Evaluation of the CRISPR/Cas9 Genetic Constructs in Efficient Disruption of Porcine Genes for Xenotransplantation Purposes Along with an Assessment of the Off-Target Mutation Formation

The increasing life expectancy of humans has led to an increase in the number of patients with chronic diseases and organ failure. However, the imbalance between the supply and the demand for human organs is a serious problem in modern transplantology. One of many solutions to overcome this problem is the use of xenotransplantation. The domestic pig (Sus scrofa domestica) is currently considered as the most suitable for human organ procurement. However, there are discrepancies between pigs and humans that lead to the creation of immunological barriers preventing the direct xenograft. The introduction of appropriate modifications to the pig genome to prevent xenograft rejection is crucial in xenotransplantation studies. In this study, porcine GGTA1, CMAH, β4GalNT2, vWF, ASGR1 genes were selected to introduce genetic modifications. The evaluation of three selected gRNAs within each gene was obtained, which enabled the selection of the best site for efficient introduction of changes. Modifications were examined after nucleofection of porcine primary kidney fibroblasts with CRISPR/Cas9 system genetic constructs, followed by the tracking of indels by decomposition (TIDE) analysis. In addition, off-target analysis was carried out for selected best gRNAs using the TIDE tool, which is new in the research conducted so far and shows the utility of this tool in these studies.

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Characterization of Resistance to the Nonnucleoside NS5B Inhibitor Filibuvir in Hepatitis C Virus-Infected Patients

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.05611-11 ◽

2011 ◽

Vol 56 (3) ◽

pp. 1331-1341 ◽

Cited By ~ 27

Author(s):

Philip J. F. Troke ◽

Marilyn Lewis ◽

Paul Simpson ◽

Katrina Gore ◽

Jennifer Hammond ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Amino Acid ◽

Site Directed Mutagenesis ◽

Phenotypic Resistance ◽

Number Of Patients ◽

Chronic Hcv ◽

Selection Of

ABSTRACTFilibuvir (PF-00868554) is an investigational nonnucleoside inhibitor of the hepatitis C virus (HCV) nonstructural 5B (NS5B) RNA-dependent RNA polymerase currently in development for treating chronic HCV infection. The aim of this study was to characterize the selection of filibuvir-resistant variants in HCV-infected individuals receiving filibuvir as short (3- to 10-day) monotherapy. We identified amino acid M423 as the primary site of mutation arising upon filibuvir dosing. Through bulk cloning of clinical NS5B sequences into a transient-replicon system, and supported by site-directed mutagenesis of the Con1 replicon, we confirmed that mutations M423I/T/V mediate phenotypic resistance. Selection in patients of an NS5B mutation at M423 was associated with a reduced replicative capacityin vitrorelative to the pretherapy sequence; consistent with this, reversion to wild-type M423 was observed in the majority of patients following therapy cessation. Mutations at NS5B residues R422 and M426 were detected in a small number of patients at baseline or the end of therapy and also mediate reductions in filibuvir susceptibility, suggesting these are rare but clinically relevant alternative resistance pathways. Amino acid variants at position M423 in HCV NS5B polymerase are the preferred pathway for selection of viral resistance to filibuvirin vivo.

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Development and validation of a sequential two-step algorithm for the screening of individuals with potential polycythaemia vera

Scientific Reports ◽

10.1038/s41598-020-80459-y ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Miguel Piris-Villaespesa ◽

Alberto Álvarez-Larrán ◽

Adolfo Saez-Marín ◽

Claudia Nuñez-Torrón ◽

Gloria Muñoz-Martin ◽

...

