Search

Defective glucose-stimulated insulin secretion (GSIS) and β-cell senescence are hallmarks in diabetes. The mitochondrial enzyme pyruvate carboxylase (PC) has been shown to promote GSIS and β-cell proliferation in the clonal β-cell lines, yet its physiological relevance remains unknown. Here, we provide animal and human data showing a role of PC in protecting β-cells against senescence and maintaining GSIS under different physiological and pathological conditions. β-cell-specific deletion of PC impaired GSIS and induced β-cell senescence in the mouse models under either a standard chow diet or prolonged high-fat diet feeding. Transcriptomic analysis indicated that p53-related senescence and cell cycle arrest are activated in PC-deficient islets. Overexpression of PC inhibited hyperglycemia- and aging-induced p53-related senescence in human and mouse islets as well as INS-1E β-cells, whereas knockdown of PC provoked senescence. Mechanistically, PC interacted with MDM2 to prevent its degradation via the MDM2 binding motif, which in turn restricts the p53-dependent senescent program in β-cells. On the contrary, the regulatory effects of PC on GSIS and the tricarboxylic acid (TCA) anaplerotic flux are p53-independent. We illuminate a function of PC in controlling β-cell senescence through the MDM2-p53 axis., Competing Interests: Competing interests statement:The authors declare no competing interest.

Overexpression or activation of oncogenes or loss of tumor-suppressor genes can induce cellular senescence as a defense mechanism against tumor development, thereby maintaining cellular homeostasis. However, cancer cells can circumvent this senescent state and continue to spread. Myosin light chain kinase (MLCK) is downregulated in many breast cancers. Here we report that downregulation of MLCK in normal breast epithelial cells induces a senescence-associated secretory phenotype and stimulates migration. The reduction of MLCK results in increased p21 Cip1 expression, dependent on p53 and the AKT-mammalian target of rapamycin pathway. Subsequently, p21 Cip1 promotes the secretion of soluble ICAM-1, IL-1α, IL-6 and IL-8, thereby enhancing collective cell migration in a non-cell-autonomous manner. These findings provide new mechanistic insights into the role of MLCK in cellular senescence and cancer progression., (© 2024. The Author(s).)

Ethnopharmacological Relevance: Ecliptea herba, a traditional Chinese herbal medicine for hair loss, was first recorded in the Tang Dynasty's 'Qian Jin Yue Ling', of which the active ingredients and mechanisms of action in the treatment of chemotherapy-induced hair loss remain poorly investigated., Aim of the Study: To investigate the effects of the petroleum ether extract of Eclipta (PEE) on alopecia and follicle damage and elucidate its potential therapeutic mechanisms using the integration of network pharmacology, bioinformatics, and experimental validation., Materials and Methods: UPLC-MS was used to analyse the chemical composition of PEE. A network pharmacology approach was employed to establish the 'components-targets-pathways' network of PEE to explore potential therapeutic pathways and targets. Molecular docking was used for validation, and the mechanism of PEE in treating chemotherapy-induced alopecia (CIA) was elucidated using in vitro and in vivo on CIA models., Results: UPLC-MS analysis of PEE revealed 185 components, while network pharmacology and molecular docking analyses revealed potential active compounds and their target molecules, suggesting the involvement of core genes, such as TP53, ESR1, AKT1, IL6, TNF, and EGFR. The key components included wedelolactone, dimethyl-wedelolactone, luteoloside, linarin, and hispidulin. In vivo, PEE promoted hair growth, restored the number of hair follicles, and reduced follicle apoptosis. Conversely, in vitro, PEE enhanced cell viability, reduced apoptosis, and protected HaCaT cells from damage induced by 4-hydroperoxycyclophosphamide (4-HC)., Conclusions: PEE alleviated hair follicle damage in CIA mice by inhibiting the P53/Fas pathway, which may be associated with inhibiting hair follicle cell apoptosis. This study provides a novel therapeutic strategy for treating cyclophosphamide-induced hair loss., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The pluripotent mouse embryonic stem cell (mESCs) can transit into the totipotent-like state, and the transcription factor DUX is one of the master regulators of this transition. Intriguingly, this transition in mESCs is accompanied by massive cell death, which significantly impedes the establishment and maintenance of totipotent cells in vitro, yet the underlying mechanisms of this cell death remain largely elusive. In this study, we found that the totipotency transition in mESCs triggered cell death through the upregulation of DUX. Specifically, R-loops are accumulated upon DUX induction, which subsequently lead to DNA replication stress (RS) in mESCs. This RS further activates p53 and PMAIP1, ultimately leading to Caspase-9/7-dependent intrinsic apoptosis. Notably, inhibiting this intrinsic apoptosis not only mitigates cell death but also enhances the efficiency of the totipotency transition in mESCs. Our findings thus elucidate one of the mechanisms underlying cell apoptosis during the totipotency transition in mESCs and provide a strategy for optimizing the establishment and maintenance of totipotent cells in vitro., (© 2024. The Author(s).)

