Efficient single copy integration via homology-directed repair (scHDR) by 5'modification of large DNA donor fragments in mice.

CRISPR/Cas-based approaches have largely replaced conventional gene targeting strategies. However, homology-directed repair (HDR) in the mouse genome is not very efficient, and precisely inserting longer sequences using HDR remains challenging given that donor constructs preferentially integrate as concatemers. Here, we showed that injecting 5' biotinylated donor DNA into mouse embryos at the two-cell stage led to efficient single-copy HDR (scHDR) allele generation. Our dedicated genotyping strategy showed that these alleles occurred with frequencies of 19%, 20%, and 26% at three independent gene loci, indicating that scHDR was dramatically increased by 5' biotinylation. Thus, we suggest that the combination of a 5' biotinylated donor and diligent analysis of concatemer integration are prerequisites for efficiently and reliably generating conditional alleles or other large fragment knock-ins in the mouse genome.

Dose-intensified stereotactic body radiotherapy for painful vertebral metastases: A randomized phase 3 trial.

BACKGROUND: The purpose of this randomised study was to determine whether dose-intensified stereotactic body radiotherapy (SBRT) for painful vertebral metastases results in increased rates of pain improvement compared with conventional external beam radiotherapy (cEBRT) (control) 6 months after treatment. METHODS: This randomized, controlled phase 3 trial was conducted between November 2016 and January 2023, when it was stopped early. Patients were eligible if they were aged 18 years or older; had one or two painful, stable, or potentially unstable vertebral metastases; and had a life expectancy of 1 year or longer according to the investigator's estimates. Patients received 48.5 grays (Gy) in 10 fractions (with epidural involvement) or 40 Gy in five fractions (without epidural involvement) in the SBRT group and 30 Gy in 10 fractions or 20 Gy in five fractions in the cEBRT group, respectively. The primary end point was an improvement in the pain score at the treated site by at least 2 points (on a visual analog scale from 0 to 10 points) at 6-month follow-up. Data were analyzed on an intention-to-treat and per-protocol basis. RESULTS: Of 214 patients who were screened for eligibility, 63 were randomized 1:1 between SBRT (33 patients with 36 metastases) and cEBRT (30 patients with 31 metastases). The median age of all patients was 66 years, and 40 patients were men (63.5%). In the intention-to-treat analysis, the 6-month proportion of patients who had metastases with pain reduction by 2 or more points was significantly higher in the SBRT group versus the control group (69.4% vs. 41.9%, respectively; two-sided p = .02). Changes in opioid medication intake relative to baseline were nonsignificant between the groups. No differences were observed in vertebral compression fracture or adverse event rates between the groups. CONCLUSIONS: Dose-intensified SBRT improved pain score more effectively than cEBRT at 6 months.

Impact of Imaging-Guided Localization on Performance of Tailored Axillary Surgery in Patients with Clinically Node-Positive Breast Cancer: Prospective Cohort Study Within TAXIS (OPBC-03, SAKK 23/16, IBCSG 57-18, ABCSG-53, GBG 101).

BACKGROUND: Tailored axillary surgery (TAS) is a novel surgical concept for clinical node-positive breast cancer. It consists of the removal of the sentinel lymph nodes (LNs), as well as palpably suspicious nodes. The TAS technique can be utilized in both the upfront and neoadjuvant chemotherapy (NACT) setting. This study assessed whether/how imaging-guided localization (IGL) influenced TAS. PATIENTS AND METHODS: This was a prospective observational cohort study preplanned in the randomized phase-III OPBC-03/TAXIS trial. IGL was performed at the surgeon's discretion for targeted removal of LNs during TAS. Immediate back-up axillary lymph node dissection (ALND) followed TAS according to TAXIS randomization. RESULTS: Five-hundred patients were included from 44 breast centers in six countries, 151 (30.2%) of whom underwent NACT. IGL was performed in 84.4% of all patients, with significant variation by country (77.6-100%, p < 0.001). No difference in the median number of removed (5 vs. 4, p = 0.3) and positive (2 vs. 2, p = 0.6) LNs by use of IGL was noted. The number of LNs removed during TAS with IGL remained stable over time (p = 0.8), but decreased significantly without IGL, from six (IQR 4-6) in 2019 to four (IQR 3-4) in 2022 (p = 0.015). An ALND was performed in 249 patients, removing another 12 (IQR 9-17) LNs, in which a median number of 1 (IQR 0-4) was positive. There was no significant difference in residual nodal disease after TAS with or without IGL (68.0% vs. 57.6%, p = 0.2). CONCLUSIONS: IGL did not significantly change either the performance of TAS or the volume of residual nodal tumor burden. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03513614.

