bims-dresag 2021-07-25 papers

bims-dresag

Biomed News

on DNA damage and repair, cellular senescence and aging, gene therapy

Issue of 2021‒07‒25
eight papers selected by
Pengyi Yan
Shanghai Jiao Tong University

Understanding and overcoming resistance to PARP inhibitors in cancer therapy.
Opportunities for Utilization of DNA Repair Inhibitors in Homologous Recombination Repair-Deficient and Proficient Pancreatic Adenocarcinoma.
Sirtuin 6: linking longevity with genome and epigenome stability.
CRISPRi/a Screening with Human iPSCs.
Generation of Efficient Knock-in Mouse and Human Pluripotent Stem Cells Using CRISPR-Cas9.
CRISPR/Cas9-based inactivation of human papillomavirus oncogenes E6 or E7 induces senescence in cervical cancer cells.
The aetiology and molecular landscape of insulin resistance.
A Case-Based Clinical Approach to the Investigation, Management and Screening of Families with BRCA2 Related Prostate Cancer.

Nat Rev Clin Oncol. 2021 Jul 20.

Understanding and overcoming resistance to PARP inhibitors in cancer therapy.

Mariana Paes Dias, Sarah C Moser, Shridar Ganesan, Jos Jonkers.

Developing novel targeted anticancer therapies is a major goal of current research. The use of poly(ADP-ribose) polymerase (PARP) inhibitors in patients with homologous recombination-deficient tumours provides one of the best examples of a targeted therapy that has been successfully translated into the clinic. The success of this approach has so far led to the approval of four different PARP inhibitors for the treatment of several types of cancers and a total of seven different compounds are currently under clinical investigation for various indications. Clinical trials have demonstrated promising response rates among patients receiving PARP inhibitors, although the majority will inevitably develop resistance. Preclinical and clinical data have revealed multiple mechanisms of resistance and current efforts are focused on developing strategies to address this challenge. In this Review, we summarize the diverse processes underlying resistance to PARP inhibitors and discuss the potential strategies that might overcome these mechanisms such as combinations with chemotherapies, targeting the acquired vulnerabilities associated with resistance to PARP inhibitors or suppressing genomic instability.

DOI: https://doi.org/10.1038/s41571-021-00532-x
Clin Cancer Res. 2021 Jul 20. pii: clincanres.1367.2021. [Epub ahead of print]

Opportunities for Utilization of DNA Repair Inhibitors in Homologous Recombination Repair-Deficient and Proficient Pancreatic Adenocarcinoma.

James M Cleary, Brian M Wolpin, Stephanie K Dougan, Srivatsan Raghavan, Harshabad Singh, Brandon Huffman, Nilay S Sethi, Jonathan A Nowak, Geoffrey I Shapiro, Andrew J Aguirre, Alan D D'Andrea.

Pancreatic cancer is rapidly progressive and notoriously difficult to treat with cytotoxic chemotherapy and targeted agents. Recent demonstration of the efficacy of maintenance PARP inhibition in germline BRCA mutated pancreatic cancer has raised hopes that increased understanding of the DNA damage response pathway will lead to new therapies in both homologous recombination (HR) repair-deficient and proficient pancreatic cancer. Here, we review the potential mechanisms of exploiting HR deficiency, replicative stress, and DNA damage-mediated immune activation through targeted inhibition of DNA repair regulatory proteins.

DOI: https://doi.org/10.1158/1078-0432.CCR-21-1367
Trends Cell Biol. 2021 Jul 17. pii: S0962-8924(21)00125-2. [Epub ahead of print]

Sirtuin 6: linking longevity with genome and epigenome stability.

Anatoly Korotkov, Andrei Seluanov, Vera Gorbunova.