Cells that lack p53 signaling frequently occur in ulcerative colitis (UC) and are considered early drivers in UC-associated colorectal cancer (CRC). Epithelial injury during colitis is associated with transient stem cell reprogramming from the adult, homeostatic to a "fetal-like" regenerative state. Here, we use murine and organoid-based models to study the role of Trp53 during epithelial reprogramming. We find that p53 signaling is silent and dispensable during homeostasis but strongly up-regulated in the epithelium upon DSS-induced colitis. While in WT cells this causes termination of the regenerative state, crypts that lack Trp53 remain locked in the highly proliferative, regenerative state long-term. The regenerative state in WT cells requires high Wnt signaling to maintain elevated levels of glycolysis. Instead, Trp53 deficiency enables Wnt-independent glycolysis due to overexpression of rate-limiting enzyme PKM2. Our study reveals the context-dependent relevance of p53 signaling specifically in the injury-induced regenerative state, explaining the high abundance of clones lacking p53 signaling in UC and UC-associated CRC.

The master regulator of the DNA damage response, the transcription factor p53, orchestrates multiple downstream responses and coordinates repair processes. In response to double-strand DNA breaks, p53 exhibits pulses of expression, but how it achieves temporal coordination of downstream responses remains unclear. Here, we show that p53's posttranslational modification state is altered between its first and second pulses of expression. We show that acetylations at two sites, K373 and K382, were reduced in the second pulse, and these acetylations differentially affected p53 target genes, resulting in changes in gene expression programs over time. This interplay between dynamics and modification may offer a strategy for cellular hubs like p53 to temporally organize multiple processes in individual cells.