Trends in use of neoadjuvant systemic therapy in patients with clinically node-positive breast cancer in Europe: prospective TAXIS study (OPBC-03, SAKK 23/16, IBCSG 57-18, ABCSG-53, GBG 101).

PURPOSE: The aim of this study was to evaluate clinical practice heterogeneity in use of neoadjuvant systemic therapy (NST) for patients with clinically node-positive breast cancer in Europe. METHODS: The study was preplanned in the international multicenter phase-III OPBC-03/TAXIS trial (ClinicalTrials.gov Identifier: NCT03513614) to include the first 500 randomized patients with confirmed nodal disease at the time of surgery. The TAXIS study's pragmatic design allowed both the neoadjuvant and adjuvant setting according to the preferences of the local investigators who were encouraged to register eligible patients consecutively. RESULTS: A total of 500 patients were included at 44 breast centers in six European countries from August 2018 to June 2022, 165 (33%) of whom underwent NST. Median age was 57 years (interquartile range [IQR], 48-69). Most patients were postmenopausal (68.4%) with grade 2 and 3 hormonal receptor-positive and human epidermal growth factor receptor 2-negative breast cancer with a median tumor size of 28 mm (IQR 20-40). The use of NST varied significantly across the countries (p < 0.001). Austria (55.2%) and Switzerland (35.8%) had the highest percentage of patients undergoing NST and Hungary (18.2%) the lowest. The administration of NST increased significantly over the years (OR 1.42; p < 0.001) and more than doubled from 20 to 46.7% between 2018 and 2022. CONCLUSION: Substantial heterogeneity in the use of NST with HR+/HER2-breast cancer exists in Europe. While stringent guidelines are available for its use in triple-negative and HER2+ breast cancer, there is a need for the development of and adherence to well-defined recommendations for HR+/HER2-breast cancer.

Patterns of Electrocardiographic Abnormalities in Children with Hypertrophic Cardiomyopathy.

Hypertrophic cardiomyopathy (HCM), a common cardiomyopathy in children, is an important cause of morbidity and mortality. Early recognition and appropriate management are important. An electrocardiogram (ECG) is often used as a screening tool in children to detect heart disease. The ECG patterns in children with HCM are not well described.ECGs collected from an international cohort of children, and adolescents (≤ 21 years) with HCM were reviewed. 482 ECGs met inclusion criteria. Age ranged from 1 day to 21 years, median 13 years. Of the 482 ECGs, 57 (12%) were normal. The most common abnormalities noted were left ventricular hypertrophy (LVH) in 108/482 (22%) and biventricular hypertrophy (BVH) in 116/482 (24%) Of the patients with LVH/BVH (n = 224), 135 (60%) also had a strain pattern (LVH in 83, BVH in 52). Isolated strain pattern (in the absence of criteria for hypertrophy) was seen in 43/482 (9%). Isolated pathologic Q waves were seen in 71/482 (15%). Pediatric HCM, 88% have an abnormal ECG. The most common ECG abnormalities were LVH or BVH with or without strain. Strain pattern without hypertrophy and a pathologic Q wave were present in a significant proportion (24%) of patients. Thus, a significant number of children with HCM have ECG abnormalities that are not typical for "hypertrophy". The presence of the ECG abnormalities described above in a child should prompt further examination with an echocardiogram to rule out HCM.

Preventing suicide in post-secondary students: a scoping review of suicide prevention programs.

