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RNF168 orchestrates a ubiquitin-dependent DNA damage response to regulate the recruitment of repair factors, such as 53BP1 to DNA double-strand breaks (DSBs). In addition to its canonical functions in DSB signaling, RNF168 may facilitate DNA replication fork progression. However, the precise role of RNF168 in DNA replication remains unclear. Here, we demonstrate that RNF168 is recruited to DNA replication factories in a manner that is independent of the canonical DSB response pathway regulated by Ataxia-Telangiectasia Mutated (ATM) and RNF8. We identify a degenerate Proliferating Cell Nuclear Antigen (PCNA)-interacting peptide (DPIP) motif in the C-terminus of RNF168, which together with its Motif Interacting with Ubiquitin (MIU) domain mediates binding to mono-ubiquitylated PCNA at replication factories. An RNF168 mutant harboring inactivating substitutions in its DPIP box and MIU1 domain (termed RNF168 ΔDPIP/ΔMIU1) is not recruited to sites of DNA synthesis and fails to support ongoing DNA replication. Notably, the PCNA interaction-deficient RNF168 ΔDPIP/ΔMIU1 mutant fully rescues the ability of RNF168-/- cells to form 53BP1 foci in response to DNA DSBs. Therefore, RNF168 functions in DNA replication and DSB signaling are fully separable. Our results define a new mechanism by which RNF168 promotes DNA replication independently of its canonical functions in DSB signaling., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Standard initial systemic treatment for patients with metastatic prostate cancer includes agents that target androgen receptor (AR) signaling. Despite an initial positive response to these AR pathway inhibitors (ARPIs), acquired resistance remains a significant challenge. We show that treatment of AR-positive prostate cancer cells with the frontline ARPI enzalutamide induces DNA replication stress. Such stress is exacerbated by suppression of translesion DNA synthesis (TLS), leading to aberrant accumulation of single-stranded DNA (ssDNA) gaps and persistent DNA damage biomarkers. We further demonstrate that the TLS inhibitor, JH-RE-06, markedly sensitizes AR-positive prostate cancer cells, but not AR-negative benign cells, to enzalutamide in vitro. Combination therapy with enzalutamide and JH-RE-06 significantly suppresses cancer growth in a syngeneic murine tumor model over vehicle control or individual treatment groups. These findings suggest that AR inhibition broadly triggers DNA replication stress in hormone-sensitive prostate cancer, thereby exposing a unique vulnerability that can be exploited by a TLS-disrupting adjuvant for targeted therapy.

Almost all Glioblastoma (GBM) are either intrinsically resistant to the chemotherapeutical drug temozolomide (TMZ) or acquire therapy-induced mutations that cause chemoresistance and recurrence. The genome maintenance mechanisms responsible for GBM chemoresistance and hypermutation are unknown. We show that the E3 ubiquitin ligase RAD18 (a proximal regulator of TLS) is activated in a Mismatch repair (MMR)-dependent manner in TMZ-treated GBM cells, promoting post-replicative gap-filling and survival. An unbiased CRISPR screen provides an aerial map of RAD18-interacting DNA damage response (DDR) pathways deployed by GBM to tolerate TMZ genotoxicity. Analysis of mutation signatures from TMZ-treated GBM reveals a role for RAD18 in error-free bypass of O 6 mG (the most toxic TMZ-induced lesion), and error-prone bypass of other TMZ-induced lesions. Our analyses of recurrent GBM patient samples establishes a correlation between low RAD18 expression and hypermutation. Taken together we define molecular underpinnings for the hallmark tumorigenic phenotypes of TMZ-treated GBM., (© 2024. The Author(s).)

The functional status of the tumor suppressor p53 is a critical component in determining the sensitivity of cancer cells to many chemotherapeutic agents. DNA topoisomerase II (Top2) plays essential roles in DNA metabolism and is the target of FDA approved chemotherapeutic agents. Topoisomerase targeting drugs convert the enzyme into a DNA damaging agent and p53 influences cellular responses to these agents. We assessed the impact of the loss of p53 function on the formation of DNA damage induced by the Top2 poison etoposide. Using human HCT116 cells, we found resistance to etoposide in cell growth assays upon the functional loss of p53. Nonetheless, cells lacking fully functional p53 were etoposide hypersensitive in clonogenic survival assays. This complex role of p53 led us to directly examine the effects of p53 status on topoisomerase-induced DNA damage. A deficiency in functional p53 resulted in elevated levels of the Top2 covalent complexes (Top2cc) in multiple cell lines. Employing genome-wide siRNA screens, we identified a set of genes for which reduced expression resulted in enhanced synthetic lethality upon etoposide treatment of p53 defective cells. We focused on one hit from this screen, ATR, and showed that decreased expression sensitized the p53-defective cells to etoposide in all assays and generated elevated levels of Top2cc in both p53 proficient and deficient cells. Our findings suggest that a combination of etoposide treatment with functional inactivation of DNA repair in p53 defective cells could be used to enhance the therapeutic efficacy of Top2 targeting agents., Competing Interests: CONFLICTS OF INTEREST Authors have no conflicts of interest to declare., (Copyright: © 2022 Menendez et al.)

