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Background Chronic limb-threatening ischaemia with ischaemic pain and/or tissue loss. Objective To examine the clinical and cost-effectiveness of a vein bypass-first compared to a best endovascular treatment-first revascularisation strategy in preventing major amputation or death. Design Superiority, open, pragmatic, multicentre, phase III randomised trial. Setting Thirty-nine vascular surgery units in the United Kingdom, and one each in Sweden and Denmark. Participants Patients with chronic limb-threatening ischaemia due to atherosclerotic peripheral arterial disease who required an infra-popliteal revascularisation, with or without an additional more proximal infra-inguinal revascularisation procedure, to restore limb perfusion. Interventions A vein bypass-first or a best endovascular treatment-first infra-popliteal, with or without an additional more proximal infra-inguinal revascularisation strategy. Main outcome measures The primary outcome was amputation-free survival. Secondary outcomes included overall survival, major amputation, further revascularisation interventions, major adverse limb event, health-related quality of life and serious adverse events. Methods Participants were randomised to a vein bypass-first or a best endovascular treatment-first revascularisation strategy. The original sample size of 600 participants (247 events) was based on a hazard ratio of 0.66 with amputation-free survival rates of 0.72, 0.62, 0.53, 0.47 and 0.35 in years 1–5 in the best endovascular treatment-first group with 90% power and alpha at p = 0.05. The sample size was revised to an event-based approach as a result of increased follow-up time due to slower than anticipated recruitment rates. Participants were followed up for a minimum of 2 years. A cost-effectiveness analysis was employed to estimate differences in total hospital costs and amputation-free survival between the groups. Additionally, a cost–utility analysis was carried out and the total cost and quality-adjusted life-years, 2 and 3 years after randomisation were used. Results Between 22 July 2014 and 30 November 2020, 345 participants were randomised, 172 to vein bypass-first and 173 to best endovascular treatment-first. Non-amputation-free survival occurred in 108 (63%) of 172 patients in the vein bypass-first group and 92 (53%) of 173 patients in the best endovascular treatment-first group [adjusted hazard ratio 1.35 (95% confidence interval 1.02 to 1.80); p = 0.037]. Ninety-one (53%) of 172 patients in the vein bypass-first group and 77 (45%) of 173 patients in the best endovascular treatment-first group died [adjusted hazard ratio 1.37 (95% confidence interval 1.00 to 1.87)]. Over follow-up, the economic evaluation discounted results showed that best endovascular treatment-first was associated with £1690 less hospital costs compared to vein bypass-first. The cost utility analysis showed that compared to vein bypass-first, best endovascular treatment-first was associated with £224 and £2233 less discounted hospital costs and 0.016 and 0.085 discounted quality-adjusted life-year gain after 2 and 3 years from randomisation. Limitations Recruiting patients to the Bypass versus Angioplasty in Severe Ischaemia of the Leg Trial-2 trial was difficult and the target number of events was not achieved. Conclusions A best endovascular treatment-first revascularisation strategy was associated with better amputation-free survival, which was largely driven by fewer deaths. Overall, the economic evaluation results suggest that best endovascular treatment-first dominates vein bypass-first in the cost-effectiveness analysis and cost–utility analysis as it was less costly and more effective than a vein bypass-first strategy. Future work The Bypass versus Angioplasty in Severe Ischaemia of the Leg Trial-2 investigators have a data sharing agreement with the BEst Surgical Therapy in patients with Chronic Limb threatening Ischaemia investigators. One output of this collaboration will be an individual patient data meta-analysis. Study registration Current Controlled Trials ISRCTN27728689. Funding This award was funded by the National Institute for Health and Care Research (NIHR) Health Technology Assessment programme (NIHR award ref: 12/35/45) and is published in full in Health Technology Assessment; Vol. 28, No. 65. See the NIHR Funding and Awards website for further award information. Plain language summary Atherosclerosis, or narrowing of the arteries, can occur as a result of smoking, high blood pressure, diabetes, or high cholesterol in the blood. Atherosclerosis can affect any artery, including those supplying the legs, where the condition is called peripheral arterial disease. The most severe form of peripheral arterial disease is chronic limb-threatening ischaemia which can cause severe pain in the foot as well as ulcers and gangrene. Unless the blood supply to the leg and foot is improved, by a process called revascularisation, people with chronic limb-threatening ischaemia are at high risk of amputation and death. The blood supply can be improved by using a