Suicide among students enrolled in post-secondary education, including university or college, is a major public health concern. Previous research has examined the effectiveness of suicide prevention programs for this population. However, the effective elements of these interventions remain unknown. This study reviewed the literature on suicide prevention programs for post-secondary students, exploring and identifying those elements likely contributing to their effectiveness. A scoping review process was undertaken exploring suicide prevention programs for post-secondary students. Methodological quality of the articles was assessed, and content analysis was used to explore the programs and their effective elements. Twenty seven articles were included in this review, covering a variety of approaches. Gatekeeper training programs were the most common type of suicide prevention program. Programs for post-secondary students may be effective in improving student rates of engagement with mental health services and were associated with greater knowledge, and help-seeking attitudes and behaviors, and gatekeeper-related outcomes. While evidence was found supporting the effectiveness of some interventions such as gatekeeper programs to influence suicide-related knowledge, attitudes and behaviour, further and more rigorous research surrounding suicide prevention programs for post-secondary students is required, with a particular emphasis on student outcomes.

The efficacy of the "Talk-to-Me" suicide prevention and mental health education program for tertiary students: a crossover randomised control trial.

Despite suicide ideation being one of the most frequently reported health issues impacting tertiary students, there is a paucity of research evaluating the efficacy of preventive interventions aimed at improving mental health outcomes for students studying at two tertiary institutes. The current study evaluated the efficacy of the "Talk-to-Me" Mass Open Online Course (MOOC) in improving tertiary students' abilities to support the mental health of themselves and their peers via a randomised controlled trial design, comparing them to a waitlist control group. Overall, 129 tertiary students (M = 25.22 years, SD = 7.43; 80% female) undertaking a health science or education course at two Western Australian universities were randomly allocated to either "Talk-to-Me" (n = 66) or waitlist control (n = 63) groups. The participants' responses to suicidal statements (primary outcome), knowledge of mental health, generalised self-efficacy, coping skills, and overall utility of the program (secondary outcomes) were collected at three timepoints (baseline 10-weeks and 24-weeks from baseline). Assessment time and group interaction were explored using a random-effects regression model, examining changes in the primary and secondary outcomes. Intention-to-treat analysis (N = 129) at 10-weeks demonstrated a significant improvement in generalised self-efficacy for "Talk-to-Me" compared to the control group (ES = 0.36, p = .04), with only the "Talk-to-Me" participants reporting increased knowledge in responding to suicidal ideation (primary outcome). This change was sustained for 24 weeks. Findings provide preliminary evidence suggesting that the "Talk-to-Me" MOOC can effectively improve tertiary students' mental health and knowledge of how to support themselves and others in distress. ACTRN12619000630112, registered 18-03-2019, anzctr.org.au.

[Smarter Medicine in Radiation Oncology - current and proven treatment concepts offering the greatest possible benefit to cancer patients in clinical practice]. / Smarter Medicine in der Radio-Onkologie.

Smarter Medicine in Radiation Oncology - current and proven treatment concepts offering the greatest possible benefit to cancer patients in clinical practice Abstract. Modern procedures in radiation therapy with respect to indication, therapy planning and implementation of shorter treatment regimens result in high local tumor control and excellent quality of life in frequent and clinically relevant disease entities including breast and prostate cancer as well as bone metastases. In the adjuvant therapy for early breast cancer, a careful comparative analysis of the use of endocrine adjuvant treatment over five years versus postoperative radiation therapy over less than four weeks may result in omission of radiation treatment in a selected patient population. Partial breast irradiation, while treating the tumor bed only in low risk situations, and hypofractionated whole breast radiotherapy for patients with a higher risk profile for local recurrence, halved treatment time compared to 10 years ago, making adjuvant radiotherapy for patients with breast cancer more convenient. In patients with localized prostate cancer, the introduction of hypofractionated treatment regimens halved the number of daily fractions and current developments point towards a one-week outpatient therapy for locally limited disease using high precision stereotactic techniques and accurate therapy planning based on multifunctional imaging. Patients with bone metastases are more and more treated with a single fraction, high precision stereotactic radiotherapy (radiosurgery) with high analgesic potential and achieving long lasting local tumor control. These modern, often significantly shorter, radiotherapy treatment regimens not only result in an excellent treatment response and better quality of life for patients, but also include a better utilization of radiotherapy resources needing less treatment machines and thus helping to reduce costs in the Swiss healthcare system.

Saraf-dependent activation of mTORC1 regulates cardiac growth.

