Genome-scale clustered regularly interspaced short palindromic repeats screen identifies nucleotide metabolism as an actionable therapeutic vulnerability in diffuse large B-cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is the most common malignancy that develops in patients with ataxia-telangiectasia, a cancer-predisposing inherited syndrome characterized by inactivating germline ATM mutations. ATM is also frequently mutated in sporadic DLBCL. To investigate lymphomagenic mechanisms and lymphoma-specific dependencies underlying defective ATM, we applied ribonucleic acid (RNA)-seq and genome-scale loss-offunction clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 screens to systematically interrogate B-cell lymphomas arising in a novel murine model (Atm-/-nu-/-) with constitutional Atm loss, thymic aplasia but residual T-cell populations. Atm-/-nu-/-lymphomas, which phenotypically resemble either activated B-cell-like or germinal center Bcell-like DLBCL, harbor a complex karyotype, and are characterized by MYC pathway activation. In Atm-/-nu-/-lymphomas, we discovered nucleotide biosynthesis as a MYCdependent cellular vulnerability that can be targeted through the synergistic nucleotidedepleting actions of mycophenolate mofetil (MMF) and the WEE1 inhibitor, adavosertib (AZD1775). The latter is mediated through a synthetically lethal interaction between RRM2 suppression and MYC dysregulation that results in replication stress overload in Atm-/-nu-/-lymphoma cells. Validation in cell line models of human DLBCL confirmed the broad applicability of nucleotide depletion as a therapeutic strategy for MYC-driven DLBCL independent of ATM mutation status. Our findings extend current understanding of lymphomagenic mechanisms underpinning ATM loss and highlight nucleotide metabolism as a targetable therapeutic vulnerability in MYC-driven DLBCL.

Disruption of CTCF-YY1-dependent looping of the human papillomavirus genome activates differentiation-induced viral oncogene transcription.

The complex life cycle of oncogenic human papillomavirus (HPV) initiates in undifferentiated basal epithelial keratinocytes where expression of the E6 and E7 oncogenes is restricted. Upon epithelial differentiation, E6/E7 transcription is increased through unknown mechanisms to drive cellular proliferation required to support virus replication. We report that the chromatin-organising CCCTC-binding factor (CTCF) promotes the formation of a chromatin loop in the HPV genome that epigenetically represses viral enhancer activity controlling E6/E7 expression. CTCF-dependent looping is dependent on the expression of the CTCF-associated Yin Yang 1 (YY1) transcription factor and polycomb repressor complex (PRC) recruitment, resulting in trimethylation of histone H3 at lysine 27. We show that viral oncogene up-regulation during cellular differentiation results from YY1 down-regulation, disruption of viral genome looping, and a loss of epigenetic repression of viral enhancer activity. Our data therefore reveal a key role for CTCF-YY1-dependent looping in the HPV life cycle and identify a regulatory mechanism that could be disrupted in HPV carcinogenesis.

End User and Implementer Experiences of mHealth Technologies for Noncommunicable Chronic Disease Management in Young Adults: Systematic Review.

BACKGROUND: Chronic noncommunicable diseases (NCDs) such as asthma, diabetes, cancer, and persistent musculoskeletal pain impose an escalating and unsustainable burden on young people, their families, and society. Exploring how mobile health (mHealth) technologies can support management for young people with NCDs is imperative. OBJECTIVE: The aim of this study was to identify, appraise, and synthesize available qualitative evidence on users' experiences of mHealth technologies for NCD management in young people. We explored the perspectives of both end users (young people) and implementers (health policy makers, clinicians, and researchers). METHODS: A systematic review and meta-synthesis of qualitative studies. Eligibility criteria included full reports published in peer-reviewed journals from January 2007 to December 2016, searched across databases including EMBASE, MEDLINE (PubMed), Scopus, and PsycINFO. All qualitative studies that evaluated the use of mHealth technologies to support young people (in the age range of 15-24 years) in managing their chronic NCDs were considered. Two independent reviewers identified eligible reports and conducted critical appraisal (based on the Joanna Briggs Institute Qualitative Assessment and Review Instrument: JBI-QARI). Three reviewers independently, then collaboratively, synthesized and interpreted data through an inductive and iterative process to derive emergent themes across the included data. External validity checking was undertaken by an expert clinical researcher and for relevant content, a health policy expert. Themes were subsequently subjected to a meta-synthesis, with findings compared and contrasted between user groups and policy and practice recommendations derived. RESULTS: Twelve studies met our inclusion criteria. Among studies of end users (N=7), mHealth technologies supported the management of young people with diabetes, cancer, and asthma. Implementer studies (N=5) covered the management of cognitive and communicative disabilities, asthma, chronic self-harm, and attention deficit hyperactivity disorder. Quality ratings were higher for implementer compared with end user studies. Both complementary and unique user themes emerged. Themes derived for end users of mHealth included (1) Experiences of functionality that supported self-management, (2) Acceptance (technical usability and feasibility), (3) Importance of codesign, and (4) Perceptions of benefit (self-efficacy and empowerment). For implementers, derived themes included (1) Characteristics that supported self-management (functional, technical, and behavior change); (2) Implementation challenges (systems level, service delivery level, and clinical level); (3) Adoption considerations for specific populations (training end users; specific design requirements); and (4) Codesign and tailoring to facilitate uptake and person-centered care. CONCLUSIONS: Synthesizing available data revealed both complementary and unique user perspectives on enablers and barriers to designing, developing, and implementing mHealth technologies to support young people's management of their chronic NCDs. TRIAL REGISTRATION: PROSPERO CRD42017056317; http://www.crd.york.ac.uk/PROSPERO/display_record.asp?ID=CRD 42017056317 (Archived by WebCite at http://www.webcitation.org/6vZ5UkKLp).