Recurring lung injury, chronic inflammation, aberrant tissue repair and impaired tissue remodelling contribute to the pathogenesis of pulmonary fibrosis (PF). Neutrophil extracellular traps (NETs) are released by activated neutrophils to trap, immobilise and kill invading pathogen and is facilitated by peptidyl arginine deiminase-4 (PAD-4). Dysregulated NETs release and abnormal PAD-4 activation plays a crucial role in activating pro-fibrotic events in PF. Developmental endothelial locus-1 (Del-1), expressed by the endothelial cells of lungs and brain acts as an endogenous inhibitor of inflammation and fibrosis. We have hypothesised that PAD-4 inhibitor exerts anti-inflammatory and anti-fibrotic effects in mice model of PF. We have also hypothesised by PAD-4 regulated the transcription of Del-1 through co-repression and its inhibition potentiates anti-fibrotic effects of Del-1. In our study, the PAD-4 inhibitor chloro-amidine (CLA) demonstrated anti-NETotic and anti-inflammatory effects in vitro in differentiated HL-60 cells. In a bleomycin-induced PF mice model, CLA administration in two doses (3 mg/kg, I.P and 10 mg/kg, I.P) improved lung function, normalized bronchoalveolar lavage fluid parameters, and attenuated fibrotic events, including markers of extracellular matrix and epithelial-mesenchymal transition. Histological analyses confirmed the restoration of lung architecture and collagen deposition with CLA treatment. ELISA, IHC, IF, RT-PCR, and immunoblot analysis supported the anti-NETotic effects of CLA. Furthermore, BLM-induced PF reduced Del-1 and p53 expression, which was normalized by CLA treatment. These findings suggest that inhibition of PAD-4 results in amelioration of PF in animal model and may involve modulation of Del-1 and p53 pathways, warranting further investigation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The most common type of non-small cell lung cancer is lung adenocarcinoma (LUAD), which is characterized by high morbidity and poor survival. Up-regulation of SASS6 expression can lead to the progression of various malignant tumors. However, there are no relevant studies on the role of SASS6 in LUAD. SASS6 was highly expressed in most tumors, reflecting a good diagnostic value, and its overexpression in LUAD indicated discouraging overall prognosis. Functional enrichment analysis suggested that SASS6 was associated with cell cycle in LUAD. In addition, patients with high SASS6 expression had worse immune infiltration, but higher TMB and immune checkpoint, and higher sensitivity to multiple targeted drugs such as osimertinib. Cell experiments confirmed that knockdown of SASS6 could inhibit the viability of tumor cells.SASS6 has important value in the diagnosis of cancer. In particular, SASS6 is a crucial factor in the progression of LUAD, and has important clinical value, especially in the diagnosis, prognosis and treatment., (© 2024. The Author(s).)

Human skin aging, a complex process influenced by intrinsic aging and extrinsic photoaging, is marked by the accumulation of reactive oxygen species (ROS) that cause DNA damage, impaired dermal fibroblast function, and wrinkle formation. External stressors, such as ultraviolet (UV) radiation, can trigger cellular senescence. Sirtuin-1 (SIRT1), an NAD + -dependent enzyme in the sirtuin family, plays a crucial role in deacetylating p53, thereby inhibiting its nuclear translocation and reducing skin senescence. Galangin, a flavonoid found in honey and Alpinia officinarum root, has antioxidant and anti-inflammatory properties. This study investigates the protective mechanism of galangin against UVB-induced senescence in human dermal fibroblasts (HDFs) by examining its effects on SIRT1 and its target, acetylated-p53. An in vitro model of UVB-induced senescence using HDFs and an in vivo model using nude mice were employed to assess the dermal protective effects of galangin. The results demonstrate that while UVB exposure does not decrease SIRT1 protein levels, it impairs its enzymatic function. However, galangin treatment counteracts these adverse effects. Additionally, UVB exposure significantly reduces cell viability and upregulates senescence markers like p16, p21, and p53 nuclear transactivation. An increase in senescence-associated β-galactosidase (SA-β-gal) positive cells was observed in UVB-exposed dermal fibroblasts. Galangin treatment mitigates UVB-induced cellular senescence by enhancing SIRT1-mediated p53 deacetylation, thereby inhibiting nuclear translocation and reducing dermal senescence. These findings suggest that galangin is a promising agent for alleviating UVB-induced skin aging and could be a potential component in antiaging cosmetic formulations.

