Twice-Weekly Outpatient Rehabilitation Intervention for Young Children With Spinal Muscular Atrophy Treated With Genetic-Based Therapies: Protocol for a Feasibility Study.

BACKGROUND: Spinal muscular atrophy (SMA) is a progressive neuromuscular disorder that causes muscle weakness and is the leading genetic cause of infant mortality worldwide. While no definitive cure exists, the approval of 3 genetic-based therapies in Canada since 2018 has led to significant improvements in muscle function for children with SMA. With that, there are no evidence-based rehabilitation interventions and minimal evidence on the combined effects of genetic-based therapies and rehabilitation. OBJECTIVE: This protocol describes the methodology to assess the feasibility of a twice-weekly outpatient rehabilitation intervention focusing on gross and fine motor function to inform the methodology and sample size of a definitive clinical trial. METHODS: We will conduct a single-center nonrandomized pilot and feasibility trial to explore an outpatient rehabilitation intervention for children aged 6 months to 3 years with SMA treated with genetic-based therapies. Participation in the study will occur over a 25-week period, with a baseline assessment visit followed by a 12-week intervention period and a 12-week nonintervention period. The rehabilitation intervention comprises weekly physical and occupational therapy for 11 weeks. Assessments will occur at baseline (week 0), end of intervention or early withdrawal (week 12), and follow-up (week 24). Predetermined feasibility indicators will evaluate study feasibility across process (recruitment rates, eligibility criteria, adherence rates, retention rates, questionnaire suitability, and acceptability), resource (time, implementation, and execution), management (materials and data), and scientific (safety, tolerability, and preliminary efficacy) domains. RESULTS: This project was funded in March 2022, and data will be collected between March 2023 and December 2023. Data analysis will occur between January 2024 and March 2024, with publication expected in the fall of 2024. The protocol for the feasibility trial will be considered successful if it meets the success criteria set out for the feasibility indicators. Indicators of specific interest include all process indicators, as well as time. Exploratory indicators will be reported. Pragmatically, the results of the feasibility trial will inform changes to the protocol and the start-up of a definitive multisite trial. CONCLUSIONS: This novel twice-weekly outpatient rehabilitation intervention will be the first step toward filling the need for an evidence-based rehabilitation intervention for children with SMA treated with genetic-based therapies. It is expected that consistent and intensive rehabilitation therapy will augment functional gains being observed in this population. In the future, a definitive trial will measure the efficacy of the intervention. TRIAL REGISTRATION: ClinicalTrials.gov NCT05638750; https://clinicaltrials.gov/study/NCT05638750. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/46363.

Tough transitions: Family caregiver experiences with a pediatric long-term ventilation discharge pathway.

OBJECTIVES: Discharging a child home on long-term ventilation (LTV) via tracheostomy is complex and involves multiple healthcare providers across healthcare sectors. To date, there has been a paucity of data with respect to the experiences of families transitioning a child home on LTV. Our objective was to explore the perceptions of family caregivers (FCs) who have completed a newly developed LTV discharge pathway as they transitioned home. METHODS: We conducted 11 semi-structured interviews with FCs. Interviews focused on FC's experience with the training process, perception of competency from a knowledge and skill perspective, and opportunities for improvement. Interviews were audiotaped, transcribed verbatim, coded, and analyzed using an inductive thematic analysis approach. RESULTS: Eight mothers and three fathers of ten children participated. Six primary themes were identified: (1) making an informed decision, (2) transitioning to rehabilitation, (3) building capacity for self-care, (4) coordinating case management, (5) readying for discharge home, and (6) experiencing home care. CONCLUSION: Overall, FCs felt that the preparation and transition support obtained through the application of a standardized LTV discharge pathway allowed successful attainment of new knowledge and skills necessary to care for their child with LTV at home.

Association Analysis of a Microsatellite Repeat in the TRIB1 Gene With Prostate Cancer Risk, Aggressiveness and Survival.