vein from the leg to bypass around the blockages (vein bypass) or by using a balloon (angioplasty) or small metal tubes (stents) to reopen the blocked arteries (best endovascular treatment). There is debate about which type of revascularisation is best in terms of preventing amputation and death, especially in people who need revascularisation of the arteries below the knee. Bypass versus Angioplasty in Severe Ischaemia of the Leg Trial-2 is the first randomised controlled trial to compare vein bypass-first and best endovascular treatment-first in this group of patients. Bypass versus Angioplasty in Severe Ischaemia of the Leg Trial-2 found that people randomised to a vein bypass-first revascularisation strategy were 35% more likely to require a major amputation or die than those randomised to a best endovascular treatment-first strategy. Most of this difference in favour of best endovascular treatment-first was due to a higher number of patients dying in the vein bypass-first group. Best endovascular treatment-first was also cheaper for the National Health Service. The results of this study suggest that in patients with chronic limb-threatening ischaemia due to peripheral arterial disease in the arteries below the knee, who are suitable for both vein bypass and best endovascular treatment and where there is uncertainty as to which is best, best endovascular treatment should be offered first rather than vein bypass. Scientific summary Background Chronic limb-threatening ischaemia (CLTI) is the severest manifestation of peripheral arterial disease (PAD) and presents with ischaemic pain at rest or tissue loss (ulceration, gangrene or both) or both. The researchers compared the effectiveness of a vein bypass (VB)-first with a best endovascular treatment (BET)-first revascularisation strategy in terms of preventing major amputation and death in patients with CLTI who required an infra-popliteal (IP) revascularisation, with or without an additional more proximal infra-inguinal revascularisation procedure, to restore limb perfusion. Objectives The primary objective of the study was to examine the effectiveness and cost-effectiveness of using a VB-first compared to a BET-first revascularisation strategy in terms of preventing major (above the ankle) amputation or death from any cause [amputation-free survival (AFS)] in patients with CLTI who required an IP, with or without an additional more proximal infra-inguinal revascularisation procedure, to restore limb perfusion. Design Superiority, open-label, pragmatic, multicentre, phase III randomised trial. Setting Thirty-nine vascular surgery units in the United Kingdom, and one each in Sweden and Denmark. Participants Those with CLTI due to atherosclerotic PAD and who required an IP revascularisation, with or without an additional more proximal infra-inguinal revascularisation procedure, to restore limb perfusion. Interventions A VB-first or a BET-first IP, with or without an additional more proximal infra-inguinal revascularisation strategy, to restore limb perfusion. Most VBs were constructed with great saphenous vein and originated from the common or superficial femoral arteries. Most endovascular interventions comprised plain balloon angioplasty with selective use of plain or drug-eluting stents. Outcome measures The primary outcome was AFS defined as time to first major (above the ankle) amputation or death from any cause. Secondary outcomes included: time to death from any cause (overall survival); time to major amputation of the trial leg; major adverse limb event (defined as major amputation of the trial leg, or any further major revascularisation intervention to the trial leg, following the first revascularisation intervention); major adverse cardiac event (defined as chronic limb-threatening ischaemia and/or major amputation affecting the non-trial leg, myocardial infarction, transient ischaemic attack or stroke); 30-day morbidity and mortality; relief of ischaemic pain as determined by visual analogue scale and opiate usage; health- related quality of life using generic [EuroQol-5 Dimensions, five-level version (EQ-5D-5L), Short Form questionnaire-12 items, ICEpop CAPability measure for Older people] and disease-specific (the Vascular Quality of Life Questionnaire) tools; further major revascularisation intervention to the trial leg (following the first revascularisation intervention); re-intervention and crossover intervention (where re-intervention is defined as the same, and a crossover procedure is defined as an alternative, revascularisation procedure to the first revascularisation procedure post-randomisation); healing of tissue loss (ulcers, gangrene) at or below the ankle presumed to be caused by PAD as assessed by the perfusion, extent, depth, infection and sensation score, the Wound Ischaemia and foot Infection tool; and haemodynamic measurements (ankle–brachial pressure index, and toe brachial pressure index). Serious adverse events were collected up to 30 days post first revascularisation. Economic evaluation analyses in the form of cost-effectiveness and cost–utility analysis (CUA) were conducted