Adaptive DNA amplification of synthetic gene circuit opens a way to overcome cancer chemoresistance.

Drug resistance continues to impede the success of cancer treatments, creating a need for experimental model systems that are broad, yet simple, to allow the identification of mechanisms and novel countermeasures applicable to many cancer types. To address these needs, we investigated a set of engineered mammalian cell lines with synthetic gene circuits integrated into their genome that evolved resistance to Puromycin. We identified DNA amplification as the mechanism underlying drug resistance in 4 out of 6 replicate populations. Triplex-forming oligonucleotide (TFO) treatment combined with Puromycin could efficiently suppress the growth of cell populations with DNA amplification. Similar observations in human cancer cell lines suggest that TFOs could be broadly applicable to mitigate drug resistance, one of the major difficulties in treating cancer.

Monitoring Genomic Structural Rearrangements Resulting from Gene Editing.

The cytogenomics-based methodology of directional genomic hybridization (dGH) enables the detection and quantification of a more comprehensive spectrum of genomic structural variants than any other approach currently available, and importantly, does so on a single-cell basis. Thus, dGH is well-suited for testing and/or validating new advancements in CRISPR-Cas9 gene editing systems. In addition to aberrations detected by traditional cytogenetic approaches, the strand specificity of dGH facilitates detection of otherwise cryptic intra-chromosomal rearrangements, specifically small inversions. As such, dGH represents a powerful, high-resolution approach for the quantitative monitoring of potentially detrimental genomic structural rearrangements resulting from exposure to agents that induce DNA double-strand breaks (DSBs), including restriction endonucleases and ionizing radiations. For intentional genome editing strategies, it is critical that any undesired effects of DSBs induced either by the editing system itself or by mis-repair with other endogenous DSBs are recognized and minimized. In this paper, we discuss the application of dGH for assessing gene editing-associated structural variants and the potential heterogeneity of such rearrangements among cells within an edited population, highlighting its relevance to personalized medicine strategies.

A reference human induced pluripotent stem cell line for large-scale collaborative studies.

Human induced pluripotent stem cell (iPSC) lines are a powerful tool for studying development and disease, but the considerable phenotypic variation between lines makes it challenging to replicate key findings and integrate data across research groups. To address this issue, we sub-cloned candidate human iPSC lines and deeply characterized their genetic properties using whole genome sequencing, their genomic stability upon CRISPR-Cas9-based gene editing, and their phenotypic properties including differentiation to commonly used cell types. These studies identified KOLF2.1J as an all-around well-performing iPSC line. We then shared KOLF2.1J with groups around the world who tested its performance in head-to-head comparisons with their own preferred iPSC lines across a diverse range of differentiation protocols and functional assays. On the strength of these findings, we have made KOLF2.1J and its gene-edited derivative clones readily accessible to promote the standardization required for large-scale collaborative science in the stem cell field.

The Impact of COVID-19 on Patients With ADPKD.

PURPOSE OF REVIEW: Patients with autosomal dominant polycystic kidney disease (ADPKD) have kidney cysts and kidney enlargement decades before progressing to advanced chronic kidney disease (CKD), meaning patients live most of their adult life with a chronic medical condition. The coronavirus disease 2019 (COVID-19) pandemic has created common questions among patients with ADPKD. In this review, we discuss COVID-19 concerns centered around a patient with a common clinical vignette. SOURCES OF INFORMATION: We performed PubMed and Google scholar searches for English, peer-reviewed studies related to "COVID-19," "ADPKD," "CKD," "tolvaptan," "angiotensin-converting enzyme inhibitors" (ACEi), "angiotensin receptor blockers" (ARB), and "vaccination." We also evaluated transplant data provided by the Ontario Trillium Gift of Life Network. METHODS: Following an assessment of available literature, this narrative review addresses common questions of patients with ADPKD in the context of the COVID-19 pandemic. KEY FINDINGS: Data regarding the risk of developing COVID-19 and the risk of adverse COVID-19 outcomes in patients with ADPKD remain limited, but patients with ADPKD with impaired estimated glomerular filtration rate (eGFR), kidney transplants, or on dialysis are likely at similar increased risk as those with generally defined CKD. We provide strategies to improve virtual care, which is likely to persist after the pandemic. Current evidence suggests ACEi, ARB, and tolvaptan treatment should be continued unless contraindicated due to severe illness. When available, and in the absence of a severe allergy, vaccination is recommended for all patients with ADPKD. LIMITATIONS: This narrative review is limited by a paucity of high-quality data on COVID-19 outcomes in patients specifically with ADPKD. IMPLICATIONS: Patients with ADPKD who have developed advanced CKD, require dialysis, or who have received a kidney transplant are at elevated risk of COVID-19 complications.