  Sirtuin 6 (SIRT6) has been in the spotlight of aging research because progeroid phenotypes are associated with SIRT6 deficiency. SIRT6 has multiple molecular functions, including DNA repair and heterochromatin regulation, which position SIRT6 as a hub that regulates genome and epigenome stability. Genomic instability caused by persistent DNA damage and accumulating mutations, together with alterations in the epigenetic landscape and derepression of repetitive genetic elements, have emerged as mechanisms driving organismal aging. Enhanced levels of SIRT6 expression or activity provide avenues for rejuvenation strategies. This review focuses on the role of SIRT6 in the maintenance of genome and epigenome stability and its link to longevity. We propose a model where SIRT6 together with lamins control aging and rejuvenation by maintaining epigenetic silencing of repetitive elements.

Keywords:  DNA damage repair; SIRT6; epigenome; healthspan; longevity; rejuvenation

DOI:  https://doi.org/10.1016/j.tcb.2021.06.009
Methods Mol Biol. 2021 ;2320 261-281

CRISPRi/a Screening with Human iPSCs.

Masataka Nishiga, Lei S Qi, Joseph C Wu.

  Identifying causative genes in a given phenotype or disease model is important for biological discovery and drug development. The recent development of the CRISPR/Cas9 system has enabled unbiased and large-scale genetic perturbation screens to identify causative genes by knocking out many genes in parallel and selecting cells with desired phenotype of interest. However, compared to cancer cell lines, human somatic cells including cardiomyocytes (CMs), neuron cells, and endothelial cells are not easy targets of CRISPR screens because CRISPR screens require a large number of isogenic cells to be cultured and thus primary cells from patients are not ideal. The combination of CRISPR screens with induced pluripotent stem cell (iPSC) technology would be a powerful tool to identify causative genes and pathways because iPSCs can be expanded easily and differentiated to any cell type in principle. Here we describe a robust protocol for CRISPR screening using human iPSCs. Because each screening is different and needs to be customized depending on the cell types and phenotypes of interest, we show an example of CRISPR knockdown screening using CRISPRi system to identify essential genes to differentiate iPSCs to CMs.

Keywords:  CRISPR/Cas9; Cardiomyocytes; Genome editing; Induced pluripotent stem cells

DOI:  https://doi.org/10.1007/978-1-0716-1484-6_23
Methods Mol Biol. 2021 ;2320 247-259

Generation of Efficient Knock-in Mouse and Human Pluripotent Stem Cells Using CRISPR-Cas9.

Tatsuya Anzai, Hiromasa Hara, Nawin Chanthra, Taketaro Sadahiro, Masaki Ieda, Yutaka Hanazono, Hideki Uosaki.

  A knock-in can generate fluorescent or Cre-reporter under the control of an endogenous promoter. It also generates knock-out or tagged-protein with fluorescent protein and short tags for tracking and purification. Recent advances in genome editing with clustered regularly interspaced short palindromic repeat (CRISPR) and CRISPR-associated protein 9 (Cas9) significantly increased the efficiencies of making knock-in cells. Here we describe the detailed protocols of generating knock-in mouse and human pluripotent stem cells (PSCs) by electroporation and lipofection, respectively.

Keywords:  CRISPR-Cas9; Genome editing; Homology-directed repair; Knock-in; Pluripotent stem cells

DOI:  https://doi.org/10.1007/978-1-0716-1484-6_22
Virology. 2021 Jul 14. pii: S0042-6822(21)00152-5. [Epub ahead of print]562 92-102

CRISPR/Cas9-based inactivation of human papillomavirus oncogenes E6 or E7 induces senescence in cervical cancer cells.

Raviteja Inturi, Per Jemth.