from the perspective of the UK NHS alongside the trial. The base case analyses considered only hospital costs and an additional scenario and subgroup analyses were carried out. Sample size The original sample size was based on a time-to-event analysis to be undertaken 2 years after completion of recruitment. It was anticipated that recruitment would take place over 3 years: 20% of patients recruited in year 1, 40% in year 2 and 40% in year 3. Based on the Bypass versus Angioplasty in Severe Ischaemia of the Leg Trial (BASIL-1) trial. AFS rates were assumed to be 0.72 in year 1, 0.62 in year 2, 0.53 in year 3, 0.47 in year 4 and 0.35 in year 5. Allowing for a 10% attrition rate and based on the survival estimates calculated using the BASIL-1 data, a population of 600 participants (247 primary outcome events) would have 90% power to detect a reduction in AFS of one-third [hazard ratio (HR) 0.66] at the 5% significance level. The initial assumptions made in this trial concerning recruitment rates were not achieved; therefore, recruitment continued beyond year 3. As a result, the median length of follow-up was longer than originally planned. Therefore, the number of randomised participants required to observe 247 events (as per the original sample size target) was reduced due to the increased exposure time. With support of the funder and independent oversight from the Data Monitoring Committee, recruitment rates, length of follow-up, and pooled event rates over time were modelled to predict the number of participants needed to reach 247 events, with 24 months minimum follow-up in each participant. The modelling was updated approximately every 6 months based on emerging data. Results Between 22 July 2014 and 30 November 2020, 345 participants were randomised, 172 to the VB-first group and 173 to the BET-first group. The baseline characteristics of the two groups were similar. Major amputation or death occurred in 108 (63%) of 172 patients in the VB-first group and 92 (53%) of 173 patients in the BET-first group {adjusted HR 1.35 [95% confidence interval (CI) 1.02 to 1.80]; p = 0.037}. Ninety-one (53%) of 172 patients in the VB-first group and 77 (45%) of 173 patients in the BET-first group died [adjusted HR 1.37 (95% CI 1.00 to 1.87)]. In both groups the commonest causes of morbidity and mortality, including those occurring within 30 days of their first revascularisation, were cardiovascular and respiratory events. Over follow-up, the economic evaluation results showed that BET-first was associated with £1690 less hospital costs compared to VB-first with additional survival without amputation. The cost utility analysis showed that compared to VB-first, BET-first was associated with £2524 and £2233 less hospital costs and 0.016 and 0.085 QALY gain after 2 and 3 years from randomisation. Conclusions In the BASIL-2 trial, a BET-first revascularisation strategy was associated with a better AFS, which was largely driven by fewer deaths in the BET-first group. In both the CUA and cost-effectiveness analysis (CEA) analyses and from a NHS and societal perspective BET-first dominated VB-first and is therefore a highly cost-effective intervention for the NHS. These data suggest a greater role for BET in the management of patients with CLTI who require an IP revascularisation to restore limb perfusion. In patients with CLTI who required an IP, with or without an additional more proximal infra-inguinal revascularisation procedure, to restore limb perfusion, a BET-first revascularisation strategy was associated with reduced hospital costs (£1690) and improved AFS (0.429 years), out to 7 years following randomisation. BET-first therefore dominated VB-first in the CEA. Similarly, in the health-related quality of life the CUA, BET-first was cost-saving with improved QALYs (£2524 and £2233 less hospital costs and 0.016 and 0.085 more QALYs at 2 and 3 years, respectively) and so dominated VB-first. This economic analysis therefore shows that BET-first is a cost-effective option from an NHS and societal perspective. The sensitivity analysis supported the base-case analysis and BET-first was found to be cost-effective at different willingness-to-pay thresholds. Similar findings were found in all other scenario analyses when considering costs of primary and other hospital healthcare services, taking a broader societal perspective, which includes out-of-pocket expenditure and the costs associated with productivity loss, patient’s adherence to study protocol, the impact of imputation by focusing on participants with complete hospital cost and EQ-5D-5L data only. However, these findings should be interpreted cautiously given the large number of imputed cost values and the substantial probability of a very small QALY difference. Study registration Current Controlled Trials ISRCTN27728689. Funding This award was funded by the National Institute for Health and Care Research (NIHR) Health Technology Assessment programme (NIHR award ref: 12/35/45) and is published in full in Health Technology Assessment; Vol. 28, No. 65. See the NIHR Funding and Awards website for further award information.

