GMP-compliant iPS cell lines show widespread plasticity in a new set of differentiation workflows for cell replacement and cancer immunotherapy.

Cell therapeutic applications based on induced pluripotent stem cells (iPSCs) appear highly promising and challenging at the same time. Good manufacturing practice (GMP) regulations impose necessary yet demanding requirements for quality and consistency when manufacturing iPSCs and their differentiated progeny. Given the scarcity of accessible GMP iPSC lines, we have established a corresponding production workflow to generate the first set of compliant cell banks.

Identification of genome edited cells using CRISPRnano.

Genome engineering-induced cleavage sites can be resolved by non-homologous end joining (NHEJ) or homology-directed repair (HDR). Identifying genetically modified clones at the target locus remains an intensive and laborious task. Different workflows and software that rely on deep sequencing data have been developed to detect and quantify targeted mutagenesis.

Impaired p53-Mediated DNA Damage Response Contributes to Microcephaly in Nijmegen Breakage Syndrome Patient-Derived Cerebral Organoids.

Nijmegen Breakage Syndrome (NBS) is a rare autosomal recessive genetic disorder caused by mutations within nibrin (), a DNA damage repair protein. Hallmarks of NBS include chromosomal instability and clinical manifestations such as growth retardation, immunodeficiency, and progressive microcephaly. We employed induced pluripotent stem cell-derived cerebral organoids from two NBS patients to study the etiology of microcephaly.

A call for consensus guidelines on monitoring the integrity of nuclear and mitochondrial genomes in human pluripotent stem cells.

Despite the widespread use of human pluripotent stem cells (hPSCs), there is no general consensus with respect to monitoring the integrity of nuclear and mitochondrial genome of hPSCs. We argue that clear guidelines should be in place, and we have identified frequently asked questions that can be used to develop a practical guide for hPSCs users.

Generation of an induced pluripotent stem cell line (IUFi001) from a Cockayne syndrome patient carrying a mutation in the ERCC6 gene.

Human fibroblasts from a Cockayne Syndrome (CS) patient carrying the compound heterozygous c.1131 A > T and c.2571C > T within ERCC Excision Repair 6 (ERCC6) were reprogramed to generate integration-free induced pluripotent stem cells (iPSCs).

Fast but not furious: a streamlined selection method for genome-edited cells.

In the last decade, transcription activator-like effector nucleases and CRISPR-based genome engineering have revolutionized our approach to biology. Because of their high efficiency and ease of use, the development of custom knock-out and knock-in animal or cell models is now within reach for almost every laboratory. Nonetheless, the generation of genetically modified cells often requires a selection step, usually achieved by antibiotics or fluorescent markers.

Generation of a Crigler-Najjar Syndrome Type I patient-derived induced pluripotent stem cell line CNS705 (HHUUKDi005-A).

Human fibroblasts cells from a Crigler-Najjar Syndrome (CNS) patient were used to generate integration-free induced pluripotent stem cells (iPSCs) by over-expressing episomal-based plasmids expressing OCT4, SOX2, NANOG, KLF4, c-MYC and LIN28. The derived CNS705-iPSC line is homozygous for the UGT1A1 c.877_890delTACATTAATGCTTCinsA mutation.

A stem cell based model of NAFLD enables the analysis of patient specific individual metabolic adaptations in response to a high fat diet and AdipoRon interference.

Non-alcoholic fatty liver disease (NAFLD) is a multifactorial disease. Its development and progression depend on genetically predisposed susceptibility of the patient towards several 'hits' that induce fat storage first and later inflammation and fibrosis. Here, we differentiated induced pluripotent stem cells (iPSCs) derived from four distinct donors with varying disease stages into hepatocyte like cells (HLCs) and determined fat storage as well as metabolic adaptations after stimulations with oleic acid.

Matrix Metalloproteinase 14 Mediates APP Proteolysis and Lysosomal Alterations Induced by Oxidative Stress in Human Neuronal Cells.

The alteration of amyloid precursor protein (APP) proteolysis is a hallmark of Alzheimer's disease (AD). Recent studies have described noncanonical pathways of APP processing that seem partly executed by lysosomal enzymes. Our laboratory's human SK-N-MC model has shown that oxidative stress (OS) alters the lysosomal degradation pathway and the processing/metabolism of APP.