Micronuclei (MN), defined as small extra-nuclear chromatin bodies enclosed by a nuclear envelope, serve as noticeable markers of chromosomal instability (CIN). The MN have been used for breast cancer (BC) screening, diagnosis, and prognosis. However, more recently they have gained attention as seats for active chromosomal rearrangements. BC subtypes exhibit differential CIN levels and aggressiveness. This study aimed to investigate MN chromosomal contents across BC subtypes, exploring its potential role in aggressiveness and pathogenesis. Immunostaining of BC cells was performed with anti-centromeric antibody followed by confocal microscopy. Further, fluorescence in situ hybridization (FISH) was done to check the presence of specific chromosomes in the MN. The real time PCR was also done from the RNA isolated from MN to check the expression of TP53 gene. BC cell lines (CLs) showed the presence of both centromere-positive ( +) and -negative ( -) MN, with significant variation in frequency among hormone and human epidermal growth factor receptor positive and triple-negative (TN) BC cells. FISH targeting chromosomes 1, 3, 8, 11, and 17 detected centromeric signals for all the above chromosomes in MN with a relatively higher prevalence of chromosome 17 in all the CLs. Out of all the CLs, TNBC cells demonstrated the highest frequency of centromere + and chromosome 17 + MN. TP53 expression could also be demonstrated inside the MN by FISH and real time PCR. Patient sample imprints also confirmed the presence of chromosome 17 in MN with polysomy of the same in corresponding nuclei. The high prevalence of chromosome 17 in BC MN may connote the importance of its rearrangements in the pathogenesis of BC. Further, the higher prevalence of chromosome 17 and 1 signals in TNBC MN point towards the significance of pathogenetic events involving the genes located in these chromosomes in evolution of this more aggressive phenotype., (© 2024. The Author(s) under exclusive licence to Japan Human Cell Society.)

Extranodal NK/T-cell lymphoma (ENKTCL) is an Epstein-Barr virus (EBV)-related neoplasm preferentially involving the upper aerodigestive tract. Here we show that NK-cell-specific Trp53 disruption in mice leads to the development of NK-cell lymphomas after long latency, which involve not only the hematopoietic system but also the salivary glands. Before tumor onset, Trp53 knockout causes extensive gene expression changes, resulting in immature NK-cell expansion, exclusively in the salivary glands. Both human and murine NK-cell lymphomas express tissue-resident markers, suggesting tissue-resident NK cells as their cell-of-origin. Murine NK-cell lymphomas show recurrent Myc amplifications and upregulation of MYC target gene signatures. EBV-encoded latent membrane protein 1 expression accelerates NK-cell lymphomagenesis and causes diverse microenvironmental changes, particularly myeloid propagation, through interferon-γ signaling. In turn, myeloid cells support tumor cells via CXCL16-CXCR6 signaling and its inhibition is effective against NK-cell tumors in vivo. Remarkably, KLRG1-expressing cells expand in the tumor and are capable of repopulating tumors in secondary recipients. Furthermore, targeting KLRG1 alone or combined with MYC inhibition using an eIF4 inhibitor is effective against NK-cell tumors. Therefore, our observations provide insights into the pathogenesis and highlight potential therapeutic targets, including CXCL16, KLRG1, and MYC, in ENKTCL, which can help improve its diagnostic and therapeutic strategies., (© 2024. The Author(s).)

Epstein-Barr virus (EBV) co-infections with human papillomavirus (HPV) have been observed in oropharyngeal squamous cell carcinoma. Modeling EBV/HPV co-infection in organotypic epithelial raft cultures revealed that HPV16 E7 inhibited EBV productive replication through the facilitated degradation of the retinoblastoma protein pRb/p105. To further understand how pRb is required for EBV productive replication, we generated CRISPR-Cas9 pRb knockout (KO) normal oral keratinocytes (NOKs) in the context of wild-type and mutant K120E p53. EBV replication was examined in organotypic rafts as a physiological correlate for epithelial differentiation. In pRb KO rafts, EBV DNA copy number was statistically decreased compared to vector controls, regardless of p53 context. Loss of pRb did not affect EBV binding or internalization of calcium-treated NOKs or early infection of rafts. Rather, the block in EBV replication correlated with impaired immediate early gene expression. An EBV infection time course in rafts with mutant p53 demonstrated that pRb-positive basal cells were initially infected with delayed replication occurring in differentiated layers. Loss of pRb showed increased S-phase progression makers and elevated activator E2F activity in raft tissues. Complementation with a panel of pRb/E2F binding mutants showed that wild type or pRb∆685 mutant capable of E2F binding reduced S-phase marker gene expression, rescued EBV DNA replication, and restored BZLF1 expression in pRb KO rafts. However, pRb KO complemented with pRb661W mutant, unable to bind E2Fs, failed to rescue EBV replication in raft culture. These findings suggest that EBV productive replication in differentiated epithelium requires pRb inhibition of activator E2Fs to restrict S-phase progression.IMPORTANCEA subset of human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma is co-positive for Epstein-Barr virus (EBV). Potential oncogenic viral interactions revealed that HPV16 E7 inhibited productive EBV replication within the differentiated epithelium. As E7 mediates the degradation of pRb, we aimed to establish how pRb is involved in EBV replication. In the context of differentiated epithelium using organotypic raft culture, we evaluated how the loss of pRb affects EBV lytic replication to better comprehend EBV contributions to carcinogenesis. In this study, ablation of pRb interfered with EBV replication at the level of immediate early gene expression. Loss of pRb increased activator E2Fs and associated S-phase gene expression throughout the differentiated epithelium. Complementation studies showed that wild type and pRb mutant capable of binding to E2F rescued EBV replication, while pRb mutant lacking E2F binding did not. Altogether, these studies support that in differentiated tissues, HPV16 E7-mediated degradation of pRb inhibits EBV replication through unregulated E2F activity., Competing Interests: The authors declare no conflict of interest.