Pathological cardiac hypertrophy is an independent risk for heart failure (HF) and sudden death. Deciphering signaling pathways regulating intracellular Ca2+ homeostasis that control adaptive and pathological cardiac growth may enable identification of novel therapeutic targets. The objective of the present study is to determine the role of the store-operated calcium entry-associated regulatory factor (Saraf), encoded by the Tmem66 gene, on cardiac growth control in vitro and in vivo. Saraf is a single-pass membrane protein located at the sarco/endoplasmic reticulum and regulates intracellular calcium homeostasis. We found that Saraf expression was upregulated in the hypertrophied myocardium and was sufficient for cell growth in response to neurohumoral stimulation. Increased Saraf expression caused cell growth, which was associated with dysregulation of calcium-dependent signaling and sarcoplasmic reticulum calcium content. In vivo, Saraf augmented cardiac myocyte growth in response to angiotensin II and resulted in increased cardiac remodeling together with worsened cardiac function. Mechanistically, Saraf activated mTORC1 (mechanistic target of rapamycin complex 1) and increased protein synthesis, while mTORC1 inhibition blunted Saraf-dependent cell growth. In contrast, the hearts of Saraf knockout mice and Saraf-deficient myocytes did not show any morphological or functional alterations after neurohumoral stimulation, but Saraf depletion resulted in worsened cardiac function after acute pressure overload. SARAF knockout blunted transverse aortic constriction cardiac myocyte hypertrophy and impaired cardiac function, demonstrating a role for SARAF in compensatory myocyte growth. Collectively, these results reveal a novel link between sarcoplasmic reticulum calcium homeostasis and mTORC1 activation that is regulated by Saraf.

[Suicide postvention: what mental health professionals need to know]. / Postvention bei Suizid: Was man als Kinder- und Jugendpsychiater und -therapeut wissen sollte.

Postvention is the term used to describe a package of interventions designed for people affected by suicide. Inpatient suicides in Departments of Child and Adolescent Psychiatry are rare, but nevertheless constitute an immense burden to family members, friends, and fellow patients, as well as to medical staff members. Besides including an effective management of the acute crisis situation with well-defined communication rules, appropriate interaction with the bereaved, and prevention of copycat suicides in other patients, reliable guidelines for dealing with inpatient suicide must also contain a precise procedure on how to handle mental stress in physicians, psychotherapists, and other staff members. Furthermore, postvention should be an important part of the professional training of physicians as well as psychotherapists who work in child and adolescent psychiatry departments or outpatient clinics.

Haemoglobin and creatinine values as prognostic factors for outcome of concurrent radiochemotherapy in locally advanced head and neck cancers : Secondary results of two European randomized phase III trials (ARO 95-06, SAKK 10/94).

BACKGROUND: To determine the influence of baseline laboratory values on treatment outcome in patients with locally advanced head and neck cancer (HNSCC). METHODS: Data of the randomized trials ARO 95 -06 (n = 384) and SAKK 10 /94 (n = 224) were pooled for a total sample size of 608 patients. Haemoglobin (Hb) and creatinine (Cr) were available at baseline and their association with locoregional recurrence-free survival (LRRFS), distant metastasis-free survival (DMFS), cancer-specific survival (CSS), and overall survival (OS) was analyzed using univariable and multivariable Cox regression models. RESULTS: A total of 580 and 564 patients were available with baseline Hb and Cr values in the pooled analysis. Univariable analyses revealed that lower baseline Hb values were significantly associated with decreased LRRFS, DMFS, CSS and OS. This effect remained significant for OS when the treatment arms (radiotherapy [RT] alone vs. chemoradiation [CRT]) were analyzed separately. Higher baseline Cr was associated with improved OS in the pooled analysis. Interestingly, the prognostic value of baseline Cr appeared to be limited to the subgroup of 284 patients who were treated with CRT. In the multivariable Cox regression model lower baseline Hb remained associated with decreased OS both in the patients who received CRT (HR 0.79, 95 % CI 0.66-0.94, p = 0.009) and in those patients who underwent RT alone (HR 0.67, 95 % CI 0.58-0.78, p < 0.001). Increased baseline Cr remained significantly associated with improved OS in patients who underwent CRT (HR 0.79, 95 % CI 0.69-0.92, p = 0.002) but not in those patients who underwent RT alone. CONCLUSIONS: An association between lower baseline Hb and inferior treatment outcome was confirmed. Baseline Cr was introduced as a prognosticator of outcome after CRT for locally advanced HNSCC.

