Mutation Patterns Predict Drug Sensitivity in Acute Myeloid Leukemia.

Purpose: The inherent genetic heterogeneity of acute myeloid leukemia (AML) has challenged the development of precise and effective therapies. The objective of this study was to elucidate the genomic basis of drug resistance or sensitivity, identify signatures for drug response prediction, and provide resources to the research community.

Experimental Design: We performed targeted sequencing, high-throughput drug screening, and single-cell genomic profiling on leukemia cell samples derived from patients with AML.

Multiplex single-cell chemical genomics reveals the kinase dependence of the response to targeted therapy.

Chemical genetic screens are a powerful tool for exploring how cancer cells' response to drugs is shaped by their mutations, yet they lack a molecular view of the contribution of individual genes to the response to exposure. Here, we present sci-Plex-Gene-by-Environment (sci-Plex-GxE), a platform for combined single-cell genetic and chemical screening at scale. We highlight the advantages of large-scale, unbiased screening by defining the contribution of each of 522 human kinases to the response of glioblastoma to different drugs designed to abrogate signaling from the receptor tyrosine kinase pathway.

Multiplex single-cell chemical genomics reveals the kinase dependence of the response to targeted therapy.

Chemical genetic screens are a powerful tool for exploring how cancer cells' response to drugs is shaped by their mutations, yet they lack a molecular view of the contribution of individual genes to the response to exposure. Here, we present sci-Plex-ene-by-nvironment (sci-Plex-x), a platform for combined single-cell genetic and chemical screening at scale. We highlight the advantages of large-scale, unbiased screening by defining the contribution of each of 522 human kinases to the response of glioblastoma to different drugs designed to abrogate signaling from the receptor tyrosine kinase pathway.

Fanconi anemia-isogenic head and neck cancer cell line pairs: A basic and translational science resource.

Fanconi anemia (FA) is a heritable malformation, bone marrow failure and cancer predisposition syndrome that confers an exceptionally high risk of squamous carcinomas. These carcinomas originate in epithelia lining the mouth, proximal esophagus, vulva and anus: their origins are not understood, and no effective ways have been identified to prevent or delay their appearance. Many FA-associated carcinomas are also therapeutically challenging: they may be multi-focal and stage-advanced at diagnosis, and most individuals with FA cannot tolerate standard-of-care systemic therapies such as DNA cross-linking drugs or ionizing radiation due to constitutional DNA damage hypersensitivity.

A functional module states framework reveals transcriptional states for drug and target prediction.

Cells are complex systems in which many functions are performed by different genetically defined and encoded functional modules. To systematically understand how these modules respond to drug or genetic perturbations, we develop a functional module states framework. Using this framework, we (1) define the drug-induced transcriptional state space for breast cancer cell lines using large public gene expression datasets and reveal that the transcriptional states are associated with drug concentration and drug targets, (2) identify potential targetable vulnerabilities through integrative analysis of transcriptional states after drug treatment and gene knockdown-associated cancer dependency, and (3) use functional module states to predict transcriptional state-dependent drug sensitivity and build prediction models for drug response.

Unsupervised discovery of dynamic cell phenotypic states from transmitted light movies.

Identification of cell phenotypic states within heterogeneous populations, along with elucidation of their switching dynamics, is a central challenge in modern biology. Conventional single-cell analysis methods typically provide only indirect, static phenotypic readouts. Transmitted light images, on the other hand, provide direct morphological readouts and can be acquired over time to provide a rich data source for dynamic cell phenotypic state identification.

High-throughput, microscope-based sorting to dissect cellular heterogeneity.

Microscopy is a powerful tool for characterizing complex cellular phenotypes, but linking these phenotypes to genotype or RNA expression at scale remains challenging. Here, we present Visual Cell Sorting, a method that physically separates hundreds of thousands of live cells based on their visual phenotype. Automated imaging and phenotypic analysis directs selective illumination of Dendra2, a photoconvertible fluorescent protein expressed in live cells; these photoactivated cells are then isolated using fluorescence-activated cell sorting.

Tumour predisposition and cancer syndromes as models to study gene-environment interactions.

