Microfluidic Digital Immunoassay for Point-of-Care Detection of NT-proBNP from Whole Blood.

The high prevalence and economic burden of heart failure remain a challenge to global health. This lifelong disease leads to a buildup of permanent heart damage, making early detection and frequent monitoring crucial for effective treatment. N-terminal proBNP (NT-proBNP) is an important biomarker for monitoring the disease state, but current commercial and research NT-proBNP assays require phlebotomy and bulky equipment or do not satisfy clinical requirements such as sensitivity and detection thresholds. Here, we report a point-of-care (POC) compatible microfluidic digital immunoassay that can quantify the NT-proBNP concentration in a small volume of whole blood. Our automated microfluidic device takes whole blood samples mixed with biotinylated detection antibodies and passes through a plasma filter to react with a capture antibody-functionalized sensor surface. Streptavidin-coated gold nanoparticles (GNPs) are then released to mark the surface-bound single NT-proBNP immunocomplexes and recorded with bright-field microscopy. NT-proBNP concentrations in the sample are quantified via a hybrid digital/analog calibration curve. Digital counts of bound GNPs are used as readout signal at low concentrations for high sensitivity detection, and GNP pixel occupancies are used at high concentrations for extended dynamic range. With this approach, we detected NT-proBNP in the range of 8.24-10 000 pg/mL from 7 µL of whole blood in 10 min, with a limit of detection of 0.94 pg/mL. Finally, the method was validated with 15 clinical serum samples, showing excellent linear correlation (r = 0.998) with Roche's Elecsys proBNP II assay. This evidence indicates that this method holds promise for decentralized monitoring of heart failure.

IL-1Ra gene transfer potentiates BMP2-mediated bone healing by redirecting osteogenesis toward endochondral ossification.

An estimated 100,000 patients each year in the United States suffer severe disability from bone defects that fail to heal, a condition where bone-regenerative therapies could provide substantial clinical benefits. Although recombinant human bone morphogenetic protein-2 (rhBMP2) is an osteogenic growth factor that is clinically approved for this purpose, it is only effective when used at exceedingly high doses that incur substantial costs, induce severe inflammation, produce adverse side effects, and form morphologically abnormal bone. Using a validated rat femoral segmental defect model, we show that bone formed in response to clinically relevant doses of rhBMP2 is accompanied by elevated expression of interleukin-1 (IL-1). Local delivery of cDNA encoding the IL-1 receptor antagonist (IL-1Ra) achieved bridging of segmental, critical size defects in bone with a 90% lower dose of rhBMP2. Unlike use of high-dose rhBMP2, bone formation in the presence of IL-1Ra occurred via the native process of endochondral ossification, resulting in improved quality without sacrificing the mechanical properties of the regenerated bone. Our results demonstrate that local immunomodulation may permit effective use of growth factors at lower doses to recapitulate more precisely the native biology of healing, leading to higher-quality tissue regeneration.

Assessing the Effect of Middle Ear Effusions on Wideband Acoustic Immittance Using Optical Coherence Tomography.

OBJECTIVES: Wideband acoustic immittance (WAI) noninvasively assesses middle ear function by measuring the sound conduction over a range of audible frequencies. Although several studies have shown the potential of WAI for detecting the presence of middle ear effusions (MEEs), determining the effects of MEE type and amount on WAI in vivo has been challenging due to the anatomical location of middle ear cavity. The purpose of this study is to correlate WAI measurements with physical characteristics of the middle ear and MEEs determined by optical coherence tomography (OCT), a noninvasive optical imaging technique. DESIGN: Sixteen pediatric subjects (average age of 7 ± 4 years) were recruited from the primary care clinic at Carle Foundation Hospital (Urbana, IL). A total of 22 ears (normal: 15 ears, otitis media with effusion: 6 ears, and acute otitis media: 1 ear, based on physician's diagnosis) were examined via standard otoscopy, tympanometry, OCT imaging, and WAI measurements in a busy, community-based clinical setting. Cross-sectional OCT images were analyzed to quantitatively assess the presence, type (relative turbidity based on the amount of scattering), and amount (relative fluid level) of MEEs. These OCT metrics were utilized to categorize subject ears into no MEE (control), biofilm without a MEE, serous-scant, serous-severe, mucoid-scant, and mucoid-severe MEE groups. The absorbance levels in each group were statistically evaluated at α = 0.05. RESULTS: The absorbance of the control group showed a similar trend when compared with a pediatric normative dataset, and the presence of an MEE generally decreased the power absorbance. The mucoid MEE group showed significantly less power absorbance from 2.74 to 4.73 kHz (p < 0.05) when compared with the serous MEE group, possibly due to the greater mass impeding the middle ear system. Similarly, the greater amount of middle ear fluid contributed to the lower power absorbance from 1.92 to 2.37 kHz (p< 0.05), when compared with smaller amounts of fluid. As expected, the MEEs with scant fluid only significantly affected the power absorbance at frequencies greater than 4.85 kHz. A large variance in the power absorbance was observed between 2 and 5 kHz, suggesting the dependence on both the type and amount of MEE. CONCLUSIONS: Physical characteristics of the middle ear and MEEs quantified from noninvasive OCT images can be helpful to understand abnormal WAI measurements. Mucoid MEEs decrease the power absorbance more than serous MEEs, and the greater amounts of MEE decreases the power absorbance, especially at higher (>2 kHz) frequencies. As both the type and amount of MEE can significantly affect WAI measurements, further investigations to correlate acoustic measurements with physical characteristics of middle ear conditions in vivo is needed.