Vemurafenib-induced radiation recall dermatitis: case report and review of the literature.

The cutaneous effects of BRAF (serine/threonine protein kinase B-raf) inhibitors such as vemurafenib remain poorly defined. Rash, squamous cell carcinoma, keratoacanthoma and photosensitivity are the most common grade 2 or 3 adverse events observed in clinical trials. We here report the case of a patient with a BRAF V600E mutated metastatic melanoma who developed severe radiation recall dermatitis 6 weeks after completing radiotherapy. Vemurafenib treatment had been initiated 1 week before the development of dermatitis because of rapidly progressing disease. Upon topical treatment of the affected skin areas, clinical symptoms regressed over a period of 2 months, although vemurafenib was continuously administered. As our case goes in line with other reports, we believe that physicians should be aware of this additional cutaneous side effect of vemurafenib and that continuation of the treatment is safe when close clinical control and interdisciplinary management can be provided.

Exposure to radio-frequency electromagnetic fields from broadcast transmitters and risk of childhood cancer: a census-based cohort study.

We investigated the association between exposure to radio-frequency electromagnetic fields (RF-EMFs) from broadcast transmitters and childhood cancer. First, we conducted a time-to-event analysis including children under age 16 years living in Switzerland on December 5, 2000. Follow-up lasted until December 31, 2008. Second, all children living in Switzerland for some time between 1985 and 2008 were included in an incidence density cohort. RF-EMF exposure from broadcast transmitters was modeled. Based on 997 cancer cases, adjusted hazard ratios in the time-to-event analysis for the highest exposure category (>0.2 V/m) as compared with the reference category (<0.05 V/m) were 1.03 (95% confidence interval (CI): 0.74, 1.43) for all cancers, 0.55 (95% CI: 0.26, 1.19) for childhood leukemia, and 1.68 (95% CI: 0.98, 2.91) for childhood central nervous system (CNS) tumors. Results of the incidence density analysis, based on 4,246 cancer cases, were similar for all types of cancer and leukemia but did not indicate a CNS tumor risk (incidence rate ratio = 1.03, 95% CI: 0.73, 1.46). This large census-based cohort study did not suggest an association between predicted RF-EMF exposure from broadcasting and childhood leukemia. Results for CNS tumors were less consistent, but the most comprehensive analysis did not suggest an association.

A progressive dopaminergic phenotype associated with neurotoxic conversion of α-synuclein in BAC-transgenic rats.

Conversion of soluble α-synuclein into insoluble and fibrillar inclusions is a hallmark of Parkinson's disease and other synucleinopathies. Accumulating evidence points towards a relationship between its generation at nerve terminals and structural synaptic pathology. Little is known about the pathogenic impact of α-synuclein conversion and deposition at nigrostriatal dopaminergic synapses in transgenic mice, mainly owing to expression limitations of the α-synuclein construct. Here, we explore whether both the rat as a model and expression of the bacterial artificial chromosome construct consisting of human full-length wild-type α-synuclein could exert dopaminergic neuropathological effects. We found that the human promoter induced a pan-neuronal expression, matching the rodent α-synuclein expression pattern, however, with prominent C-terminally truncated fragments. Ageing promoted conversion of both full-length and C-terminally truncated α-synuclein species into insolube and proteinase K-resistant fibres, with strongest accumulation in the striatum, resembling biochemical changes seen in human Parkinson's disease. Transgenic rats develop early changes in novelty-seeking, avoidance and smell before the progressive motor deficit. Importantly, the observed pathological changes were associated with severe loss of the dopaminergic integrity, thus resembling more closely the human pathology.

Understanding the utility of "Talk-to-Me" an online suicide prevention program for Australian university students.

