP53 cohort and 9 (27.3%) of 33 in the elderly cohort achieved a complete response. Median minimal residual disease (MRD) in peripheral blood was 3.8 × 10−2 at 4 years, with MRD-negative (<10−4) remissions in 5 (10.2%) patients. In the TP53 cohort, the estimated 5-year progression-free survival (PFS) was 74.4% in TN-CLL compared with 19.4% in RR-CLL (P = .0002), and overall survival (OS) was 85.3% vs 53.7%, respectively (P = .023). In the elderly cohort, the estimated 5-year PFS and OS in RR-CLL were 64.8% and 71.6%, respectively, and no event occurred in TN-CLL. Long-term administration of ibrutinib was well tolerated and provided durable disease control for most patients. This trial was registered at www.clinicaltrials.gov as #NCT01500733.
•With 5-year median follow-up, continuous single-agent ibrutinib therapy was well tolerated with deepening of response.•Previously untreated patients, even those with TP53 aberration, achieved durable responses.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
The Bruton tyrosine kinase inhibitor ibrutinib induces high rates of clinical response in chronic lymphocytic leukemia (CLL). However, there remains a need for adjunct treatments to deepen response ...and to overcome drug resistance. Blinatumomab, a CD19/CD3 bispecific antibody (bsAb) designed in the BiTE (bispecific T-cell engager) format, is approved by the US Food and Drug Administration for the treatment of relapsed or refractory B-cell precursor acute lymphoblastic leukemia. Because of its short half-life of 2.1 hours, blinatumomab requires continuous intravenous dosing for efficacy. We developed a novel CD19/CD3 bsAb in the single-chain Fv-Fc format (CD19/CD3-scFv-Fc) with a half-life of ∼5 days. In in vitro experiments, both CD19/CD3-scFv-Fc and blinatumomab induced >90% killing of CLL cells from treatment-naïve patients. Antileukemic activity was associated with increased autologous CD8 and CD4 T-cell proliferation, activation, and granzyme B expression. In the NOD/SCID/IL2Rγnull patient-derived xenograft mouse model, once-weekly treatment with CD19/CD3-scFv-Fc eliminated >98% of treatment-naïve CLL cells in blood and spleen. By contrast, blinatumomab failed to induce a response, even when administered daily. We next explored the activity of CD19/CD3-scFv-Fc in the context of ibrutinib treatment and ibrutinib resistance. CD19/CD3-scFv-Fc induced more rapid killing of CLL cells from ibrutinib-treated patients than those from treatment-naïve patients. CD19/CD3-scFv-Fc also demonstrated potent activity against CLL cells from patients with acquired ibrutinib-resistance harboring BTK and/or PLCG2 mutations in vitro and in vivo using patient-derived xenograft models. Taken together, these data support investigation of CD19/CD3 bsAb's and other T cell-recruiting bsAb's as immunotherapies for CLL, especially in combination with ibrutinib or as rescue therapy in ibrutinib-resistant disease.
•A CD19/CD3 single-chain Fv-Fc bsAb mediated potent killing of CLL cells by autologous T cells in vitro and in vivo.•bsAb-mediated cytotoxicity was enhanced by prior therapy with ibrutinib and extended to ibrutinib-resistant disease.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Point-of-care COVID-19 assays that are more sensitive than the current RT-PCR (reverse transcription polymerase chain reaction) gold standard assay are needed to improve disease control efforts. We ...describe the development of a portable, ultrasensitive saliva-based COVID-19 assay with a 15-min sample-to-answer time that does not require RNA isolation or laboratory equipment. This assay uses CRISPR-Cas12a activity to enhance viral amplicon signal, which is stimulated by the laser diode of a smartphone-based fluorescence microscope device. This device robustly quantified viral load over a broad linear range (1 to 10
copies/μl) and exhibited a limit of detection (0.38 copies/μl) below that of the RT-PCR reference assay. CRISPR-read SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) RNA levels were similar in patient saliva and nasal swabs, and viral loads measured by RT-PCR and the smartphone-read CRISPR assay demonstrated good correlation, supporting the potential use of this portable assay for saliva-based point-of-care COVID-19 diagnosis.