Contribution of Implanted, Genetically Modified Muscle Progenitor Cells Expressing BMP-2 to New Bone Formation in a Rat Osseous Defect.

Because muscle contains osteoprogenitor cells and has a propensity to form bone, we have explored its utility in healing large osseous defects. Healing is achieved by the insertion of muscle fragments transduced with adenovirus encoding BMP-2 (Ad.BMP-2). However, it is not known whether the genetically modified muscle contributes osteoprogenitor cells to healing defects or merely serves as a local source of BMP-2. This question is part of the larger debate on the fate of progenitor cells introduced into sites of tissue damage to promote regeneration. To address this issue, we harvested fragments of muscle from rats constitutively expressing GFP, transduced them with Ad.BMP-2, and implanted them into femoral defects in wild-type rats under various conditions. GFP+ cells persisted within defects for the entire 8 weeks of the experiments. In the absence of bone formation, these cells presented as fibroblasts. When bone was formed, GFP+ cells were present as osteoblasts and osteocytes and also among the lining cells of new blood vessels. The genetically modified muscle thus contributed progenitor cells as well as BMP-2 to the healing defect, a property of great significance in light of the extensive damage to soft tissue and consequent loss of endogenous progenitors in problematic fractures.

The effect of moderate-intensity exercise on nightly variability in objectively measured sleep parameters among older women.

Objective/Background: Exercise training has been demonstrated to beneficially influence mean-level measures of sleep; however, few studies have examined the impact of an exercise intervention on night-to-night variability in sleep. This study investigated whether four months of moderate-intensity exercise impacted night-to-night variability in sleep among older women. Methods: Participants (n = 49) were randomized to one of two moderate-intensity walking programs with different doses of energy expenditure: low-dose (n = 23: 8 kcal/kg of body weight per week) or high-dose (n = 26: 14 kcal/kg of body weight per week). Sleep parameters were assessed objectively via actigraphy at baseline, mid- (2 months), and postintervention (4 months). Nightly variability in each of the sleep parameters was calculated using the seven-day standard deviation (SD) and a coefficient of variation (SD/mean x 100%). Cardiorespiratory fitness (VO2peak) was measured at baseline and postintervention using a graded treadmill test. Results: Both measures of nightly variability demonstrated a borderline to significantly lower amount of night-to-night variability in wake time after sleep onset (WASO) and number of awakenings at postintervention in comparison to baseline (p ≤ 0.05). Higher VO2peak levels at baseline were associated with less time in bed and lower total sleep time variability throughout the exercise intervention (p < 0.05). Conclusion: Overall, participation in moderate-intensity exercise was observed to reduce the amount of nightly variability for WASO and number of awakenings over time in older women.

A large microRNA cluster on chromosome 19 is a transcriptional hallmark of WHO type A and AB thymomas.