Purpose/objective(s)Bridging radiation therapy (bRT) is increasingly being utilized prior to chimeric antigen receptor (CAR) T-cell therapy for large B-cell lymphoma (LBCL). It is unknown how the extent of cytoreduction during bRT impacts outcomes.Materials/methodsWe retrospectively reviewed patients with LBCL treated with bRT followed by CAR T-cell therapy. Metabolic tumor volume (MTV), maximum standardized uptake value (SUVmax), SUVmean, and total lesion glycolysis (TLG) were extracted from F18-fluorodeoxyglucose positron emission tomography (PET) scans acquired prior to bRT and between completion of bRT and CAR T-cell infusion. Delta radiomics based on changes of these values were then calculated. The association between delta radiomics and oncologic outcomes [progression-free survival (PFS), freedom from distant progression (FFDP), and local control (LC)] were then examined.ResultsThirty-three sites across 23 patients with LBCL were irradiated. All metabolically active disease was treated in 10 patients. Following bRT, median overall decreases (including unirradiated sites) in MTV, SUVmax, SUVmean, and TLG were 22.2 cc (63.1%), 8.9 (36.8%), 3.4 (31.1%), and 297.9 cc (75.8%), respectively. Median decreases in MTV, SUVmax, SUVmean, and TLG in irradiated sites were 15.6 cc (91.1%), 17.0 (74.6%), 6.8 (55.3%), and 157.0 cc (94.6%), respectively. Median follow-up was 15.2 months. A decrease in SUVmax of at least 54% was associated with improved PFS (24-month PFS: 83.3% vs. 28.1%; p = 0.037) and FFDP (24-month FFDP: 100% vs. 62.4%; p < 0.001). A decrease in MTV of at least 90% was associated with improved FFDP (24-month FFDP: 100% vs. 62.4%; p < 0.001). LC was improved in sites with decreases in SUVmax of at least 71% (24-month LC: 100% vs. 72.7%; p < 0.001). Decreases of MTV by at least 90% (100% vs. 53.3%; p = 0.038) and TLG by at least 95% (100% vs. 56.3%; p = 0.067) were associated with an improved complete response rate.ConclusionbRT led to substantial reductions in MTV, SUVmax, SUVmean, and TLG. The relative extent of these decreases correlated with improved outcomes after CAR T-cell infusion. Prospective cohorts should validate the value of interim PET following bRT for quantifying changes in disease burden and associated prognosis.