An 84-year-old man with nonmetastatic castrate-sensitive prostate cancer was referred for a 68Ga-prostate-specific membrane antigen (PSMA) PET/CT scan with a prostate-specific antigen level of 3.6 ...ng/mL for restaging. He was 22 years post-radical prostatectomy and had salvage radiation being managed with intermittent hormonal therapy. Imaging revealed a right lower lobe mass with increased PSMA uptake (SUVmax 6.2). Biopsy and subsequent immunostaining determined the mass to be diffuse large B-cell lymphoma. We report a case of diffuse large B-cell lymphoma diagnosed in the setting of PSMA positivity, highlighting awareness for oncologists and radiologists to know this possibility.
B-cell acute lymphoblastic leukemia (B-ALL) remains a hard-to-treat disease with a poor prognosis in adults. Mucosa-associated lymphoid tissue lymphoma translocation protein 1 (MALT1) is a ...para-caspase required for B-cell receptor (BCR)-mediated NF-κB activation. Inhibition of MALT1 in preclinical models has proven efficacious in many B-cell malignancies including chronic lymphocytic leukemia, mantle cell lymphoma and diffuse large B-cell lymphoma. We sought to examine the role of MALT1 in B-ALL and determine the biological consequences of its inhibition. Targeting MALT1 with both Z-VRPR-fmk and MI-2 efficiently kills B-ALL cells independent of the cell-of-origin (pro, pre, mature) or the presence of the Philadelphia chromosome, and spares normal B cells. The mechanism of cell death was through apoptotic induction, mostly in cycling cells. The proteolytic activity of MALT1 can be studied by measuring its ability to cleave its substrates. Surprisingly, with the exception of mature B-ALL, we did not detect cleavage of MALT1 substrates at baseline, nor after proteasomal inhibition or following activation of pre-BCR. To explore the possibility of a distinct role for MALT1 in B-ALL, independent of signaling through BCR, we studied the changes in gene expression profiling following a 24-hour treatment with MI-2 in 12 B-ALL cell lines. Our transcriptome analysis revealed a strong inhibitory effect on MYC-regulated gene signatures, further confirmed by Myc protein downregulation, concomitant with an increase in the Myc degrader FBXW7. In conclusion, our evidence suggests a novel role for MALT1 in B-ALL through Myc regulation and provides support for clinical testing of MALT1 inhibitors in B-ALL.
To interrogate signaling pathways activated in mantle cell lymphoma (MCL) in vivo, we contrasted gene expression profiles of 55 tumor samples isolated from blood and lymph nodes from 43 previously ...untreated patients with active disease. In addition to lymph nodes, MCL often involves blood, bone marrow, and spleen and is incurable for most patients. Recently, the Bruton tyrosine kinase (BTK) inhibitor ibrutinib demonstrated important clinical activity in MCL. However, the role of specific signaling pathways in the lymphomagenesis of MCL and the biologic basis for ibrutinib sensitivity of these tumors are unknown. Here, we demonstrate activation of B-cell receptor (BCR) and canonical NF-κB signaling specifically in MCL cells in the lymph node. Quantification of BCR signaling strength, reflected in the expression of BCR regulated genes, identified a subset of patients with inferior survival after cytotoxic therapy. Tumor proliferation was highest in the lymph node and correlated with the degree of BCR activation. A subset of leukemic tumors showed active BCR and NF-κB signaling apparently independent of microenvironmental support. In one of these samples, we identified a novel somatic mutation in RELA (E39Q). This sample was resistant to ibrutinib-mediated inhibition of NF-κB and apoptosis. In addition, we identified germ line variants in genes encoding regulators of the BCR and NF-κB pathway previously implicated in lymphomagenesis. In conclusion, BCR signaling, activated in the lymph node microenvironment in vivo, appears to promote tumor proliferation and survival and may explain the sensitivity of this lymphoma to BTK inhibitors.