BACKGROUND: Thymomas are one of the most rarely diagnosed malignancies. To better understand its biology and to identify therapeutic targets, we performed next-generation RNA sequencing. METHODS: The RNA was sequenced from 13 thymic malignancies and 3 normal thymus glands. Validation of microRNA expression was performed on a separate set of 35 thymic malignancies. For cell-based studies, a thymoma cell line was used. RESULTS: Hierarchical clustering revealed 100% concordance between gene expression clusters and WHO subtype. A substantial differentiator was a large microRNA cluster on chr19q13.42 that was significantly overexpressed in all A and AB tumours and whose expression was virtually absent in the other thymomas and normal tissues. Overexpression of this microRNA cluster activates the PI3K/AKT/mTOR pathway. Treatment of a thymoma AB cell line with a panel of PI3K/AKT/mTOR inhibitors resulted in marked reduction of cell viability. CONCLUSIONS: A large microRNA cluster on chr19q13.42 is a transcriptional hallmark of type A and AB thymomas. Furthermore, this cluster activates the PI3K pathway, suggesting the possible exploration of PI3K inhibitors in patients with these subtypes of tumour. This work has led to the initiation of a phase II clinical trial of PI3K inhibition in relapsed or refractory thymomas (http://clinicaltrials.gov/ct2/show/NCT02220855).

Evaluation of the Effectiveness of the H.A.N.D.S.SM Program: A School Nurse Diabetes Management Education Program.

The purpose of this project was to determine the effectiveness of the Helping Administer to the Needs of the Student with Diabetes in Schools (H.A.N.D.S.(sm)) continuing education program in improving the level of experience and competence in performing services associated with diabetes care. This program is a live course for school nurses providing clinical information about diabetes management and their professional role in the care of students with diabetes. Pre- and post-surveys were administered via e-mail to assess their level of experience and competence in diabetes care. A total of 105 nurses completed both surveys and were included in the analysis. The changes between pre- and post-survey questions were assessed. The H.A.N.D.S. participants' levels of experience and competence for each of the four categories of diabetes care improved significantly, and a greater number of nurses reported being able to perform the services independently and having the ability to teach others.

Inflammatory cytokines induce a unique mineralizing phenotype in mesenchymal stem cells derived from human bone marrow.

Bone marrow contains mesenchymal stem cells (MSCs) that can differentiate along multiple mesenchymal lineages. In this capacity they are thought to be important in the intrinsic turnover and repair of connective tissues while also serving as a basis for tissue engineering and regenerative medicine. However, little is known of the biological responses of human MSCs to inflammatory conditions. When cultured with IL-1ß, marrow-derived MSCs from 8 of 10 human subjects deposited copious hydroxyapatite, in which authenticity was confirmed by Fourier transform infrared spectroscopy. Transmission electron microscopy revealed the production of fine needles of hydroxyapatite in conjunction with matrix vesicles. Alkaline phosphatase activity did not increase in response to inflammatory mediators, but PPi production fell, reflecting lower ectonucleotide pyrophosphatase activity in cells and matrix vesicles. Because PPi is the major physiological inhibitor of mineralization, its decline generated permissive conditions for hydroxyapatite formation. This is in contrast to MSCs treated with dexamethasone, where PPi levels did not fall and mineralization was fuelled by a large and rapid increase in alkaline phosphatase activity. Bone sialoprotein was the only osteoblast marker strongly induced by IL-1ß; thus these cells do not become osteoblasts despite depositing abundant mineral. RT-PCR did not detect transcripts indicative of alternative mesenchymal lineages, including chondrocytes, myoblasts, adipocytes, ligament, tendon, or vascular smooth muscle cells. IL-1ß phosphorylated multiple MAPKs and activated nuclear factor-κB (NF-κB). Certain inhibitors of MAPK and PI3K, but not NF-κB, prevented mineralization. The findings are of importance to soft tissue mineralization, tissue engineering, and regenerative medicine.

The Beneficial Effects of Soluble Silicon Fertilizer in Dendrobium Orchids: Silicon-Augmented Resistance against Damage by Insect Pests and Fungal Pathogens.