HIV-1 Capsid Rapidly Induces Long-Lived CPSF6 Puncta in Non-Dividing Cells, but Similar Puncta Already Exist in Uninfected T-Cells.

The HIV-1 capsid (CA) protein forms the outer shell of the viral core that is released into the cytoplasm upon infection. CA binds various cellular proteins, including CPSF6, that direct HIV-1 integration into speckle-associated domains in host chromatin. Upon HIV-1 infection, CPSF6 forms puncta in the nucleus. Here, we characterised these CPSF6 puncta further in HeLa cells, T-cells and macrophages and confirmed that integration and reverse transcription are not required for puncta formation. Indeed, we found that puncta formed very rapidly after infection, correlating with the time that CA entered the nucleus. In aphidicolin-treated HeLa cells and macrophages, puncta were detected for the length of the experiment, suggesting that puncta are only lost upon cell division. CA still co-localised with CPSF6 puncta at the latest time points, considerably after the peak of reverse transcription and integration. Intriguingly, the number of puncta induced in macrophages did not correlate with the MOI or the total number of nuclear speckles present in each cell, suggesting that CA/CPSF6 is only directed to a few nuclear speckles. Furthermore, we found that CPSF6 already co-localised with nuclear speckles in uninfected T-cells, suggesting that HIV-1 promotes a natural behaviour of CPSF6.

The anti-tumour activity of DNA methylation inhibitor 5-aza-2'-deoxycytidine is enhanced by the common analgesic paracetamol through induction of oxidative stress.

The DNA demethylating agent 5-aza-2'-deoxycytidine (DAC, decitabine) has anti-cancer therapeutic potential, but its clinical efficacy is hindered by DNA damage-related side effects and its use in solid tumours is debated. Here we describe how paracetamol augments the effects of DAC on cancer cell proliferation and differentiation, without enhancing DNA damage. Firstly, DAC specifically upregulates cyclooxygenase-2-prostaglandin E2 pathway, inadvertently providing cancer cells with survival potential, while the addition of paracetamol offsets this effect. Secondly, in the presence of paracetamol, DAC treatment leads to glutathione depletion and finally to accumulation of ROS and/or mitochondrial superoxide, both of which have the potential to restrict tumour growth. The benefits of combined treatment are demonstrated here in head and neck squamous cell carcinoma (HNSCC) and acute myeloid leukaemia cell lines, further corroborated in a HNSCC xenograft mouse model and through mining of publicly available DAC and paracetamol responses. The sensitizing effect of paracetamol supplementation is specific to DAC but not its analogue 5-azacitidine. In summary, the addition of paracetamol could allow for DAC dose reduction, widening its clinical usability and providing a strong rationale for consideration in cancer therapy.

Epigenetic regulation of human papillomavirus transcription in the productive virus life cycle.

Human papillomaviruses (HPV) are a large family of viruses which contain a circular, double-stranded DNA genome of approximately 8000 base pairs. The viral DNA is chromatinized by the recruitment of cellular histones which are subject to host cell-mediated post-translational epigenetic modification recognized as an important mechanism of virus transcription regulation. The HPV life cycle is dependent on the terminal differentiation of the target cell within epithelia-the keratinocyte. The virus life cycle begins in the undifferentiated basal compartment of epithelia where the viral chromatin is maintained in an epigenetically repressed state, stabilized by distal chromatin interactions between the viral enhancer and early gene region. Migration of the infected keratinocyte towards the surface of the epithelium induces cellular differentiation which disrupts chromatin looping and stimulates epigenetic remodelling of the viral chromatin. These epigenetic changes result in enhanced virus transcription and activation of the virus late promoter facilitating transcription of the viral capsid proteins. In this review article, we discuss the complexity of virus- and host-cell-mediated epigenetic regulation of virus transcription with a specific focus on differentiation-dependent remodelling of viral chromatin during the HPV life cycle.