Cell division and organismal development are exquisitely orchestrated and regulated processes. The dysregulation of the molecular mechanisms underlying these processes may cause cancer, a consequence of cell-intrinsic and/or cell-extrinsic events. Cellular DNA can be damaged by spontaneous hydrolysis, reactive oxygen species, aberrant cellular metabolism or other perturbations that cause DNA damage.

New Human Chromosomal Sites with "Safe Harbor" Potential for Targeted Transgene Insertion.

This study identified 35 new sites for targeted transgene insertion that have the potential to serve as new human genomic "safe harbor" sites (SHS). SHS potential for these 35 sites, located on 16 chromosomes, including both arms of the human X chromosome, and for the existing human SHS , , and was assessed using eight different desirable, widely accepted criteria for SHS verifiable with human genomic data. Three representative newly identified sites on human chromosomes 2 and 4 were then experimentally validated by and cleavage-sensitivity tests, and analyzed for population-level and cell line-specific sequence variants that might confound site targeting.

An RNAi screen in human cell lines reveals conserved DNA damage repair pathways that mitigate formaldehyde sensitivity.

Formaldehyde is a ubiquitous DNA damaging agent, with human exposures occurring from both exogenous and endogenous sources. Formaldehyde exposure can result in multiple types of DNA damage, including DNA-protein crosslinks and thus, is representative of other exposures that induce DNA-protein crosslinks such as cigarette smoke, automobile exhaust, wood smoke, metals, ionizing radiation, and certain chemotherapeutics. Our objective in this study was to identify the genes necessary to mitigate formaldehyde toxicity following chronic exposure in human cells.

Genomic and Molecular Landscape of DNA Damage Repair Deficiency across The Cancer Genome Atlas.

DNA damage repair (DDR) pathways modulate cancer risk, progression, and therapeutic response. We systematically analyzed somatic alterations to provide a comprehensive view of DDR deficiency across 33 cancer types. Mutations with accompanying loss of heterozygosity were observed in over 1/3 of DDR genes, including TP53 and BRCA1/2.

Werner syndrome (WRN) gene variants and their association with altered function and age-associated diseases.

Werner syndrome (WS) is a heritable autosomal recessive human disorder characterized by the premature onset of several age-associated pathologies including cancer. The protein defective in WS patients, WRN, is encoded by a member of the human RECQ gene family that contains both a DNA exonuclease and a helicase domain. WRN has been shown to participate in several DNA metabolic pathways including DNA replication, recombination and repair, as well as telomere maintenance and transcription modulation.

Homozygosity for the WRN Helicase-Inactivating Variant, R834C, does not confer a Werner syndrome clinical phenotype.

Loss-of-function mutations in the WRN helicase gene cause Werner syndrome- a progeroid syndrome with an elevated risk of cancer and other age-associated diseases. Large numbers of single nucleotide polymorphisms have been identified in WRN. We report here the organismal, cellular, and molecular phenotypes of variant rs3087425 (c.

Human RECQ Helicase Pathogenic Variants, Population Variation and "Missing" Diseases.

Heritable loss of function mutations in the human RECQ helicase genes BLM, WRN, and RECQL4 cause Bloom, Werner, and Rothmund-Thomson syndromes, cancer predispositions with additional developmental or progeroid features. In order to better understand RECQ pathogenic and population variation, we systematically analyzed genetic variation in all five human RECQ helicase genes. A total of 3,741 unique base pair-level variants were identified, across 17,605 potential mutation sites.

Metformin improves defective hematopoiesis and delays tumor formation in Fanconi anemia mice.

Fanconi anemia (FA) is an inherited bone marrow failure disorder associated with a high incidence of leukemia and solid tumors. Bone marrow transplantation is currently the only curative therapy for the hematopoietic complications of this disorder. However, long-term morbidity and mortality remain very high, and new therapeutics are badly needed.

Werner syndrome through the lens of tissue and tumour genomics.

Werner syndrome (WS) is the canonical adult human progeroid ('premature aging') syndrome. Patients with this autosomal recessive Mendelian disorder display constitutional genomic instability and an elevated risk of important age-associated diseases including cancer. Remarkably few analyses of WS patient tissue and tumors have been performed to provide insight into WS disease pathogenesis or the high risk of neoplasia.