The effects of soluble silicon fertilization on monocots and dicots have been widely studied. However, little is known regarding its effects on protecting epiphytes against insect and fungal pests. The efficacy of silicon fertilizer to reduce damage by thrips pest complexes, namely: Thrips palmi Karny, Frankliniella occidentalis Pergande, Chaetanaphothrips orchidii Moulton, and Chaetanaphothrips signipennis Bagnall (Thysanoptera: Thripidae), and the fungal pathogens: Botrytis cinerea Persoon (Helotiales: Sclerotiniaceae) and Fusarium spp. Link (Hypocreales: Nectriaceae) was examined during a nine-month greenhouse trial in Hawaii. The trial assessed yield, quality, and pest damage across three common varieties of dendrobiums. All replicates received additional soluble silicon fertilizer applications alternating weekly between soil drench and foliar (50 mg Si/plant) applications. Yield, quality, and spray length, pest damage, plant vigor, SPAD, and leaf temperature were measured. Data were analyzed using a generalized linear model (glm) with repeated measures followed by post-hoc pair-wise comparisons in R, version 4.3.1. Treatment effects were significant at p < 0.001 for the majority of the explanatory variables including: marketable yield, spray length, thrips damage, and fungal damage. Overall, the lavender variety ('Uniwai Supreme') benefited the most from silicon applications with a 73.0% increase in marketable yield, compared to the white variety ('Uniwai Mist'), which had an increase of 50.6% marketable sprays in contrast to its untreated control. Si benefits conferred to the purple variety ('Uniwai Royale') were intermediate to the lavender and white varieties. Although the magnitude of Si benefits varied among the varieties, all dendrobium varieties significantly benefited from silicon fertilization.

Blastomyces Induced Otomastoiditis With Local Soft Tissue Invasion: A Case Report.

The aim of this report is to document a very rare case of Blastomycosis dermatitidis mastoiditis with extension into the retromastoid soft tissue and surrounding muscle. Blastomycosis dermatitidis is a dimorphic fungus of endemic areas which classically infiltrates the lungs; however, dissemination presenting as otomastoiditis is exceedingly rare. The patient was an immunocompetent 27-year-old male with no significant preexisting health conditions. He had significant work exposure to dust and soil and was referred to our department for evaluation of otalgia with headaches, hearing loss, and intermittent facial paralysis. Initially, the extent of the infection was unknown. Based on extensive disease on magnetic resonance imaging, the patient was scheduled for urgent tympanoplasty and mastoidectomy. Postoperative treatment with itraconazole resolved any further manifestations and halted further soft tissue invasion. It is important to consider uncommon fungal infections in the workup of persistent otalgia, especially when presenting with facial paralysis and a history of environmental exposure to soil and dust. This type of infection should be considered regardless of immunodeficiency status. Early detection may prevent hearing loss and local invasion into surrounding structures.

A Pilot Study of the Effect of a Non-Contact Boxing Exercise Intervention on Respiratory Pressure and Phonation Aerodynamics in People with Parkinson's Disease.

This study investigated the effects of a non-contact boxing exercise program on maximum expiratory pressure and aerodynamic voice measurements. METHODS: Eight adult males diagnosed with Parkinson's disease participated in the study. Individuals participated in twice-weekly exercise classes lasting one hour across 12-months. Dependent variables were measured on three baseline days and then at six additional time points. A pressure meter acquired maximum expiratory pressure, and a pneumotachograph system acquired transglottal airflow and subglottal air pressure. RESULTS: Measures of average maximum expiratory pressure significantly increased after 9- and 12- months of exercise when compared to baseline. There was an increasing trend for these measures in all participants, with a corresponding large effect size. Measures of transglottal airflow and subglottal pressure did not change over the course of 9- or 12-months, although their stability may indicate that the exercise program influenced maintenance of respiratory-phonatory coordination during voicing. CONCLUSIONS: A non-contact boxing exercise program had a significant effect on maximum expiratory pressure in people with Parkinson's disease. The aerobic nature of the program and challenges to the respiratory muscles potentially explain the "ingredient" causing this effect. The small sample size of this pilot study necessitates future research incorporating larger and more diverse participants.

Versatility of mesenchymal stem cell-derived extracellular vesicles in tissue repair and regenerative applications.

Mesenchymal stem/stromal cells (MSCs) are multipotent somatic cells that have been widely explored in the field of regenerative medicine. MSCs possess the ability to secrete soluble factors as well as lipid bound extracellular vesicles (EVs). MSCs have gained increased interest and attention as a result of their therapeutic properties, which are thought to be attributed to their secretome. However, while the use of MSCs as whole cells pose heterogeneity concerns and survival issues post-transplantation, such limitations are absent in cell-free EV-based treatments. EVs derived from MSCs are promising therapeutic agents for a range of clinical conditions and disorders owing to their immunomodulatory, pro-regenerative, anti-inflammatory, and antifibrotic activity. Recent successes with preclinical studies using EVs for repair and regeneration of damaged tissues such as cardiac tissue, lung, liver, pancreas, bone, skin, cornea, and blood diseases are discussed in this review. We also discuss delivery strategies of EVs using biomaterials as delivery vehicles through systemic or local administration. Despite its effectiveness in preclinical investigations, the application of MSC-EV in clinical settings will necessitate careful consideration surrounding issues such as: i) scalability and isolation, ii) biodistribution, iii) targeting specific tissues, iv) quantification and characterization, and v) safety and efficacy of dosage. The future of EVs in regenerative medicine is promising yet still needs further investigation on enhancing the efficacy, scalability, and potency for clinical applications.

