Regulation of AMPK activation by extracellular matrix stiffness in pancreatic cancer.

The adenosine monophosphate (AMP)-activated protein kinase (AMPK) sits at a central node in the regulation of energy metabolism and tumor progression. AMPK is best known to sense high cellular ADP or AMP levels, which indicate the depletion of energy stores. Previous studies have shown that the low expression of phosphorylated AMPK is associated with a poor prognosis of pancreatic cancer. In this study, we report that AMPK is also highly sensitive to extracellular matrix (ECM) stiffness. We found that AMPK is activated in cells when cultured under low ECM stiffness conditions and is functionally required for the metabolic switch induced by ECM stiffness. This regulation of AMPK requires the Hippo kinases but not LKB1/CaMKKß. Hippo kinases directly phosphorylate AMPKα at Thr172 to activate AMPK at low ECM stiffness. Furthermore, we found AMPK activity is inhibited in patients with pancreatic ductal adenocarcinoma (PDAC) with high ECM stiffness and is associated with a poor survival outcome. The activation of Hippo kinases by ROCK inhibitor Y-27632 in combination with the mitochondrial inhibitor metformin synergistically activates AMPK and dramatically inhibits PDAC growth. Together, these findings establish a novel model for AMPK regulation by the mechanical properties of ECMs and provide a rationale for simultaneously targeting the ECM stiffness-Hippo kinases-AMPK signaling and low glucose-LKB1-AMPK signaling pathways as an effective therapeutic strategy against PDAC.

A ribosomal gene panel predicting a novel synthetic lethality in non-BRCAness tumors.

Poly (ADP-ribose) polymerase (PARP) inhibitors are one of the most exciting classes of targeted therapy agents for cancers with homologous recombination (HR) deficiency. However, many patients without apparent HR defects also respond well to PARP inhibitors/cisplatin. The biomarker responsible for this mechanism remains unclear. Here, we identified a set of ribosomal genes that predict response to PARP inhibitors/cisplatin in HR-proficient patients. PARP inhibitor/cisplatin selectively eliminates cells with high expression of the eight genes in the identified panel via DNA damage (ATM) signaling-induced pro-apoptotic ribosomal stress, which along with ATM signaling-induced pro-survival HR repair constitutes a new model to balance the cell fate in response to DNA damage. Therefore, the combined examination of the gene panel along with HR status would allow for more precise predictions of clinical response to PARP inhibitor/cisplatin. The gene panel as an independent biomarker was validated by multiple published clinical datasets, as well as by an ovarian cancer organoids library we established. More importantly, its predictive value was further verified in a cohort of PARP inhibitor-treated ovarian cancer patients with both RNA-seq and WGS data. Furthermore, we identified several marketed drugs capable of upregulating the expression of the genes in the panel without causing HR deficiency in PARP inhibitor/cisplatin-resistant cell lines. These drugs enhance PARP inhibitor/cisplatin sensitivity in both intrinsically resistant organoids and cell lines with acquired resistance. Together, our study identifies a marker gene panel for HR-proficient patients and reveals a broader application of PARP inhibitor/cisplatin in cancer therapy.

DNA Repair Deficiency Regulates Immunity Response in Cancers: Molecular Mechanism and Approaches for Combining Immunotherapy.

Defects in DNA repair pathways can lead to genomic instability in multiple tumor types, which contributes to tumor immunogenicity. Inhibition of DNA damage response (DDR) has been reported to increase tumor susceptibility to anticancer immunotherapy. However, the interplay between DDR and the immune signaling pathways remains unclear. In this review, we will discuss how a deficiency in DDR affects anti-tumor immunity, highlighting the cGAS-STING axis as an important link. We will also review the clinical trials that combine DDR inhibition and immune-oncology treatments. A better understanding of these pathways will help exploit cancer immunotherapy and DDR pathways to improve treatment outcomes for various cancers.

ASTE1 promotes shieldin-complex-mediated DNA repair by attenuating end resection.

The shieldin complex functions as the downstream effector of 53BP1-RIF1 to promote DNA double-strand break end-joining by restricting end resection. The SHLD2 subunit binds to single-stranded DNA ends and blocks end resection through OB-fold domains. Besides blocking end resection, it is unclear how the shieldin complex processes SHLD2-bound single-stranded DNA and promotes non-homologous end-joining. Here, we identify a downstream effector of the shieldin complex, ASTE1, as a structure-specific DNA endonuclease that specifically cleaves single-stranded DNA and 3' overhang DNA. ASTE1 localizes to DNA damage sites in a shieldin-dependent manner. Loss of ASTE1 impairs non-homologous end-joining, leads to hyper-resection and causes defective immunoglobulin class switch recombination. ASTE1 deficiency also causes resistance to poly(ADP-ribose) polymerase inhibitors in BRCA1-deficient cells owing to restoration of homologous recombination. These findings suggest that ASTE1-mediated 3' single-stranded DNA end cleavage contributes to the control of DSB repair choice by 53BP1, RIF1 and shieldin.