Cancer is caused by an accumulation of somatic mutations and copy number alterations (CNAs). Besides mutations, these copy number changes are key characteristics of cancer development. Nonetheless, some tumors show hardly any CNAs, a remarkable phenomenon in oncogenesis. Head and neck squamous cell carcinomas (HNSCCs) arise by either exposure to carcinogens, or infection with the human papillomavirus (HPV). HPV-negative HNSCCs are generally characterized by many CNAs and frequent mutations in CDKN2A, TP53, FAT1, and NOTCH1. Here, we present the hallmarks of the distinct subgroup of HPV-negative HNSCC with no or few CNAs (CNA-quiet) by genetic profiling of 802 oral cavity squamous cell carcinomas (OCSCCs). In total, 73 OCSCC (9.1%) are classified as CNA-quiet and 729 as CNA-other. The CNA-quiet group is characterized by wild-type TP53, frequent CASP8 and HRAS mutations, and a less immunosuppressed tumor immune microenvironment with lower density of regulatory T cells. Patients with CNA-quiet OCSCC are older, more often women, less frequently current smokers, and have a better 5-year overall survival compared to CNA-other OCSCC. This study demonstrates that CNA-quiet OCSCC should be considered as a distinct, clinically relevant subclass. Given the clinical characteristics, the patient group with these tumors will rapidly increase in the aging population., (© 2024. The Author(s).)

Background: Ischemic stroke (IS) is known for its high morbidity, disability and mortality rates, and studies designed to explore its pathophysiological mechanisms and identify novel therapeutic strategies are urgently needed. We aimed to probe the effects of the deubiquitinase OTUD3-IRP2-p53/PTGS2 pathway on cerebral ischemia‒reperfusion (I/R) injury and hippocampal neuron ferroptosis., Methods: A cerebral I/R mouse model was established. Furthermore, lentiviral vectors that overexpressed OTUD3 and knocked down IRP2 were constructed, and a series of assays were performed to probe the OTUD3/IRP2/p53/PTGS2 mechanism. An oxygen‒glucose deprivation and reoxygenation (OGD/R) model of mouse hippocampal neurons was constructed. Then, OTUD3 and IRP2 were knocked down and overexpressed, and p53 was overexpressed to explore the mechanism of the OTUD3/IRP2/p53/PTGS2 pathway., Results: OTUD3 and IRP2 were expressed at low levels in cerebral I/R models. OTUD3 promoted IRP2 expression to protect damaged hippocampal neurons. Moreover, IRP2 affected ferroptosis in hippocampal neurons. In addition, IRP2 inhibited p53. After IRP2 and p53 were overexpressed, IRP2 regulated the p53/PTGS2 pathway and affected ferroptosis in hippocampal neurons. In vivo, after overexpressing OTUD3 and knocking down IRP2, we found that overexpression of OTUD3 promoted IRP2 expression to reduce ferroptosis in hippocampal neurons and improve cerebral I/R injury via the inhibition of the p53/PTGS2 pathway., Conclusions: The deubiquitinase OTUD3 stabilized IRP2 expression to reduce hippocampal neuron ferroptosis via the p53/PTGS2 pathway to subsequently ameliorate cerebral I/R injury., (© 2024. The Author(s).)