Nutrient Adequacy in Endurance Athletes.

Proper nutrition is critical for optimal performance in endurance athletes. However, it is unclear if endurance athletes are meeting all their energy and nutrient needs. We examined if endurance athletes are meeting their nutritional requirements and if this differed by sex. Ninety-five endurance athletes (n = 95; 50.5% men; 34.9 ± 12.9 y) participated in the study. Dietary intake was evaluated using the 24 h dietary recall method. Energy and nutrient intakes were calculated using the ESHA Food Processor Diet Analysis Software and compared against reference nutrient intakes. Endurance athletes did not consume the recommended amount of energy (76.8% of athletes), carbohydrates (95.8%), linoleic acid (75.8%), α-linolenic acid (ALA) (77.9%), eicosatetraenoic and docosahexaenoic acid (96.8%), dietary fiber (49.5%), vitamins D (93.7%), E (71.6%), and K (54.7%), folate (54.7%), pantothenic acid (70.5%), biotin (83.2%), manganese (58.9%), magnesium (56.8%), chromium (91.6%), molybdenum (93.7%), choline (85.3%), and potassium (56.8%), and consumed too much saturated fat (50.5%) and sodium (94.7%) than recommended. Fisher's Exact test showed that the requirements for dietary fiber (70.8% vs. 27.7%), ALA (87.5% vs. 68.1%), and total water (70.8% vs. 44.7%) were not met by more men versus women (p < 0.05). The needs for protein (70.2% vs. 25%) and vitamin B12 (46.8% vs. 22.9%) were not met by more women compared to men (p < 0.05). These findings need to be confirmed by a larger study.

Effect of Exercise Training on Lipoprotein Subclass Particle Concentrations and Sizes in Older Women: Results from a Randomized Controlled Trial.

BACKGROUND: Evidence suggests that lipoprotein subclass particles are critical markers of cardiovascular disease (CVD) risk. Older women have increased CVD risk related to age. The purpose of this study was to determine whether low and moderate doses of exercise influence lipoprotein subclasses. METHODS: Women (60-75 years) were randomized into groups for 16 weeks of moderate-intensity exercise training at a low or moderate dose (33.6 and 58.8 kJ/kg body weight weekly, respectively). Lipoprotein subclasses were determined by nuclear magnetic resonance spectroscopy before and after the training. RESULTS: The average weekly exercise duration was 109 and 164 min, for low- and moderate-dose groups, respectively. In the low-dose group, high-density lipoprotein particle (HDL-P) concentration decreased (Δ = -1.9 ± 3.1 µmol/L, mean ± SD, p = 0.002) and mean HDL-P size increased (Δ = 0.1 ± 0.3 nm, p = 0.028). In the moderate-dose group, mean HDL-P size (Δ = 0.1 ± 0.2 nm; p = 0.024) and low-density lipoprotein particle size increased (Δ = 0.4 ± 3.9 nm; p = 0.007). Baseline body mass index, peak oxygen consumption and age were associated with changes in a few lipoprotein subclasses. CONCLUSIONS: In this sample of inactive older women, moderate-intensity exercise training at a dose equivalent to or even lower than the minimally recommended level by public health agencies induced changes in lipoprotein subclasses in line with reduced CVD risk. However, higher doses are encouraged for greater health benefits.

The NAD salvage pathway in mesenchymal cells is indispensable for skeletal development in mice.