•Activation of BCR and canonical NF-κB signaling in the lymph node correlates with survival in MCL.•Mutations and polymorphisms in BCR and NF-κB pathways may confer cell autonomous signaling and affect response to ibrutinib.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
IMPORTANCE: Cutaneous disease in dermatomyositis has no standardized treatment approach and so presents a challenging task for patients and clinicians. OBJECTIVE: To study the efficacy and safety of ...apremilast as an add-on therapy in patients with recalcitrant cutaneous dermatomyositis. DESIGN, SETTING, AND PARTICIPANTS: This phase 2a, open-label, single-arm nonrandomized controlled trial was conducted at a single center from June 2018 to June 2021. Participants were 8 patients with recalcitrant cutaneous dermatomyositis, defined by a cutaneous disease activity severity index (CDASI) score greater than 5 despite treatment with steroids, steroid-sparing agents, or both. Data were analyzed from June 2018 to June 2021. INTERVENTIONS: Apremilast 30 mg orally twice daily was added to ongoing treatment regimens. MAIN OUTCOMES AND MEASURES: The primary outcome was the overall response rate (ORR) at 3 months. Key secondary outcomes were the safety and toxicity of apremilast and the durability of response at 6 months. The CDASI, muscle score, dermatology life quality index (DLQI), and depression assessments were performed at baseline and regularly until month 7. Skin biopsies were performed at baseline and 3 months after apremilast (defined as 3 months into active apremilast therapy) and tested for gene expression profiling and immunohistochemical stains. Adverse events were assessed using the Common Terminology Criteria for Adverse Events version 5.0. RESULTS: Among 8 patients with recalcitrant cutaneous dermatomyositis (all women; mean SD age, 54 15.9 years), a response was found at 3 months after apremilast among 7 patients (ORR, 87.5%). The mean (SD) decrease in CDASI was 12.9 (6.3) points at 3 months (P < .001). Apremilast was well tolerated, with no grade 3 or higher adverse events. Sequencing of RNA was performed on skin biopsies taken from 7 patients at baseline and at 3 months after therapy. Appropriate negative (ie, no primary antibody) and positive (ie, tonsil and spleen) controls were stained in parallel with each set of slides studied. Of 39 076 expressed genes, there were 195 whose expression changed 2-fold or more at P < .01 (123 downregulated and 72 upregulated genes). Gene set enrichment analysis identified 13 pathways in which apremilast was associated with downregulated expression, notably signal transducers and activators of transcription 1 (STAT1), STAT3, interleukin 4 (IL-4), IL-6, IL-12, IL-23, interferon γ (IFNγ), and tumor necrosis factor α (TNFα) pathways. In immunohistochemical staining, there was a mean (SD) decrease in phosphorylation levels STAT1 (22.3% 28.3% positive cells) and STAT3 (13.4% 11.6% positive cells) at the protein level, a downstream signaling pathway for the downregulated cytokines. CONCLUSIONS AND RELEVANCE: These findings suggest that apremilast was a safe and efficacious add-on treatment in recalcitrant dermatomyositis, with an overall response rate of 87.5% and associations with downregulation of multiple inflammatory pathways. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03529955
The clinical efficacy displayed by ibrutinib in chronic lymphocytic leukemia (CLL) has been challenged by the frequent emergence of resistant clones. The ibrutinib target, Bruton's tyrosine kinase ...(BTK), is essential for B-cell receptor signaling, and most resistant cases carry mutations in
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, a downstream effector target of BTK. Recent findings show that MI-2, a small molecule inhibitor of the para-caspase MALT1, is effective in preclinical models of another type of BCR pathway-dependent lymphoma. We therefore studied the activity of MI-2 against CLL and ibrutinib-resistant CLL. Treatment of CLL cells
with MI-2 inhibited MALT1 proteolytic activity reduced BCR and NF-κB signaling, inhibited nuclear translocation of RelB and p50, and decreased Bcl-xL levels. MI-2 selectively induced dose and time-dependent apoptosis in CLL cells, sparing normal B lymphocytes. Furthermore, MI-2 abrogated survival signals provided by stromal cells and BCR cross-linking and was effective against CLL cells harboring features associated with poor outcomes, including 17p deletion and unmutated
Notably, MI-2 was effective against CLL cells collected from patients harboring mutations conferring resistance to ibrutinib. Overall, our findings provide a preclinical rationale for the clinical development of MALT1 inhibitors in CLL, in particular for ibrutinib-resistant forms of this disease.
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