Patients with relapsed/refractory (R/R) transformed diffuse large B-cell lymphoma (DLBCL) from indolent B-cell lymphomas, including Richter transformation (RT), have a poor prognosis. PD-1/PD-L1 antibodies produce modest objective and complete response rates (ORR and CRR) in B-NHL as monotherapy but may synergize with immunogenic chemotherapies like gemcitabine and oxaliplatin (GemOx). Thus, we evaluated the safety and efficacy of atezolizumab plus rituximab and GemOx (R-GemOx+Atezo) in R/R transformed DLBCL, including RT. We conducted a phase I trial including patients with transformed DLBCL after ≥1 prior therapy. Patients received up to 4 cycles of R-GemOx-+Atezo. Patients in CR could then proceed to Ratezo maintenance until progression. A safety lead-in with dose-limiting toxicity (DLT) evaluation was enrolled to confirm the recommended phase 2 dose (RP2D), followed by 2 expansion cohorts: one for transformed follicular lymphoma (FL) and another for non-FL transformed DLBCL, including RT. Twenty-seven patients were enrolled. One of the 6 safety lead-in patients had a DLT attributed to atezolizumab, a grade 4 Stevens-Johnson syndrome (SJS). The most common grade ≥3 events were neutropenia (18.5%), lymphopenia (18.5%), and thrombocytopenia (14.8%). The overall and complete response rates (ORR and CRR) were 59% and 33%, respectively. The ORR and CRR in transformed FL were 79% and 43%, and 38% and 23% in transformed non-FL, respectively. The median PFS and OS of the total population were 4.2 and 7.7 months, respectively. R-GemOx+Atezo was well tolerated and demonstrated promising preliminary efficacy in patients with R/R transformed DLBCL.

Aberrant expression of the double homeobox 4 (DUX4) gene in skeletal muscle predominantly drives the pathogenesis of facioscapulohumeral muscular dystrophy (FSHD). We recently demonstrated that berberine, an herbal extract known for its ability to stabilize guanine–quadruplex structures, effectively downregulates DUX4 expression in FSHD patient-derived myoblasts and in mice overexpressing exogenous DUX4 after viral vector-based treatment. Here, we sought to confirm berberine’s inhibitory efficacy on DUX4 in the widely used FSHD-like transgenic mouse model, ACTA1-MCM/FLExDUX4, where DUX4 is induced at pathogenic levels using tamoxifen. Animals repeatedly treated with berberine via intraperitoneal injections for 4 weeks exhibited significant reductions in both mRNA and protein levels of DUX4, and in mRNA expression of murine DUX4-related genes. This inhibition translated into improved forelimb muscle strength and positive alterations in important FSHD-relevant cellular pathways, although its impact on muscle mass and histopathology was less pronounced. Collectively, our data confirm the efficacy of berberine in downregulating DUX4 expression in the most relevant FSHD mouse model. However, further optimization of dosing regimens and new studies to enhance the bioavailability of berberine in skeletal muscle are warranted to fully leverage its therapeutic potential for FSHD treatment.

Periostin, a multifunctional 90 kDa protein, plays a pivotal role in the pathogenesis of fibrosis across various tissues, including skeletal muscle. It operates within the transforming growth factor beta 1 (Tgf-β1) signalling pathway and is upregulated in fibrotic tissue. Alternative splicing of Periostin’s C-terminal region leads to six protein-coding isoforms. This study aimed to elucidate the contribution of the isoforms containing the amino acids encoded by exon 17 (e17+ Periostin) to skeletal muscle fibrosis and investigate the therapeutic potential of manipulating exon 17 splicing. We identified distinct structural differences between e17+ Periostin isoforms, affecting their interaction with key fibrotic proteins, including Tgf-β1 and integrin alpha V. In vitro mouse fibroblast experimentation confirmed the TGF-β1-induced upregulation of e17+ Periostin mRNA, mitigated by an antisense approach that induces the skipping of exon 17 of the Postn gene. Subsequent in vivo studies in the D2.mdx mouse model of Duchenne muscular dystrophy (DMD) demonstrated that our antisense treatment effectively reduced e17+ Periostin mRNA expression, which coincided with reduced full-length Periostin protein expression and collagen accumulation. The grip strength of the treated mice was rescued to the wild-type level. These results suggest a pivotal role of e17+ Periostin isoforms in the fibrotic pathology of skeletal muscle and highlight the potential of targeted exon skipping strategies as a promising therapeutic approach for mitigating fibrosis-associated complications.

