Anti-CD19 chimeric antigen receptor T-cell therapy in older patients with relapsed or refractory large B-cell lymphoma: A multicenter study.

Chimeric antigen receptor T-cell (CAR-T) therapy, despite being a potentially curative therapy in relapsed or refractory (RR) large B-cell lymphoma (LBCL), remains underutilized in older patients due to limited clinical data. We therefore studied the safety and efficacy of CAR-T therapy in older patients with RR LBCL in the real-world setting. Patients aged ≥65 years with RR LBCL, treated with anti-CD19 CAR-T therapy at 7 US institutions were included in this multicenter, retrospective, observational study. In total, 226 patients were included. Median age at infusion was 71 years (range 65-89). Best objective and complete response rates were 86% and 62%, respectively. Median follow-up after infusion was 18.3 months. The median progression-free survival (PFS) was 6.9 months, with 6- and 12-month PFS estimates of 54% and 44%, respectively. The nonrelapse mortality (NRM) rate was 10.9% at day 180, primarily due to infections, and not impacted by the age groups. Grade ≥3 cytokine release syndrome and neurotoxicity occurred in 7% and 26%, respectively. In univariate analysis, no significant difference in PFS was seen regardless of the age groups or CAR-T type, whereas ECOG PS ≥2, elevated LDH, bulky disease, advanced stage, extranodal involvement, the need for bridging therapy, and prior bendamustine exposure were associated with shorter PFS. These findings support the use of CAR-T in older patients, including those aged ≥80 years. The age at CAR-T therapy did not influence safety, survival, and NRM outcomes. Older patients should not be excluded from receiving CAR-T therapy solely based on their chronological age.

CRISPR-based assay reveals SARS-CoV-2 RNA dynamic changes and redistribution patterns in non-human primate model.

Mounting evidence indicates that SARS-CoV-2 can infect multiple systemic tissues, but few studies have evaluated SARS-CoV-2 RNA dynamics in multiple specimen types due to their reduced accessibility and diminished performance of RT-qPCR with non-respiratory specimens. Here, we employed an ultrasensitive CRISPR-RT-PCR assay to analyze longitudinal mucosal (nasal, buccal, pharyngeal, and rectal), plasma, and breath samples from SARS-CoV-2-infected non-human primates (NHPs) to detect dynamic changes in SARS-CoV-2 RNA level and distribution among these specimens. We observed that CRISPR-RT-PCR results consistently detected SARS-CoV-2 RNA in all sample types at most time points post-infection, and that SARS-CoV-2 infection dose and administration route did not markedly affect the CRISPR-RT-PCR signal detected in most specimen types. However, consistent RT-qPCR positive results were restricted to nasal, pharyngeal, and rectal swab samples, and tended to decrease earlier than CRISPR-RT-PCR results, reflecting lower assay sensitivity. SARS-CoV-2 RNA was detectable in both pulmonary and extrapulmonary specimens from early to late infection by CRISPR-RT-PCR, albeit with different abundance and kinetics, with SARS-CoV-2 RNA increases detected in plasma and rectal samples trailing those detected in upper respiratory tract samples. CRISPR-RT-PCR assays for SARS-CoV-2 RNA in non-respiratory specimens may thus permit direct diagnosis of suspected COVID-19 cases missed by RT-PCR, while tracking SARS-CoV-2 RNA in minimally invasive alternate specimens may better evaluate the progression and resolution of SARS-CoV-2 infections.

A smartphone-read ultrasensitive and quantitative saliva test for COVID-19.

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 105 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.

Liposome-mediated detection of SARS-CoV-2 RNA-positive extracellular vesicles in plasma.

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.

Sensitive tracking of circulating viral RNA through all stages of SARS-CoV-2 infection.

BACKGROUNDCirculating severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA may represent a more reliable indicator of infection than nasal RNA, but quantitative reverse transcription PCR (RT-qPCR) lacks diagnostic sensitivity for blood samples.METHODSA CRISPR-augmented RT-PCR assay that sensitively detects SARS-CoV-2 RNA was employed to analyze viral RNA kinetics in longitudinal plasma samples from nonhuman primates (NHPs) after virus exposure; to evaluate the utility of blood SARS-CoV-2 RNA detection for coronavirus disease 2019 (COVID-19) diagnosis in adults cases confirmed by nasal/nasopharyngeal swab RT-PCR results; and to identify suspected COVID-19 cases in pediatric and at-risk adult populations with negative nasal swab RT-qPCR results. All blood samples were analyzed by RT-qPCR to allow direct comparisons.RESULTSCRISPR-augmented RT-PCR consistently detected SARS-CoV-2 RNA in the plasma of experimentally infected NHPs from 1 to 28 days after infection, and these increases preceded and correlated with rectal swab viral RNA increases. In a patient cohort (n = 159), this blood-based assay demonstrated 91.2% diagnostic sensitivity and 99.2% diagnostic specificity versus a comparator RT-qPCR nasal/nasopharyngeal test, whereas RT-qPCR exhibited 44.1% diagnostic sensitivity and 100% specificity for the same blood samples. This CRISPR-augmented RT-PCR assay also accurately identified patients with COVID-19 using one or more negative nasal swab RT-qPCR results.CONCLUSIONResults of this study indicate that sensitive detection of SARS-CoV-2 RNA in blood by CRISPR-augmented RT-PCR permits accurate COVID-19 diagnosis, and can detect COVID-19 cases with transient or negative nasal swab RT-qPCR results, suggesting that this approach could improve COVID-19 diagnosis and the evaluation of SARS-CoV-2 infection clearance, and predict the severity of infection.TRIAL REGISTRATIONClinicalTrials.gov. NCT04358211.FUNDINGDepartment of Defense, National Institute of Allergy and Infectious Diseases, National Institute of Child Health and Human Development, and the National Center for Research Resources.

Circulating-tumor DNA as predictor of enzalutamide response post-abiraterone treatment in metastatic castration-resistant prostate cancer.

BACKGROUND: The crossover from abiraterone acetate (AA) to enzalutamide (ENZA) is a frequent approach in clinical practice. Our aim was to explore the role of genomic alterations as putative biomarkers of response to sequential AA followed by ENZA in mCRPC and their association with clinical outcomes. PATIENTS AND METHODS: This was a multi-center, retrospective analysis of mCRPC patients with circulating-tumor DNA (ctDNA) post-AA and prior to ENZA treatment. Objectives of this analysis were to assess PSA response, time to PSA progression (TTP) and overall survival (OS) in mCRPC patients treated with ENZA following progression on AA with respect to genomic aberrations detected by ctDNA. RESULTS: A total of 28 patients with mCRPC were identified. Median time between AA and ENZA was 3.1 months and median initial PSA prior to ENZA was 35.0 ng/mL. Nine patients (32.1%) achieved PSA responses to ENZA. Most patients (79.0%) achieved confirmed PSA progression with median TTP of 1.6 months (95% CI, 0.7-2.4). Somatic alterations in AR genes were detected in 36.0% of patients with other common alterations detected including 39.0% TP53, 11.0% DNA repair, and 11.0% PTEN. A lack of AR alterations was associated with better PSA response to ENZA (p = 0.04). CONCLUSION: While lack of AR alterations in ctDNA was associated with more favorable outcomes, the present dataset is insufficient to recommend the use of ctDNA to impact clinical decision-making in this setting. Further understanding of the implications of the genomic phenotype in ctDNA of castration-resistant tumors and the potential therapeutic implications is required.