NAD is an essential co-factor for cellular energy metabolism and multiple other processes. Systemic NAD+ deficiency has been implicated in skeletal deformities during development in both humans and mice. NAD levels are maintained by multiple synthetic pathways but which ones are important in bone forming cells is unknown. Here, we generate mice with deletion of Nicotinamide Phosphoribosyltransferase (Nampt), a critical enzyme in the NAD salvage pathway, in all mesenchymal lineage cells of the limbs. At birth, NamptΔPrx1 exhibit dramatic limb shortening due to death of growth plate chondrocytes. Administration of the NAD precursor nicotinamide riboside during pregnancy prevents the majority of in utero defects. Depletion of NAD post-birth also promotes chondrocyte death, preventing further endochondral ossification and joint development. In contrast, osteoblast formation still occurs in knockout mice, in line with distinctly different microenvironments and reliance on redox reactions between chondrocytes and osteoblasts. These findings define a critical role for cell-autonomous NAD homeostasis during endochondral bone formation.

In Vivo Optical Characterization of Middle Ear Effusions and Biofilms During Otitis Media.

Otitis media (OM), a common ear infection, is characterized by the presence of an accumulated middle ear effusion (MEE) in a normally air-filled middle ear cavity. While assessing the MEE plays a critical role in the overall management of OM, identifying and examining the MEE is challenging with the current diagnostic tools since the MEE is located behind the semi-opaque eardrum. The objective of this cross-sectional, observational study is to non-invasively visualize and characterize MEEs and bacterial biofilms in the middle ear. A portable, handheld, otoscope-integrated optical coherence tomography (OCT) system combined with novel analytical methods has been developed. In vivo middle ear OCT images were acquired from 53 pediatric subjects (average age of 3.9 years; all awake during OCT imaging) diagnosed with OM and undergoing a surgical procedure (ear tube surgery) to aspirate the MEE and aerate the middle ear. In vivo middle ear OCT acquired prior to the surgery was compared with OCT of the freshly extracted MEEs, clinical diagnosis, and post-operative evaluations. Among the subjects who were identified with the presence of MEEs, 89.6% showed the presence of the TM-adherent biofilm in in vivo OCT. This study provides an atlas of middle ear OCT images exhibiting a range of depth-resolved MEE features, which can only be visualized and assessed non-invasively through OCT. Quantitative metrics of OCT images acquired prior to the surgery were statistically correlated with surgical evaluations of MEEs. Measurements of MEE characteristics will provide new readily available information that can lead to improved diagnosis and management strategies for the highly prevalent OM in children.

High-affinity binding to the SARS-CoV-2 spike trimer by a nanostructured, trivalent protein-DNA synthetic antibody.

Multivalency enables nanostructures to bind molecular targets with high affinity. Although antibodies can be generated against a wide range of antigens, their shape and size cannot be tuned to match a given target. DNA nanotechnology provides an attractive approach for designing customized multivalent scaffolds due to the addressability and programmability of the nanostructure shape and size. Here, we design a nanoscale synthetic antibody ("nano-synbody") based on a three-helix bundle DNA nanostructure with one, two, or three identical arms terminating in a mini-binder protein that targets the SARS-CoV-2 spike protein. The nano-synbody was designed to match the valence and distance between the three receptor binding domains (RBDs) in the spike trimer, in order to enhance affinity. The protein-DNA nano-synbody shows tight binding to the wild-type, Delta, and several Omicron variants of the SARS-CoV-2 spike trimer, with affinity increasing as the number of arms increases from one to three. The effectiveness of the nano-synbody was also verified using a pseudovirus neutralization assay, with the three-arm nanostructure inhibiting two Omicron variants against which the structures with only one or two arms are ineffective. The structure of the three-arm nano-synbody bound to the Omicron variant spike trimer was solved by negative-stain transmission electron microscopy reconstruction, and shows the protein-DNA nanostructure with all three arms attached to the RBD domains, confirming the intended trivalent attachment. The ability to tune the size and shape of the nano-synbody, as well as its potential ability to attach two or more different binding ligands, will enable the high-affinity targeting of a range of proteins not possible with traditional antibodies.

Zanubrutinib in patients with previously treated B-cell malignancies intolerant of previous Bruton tyrosine kinase inhibitors in the USA: a phase 2, open-label, single-arm study.