Abstract Signal regulatory protein alpha (SIRPα) is the receptor for cluster of differentiation (CD)47, a potent “don't eat me” signal for macrophages. Disruption of CD47‐SIRPα signaling in the presence of prophagocytic signals can lead to enhanced phagocytosis of tumor cells, resulting in a direct antitumor effect; agents targeting this pathway have shown efficacy in non‐Hodgkin lymphoma (NHL) and other tumor types. GS‐0189 is a novel anti‐SIRPα humanized monoclonal antibody. Here we report: (1) clinical safety, preliminary activity, and pharmacokinetics of GS‐0189 as monotherapy and in combination with rituximab from a phase 1 clinical trial in patients with relapsed/refractory NHL (NCT04502706, SRP001); (2) in vitro characterization of GS‐0189 binding to SIRPα; and (3) in vitro phagocytic activity. Clinically, GS‐0189 was well tolerated in patients with relapsed/refractory NHL with evidence of clinical activity in combination with rituximab. Receptor occupancy (RO) of GS‐0189 was highly variable in NHL patients; binding affinity studies showed significantly higher affinity for SIRPα variant 1 than variant 2, consistent with RO in patient and healthy donor samples. In vitro phagocytosis induced by GS‐0189 was also SIRPα variant–dependent. Although clinical development of GS‐0189 was discontinued, the CD47‐SIRPα signaling pathway remains a promising therapeutic target and should continue to be explored.

PurposeBased on the potential for ipilimumab (I) to augment T-cell activation, we hypothesize that ipilimumab would augment the efficacy of rituximab (R) in patients with relapsed/refractory (R/R) CD20+non-Hodgkin's lymphoma (NHL). This phase I study aimed to identify a recommended phase 2 dose, document toxicities, and preliminarily assess efficacy and potential predictive biomarkers.Patients and methodsThirty-three patients with R/R CD20+B-cell lymphoma received R at 375 mg/m2weekly for 4 weeks and I at 3 mg/kg on day 1 and every 3 weeks for four doses. Responding patients went on to maintenance with each agent given every 12 weeks. To facilitate correlative analysis, the expansion phase randomized patients to simultaneous R+I versus R with I delayed 2 weeks.ResultsToxicity was manageable; no dose-limiting toxicity was observed at the doses studied. When considering the entire cohort, efficacy was modest, with an objective response rate (ORR) of 24% and median progression-free survival (PFS) of 2.6 months. However, in follicular lymphoma patients, the ORR was 58% with a median PFS of 5.6 months. The randomized comparison of R with R+I demonstrated that R+I resulted in more effective B-cell depletion (BCD). Both B-cell depletion and the ratio of CD45RA-regulatory T cell (Treg) to Treg were associated with response at all time points.ConclusionsThe combination of R+I has manageable toxicity and encouraging efficacy in R/R follicular lymphoma. The ratio of CD45RA-Tregs to total Tregs, and peripheral BCD should be studied further as potential predictors of response.

Skin inflammation is often accompanied by systemic disease, yet the pathways that regulate this escalation are little known. Here authors show that transgenic expression of human CD1a in mice leads to the escalation of experimental skin inflammation and systemic inflammatory disease, and the generalized symptoms could be alleviated by blocking antibodies developed against CD1a.