With an estimated 1.1 million men worldwide diagnosed with prostate cancer yearly, effective and more specific biomarkers for early diagnosis could lead to better patient outcome. As such, novel genetic markers are sought for this purpose. The tribbles homologue 1 gene (TRIB1) has recently shown to have a role in prostate tumorigenesis and data-mining of prostate cancer expression data confirmed clinical significance of TRIB1 in prostate cancer. For the first time, a polymorphic microsatellite in this gene was studied for its potential association with prostate cancer risk and aggressiveness. Genomic DNA was extracted from a cohort of 1,152 prostate cancer patients and 1,196 cancer-free controls and the TTTTG-TRIB1 microsatellite was genotyped. The socio-demographic and clinical characteristics were analyzed using the non-parametric t-test and two-way ANOVA. Association of the TTTTG-TRIB1 microsatellite and prostate cancer risk and aggressiveness were analyzed by binary logistic regression and confirmed by bootstrapping. Total and prostate cancer mortality was analyzed using the Kaplan Meier test. Genotype and allele correlation with TRIB1 mRNA levels was analyzed using the non-parametric Kolmogorov-Smirnov test. To predict the effect that the TTTTG-TRIB1 polymorphisms had on the mRNA structure, the in silico RNA folding predictor tool, mfold, was used. By analyzing the publicly available data, we confirmed a significant over-expression of TRIB1 in prostate cancer compared to other cancer types, and an over-expression in prostate cancerous tissue compared to adjacent benign. Three alleles (three-five repeats) were observed for TTTTG-TRIB1. The three-repeat allele was associated with prostate cancer risk at the allele (OR = 1.16; P = 0.044) and genotypic levels (OR = 1.70; P = 0.006) and this association was age-independent. The four-repeat allele was inversely associated with prosatet cancer risk (OR = 0.57; P < 0.0001). TRIB1 expression was upregulated in tumors when compared to adjacent cancer-free tissue but was not allele specific. In silico analysis suggested that the TTTTG-TRIB1 alleles may alter TRIB1 mRNA structure. In summary, the three-repeat allele was significantly associated with prostate cancer risk, suggesting a biomarker potential for this microsatellite to predict prostate cancer. Further studies are needed to elucidate the functional role of this microsatellite in regulating TRIB1 expression, perhaps by affecting the TRIB1 mRNA structure and stability.

A microsatellite repeat in PCA3 long non-coding RNA is associated with prostate cancer risk and aggressiveness.

Short tandem repeats (STRs) are repetitive sequences of a polymorphic stretch of two to six nucleotides. We hypothesized that STRs are associated with prostate cancer development and/or progression. We undertook RNA sequencing analysis of prostate tumors and adjacent non-malignant cells to identify polymorphic STRs that are readily expressed in these cells. Most of the expressed STRs in the clinical samples mapped to intronic and intergenic DNA. Our analysis indicated that three of these STRs (TAAA-ACTG2, TTTTG-TRIB1, and TG-PCA3) are polymorphic and differentially expressed in prostate tumors compared to adjacent non-malignant cells. TG-PCA3 STR expression was repressed by the anti-androgen drug enzalutamide in prostate cancer cells. Genetic analysis of prostate cancer patients and healthy controls (N > 2,000) showed a significant association of the most common 11 repeat allele of TG-PCA3 STR with prostate cancer risk (OR = 1.49; 95% CI 1.11-1.99; P = 0.008). A significant association was also observed with aggressive disease (OR = 2.00; 95% CI 1.06-3.76; P = 0.031) and high mortality rates (HR = 3.0; 95% CI 1.03-8.77; P = 0.045). We propose that TG-PCA3 STR has both diagnostic and prognostic potential for prostate cancer. We provided a proof of concept to be applied to other RNA sequencing datasets to identify disease-associated STRs for future clinical exploratory studies.

Children with medical complexities: Addressing the gaps in respite care during transition from paediatrics to adult health care in Ontario.

Respite care is one integral component of the transition process from paediatric to adult health care, and is of particular importance to individuals with medical complexities. Numerous gaps that exist within the current system limit a child and family's ability to access quality respite care during a time when it is often most needed. Identifying and addressing these gaps in a systematic and collaborative way presents an opportunity to improve the quality of life for this vulnerable, ever growing population and their families.

Adding In Silico Assessment of Potential Splice Aberration to the Integrated Evaluation of BRCA Gene Unclassified Variants.

Clinical mutation screening of the cancer susceptibility genes BRCA1 and BRCA2 generates many unclassified variants (UVs). Most of these UVs are either rare missense substitutions or nucleotide substitutions near the splice junctions of the protein coding exons. Previously, we developed a quantitative method for evaluation of BRCA gene UVs-the "integrated evaluation"-that combines a sequence analysis-based prior probability of pathogenicity with patient and/or tumor observational data to arrive at a posterior probability of pathogenicity. One limitation of the sequence analysis-based prior has been that it evaluates UVs from the perspective of missense substitution severity but not probability to disrupt normal mRNA splicing. Here, we calibrated output from the splice-site fitness program MaxEntScan to generate spliceogenicity-based prior probabilities of pathogenicity for BRCA gene variants; these range from 0.97 for variants with high probability to damage a donor or acceptor to 0.02 for exonic variants that do not impact a splice junction and are unlikely to create a de novo donor. We created a database http://priors.hci.utah.edu/PRIORS/ that provides the combined missense substitution severity and spliceogenicity-based probability of pathogenicity for BRCA gene single-nucleotide substitutions. We also updated the BRCA gene Ex-UV LOVD, available at http://hci-exlovd.hci.utah.edu, with 77 re-evaluable variants.