Keyword(s):

Mutation Screening ◽

Roc Curves ◽

Polycythaemia Vera ◽

Normal Ranges ◽

Screening Algorithm ◽

Number Of Patients ◽

Step Algorithm ◽

Development And Validation ◽

Diagnostic Criterion ◽

Selection Of

AbstractIn 2016, the WHO included haemoglobin values within normal ranges as a diagnostic criterion for Polycythaemia Vera (PV). Since then, concerns have arisen that a large number of patients are undergoing unnecessary screening for PV. To address this issue, we estimated the prevalence of JAK2 p.V617F in individuals with elevated haemoglobin or haematocrit and developed and validated a screening algorithm for PV. A total of 15,366 blood counts performed in seven non-consecutive days were reviewed, of which 1001 were selected for subsequent JAK2 p.V617F mutation screening. Eight (0.8%) new JAK2 p.V617F-mutated cases were detected. From ROC curves, a two-step algorithm was developed based on the optimal cut-off for the detection of the JAK2 p.V617F mutation. The algorithm was prospectively validated in an independent cohort of 15,298 blood counts. A total of 1595 (10.4%) cases met the criterion for haemoglobin or haematocrit, of whom 581 passed to step 2 (3.8% of the total). The JAK2 p.V617F mutation was detected in 7 of the 501 patients tested, which accounts for 0.04% of the total cohort and 0.4% of patients with erythrocytosis. In conclusion, this data show that our two-step algorithm improves the selection of candidates for JAK2 p.V617F testing.

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Cytotoxic Activity of Antineoplastic Agents on Fertility: A Systematic Review

Revista Brasileira de Ginecologia e Obstetrícia / RBGO Gynecology and Obstetrics ◽

10.1055/s-0040-1713911 ◽

2020 ◽

Vol 42 (11) ◽

pp. 759-768

Author(s):

Gabriel Acácio de Moura ◽

Paula Bruno Monteiro

Keyword(s):

Antineoplastic Agents ◽

Site Effect ◽

Cytotoxic Action ◽

Long Term Effects ◽

Treatment Type ◽

Number Of Patients ◽

Serum Hormone ◽

Hormonal Modulation ◽

Selection Of

Abstract Objective To analyze the long-term effects of antineoplastic treatments on patient fertility. Selection of Studies The studies were selected through the New PubMed, Scielo and Lilacs databases along with references used for the creation of the present work. For the selection of studies, articles published between the periods from January 1, 2015 to April 6, 2020 in the English, Portuguese and Spanish languages were used. As inclusion criteria: cohort studies and studies conducted in vitro. As exclusion criteria: review articles, reported cases, studies that do not address thematic reproduction, studies that do not address the cancer theme, articles that used animals, articles that address the preservation of fertility and articles in duplicate in the bases. Data Collection The collected data included: age of the patient at the beginning of treatment, type of neoplasm, type of antineoplastic treatment, chemotherapy used, radiotherapy dosage, radiotherapy site, effect of antineoplastic agents on fertility and number of patients in the study. Data Synthesis Thirty studies were evaluated, antineoplastic chemotherapy agents and radiotherapy modulate serum hormone levels, reduces germ cell quantities and correlated with an increase in sterility rates. The effects mentioned occur in patients in the prepubertal and postpubertal age. Conclusion Antineoplastic treatments have cytotoxic effects on the germ cells leading to hormonal modulation, and pubertal status does not interfere with the cytotoxic action of therapies.

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Enriching CBT by Neuroscience: Novel Avenues to Achieve Personalized Treatments

International Journal of Cognitive Therapy ◽

10.1007/s41811-020-00089-0 ◽

2020 ◽

Author(s):

Kristoffer N T Månsson ◽

Ulrike Lueken ◽

Andreas Frick

Keyword(s):