  Human papillomaviruses (HPVs) such as HPV16 and HPV18 can cause cancers of the cervix, anogenital and oropharyngeal sites. Continuous expression of the HPV oncoproteins E6 and E7 are essential for transformation and maintenance of cancer cells. Therefore, therapeutic targeting of E6 or E7 genes can potentially treat HPV-related cancers. Here we report that CRISPR/Cas9-based knockout of E6 or E7 can trigger cellular senescence in HPV18 immortalized HeLa cells. Specifically, E6 or E7-inactivated HeLa cells exhibited characteristic senescence markers like enlarged cell surface area, increased β-galactosidase expression and loss of lamin B1. Since E6 and E7 are bicistronic transcripts, inactivation of HPV18 E6 resulted in knockout of both E6 and E7 and increasing levels of p53/p21 and pRb/p21, respectively. Knockout of HPV18 E7 resulted in decreased E6 expression with activation of pRb/p21 pathway. Taken together, our study demonstrates cellular senescence as an alternative outcome of HPV oncogene inactivation by CRISPR/Cas9.

Keywords:  CRISPR/Cas9; Cervical cancer; DNA virus; Human papillomavirus; Oncogenes; Senescence

DOI:  https://doi.org/10.1016/j.virol.2021.07.005
Nat Rev Mol Cell Biol. 2021 Jul 20.

The aetiology and molecular landscape of insulin resistance.

David E James, Jacqueline Stöckli, Morris J Birnbaum.

Insulin resistance, defined as a defect in insulin-mediated control of glucose metabolism in tissues - prominently in muscle, fat and liver - is one of the earliest manifestations of a constellation of human diseases that includes type 2 diabetes and cardiovascular disease. These diseases are typically associated with intertwined metabolic abnormalities, including obesity, hyperinsulinaemia, hyperglycaemia and hyperlipidaemia. Insulin resistance is caused by a combination of genetic and environmental factors. Recent genetic and biochemical studies suggest a key role for adipose tissue in the development of insulin resistance, potentially by releasing lipids and other circulating factors that promote insulin resistance in other organs. These extracellular factors perturb the intracellular concentration of a range of intermediates, including ceramide and other lipids, leading to defects in responsiveness of cells to insulin. Such intermediates may cause insulin resistance by inhibiting one or more of the proximal components in the signalling cascade downstream of insulin (insulin receptor, insulin receptor substrate (IRS) proteins or AKT). However, there is now evidence to support the view that insulin resistance is a heterogeneous disorder that may variably arise in a range of metabolic tissues and that the mechanism for this effect likely involves a unified insulin resistance pathway that affects a distal step in the insulin action pathway that is more closely linked to the terminal biological response. Identifying these targets is of major importance, as it will reveal potential new targets for treatments of diseases associated with insulin resistance.

DOI: https://doi.org/10.1038/s41580-021-00390-6
Appl Clin Genet. 2021 ;14 255-266

A Case-Based Clinical Approach to the Investigation, Management and Screening of Families with BRCA2 Related Prostate Cancer.

Bradley King, Jana McHugh, Katie Snape.

  BRCA2 is the most commonly implicated DNA damage repair gene associated with inherited prostate cancer. BRCA2 deficient prostate cancer typically presents at a younger age, is more poorly differentiated, and is associated with worse survival outcomes than non-BRCA2 associated prostate cancer. Despite these unfavourable prognostic implications, poly-ADP ribose polymerase inhibitors and platinum-based chemotherapy have been identified as potent targeted therapeutic agents towards BRCA1/2 deficient cancer cells. This review article explores the literature surrounding BRCA2-related prostate cancer through a familial clinical scenario. The investigation, diagnosis and management of BRCA2 deficient prostate cancer will be explored, alongside the implications of the identification of a germline pathogenic BRCA2 variant within a family, cascade screening and prostate cancer surveillance in unaffected male BRCA2 carriers. A greater understanding of the molecular pathogenesis of DNA damage repair gene deficient prostate cancer, coupled with new treatment paradigms and widened access to both somatic and germline genetic analysis for prostate cancer patients and their families will hopefully enable the robust implementation of high quality evidence-based clinical pathways for both the management and identification of BRCA2 deficient prostate cancer and improved screening, early detection and prevention strategies for individuals at increased genetic risk of prostate cancer.

Keywords:  BRCA; PARP inhibitors; clinical management; genomics; prostate cancer

DOI:  https://doi.org/10.2147/TACG.S261737