Fusion transcript loci share many genomic features with non-fusion loci.

BACKGROUND: Fusion transcripts are found in many tissues and have the potential to create novel functional products. Here, we investigate the genomic sequences around fusion junctions to better understand the transcriptional mechanisms mediating fusion transcription/splicing. We analyzed data from prostate (cancer) cells as previous studies have shown extensively that these cells readily undergo fusion transcription. RESULTS: We used the FusionMap program to identify high-confidence fusion transcripts from RNAseq data. The RNAseq datasets were from our (N = 8) and other (N = 14) clinical prostate tumors with adjacent non-cancer cells, and from the LNCaP prostate cancer cell line that were mock-, androgen- (DHT), and anti-androgen- (bicalutamide, enzalutamide) treated. In total, 185 fusion transcripts were identified from all RNAseq datasets. The majority (76%) of these fusion transcripts were 'read-through chimeras' derived from adjacent genes in the genome. Characterization of sequences at fusion loci were carried out using a combination of the FusionMap program, custom Perl scripts, and the RNAfold program. Our computational analysis indicated that most fusion junctions (76%) use the consensus GT-AG intron donor-acceptor splice site, and most fusion transcripts (85%) maintained the open reading frame. We assessed whether parental genes of fusion transcripts have the potential to form complementary base pairing between parental genes which might bring them into physical proximity. Our computational analysis of sequences flanking fusion junctions at parental loci indicate that these loci have a similar propensity as non-fusion loci to hybridize. The abundance of repetitive sequences at fusion and non-fusion loci was also investigated given that SINE repeats are involved in aberrant gene transcription. We found few instances of repetitive sequences at both fusion and non-fusion junctions. Finally, RT-qPCR was performed on RNA from both clinical prostate tumors and adjacent non-cancer cells (N = 7), and LNCaP cells treated as above to validate the expression of seven fusion transcripts and their respective parental genes. We reveal that fusion transcript expression is similar to the expression of parental genes. CONCLUSIONS: Fusion transcripts maintain the open reading frame, and likely use the same transcriptional machinery as non-fusion transcripts as they share many genomic features at splice/fusion junctions.

Chronic Pain Assessment Tools for Cerebral Palsy: A Systematic Review.

BACKGROUND AND OBJECTIVE: Chronic pain in children with cerebral palsy (CP) is underrecognized, leading to detriments in their physical, social, and mental well-being. Our objective was to identify, describe, and critique pediatric chronic pain assessment tools and make recommendations for clinical use for children with CP. Secondly, develop an evidence-informed toolbox to support clinicians in the assessment of chronic pain in children with disabilities. METHODS: Ovid Medline, Cumulative Index to Nursing and Allied Health Literature, and Embase databases were systematically searched by using key terms "chronic pain" and "clinical assessment tool" between January 2012 and July 2014. Tools from multiple pediatric health conditions were explored contingent on inclusion criteria: (1) children 1 to 18 years; (2) assessment focus on chronic pain; (3) psychometric properties reported; (4) written in English between 1980 and 2014. Pediatric chronic pain assessment tools were extracted and corresponding validation articles were sought for review. Detailed tool descriptions were composed and each tool underwent a formal critique of psychometric properties and clinical utility. RESULTS: Of the retrieved 2652 articles, 250 articles met eligibility, from which 52 chronic pain assessment tools were retrieved. A consensus among interprofessional working group members determined 7 chronic pain interference tools to be of importance. Not all tools have been validated with children with CP nor is there 1 tool to meet the needs of all children experiencing chronic pain. CONCLUSIONS: This study has systematically reviewed and recommended, through expert consensus, valid and reliable chronic pain interference assessment tools for children with disabilities.

How long does it take to initiate a child on long-term invasive ventilation? Results from a Canadian pediatric home ventilation program.