Outcomes after programmed death-1 (PD-1) blockade in B-cell lymphomas are disappointing with few durable responses. Histone deacetylase inhibitors exhibit favorable immunomodulatory effects and demonstrate synergistic anti-tumor immune responses with anti-PD-1 therapy in preclinical models. We, therefore, developed a phase I study to evaluate the safety and preliminary efficacy of pembrolizumab with vorinostat in relapsed/refractory B-cell lymphomas. Patients were treated in a dose-escalation cohort using a Rolling 6 design followed by an expansion cohort at the recommended phase II dose (R2PD). Fifty-two patients were enrolled (32 Hodgkin and 20 non-Hodgkin lymphoma [NHL]). Here, we report safety data from the dose escalation cohort, and the toxicity and efficacy within NHL patients. Vorinostat was administered twice daily on days 1-5 and 8-12 (dose-level [DL]1: 100 mg; DL2: 200 mg) and pembrolizumab (200 mg) was administered on day 1 of each 3-week cycle. Of six patients treated at DL1, one had a dose-limiting toxicity (DLT) (Stevens-Johnson syndrome [SJS]), and one of six had a DLT at DL2 (thromboembolism); therefore, DL2 was the RP2D. The patient developing SJS was treated with corticosteroids, infliximab, and cyclosporine but ultimately died of invasive fungal infection from the extensive immunosuppression used to treat the SJS. The most common adverse events were hypertension, diarrhea, and cytopenias. Of 20 NHL patients, nine had follicular lymphoma (FL) and 11 had diffuse large B-cell lymphoma (DLBCL). Five DLBCL patients had primary mediastinal B-cell lymphoma (PMBL). The complete and overall response rates (CR and ORR) were 11% and 22% for FL and 45% and 55% for all DLBCL. Amongst DLBCL, the CR and ORR was 80% and 80% for PMBL and 17% and 33% for non-PMBL. In conclusion, pembrolizumab with vorinostat was tolerable and produced responses in relapsed/refractory B-cell NHL, with particularly notable efficacy in PMBL (clinicaltrials gov. Identifier: NCT03150329).

Duchenne muscular dystrophy (DMD) is characterised by fibrotic tissue deposition in skeletal muscle. We assessed the role of periostin in fibrosis using mdx mice, an established DMD murine model, for which we conducted a thorough examination of periostin expression over a year. RNA and protein levels in diaphragm (DIA) muscles were assessed and complemented by a detailed histological analysis at 5 months of age. In dystrophic DIAs, periostin (Postn) mRNA expression significantly exceeded that seen in wildtype controls at all timepoints analysed, with the highest expression at 5 months of age (p < 0.05). We found Postn to be more consistently highly expressed at the earlier timepoints compared to established markers of fibrosis like transforming growth factor-beta 1 (Tgf-β1) and connective tissue growth factor (Ctgf). Immunohistochemistry confirmed a significantly higher periostin protein expression in 5-month-old mdx mice compared to age-matched healthy controls (p < 0.01), coinciding with a significant fibrotic area percentage (p < 0.0001). RT-qPCR also indicated an elevated expression of Tgf-β1, Col1α1 (collagen type 1 alpha 1) and Ctgf in mdx DIAs compared to wild type controls (p < 0.05) at 8- and 12-month timepoints. Accordingly, immunoblot quantification demonstrated elevated periostin (3, 5 and 8 months, p < 0.01) and Tgf-β1 (8 and 12 months, p < 0.001) proteins in the mdx muscle. These findings collectively suggest that periostin expression is a valuable marker of fibrosis in this relevant model of DMD. They also suggest periostin as a potential contributor to fibrosis development, with an early onset of expression, thereby offering the potential for timely therapeutic intervention and its use as a biomarker in muscular dystrophies.

ObjectiveTo determine patterns of respiratory medications used in neonatal intensive care unit graduates.Study designThe Prematurity Respiratory Outcomes Program enrolled 835 babies

We investigated the feasibility of utilizing an exon-skipping approach as a genotype-dependent therapeutic for neurofibromatosis type 1 (NF1) by determining which NF1 exons might be skipped while maintaining neurofibromin protein expression and GTPase activating protein (GAP)-related domain (GRD) function. Initial in silico analysis predicted exons that can be skipped with minimal loss of neurofibromin function, which was confirmed by in vitro assessments utilizing an Nf1 cDNA-based functional screening system. Skipping of exons 17 or 52 fit our criteria, as minimal effects on protein expression and GRD activity were noted. Antisense phosphorodiamidate morpholino oligomers (PMOs) were utilized to skip exon 17 in human cell lines with patient-specific pathogenic variants in exon 17, c.1885G>A, and c.1929delG. PMOs restored functional neurofibromin expression. To determine the in vivo significance of exon 17 skipping, we generated a homozygous deletion of exon 17 in a novel mouse model. Mice were viable and exhibited a normal lifespan. Initial studies did not reveal the presence of tumor development; however, altered nesting behavior and systemic lymphoid hyperplasia was noted in peripheral lymphoid organs. Alterations in T and B cell frequencies in the thymus and spleen were identified. Hence, exon skipping should be further investigated as a therapeutic approach for NF1 patients with pathogenic variants in exon 17, as homozygous deletion of exon 17 is consistent with at least partial function of neurofibromin.