IPSC-Derived Neuronal Cultures Carrying the Alzheimer's Disease Associated R47H Variant Enables the Construction of an Aβ-Induced Gene Regulatory Network.

Genes associated with immune response and inflammation have been identified as genetic risk factors for late-onset Alzheimer´s disease (LOAD). The rare R47H variant within triggering receptor expressed on myeloid cells 2 (TREM2) has been shown to increase the risk for developing Alzheimer's disease (AD) 2-3-fold. Here, we report the generation and characterization of a model of late-onset Alzheimer's disease (LOAD) using lymphoblast-derived induced pluripotent stem cells (iPSCs) from patients carrying the TREM2 R47H mutation, as well as from control individuals without dementia.

AP-2 reduces amyloidogenesis by promoting BACE1 trafficking and degradation in neurons.

Cleavage of amyloid precursor protein (APP) by BACE-1 (β-site APP cleaving enzyme 1) is the rate-limiting step in amyloid-β (Aβ) production and a neuropathological hallmark of Alzheimer's disease (AD). Despite decades of research, mechanisms of amyloidogenic APP processing remain highly controversial. Here, we show that in neurons, APP processing and Aβ production are controlled by the protein complex-2 (AP-2), an endocytic adaptor known to be required for APP endocytosis.

The FGF, TGFβ and WNT axis Modulate Self-renewal of Human SIX2 Urine Derived Renal Progenitor Cells.

Human urine is a non-invasive source of renal stem cells with regeneration potential. Urine-derived renal progenitor cells were isolated from 10 individuals of both genders and distinct ages. These renal progenitors express pluripotency-associated proteins- TRA-1-60, TRA-1-81, SSEA4, C-KIT and CD133, as well as the renal stem cell markers -SIX2, CITED1, WT1, CD24 and CD106.

Molybdenum(ii) complexes with p-substituted BIAN ligands: synthesis, characterization, biological activity and computational study.

New complexes [Mo(η3-C3H5)X(CO)2(4-Y-BIAN)] (4-Y-BIAN = bis(4-Y-phenyl)-acenaphthenequinonediimine), with X = Br and Y = H, Me, OMe, COOH and X = Cl, Y = OMe, as well as the cation with X = NCMe and Y = OMe were synthesized, expanding the scope of this family. Two single crystal X-ray structures (X = Br, Y = Me, OMe) display a less symmetric arrangement (axial isomer), where one N donor atom is trans to the allyl group and the second to one CO. DFT studies showed similar energies for the two possible isomers of the complexes, with a very small preference for the observed axial isomer.

CD81 Promotes a Migratory Phenotype in Neuronal-Like Cells.

Tetraspanins, such as CD81, can form lateral associations with each other and with other transmembrane proteins. These interactions may underlie CD81 functions in multiple cellular processes, such as adhesion, morphology, migration, and differentiation. Since CD81's role in neuronal cells' migration has not been established, we here evaluated effects of CD81 on the migratory phenotype of SH-SY5Y neuroblastoma cells.

Anti-EGFR Therapy to Treat Metastatic Colorectal Cancer: Not for All.

The development of monoclonal antibodies (mAbs) cetuximab and panitumumab, which target the transmembrane protein epidermal growth factor receptor (EGFR), mark a major step forward in the treatment of metastatic colorectal cancer (mCRC). However, this therapeutic progress proved to be effective only in a very restricted subset of patients. Although several mechanisms of resistance, both primary and acquired, have been identified, the only established predictive tumour biomarker for the treatment of mCRC patients is the RAS mutational status.

Fibroblast-derived integration-free iPSC line ISRM-NBS1 from an 18-year-old Nijmegen Breakage Syndrome patient carrying the homozygous NBN c.657_661del5 mutation.

Human fibroblasts cells from a female diagnosed with Nijmegen Breakage Syndrome (NBS) carrying the homozygous NBN c.657_661del5 mutation were used to generate integration-free induced pluripotent stem cells (iPSCs) by over-expressing episomal-based plasmids harbouring OCT4, SOX2, NANOG, KLF4, c-MYC and LIN28. The derived iPSC line - ISRM-NBS1 was defined as pluripotent based on (i) expression of pluripotency-associated markers (ii) embryoid body-based differentiation into cell types representative of the three germ layers and (iii) the similarity between the transcriptome of the iPSC line and the human embryonic stem cell line H1 with a Pearson correlation of 0.