Poly (ADP-ribose) polymerase (PARP) 1 and 2 enzymatic inhibitors (PARPi) are promising cancer treatments. But recently, their use has been hindered by unexplained severe anemia and treatment-related leukemia. In addition to enzymatic inhibition, PARPi also trap PARP1 and 2 at DNA lesions. Here we report that, unlike Parp2 -/- mice, which develop normally, mice expressing catalytically inactive Parp2 (E534A and Parp2 EA/EA ) succumb to Tp53- and Chk2-dependent erythropoietic failure in utero, mirroring Lig1 -/- mice. While DNA damage mainly activates PARP1, we demonstrate that DNA replication activates PARP2 robustly. PARP2 is selectively recruited and activated by 5'-phosphorylated nicks (5'p-nicks), including those between Okazaki fragments, resolved by ligase 1 (Lig1) and Lig3. Inactive PARP2, but not its active form or absence, impedes Lig1- and Lig3-mediated ligation, causing dose-dependent replication fork collapse, which is detrimental to erythroblasts with ultra-fast forks. This PARylation-dependent structural function of PARP2 at 5'p-nicks explains the detrimental effects of PARP2 inactivation on erythropoiesis, shedding light on PARPi-induced anemia and the selection for TP53/CHK2 loss., Competing Interests: Declaration of interests D.M. is a scientific editor for Molecular Cell and therefore was not involved in the peer review or the decision-making process of this manuscript., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Abstract: Mutations in the TP53 gene, particularly multihit alterations, have been associated with unfavorable clinical features and prognosis in patients diagnosed with myelodysplastic syndrome (MDS). Despite this, the role of TP53 gene aberrations in MDS with isolated deletion of chromosome 5 [MDS-del(5q)] remains unclear. This study aimed to assess the impact of TP53 gene mutations and their allelic state in patients with MDS-del(5q). To that end, a comprehensive analysis of TP53 abnormalities, examining both TP53 mutations and allelic imbalances, in 682 patients diagnosed with MDS-del(5q) was conducted. Twenty-four percent of TP53-mutated patients exhibited multihit alterations, whereas the remaining patients displayed monoallelic mutations. TP53-multihit alterations were predictive of an increased risk of leukemic transformation. The impact of monoallelic alterations was dependent on the variant allele frequency (VAF); patients with TP53-monoallelic mutations and VAF <20% exhibited behavior similar to TP53 wild type, and those with TP53-monoallelic mutations and VAF ≥20% presented outcomes equivalent to TP53-multihit patients. This study underscores the importance of considering TP53 allelic state and VAF in the risk stratification and treatment decision-making process for patients with MDS-del(5q)., (© 2024 American Society of Hematology. Published by Elsevier Inc. All rights are reserved, including those for text and data mining, AI training, and similar technologies.)

The discovery of embryonic stem cell (ESC) mediating tumoricidal activity revealed the intimate relationship between ESCs and tumor cells, but the functional role of ESCs in tumor progression is poorly understood. To further investigate tumor cell and ESC interactions, we co-cultured mouse ESCs with mouse pancreatic cancer Pan02 cells or mouse melanoma B16-F10 cells in Transwell, and found that tumor cell invasion was significantly inhibited by ESCs. Application of ESCs to tumor-bearing mice resulted in significant inhibition of tumor metastasis in vivo. RNA-Seq analyses of tumor cell and ESC co-cultures identified TP53 and related signalling as major pathways involved in ESC-mediated inhibition of tumor cell invasion and metastasis, which indicated the potential clinical application of ESCs to treat cancer., (© 2024. The Author(s).)