Mental Disorders ◽

Common Mental Disorders ◽

Behavioral Therapy ◽

Therapy Outcomes ◽

Efficient Treatment ◽

Related Research ◽

Number Of Patients ◽

Key Topics ◽

Selection Of ◽

The Way

Abstract Although cognitive behavioral therapy (CBT) is an established and efficient treatment for a variety of common mental disorders, a considerable number of patients do not respond to treatment or relapse after successful CBT. Recent findings and approaches from neuroscience could pave the way for clinical developments to enhance the outcome of CBT. Herein, we will present how neuroscience can offer novel perspectives to better understand (a) the biological underpinnings of CBT, (b) how we can enrich CBT with neuroscience-informed techniques (augmentation of CBT), and (c) why some patients may respond better to CBT than others (predictors of therapy outcomes), thus paving the way for more personalized and effective treatments. We will introduce some key topics and describe a selection of findings from CBT-related research using tools from neuroscience, with the hope that this will provide clinicians and clinical researchers with a brief and comprehensible overview of the field.

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Appropriate selection of patients with lumbar spinal stenosis for interspinous process decompression with the X STOP device

Neurosurgical FOCUS ◽

10.3171/foc.2007.22.1.5 ◽

2007 ◽

Vol 22 (1) ◽

pp. 1-7 ◽

Cited By ~ 22

Author(s):

Carl Lauryssen

Keyword(s):

Spinal Stenosis ◽

Lumbar Spinal Stenosis ◽

Neurogenic Intermittent Claudication ◽

Anatomical Structures ◽

Conservative Care ◽

Number Of Patients ◽

Spinous Processes ◽

Lumbar Spinal ◽

Selection Of Patients ◽

Selection Of

✓With the aging of the population, the number of patients suffering from progressive lumbar spinal stenosis with symptomatic neurogenic intermittent claudication is projected to increase. Unfortunately, these patients are limited to a choice between nonsurgical conservative care and more invasive decompressive surgical procedures such as laminectomy with or without fusion. The X STOP interspinous process decompression system is a commercially available device that provides a minimally invasive alternative treatment, an intermediate option within the continuum of care for these patients. The X STOP is appropriate for patients with moderately severe functional impairment whose symptoms are exacerbated in extension and relieved in flexion. Implanted between the spinous processes without disrupting the normal anatomical structures, the X STOP limits narrowing of the spinal canal and neural foramina by reducing extension at the symptomatic level(s). In this report the author details the approved indications for use of the X STOP and discusses several illustrative cases.

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POSITIVE: Perfusion imaging selection of ischemic stroke patients for endovascular therapy

Journal of NeuroInterventional Surgery ◽

10.1136/neurintsurg-2021-017315 ◽

2021 ◽

pp. neurintsurg-2021-017315

Author(s):

J Mocco ◽

Adnan H Siddiqui ◽

David Fiorella ◽

Michael J Alexander ◽

Adam S Arthur ◽

...

Keyword(s):

Ischemic Stroke ◽

Endovascular Therapy ◽

Perfusion Imaging ◽

Medical Management ◽

Primary Outcome ◽

Functional Improvement ◽

Positive Trial ◽

Stroke Patients ◽

Number Of Patients ◽

Selection Of

BackgroundThe PerfusiOn imaging Selection of Ischemic sTroke patIents for endoVascular thErapy (POSITIVE) trial was designed to evaluate functional outcome in patients with emergent large vessel occlusion (ELVO) presenting within 0–12 hours with pre-specified bifurcated arms of early and late window presentation, who were selected for endovascular thrombectomy with non-vendor specific commercially available perfusion imaging software. Recent trials demonstrating the benefit of thrombectomy up to 16–24 hours following ELVO removed equipoise to randomize late window ELVO patients and therefore the trial was halted.MethodsUp to 200 patients were to be enrolled in this FDA-cleared, prospective, randomized, multicenter international trial to compare thrombectomy and best medical management in patients with ELVO ineligible for or refractory to treatment with IV tissue plasminogen activator (IV-tPA) selected with perfusion imaging and presenting within 0–12 hours of last seen normal. The primary outcome was 90-day clinical outcome as measured by the raw modified Rankin Scale (mRS) with scores 5 and 6 collapsed (mRS shift analysis).ResultsThe POSITIVE trial suspended enrollment with the release of results from the DAWN trial and was stopped after the release of the DEFUSE 3 trial results. Thirty-three patients were enrolled (21 for medical management and 12 for thrombectomy). Twelve of the 33 patients were enrolled in the 6–12 hour cohort. Despite the early cessation, the primary outcome demonstrated statistically significant superior clinical outcomes for patients treated with thrombectomy (P=0.0060). The overall proportion of patients achieving an mRS score of 0–2 was 75% in the thrombectomy cohort and 43% in the medical management cohort (OR 4.00, 95% CI 0.84 to 19.2).ConclusionPOSITIVE supports the already established practice of delayed thrombectomy for appropriately selected patients presenting within 0–12 hours selected by perfusion imaging from any vendor. The results of the POSITIVE trial are consistent with other thrombectomy trials. The statistically significant effect on functional improvement, despite the small number of patients, reinforces the robust benefits of thrombectomy.Clinical trial registrationNCT01852201