Lymphoblast-derived integration-free iPSC line AD-TREM2-3 from a 74 year-old Alzheimer's disease patient expressing the TREM2 p.R47H variant.

Human lymphoblast cells from a male diagnosed with Alzheimer's disease (AD) expressing the TREM2 p.R47H variant were used to generate integration-free induced pluripotent stem cells (iPSCs) by over-expressing episomal-based plasmids harbouring OCT4, SOX2, KLF4, LIN28, L-MYC and p53 shRNA. The derived iPSC line - AD-TREM2-3 was defined as pluripotent based on (i) expression of pluripotency-associated markers (ii) embryoid body-based differentiation into cell types representative of the three germ layers and (iii) the similarity between the transcriptome of the iPSC line and the human embryonic stem cell line H1 with a Pearson correlation of 0.

Lymphoblast-derived integration-free iPSC line AD-TREM2-1 from a 67year-old Alzheimer's disease patient expressing the TREM2 p.R47H variant.

Human lymphoblast cells from a male diagnosed with Alzheimer's disease (AD) expressing the TREM2 p.R47H variant were used to generate integration-free induced pluripotent stem cells (iPSCs) by over-expressing episomal-based plasmids harbouring OCT4, SOX2, NANOG, LIN28, c-MYC and L-MYC. AD-TREM2-1 was defined as pluripotent based on (i) expression of pluripotency-associated markers (ii) embryoid body-based differentiation into cell types representative of the three germ layers and (iii) the similarity between the transcriptome of the iPSC line and the human embryonic stem cell line H1 with a Pearson correlation of 0.

Lymphoblast-derived integration-free ISRM-CON9 iPS cell line from a 75year old female.

Human lymphoblast cells were used to generate integration-free induced pluripotent stem cells (iPSCs) employing episomal-based plasmids expressing OCT4, SOX2, NANOG, LIN28, c-MYC and L-MYC. The derived iPSCs were defined as pluripotent based on (i) expression of pluripotency-associated markers, (ii) embryoid body-based differentiation into cell types representative of the three germ layers and (iii) the similarity between the transcriptomes of the iPSC line and the human embryonic stem cell line H1 with a Pearson correlation of 0.95.

Nijmegen Breakage Syndrome fibroblasts and iPSCs: cellular models for uncovering disease-associated signaling pathways and establishing a screening platform for anti-oxidants.

Nijmegen Breakage Syndrome (NBS) is associated with cancer predisposition, premature aging, immune deficiency, microcephaly and is caused by mutations in the gene coding for NIBRIN (NBN) which is involved in DNA damage repair. Dermal-derived fibroblasts from NBS patients were reprogrammed into induced pluripotent stem cells (iPSCs) in order to bypass premature senescence. The influence of antioxidants on intracellular levels of ROS and DNA damage were screened and it was found that EDHB-an activator of the hypoxia pathway, decreased DNA damage in the presence of high oxidative stress.

Strong inhibition of thioredoxin reductase by highly cytotoxic gold(I) complexes. DNA binding studies.

Biological properties of a series of aminophosphine-thiolate gold(I) complexes [Au(SR)(PPh2NHpy)] [Ph2PNHpy=2-(diphenylphosphinoamino)pyridine; HSR=2-mercaptopyridine (2-HSpy) (3), 2-mercaptonicotinic acid (2-H2-mna) (4), 2-thiouracil (2-HTU) (5) or 2-thiocytosine (2-HTC) (6)] and [Au(SR){PPh2NH(Htrz)}] [Ph2PNH(Htrz)=3-(diphenylphosphinoamino)-1,2,4-triazole]; HSR=2-mercaptopyridine (2-HSpy) (7), 2-thiocytosine (2-HTC) (8) or 6-thioguanine (6-HTG) (9) have been studied. Their antitumor properties have been tested in vitro against two tumor human cell lines, HeLa (derived from cervical cancer) and MCF-7 (derived from breast cancer), using a metabolic activity test (3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide, MTT). Some of them showed excellent cytotoxic activity.