OBJECTIVE: To assess the length of stay required to initiate long-term invasive ventilation at the authors' institution, which would inform future interventional strategies to streamline the in-hospital stay for these families. METHODS: A retrospective chart review of children initiated on invasive long-term ventilation via tracheostomy at the authors' acute care centre between January 2005 and December 2013 was performed. RESULTS: Thirty-five children were initiated on long-term invasive ventilation via tracheostomy at the acute care hospital; 19 (54%) were male. The median age at time of admission was 0.52 years (interquartile range [IQR] 0.06 to 9.58 years) . Musculoskeletal disease (n=11 [31%]) was the most common reason for tracheostomy insertion. Two children died during the hospital admission. Fifteen children were discharged home directly from the acute care hospital and 18 were moved to the rehabilitation hospital. Six are current inpatients of the rehabilitation centre and were never discharged home. Combining the length of stay at the acute care and rehabilitation hospitals for the entire cohort, the median length of stay was 162.0 days (IQR 98.0 to 275.0 days) and 97.0 days (IQR 69.0 to 210.0 days), respectively, from the time of tracheostomy insertion. CONCLUSIONS: The median length of stay from the initiation of invasive long-term ventilation to discharge home from the rehabilitation hospital was somewhat long compared with other ventilation programs worldwide. Additionally, approximately 20% of the cohort never transitioned home. There is a timely need to benchmark across the country and internationally, to identify and implement strategies for cohesive, coordinated care for these children to decrease overall length of stay.

Factors impacting same-day cancellation of outpatient pediatric magnetic resonance imaging under anesthesia.

BACKGROUND: Studies of elective surgical procedures indicate that cancellation is common and preventable. Little is known about cancellation of anesthesia-supported elective diagnostic imaging. OBJECTIVE: To describe the reasons for same-day cancellation of MRI studies performed under sedation or anesthesia and identify patient characteristics associated with cancellations. MATERIALS AND METHODS: This case-control study was carried out in a university-affiliated tertiary care children's hospital. Cases were defined as elective outpatient MRI studies booked under anesthesia that were cancelled after the patient had arrived in the radiology department in 2009. Matched controls were identified by selecting the same day and time 1 week before or after the cancelled case. Main outcome measures included demographics, MRI study characteristics, and social and medical factors. RESULTS: There were 111 outpatient anesthesia-supported MRI studies cancelled on the same day as the assessment (cancellation rate: 4.5%), of which 74.6% were related to family and patient factors, while 22% were related to system factors. Cancelled cases involved patients who lived in lower median income quintile neighborhoods compared to controls (2 vs. 3; P = 0.0007; odds ratio [OR] 3.81; 95% confidence interval [CI] 1.18-12.34). Those who traveled a greater median distance (in kilometers) were less likely to be cancelled (18.8 vs. 27.1, P = 0.0035). Although cancelled patients had a lower mean number of total medical services (2.5 vs. 3.0; P = 0.03; OR = 0.78; 95% CI 0.62-0.98), current medical factors (past 12 months) did not impact cancellations. CONCLUSION: Same-day cancellations of anesthesia-supported MRI studies are not uncommon, and the main predictor of cancellation seems to be socioeconomic rather than medical.

Nucleotide binding to the human multidrug resistance protein 3, MRP3.

We used a spin-labeled ATP analog, SL-ATP, to study nucleotide binding to highly purified human multidrug resistance protein 3, MRP3, which had been expressed in the yeast Pichia pastoris. SL-ATP was shown to be a good substrate analog and is hydrolyzed by MRP3 at about 10% of the Vmax for normal ATP. ESR titrations showed that 2 mol of SL-ATP readily bound per mole of MRP3 with a dissociation constant of about 100 microM in the presence of Mg(2+) ions. The binding curve was easily fitted for a hyperbolic binding relationship. SL-ATP also bound readily to MRP3 in the absence of divalent ions and presence of EDTA. The resulting binding curve, however, could not be satisfactorily fitted using the equation for hyperbola. Analysis showed that a good fit was only obtained with the Hill equation using a Hill coefficient of 4 or close to 4. Lower Hill coefficients resulted in lower goodness of the fit. Such cooperative binding may be explained by a dimerization event triggered in the absence of divalent ions and a close communication of nucleotide binding sites of the interacting dimers. These findings may be of great importance for the overall mechanism and regulation of multidrug resistance proteins.

Conformational changes in the Escherichia coli ATP synthase b-dimer upon binding to F(1)-ATPase.

Conformational changes within the subunit b-dimer of the E. coli ATP synthase occur upon binding to the F(1) sector. ESR spectra of spin-labeled b at room temperature indicated a pivotal point in the b-structure at residue 62. Spectra of frozen b +/- F(1) and calculated interspin distances suggested that where contact between b (2) and F(1) occurs (above about residue 80), the structure of the dimer changes minimally. Between b-residues 33 and 64 inter-subunit distances in the F(1)-bound b-dimer were found to be too large to accommodate tightly coiled coil packing and therefore suggest a dissociation and disengagement of the dimer upon F(1)-binding. Mechanistic implications of this "bubble" formation in the tether domain of ATP synthase b ( 2 ) are discussed.