Background: Androgen deprivation therapy (ADT) resistance is closely associated with altered AR status. Aberrant AR expression is critical for the induction of ADT resistance, necessitating the identification of an anti-PCa target independent of AR expression., Methods: Transcriptomic data and clinical information of PRAD were obtained from TCGA database. Genes with PCa-related and AR expression-independent were screened by bioinformatics, and characterized by PPI and GO functional enrichment analyses. Candidate genes were locked by co-expression correlation and disease-free survival (DFS) analyses. A prognostic gene set was established using LASSO Cox regression algorithm. Cox proportional risk regression was performed to identify a key prognostic gene. Expression of the target protein in PCa tissues was verified by The Human Protein Atlas database. In vitro validation of cellular function and molecular mechanism by knockdown and overexpression of the target gene., Results: Two AR expression-independent genes (SLC43A1 and XRCC3) were available for the optimal prognostic model. This gene set effectively predicted PRAD patients' DFS at 1-, 3- and 5-year, where XRCC3 and tumor (T) stage were independent risk factors. XRCC3 was higher expressed in PRAD patients with T3-T4 stages and accompanied by poorer DFS. IHC staining also validated its higher expression in high-risk PCa tissues. In vitro experiments demonstrated that silencing XRCC3 significantly inhibited 22Rv1 and DU145 cell proliferation, migration and invasion, while promoted apoptosis. Further, silencing XRCC3 promoted DNA damage-induced p53/Bax signaling pathway activation, which was absent with overexpression., Conclusion: Silencing XRCC3 exerts anti-PCa effects by promoting DNA damage-induced p53/Bax signaling pathway activation in an AR expression-independent manner., (© 2024. The Author(s).)

Objective: To explore the detection of BRAF, RAS, TERT promoter, and TP53 gene mutations in solitary thyroid micronodule (TMN) specimens obtained by ultrasound-guided fine-needle aspiration (US-FNA) using next-generation sequencing (NGS) technology and assess its diagnostic value in thyroid microcarcinomas (TMC). Methods: On-site recruitment of 428 patients with single suspicious TMC who underwent thyroid ultrasound examination, US-FNA, and NGS from September 2018 to July 2021 at Beijing Tongren Hospital affiliated to Capital Medical University. A total of 147 patients were finally included. NGS was used to detect mutations in the BRAF, RAS, TERT promoter, and TP53 genes in the US-FNA specimens. Comparisons were made between patients with and without gene mutations in terms of age, gender, and the maximum diameter of nodules. The diagnostic efficiency of BRAF mutation for TMC was calculated using the receiver operating characteristic (ROC) curve, with postoperative pathology as the gold standard. Results: The age [ M ( Q 1 , Q 3 )] of the 147 patients was 43.0 (32.0, 51.0) years, and 37 were male (25.2%). Among the 147 US-FNA specimens, 97 (66.0%) were detected with BRAF gene mutations, all of which were p.V600E point mutation; 6 (4.1%) were detected with RAS gene mutation, and no TERT promoter or TP53 gene mutations were detected. Postoperative pathology confirmed that 136 cases were TMC, all of which were papillary thyroid microcarcinomas (PTMC); 11 cases (7.5%)were benign. Among 136 TMC samples, BRAF gene mutations were detected in 97 cases (71.3%). There were no statistically significant differences in age, gender, and maximum nodule diameter between patients with and without BRAF gene mutations (all P >0.05). The sensitivity and specificity of BRAF gene mutation in diagnosing TMC were 71.3% and 100.0%, respectively, with an area under the ROC curve (AUC) (95% CI ) of 0.857 (0.789-0.925). For nodules classified as Bethesda Ⅲ-Ⅴ, the sensitivity and specificity were 63.0% and 100.0%, respectively, with an AUC (95% CI ) of 0.815 (0.680-0.950). Conclusions: NGS technology can successfully detect multiple gene mutations in US-FNA specimens from TMN patients, especially BRAF gene mutation, and BRAF gene mutation has certain value in diagnosing TMC.