Use of spironolactone to treat acne in adolescent females.

BACKGROUND/OBJECTIVES: Studies assessing the utility of spironolactone for treating acne in adolescent females are lacking. Thus, we sought to examine spironolactone's role in treating this patient population. METHODS: A retrospective review was performed to determine the efficacy of spironolactone treatment in adolescent females seen at Mayo Clinic in Rochester, Minnesota, from 2007 to 2017. RESULTS: In a cohort of 80 pediatric patients with a median age of 19 years (range, 14-20 years), 64 patients (80%) experienced improvement of acne on treatment with spironolactone (median dose, 100 mg daily) with a favorable side effect profile. Approximately a quarter of patients (22.5%) had a complete response; more than half (58.8%) had a complete response or a partial response greater than 50%. Initial and maximal responses were observed at a median of 3 months and 5 months, respectively. Patients received treatment with spironolactone for a median duration of 7 months (range, 3-45 months) with limited side effects. CONCLUSIONS: Spironolactone demonstrated efficacy in treating acne in adolescent females and is a safe long-term alternative to systemic antibiotics in these patients.

Differences between Stevens-Johnson syndrome versus toxic epidermal necrolysis.

BACKGROUND: To retrospectively review the outcomes of two rare cutaneous diseases, Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN), and to question the practice of averaging the mortality rate on the assumption that they are one disease. METHODS: A retrospective chart review of all patients diagnosed with SJS and TEN by a dermatologist between January 1, 2000, and January 1, 2020, at our institution was performed. Seventy-one patients were identified (21 pediatric and 50 adults). Pathology slides from 32 adult patients (64%) were evaluated by a blinded board-certified dermatopathologist. RESULTS: Of the adult patients, 31 had SJS, two had SJS-TEN overlap, and 17 had TEN. All 21 patients in the pediatric group were diagnosed with SJS mainly caused by Mycoplasma. Mortality rates were 6.5% for SJS among adults and 35.3% for TEN. Chemotherapy-induced TEN is a trigger with 50% mortality. CONCLUSIONS: SJS was more common in adults and pediatric cases than TEN (3:1) and had a much better prognosis and outcome. Combining and averaging the mortality rates of TEN and SJS are not advised as SJS is mainly a mucocutaneous disorder with good prognosis versus TEN, a systemic toxicity of multiple organs with deep skin detachment.

Disseminated intravascular coagulopathy: a complication of Stevens-Johnson syndrome/toxic epidermal necrolysis.

BACKGROUND: The purpose of this study was to retrospectively assess clinical characteristics and mortality rate of Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN) in patients who developed disseminated intravascular coagulation (DIC). METHODS: A systematic retrospective chart review of all patients with concurrent clinical diagnosis of DIC and SJS/TEN between July 1, 2012, and January 1, 2020, at the Mayo Clinic was performed. RESULTS: The incidence of DIC in patients with SJS/TEN was 1.3% at our institution (5 of 396 DIC patients). Triggers of SJS/TEN included lamotrigine, clofarabine, antibiotics, and sepsis. Two patients diagnosed with SJS and two patients with TEN succumbed to the disease. CONCLUSION: DIC is a rare underlying risk in patients with SJS/TEN and is associated with increased mortality. Early clinician awareness and aggressive intervention is advised.

Extracellular matrix stiffness determines DNA repair efficiency and cellular sensitivity to genotoxic agents.

DNA double-strand breaks (DSBs) are highly toxic lesions that can drive genetic instability. These lesions also contribute to the efficacy of radiotherapy and many cancer chemotherapeutics. DNA repair efficiency is regulated by both intracellular and extracellular chemical signals. However, it is largely unknown whether this process is regulated by physical stimuli such as extracellular mechanical signals. Here, we report that DSB repair is regulated by extracellular mechanical signals. Low extracellular matrix (ECM) stiffness impairs DSB repair and renders cells sensitive to genotoxic agents. Mechanistically, we found that the MAP4K4/6/7 kinases are activated and phosphorylate ubiquitin in cells at low stiffness. Phosphorylated ubiquitin impairs RNF8-mediated ubiquitin signaling at DSB sites, leading to DSB repair deficiency. Our results thus demonstrate that ECM stiffness regulates DSB repair efficiency and genotoxic sensitivity through MAP4K4/6/7 kinase-mediated ubiquitin phosphorylation, providing a previously unidentified regulation in DSB-induced ubiquitin signaling.