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Nine new cases of reciprocal translocation in the domestic pig (Sus scrofa domestica L.)

Journal of Heredity ◽

10.1093/jhered/89.2.136 ◽

1998 ◽

Vol 89 (2) ◽

pp. 136-142 ◽

Cited By ~ 26

Author(s):

A. Ducos ◽

H. M. Berland ◽

A. Pinton ◽

E. Guillemot ◽

A. Seguela ◽

...

Keyword(s):

Reciprocal Translocation ◽

Sus Scrofa ◽

Domestic Pig ◽

Sus Scrofa Domestica

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A Donor Utilization Index to Assess the Utilization and Discard of Deceased Donor Kidneys Perceived as High Risk

Clinical Journal of the American Society of Nephrology ◽

10.2215/cjn.02770319 ◽

2019 ◽

Vol 14 (11) ◽

pp. 1634-1641 ◽

Cited By ~ 9

Author(s):

Corey Brennan ◽

Syed Ali Husain ◽

Kristen L. King ◽

Demetra Tsapepas ◽

Lloyd E. Ratner ◽

...

Keyword(s):

High Risk ◽

Regional Variation ◽

Organ Procurement ◽

Deceased Donor ◽

Level Variation ◽

Donor Kidney ◽

Perceptions Of Risk ◽

Deceased Donor Kidney ◽

Number Of Patients ◽

Donor Characteristics

Background and objectivesAn increasing number of patients on the waitlist for a kidney transplant indicates a need to effectively utilize as many deceased donor kidneys as possible while ensuring acceptable outcomes. Assessing regional and center-level organ utilization with regards to discard can reveal regional variation in suboptimal deceased donor kidney acceptance patterns stemming from perceptions of risk.Design, setting, participants, & measurementsWe created a weighted donor utilization index from a logistic regression model using high-risk donor characteristics and discard rates from 113,640 deceased donor kidneys procured for transplant from 2010 to 2016, and used it to examine deceased donor kidney utilization in 182 adult transplant centers with >15 annual deceased donor kidney transplants. Linear regression and correlation were used to analyze differences in donor utilization indexes.ResultsThe donor utilization index was found to significantly vary by Organ Procurement and Transplantation Network region (P<0.001), revealing geographic trends in kidney utilization. When investigating reasons for this disparity, there was no significant correlation between center volume and donor utilization index, but the percentage of deceased donor kidneys imported from other regions was significantly associated with donor utilization for all centers (rho=0.39; P<0.001). This correlation was found to be particularly strong for region 4 (rho=0.83; P=0.001) and region 9 (rho=0.82; P=0.001). Additionally, 25th percentile time to transplant was weakly associated with the donor utilization index (R2=0.15; P=0.03).ConclusionsThere is marked center-level variation in the use of deceased donor kidneys with less desirable characteristics both within and between regions. Broader utilization was significantly associated with shorter time to transplantation.

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