Aim: This descriptive study set out to explore the end-of-life care perspectives both of patients with advanced carcinoma and of health-care professionals in a tertiary health-care setting in India., Method: A descriptive approach was undertaken to assess the end-of-life care perspectives from a sample of 140 patients and 40 health-care professionals. Data was collected through structured interviews with the patients and by self-administered questionnaire from the health-care professionals., Results: There was a significant association in attitudes to artificial prolongation of life with age and religion. Patients placed a higher degree of importance on emotional, social, physical and spiritual dimensions of care compared to the health-care professionals., Conclusion: There is an urgent call for formulating an end-of-life care policy and advance directives for patients with terminal illness in India. Nurses and physicians should be proactive in offering key supportive services to ensure patient autonomy and facilitate good death.

Objective: To evaluate the efficacy of oversized corneal grafts in the pediatric age group., Design: Prospective, nonrandomized clinical trial., Participants and Intervention: Forty pediatric patients with unilateral or bilateral corneal opacification of congenital or acquired origin underwent corneal grafting surgery over a period of 2 years using donor corneal buttons oversized by 1 mm., Main Outcome Measures: The parameters evaluated were indications for keratoplasty, graft clarity, visual acuity, keratometry, spherical equivalent, anterior chamber depth, and complications., Results: Corneal ulceration was the most common cause of corneal opacification (25%), followed by trauma (20%) and sclerocornea (20%). At 1 year, clear grafts were achieved in 85% of the cohort. The average keratometry at the end of 1 year was 43.28 +/- 1.65 diopters (D) in the congenital opacity group and 43.04 +/- 2.20 D in the acquired group. The keratometric astigmatism was 3.60 +/- 2.60 D in the congenital group and 2.52 +/- 2.20 D in the acquired group. Oversized grafts provided an adequate anterior chamber depth of 2.20 +/- 0.612 mm in the congenital group and 2.36 +/- 0.302 mm in the acquired group. Visual acuity of 20/80 or better was recorded in only 30% of cases in the congenital group as opposed to 47% with acquired opacities. Nine cases had episodes of graft rejection., Conclusion: Oversizing donor buttons by 1 mm provides adequate anterior chamber depth and increases the morphologic success of corneal grafting in children.

Purpose: This study aimed to evaluate the clinical profile, microbial spectrum, management modalities, and visual outcome in cases of corneal superinfection that occurred after an epidemic of acute hemorrhagic conjunctivitis., Method: A retrospective analysis of 14 eyes of 13 patients who were referred to a tertiary eye-care center with corneal ulceration after an episode of acute hemorrhagic conjunctivitis was undertaken. The parameters analyzed were age, sex, prior use of topical medications, predisposing factors, ulcer characteristics, organisms isolated, success of medical therapy and surgery, and visual outcome., Results: A definite history of topical corticosteroid use to treat acute hemorrhagic conjunctivitis was elicited in 12 (86%) eyes. Cultures were positive in 86% (12/14) eyes. Organisms isolated were Pseudomonas aeruginosa (three eyes, 25%), Fusarium species (three eyes, 25%), Aspergillus species (two eyes, 16%), and Staphylococcus aureus (two eyes, 16%). Mixed infection occurred in two patients. After discontinuation of topical corticosteroids, all patients received antimicrobial therapy. The keratitis resolved in seven eyes. Therapeutic penetrating keratoplasty was required in five eyes. Two patients were lost to follow-up., Conclusions: Corneal superinfection may occur after acute hemorrhagic conjunctivitis. Use of topical corticosteroids to treat acute hemorrhagic conjunctivitis may predispose an already compromised cornea to develop microbial keratitis and such a practice should be discouraged.