BACKGROUND: Australian university students are at risk of experiencing poor mental health, being vulnerable to self-harm and suicidal ideation. AIM: "Talk-to-Me" is a suicide ideation prevention Massive open online course (MOOC) previously showing it can support Western Australian university students' knowledge of identifying and responding to suicide ideation in themselves and others. METHODS: A multi-site one-group pre-test/post-test design with a 12-week follow-up explored the efficacy of "Talk-to-Me" for university students Australia-wide, evaluating the influence of COVID-19 and location. Overall, 217 students (55% female; mage = 24.93 years [18, 60]) enrolled in this study from 2020 to 2021. Participants' responses to suicidal statements, mental health literacy, generalized self-efficacy, help-seeking behavior, and overall utility of the program were collected at baseline, post-MOOC (10 weeks from baseline) and 12-week follow-up. The effect of time and location interaction was explored using a random-effects regression model. RESULTS: Findings indicated significant improvement in participants' knowledge of positive mental health support strategies (ES = 0.42, p < 0.001) and recognizing appropriate responses to suicidal statements (ES = 0.37, p < 0.001) at 10-weeks, with further improvement at 12 weeks follow-up (ES = 0.47 and 0.46, p < 0.001). Students reported higher generalized self-efficacy at the 12-week follow-up compared to baseline (ES = 0.19, p = 0.03) and an increased tendency to seek professional help for mental health issues (ES = 0.22, p = 0.02). CONCLUSION: These findings provide preliminary evidence of the efficacy of the "Talk-to-Me" program in supporting university students across Australia to increase their suicide-related knowledge and skills, general self-efficacy, and overall mental fitness.

Association of Axillary Dissection With Systemic Therapy in Patients With Clinically Node-Positive Breast Cancer.

Importance: The role of axillary lymph node dissection (ALND) to determine nodal burden to inform systemic therapy recommendations in patients with clinically node (cN)-positive breast cancer (BC) is currently unknown. Objective: To address the association of ALND with systemic therapy in cN-positive BC in the upfront surgery setting and after neoadjuvant chemotherapy (NACT). Design, Setting, and Participants: This was a prospective, observational, cohort study conducted from August 2018 to June 2022. This was a preplanned study within the phase 3 randomized clinical OPBC-03/TAXIS trial. Included were patients with confirmed cN-positive BC from 44 private, public, and academic breast centers in 6 European countries. After NACT, residual nodal disease was mandatory, and a minimum follow-up of 2 months was required. Exposures: All patients underwent tailored axillary surgery (TAS) followed by ALND or axillary radiotherapy (ART) according to TAXIS randomization. TAS removed suspicious palpable and sentinel nodes, whereas imaging-guidance was optional. Systemic therapy recommendations were at the discretion of the local investigators. Results: A total of 500 patients (median [IQR] age, 57 [48-69] years; 487 female [97.4%]) were included in the study. In the upfront surgery setting, 296 of 335 patients (88.4%) had hormone receptor (HR)-positive and Erb-B2 receptor tyrosine kinase 2 (ERBB2; formerly HER2 or HER2/neu)-negative disease: 145 (49.0%) underwent ART, and 151 (51.0%) underwent ALND. The median (IQR) number of removed positive lymph nodes without ALND was 3 (1-4) nodes compared with 4 (2-9) nodes with ALND. There was no association of ALND with the proportion of patients undergoing adjuvant chemotherapy (81 of 145 [55.9%] vs 91 of 151 [60.3%]; adjusted odds ratio [aOR], 0.72; 95% CI, 0.19-2.67) and type of systemic therapy. Of 151 patients with NACT, 74 (51.0%) underwent ART, and 77 (49.0%) underwent ALND. The ratio of removed to positive nodes was a median (IQR) of 4 (3-7) nodes to 2 (1-3) nodes and 15 (12-19) nodes to 2 (1-5) nodes in the ART and ALND groups, respectively. There was no observed association of ALND with the proportion of patients undergoing postneoadjuvant systemic therapy (57 of 74 [77.0%] vs 55 of 77 [71.4%]; aOR, 0.86; 95% CI, 0.43-1.70), type of postneoadjuvant chemotherapy (eg, capecitabine: 10 of 74 [13.5%] vs 10 of 77 [13.0%]; trastuzumab emtansine-DM1: 9 of 74 [12.2%] vs 11 of 77 [14.3%]), or endocrine therapy (eg, aromatase inhibitors: 41 of 74 [55.4%] vs 36 of 77 [46.8%]; tamoxifen: 8 of 74 [10.8%] vs 6 of 77 [7.8%]). Conclusion: Results of this cohort study suggest that patients without ALND were significantly understaged. However, ALND did not inform systemic therapy recommendations.