BACKGROUND: We hypothesised that zanubrutinib, a highly selective next-generation Bruton tyrosine kinase (BTK) inhibitor, would be a safe and active treatment for patients intolerant of ibrutinib, acalabrutinib, or both. We aimed to assess whether zanubrutinib would prolong treatment duration by minimising treatment-related toxicities and discontinuations in patients with previously treated B-cell malignancies. METHODS: This ongoing, phase 2, multicentre, open-label, single-arm study was done in 20 centres in the USA. Patients aged 18 or older with previously treated B-cell malignancies (chronic lymphocytic leukaemia, small lymphocytic lymphoma, mantle cell lymphoma, Waldenström macroglobulinaemia, or marginal zone lymphoma) who became intolerant of ibrutinib, acalabrutinib, or both, were orally administered zanubrutinib 160 mg twice daily or 320 mg once daily per investigator. The primary endpoint was recurrence and change in severity of ibrutinib or acalabrutinib intolerance events based on investigator-assessed adverse events. Secondary endpoints were investigator-assessed overall response rate; duration of response; disease control rate; and progression-free survival. Analyses included all patients who received any dose of the study drug. This study is registered with ClinicalTrials.gov, NCT04116437. FINDINGS: Between Oct 14, 2019, and Sept 8, 2021, 67 patients (36 [54%] men and 31 [46%] women) who were intolerant of ibrutinib (n=57; cohort 1) or of acalabrutinib or acalabrutinib and ibrutinib (n=10; cohort 2) were enrolled. 63 (94%) patients were White, one (2%) had multiple ethnicities, and three (5%) had unreported or unknown ethnicity. Most intolerance events (81 [70%] of 115 for ibrutinib; 15 [83%] of 18 for acalabrutinib) did not recur with zanubrutinib. Of the recurring events, seven (21%) of 34 ibrutinib intolerance events and two (67%) of three acalabrutinib intolerance events recurred at the same severity with zanubrutinib; 27 (79%) ibrutinib intolerance events and one (33%) acalabrutinib intolerance event recurred at a lower severity with zanubrutinib. No events recurred at higher severity. No grade 4 intolerance events recurred. 64 (96%) of 67 patients had one or more adverse events with zanubrutinib; the most common adverse events were contusion (in 15 [22%] of 67 patients), fatigue (14 [21%]), myalgia (ten [15%]), arthralgia (nine [13%]), and diarrhoea (nine [13%]). Atrial fibrillation occurred in three (4%) patients (all grade 2). Eight (12%) of 67 patients had serious adverse events (anaemia, atrial fibrillation, bronchitis, COVID-19, COVID-19 pneumonia, febrile neutropenia, salmonella gastroenteritis, transfusion reaction, trigeminal nerve disorder, and urinary tract infection). No treatment-related deaths occurred. The median follow-up time was 12·0 months (IQR 8·2-15·6). Among the 64 efficacy-evaluable patients, disease control rate was 93·8% (60; 95% CI 84·8-98·3) and overall response rate was 64·1% (41; 95% CI 51·1-75·7). The median duration of response was not reached; the 12-month event-free duration of response rate was 95·0% (95% CI 69·5-99·3). Similarly, median progression-free survival was not reached; 18-month progression-free survival was 83·8% (95% CI 62·6-93·6). INTERPRETATION: Patients intolerant of previous BTK inhibitors have limited treatment options. These results suggest that zanubrutinib, a safe and viable treatment for patients with B-cell malignancies, might fill that unmet need for those who exhibit intolerance to ibrutinib or acalabrutinib. FUNDING: BeiGene.

Improvement of tendon repair using muscle grafts transduced with TGF-ß1 cDNA.

Tendon rupture is a common injury. Inadequate endogenous repair often leaves patients symptomatic, with tendons susceptible to re-rupture. Administration of certain growth factors improves tendon healing in animal models, but their delivery remains a challenge. Here we evaluated the delivery of TGF-ß1 to tendon defects by the implantation of genetically modified muscle grafts. Rat muscle biopsies were transduced with recombinant adenovirus encoding TGF-ß1 and grafted onto surgically transected Achilles tendons in recipient animals. Tissue regenerates were compared to those of controls by biomechanical testing as well as histochemical and immunohistochemical analyses. Healing was greatly accelerated when genetically modified grafts were implanted into tendon defects, with the resulting repair tissue gaining nearly normal histological appearance as early as 2 weeks postoperatively. This was associated with decreased deposition of type III collagen in favour of large fibre bundles indicative of type I collagen. These differences in tendon composition coincided with accelerated restoration of mechanical strength. Tendon thickness increased in gene-treated animals at weeks 1 and 2, but by week 8 became significantly lower than that of controls suggesting accelerated remodelling. Thus localised TGF-ß1 delivery via adenovirus-modified muscle grafts improved tendon healing in this rat model and holds promise for clinical application.