Microwell arrays reveal cellular heterogeneity during the clonal expansion of transformed human cells.

We developed micromolded microwell arrays to study the proliferation and senescence of single cells. Microwell arrays were designed to be compatible with conventional cell culture protocols to simplify cell loading, cell culture, and imaging. We demonstrated the utility of these arrays by measuring the proliferation and senescence of isogenic cells which expressed or had been depleted of the human Werner syndrome protein.

WRN Promoter CpG Island Hypermethylation Does Not Predict More Favorable Outcomes for Patients with Metastatic Colorectal Cancer Treated with Irinotecan-Based Therapy.

Purpose: WRN promoter CpG island hypermethylation in colorectal cancer has been reported to increase sensitivity to irinotecan-based therapies. We aimed to characterize methylation of the WRN promoter, determine the effect of WRN promoter hypermethylation upon expression, and validate a previous report that WRN promoter hypermethylation predicts improved outcomes for patients with metastatic colorectal cancer (mCRC) treated with irinotecan-based therapy.

Experimental Design: WRN methylation status was assessed using methylation-specific PCR and bisulfite sequencing assays.

Werner syndrome: Clinical features, pathogenesis and potential therapeutic interventions.

Werner syndrome (WS) is a prototypical segmental progeroid syndrome characterized by multiple features consistent with accelerated aging. It is caused by null mutations of the WRN gene, which encodes a member of the RECQ family of DNA helicases. A unique feature of the WRN helicase is the presence of an exonuclease domain in its N-terminal region.

The Werner syndrome RECQ helicase targets G4 DNA in human cells to modulate transcription.

The Werner syndrome (WS) is a prototypic adult Mendelian progeroid syndrome in which signs of premature aging are associated with genomic instability and an elevated risk of cancer. The WRN RECQ helicase protein binds and unwinds G-quadruplex (G4) DNA substrates in vitro, and we identified significant enrichment in G4 sequence motifs at the transcription start site and 5' ends of first introns (false discovery rate < 0.001) of genes down-regulated in WS patient fibroblasts.

Development of a Whole Blood Paper-Based Device for Phenylalanine Detection in the Context of PKU Therapy Monitoring.

Laboratory-based testing does not allow for the sufficiently rapid return of data to enable optimal therapeutic monitoring of patients with metabolic diseases such as phenylketonuria (PKU). The typical turn-around time of several days for current laboratory-based testing is too slow to be practically useful for effective monitoring or optimizing therapy. This report describes the development of a rapid, paper-based, point-of-care device for phenylalanine detection using a small volume (40 μL) of whole blood.

Genome Sequence and Transcriptome Analyses of Chrysochromulina tobin: Metabolic Tools for Enhanced Algal Fitness in the Prominent Order Prymnesiales (Haptophyceae).

Haptophytes are recognized as seminal players in aquatic ecosystem function. These algae are important in global carbon sequestration, form destructive harmful blooms, and given their rich fatty acid content, serve as a highly nutritive food source to a broad range of eco-cohorts. Haptophyte dominance in both fresh and marine waters is supported by the mixotrophic nature of many taxa.

"...Rewritten in the skin": clues to skin biology and aging from inherited disease.

The growing diversity of heritable skin diseases, a practical challenge to clinicians and dermato-nosologists alike, has nonetheless served as a rich source of insight into skin biology and disease mechanisms. I summarize below some key insights from the recent gene-driven phase of research on Werner syndrome, a heritable adult progeroid syndrome with prominent dermatologic features, constitutional genomic instability, and an elevated risk of cancer. I also indicate how new insights into skin biology, disease, and aging may come from unexpected sources.

Conversion of a laboratory-based test for phenylalanine detection to a simple paper-based format and implications for PKU screening in low-resource settings.

Laboratory-based testing does not reach many individuals in lower-resource settings who could benefit from access to appropriate tests for diagnosis and therapy. A critical issue is laboratory-based testing often requires an environment with a high level of resources and supporting infrastructure that is not available in many areas of the world. The current report describes the conversion of a laboratory-based test for phenylalanine detection to a simple paper-based test appropriate for use in low-resource settings.