Duchenne muscular dystrophy (DMD) is a muscle wasting disorder caused by mutations in the DMD gene. Restoration of full-length dystrophin protein in skeletal muscle would have therapeutic benefit, but lentivirally mediated delivery of such a large gene in vivo has been hindered by lack of tissue specificity, limited transduction, and insufficient transgene expression. To address these problems, we developed a lentiviral vector, which contains a muscle-specific promoter and sequence-optimized full-length dystrophin, to constrain dystrophin expression to differentiated myotubes/myofibers and enhance the transgene expression. We further explored the efficiency of restoration of full-length dystrophin in vivo, by grafting DMD myoblasts that had been corrected by this optimized lentiviral vector intramuscularly into an immunodeficient DMD mouse model. We show that these lentivirally corrected DMD myoblasts effectively reconstituted full-length dystrophin expression in 93.58% ± 2.17% of the myotubes in vitro. Moreover, dystrophin was restored in 64.4% ± 2.87% of the donor-derived regenerated muscle fibers in vivo, which were able to recruit members of the dystrophin-glycoprotein complex at the sarcolemma. This study represents a significant advance over existing cell-mediated gene therapy strategies for DMD that aim to restore full-length dystrophin expression in skeletal muscle.

Purpose/objective(s): Bridging radiation therapy (bRT) is increasingly being utilized prior to chimeric antigen receptor (CAR) T-cell therapy for large B-cell lymphoma (LBCL). It is unknown how the extent of cytoreduction during bRT impacts outcomes. Materials/ Methods: We retrospectively reviewed patients with LBCL treated with bRT followed by CAR T-cell therapy. Metabolic tumor volume (MTV), maximum standardized uptake value (SUVmax), SUVmean, and total lesion glycolysis (TLG) were extracted from F18-fluorodeoxyglucose positron emission tomography (PET) scans acquired prior to bRT and between completion of bRT and CAR Tcell infusion. Delta radiomics based on changes of these values were then calculated. The association between delta radiomics and oncologic outcomes [progression-free survival (PFS), freedom from distant progression (FFDP), and local control (LC)] were then examined. Results: Thirty-three sites across 23 patients with LBCL were irradiated. All metabolically active disease was treated in 10 patients. Following bRT, median overall decreases (including unirradiated sites) in MTV, SUVmax, SUVmean, and TLG were 22.2 cc (63.1%), 8.9 (36.8%), 3.4 (31.1%), and 297.9 cc (75.8%), respectively. Median decreases in MTV, SUVmax, SUVmean, and TLG in irradiated sites were 15.6 cc (91.1%), 17.0 (74.6%), 6.8 (55.3%), and 157.0 cc (94.6%), respectively. Median follow-up was 15.2 months. A decrease in SUVmax of at least 54% was associated with improved PFS (24-month PFS: 83.3% vs. 28.1%; p = 0.037) and FFDP (24-month FFDP: 100% vs. 62.4%; p < 0.001). A decrease in MTV of at least 90% was associated with improved FFDP (24-month FFDP: 100% vs. 62.4%; p < 0.001). LC was improved in sites with decreases in SUVmax of at least 71% (24-month LC: 100% vs. 72.7%; p < 0.001). Decreases of MTV by at least 90% (100% vs. 53.3%; p = 0.038) and TLG by at least 95% (100% vs. 56.3%; p = 0.067) were associated with an improved complete response rate. Conclusion: bRT led to substantial reductions in MTV, SUVmax, SUVmean, and TLG. The relative extent of these decreases correlated with improved outcomes after CAR T-cell infusion. Prospective cohorts should validate the value of interim PET following bRT for quantifying changes in disease burden and associated prognosis. [ABSTRACT FROM AUTHOR]