Tandem Deubiquitination and Acetylation of SPRTN Promotes DNA-Protein Crosslink Repair and Protects against Aging.

DNA-protein crosslinks (DPCs) are highly toxic DNA lesions that threaten genomic integrity. Recent findings highlight that SPRTN, a specialized DNA-dependent metalloprotease, is a central player in proteolytic cleavage of DPCs. Previous studies suggest that SPRTN deubiquitination is important for its chromatin association and activation. However, the regulation and consequences of SPRTN deubiquitination remain unclear. Here we report that, in response to DPC induction, the deubiquitinase VCPIP1/VCIP135 is phosphorylated and activated by ATM/ATR. VCPIP1, in turn, deubiquitinates SPRTN and promotes its chromatin relocalization. Deubiquitination of SPRTN is required for its subsequent acetylation, which promotes SPRTN relocation to the site of chromatin damage. Furthermore, Vcpip1 knockout mice are prone to genomic instability and premature aging. We propose a model where two sequential post-translational modifications (PTMs) regulate SPRTN chromatin accessibility to repair DPCs and maintain genomic stability and a healthy lifespan.

Author Correction: Regulation of sister chromatid cohesion by nuclear PD-L1.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

STK38 promotes ATM activation by acting as a reader of histone H4 ufmylation.

The ATM (ataxia-telangiectasia mutated) kinase is rapidly activated following DNA damage and phosphorylates its downstream targets to launch DDR signaling. Recently, we and others showed that UFM1 signaling promotes ATM activation. We further discovered that monoufmylation of histone H4 at Lys31 by UFM1-specific ligase 1 (UFL1) is an important step in the amplification of ATM activation. However, how monoufmylated H4 enhances ATM activation is still unknown. Here, we report STK38, a kinase in the Hippo pathway, serves as a reader for histone H4 ufmylation to promote ATM activation in a kinase-independent manner. STK38 contains a potential UFM1 binding motif which recognizes ufmylated H4 and recruits the SUV39H1 to the double-strand breaks, resulting in H3K9 trimethylation and Tip60 activation to promote ATM activation. Together, STK38 is a previously unknown player in DNA damage signaling and functions as a reader of monoufmylated H4 at Lys31 to promote ATM activation.

Regulation of sister chromatid cohesion by nuclear PD-L1.

Programmed death ligand-1 (PD-L1 or B7-H1) is well known for its role in immune checkpoint regulation, but its function inside the tumor cells has rarely been explored. Here we report that nuclear PD-L1 is important for cancer cell sister chromatid cohesion. We found that depletion of PD-L1 suppresses cancer cell proliferation, colony formation in vitro, and tumor growth in vivo in immune-deficient NSG mice independent of its role in immune checkpoint. Specifically, PD-L1 functions as a subunit of the cohesin complex, and its deficiency leads to formation of multinucleated cells and causes a defect in sister chromatid cohesion. Mechanistically, PD-L1 compensates for the loss of Sororin, whose expression is suppressed in cancer cells overexpressing PD-L1. PD-L1 competes with Wing Apart-Like (WAPL) for binding to PDS5B, and secures proper sister chromatid cohesion and segregation. Our findings suggest an important role for nuclear PD-L1 in cancer cells independent of its function in immune checkpoint.

The bromodomain containing protein BRD-9 orchestrates RAD51-RAD54 complex formation and regulates homologous recombination-mediated repair.

Homologous recombination (HR) is important for error-free DNA double strand break repair and maintenance of genomic stability. However, upregulated HR is also used by cancer cells to promote therapeutic resistance. Therefore, inducing HR deficiency (HRD) is a viable strategy to sensitize HR proficient cancers to DNA targeted therapies in order to overcome therapeutic resistance. A bromodomain containing protein, BRD9, was previously reported to regulate chromatin remodeling and transcription. Here, we discover that following DNA damage, the bromodomain of BRD9 binds acetylated K515 on RAD54 and facilitates RAD54's interaction with RAD51, which is essential for HR. BRD9 is overexpressed in ovarian cancer and depleting BRD9 sensitizes cancer cells to olaparib and cisplatin. In addition, inhibitor of BRD9, I-BRD9, acts synergistically with olaparib in HR-proficient cancer cells. Overall, our results elucidate a role for BRD9 in HR and identify BRD9 as a potential therapeutic target to promote synthetic lethality and overcome chemoresistance.