Background: In the past decade, the biological functions of various RNA modifications in mammals have been uncovered. N4-acetylcytidine (ac4C), a highly conserved RNA modification, has been implicated in human diseases. Despite this, the involvement of RNA ac4C modification in cardiac physiology and pathology remains incompletely understood. NAT10 (N-acetyltransferase 10) stands as the sole acetyltransferase known to catalyze RNA ac4C modification. This study aims to explore the role of NAT10 and ac4C modification in cardiac physiology and pathology., Methods and Results: Cardiac-specific knockout of NAT10, leading to reduced RNA ac4C modification, during both neonatal and adult stages resulted in severe heart failure. NAT10 deficiency induced cardiomyocyte apoptosis, a crucial step in heart failure pathogenesis, supported by in vitro data. Activation of the p53 signaling pathway was closely associated with enhanced apoptosis in NAT10-deficient cardiomyocytes. As ac4C modification on mRNA influences translational efficiency, we employed ribosome footprints coupled with RNA sequencing to explore genome-wide translational efficiency changes caused by NAT10 deficiency. We identified and validated that the translational efficiency of Kmt5a was suppressed in NAT10 knockout hearts due to reduced ac4C modification on its mRNA. This finding was consistent with the observation that Kmt5a protein levels were reduced in heart failure despite unchanged mRNA expression. Knockdown of Kmt5a in cardiomyocytes recapitulated the phenotype of NAT10 deficiency, including increased cardiomyocyte apoptosis and activated p53 signaling. Finally, overexpression of Kmt5a rescued cardiomyocyte apoptosis and p53 activation induced by NAT10 inhibition., Conclusions: Our study highlights the significance of NAT10 in cardiomyocyte physiology, demonstrating that NAT10 loss is sufficient to induce cardiomyocyte apoptosis and heart failure. NAT10 regulates the translational efficiency of Kmt5a , a key mediator, through mRNA ac4C modification during heart failure.

Biallelic loss of cyclin-dependent kinase 12 (CDK12) defines a metastatic castration-resistant prostate cancer (mCRPC) subtype. It remains unclear, however, whether CDK12 loss drives prostate cancer (PCa) development or uncovers pharmacologic vulnerabilities. Here, we show Cdk12 ablation in murine prostate epithelium is sufficient to induce preneoplastic lesions with lymphocytic infiltration. In allograft-based CRISPR screening, Cdk12 loss associates positively with Trp53 inactivation but negatively with Pten inactivation. Moreover, concurrent Cdk12/Trp53 ablation promotes proliferation of prostate-derived organoids, while Cdk12 knockout in Pten-null mice abrogates prostate tumor growth. In syngeneic systems, Cdk12/Trp53-null allografts exhibit luminal morphology and immune checkpoint blockade sensitivity. Mechanistically, Cdk12 inactivation mediates genomic instability by inducing transcription-replication conflicts. Strikingly, CDK12-mutant organoids and patient-derived xenografts are sensitive to inhibition or degradation of the paralog kinase, CDK13. We therein establish CDK12 as a bona fide tumor suppressor, mechanistically define how CDK12 inactivation causes genomic instability, and advance a therapeutic strategy for CDK12-mutant mCRPC., Competing Interests: Declaration of interests A.M.C. co-founded and serves on scientific advisory boards (SABs) of Lynx Dx, Flamingo Therapeutics, Medsyn Pharma, Oncopia Therapeutics, and Esanik Therapeutics. A.M.C. is an advisor to Aurigene Oncology Limited, Proteovant, Tempus, Rappta, and Ascentage. C.J.L. received research funding from AstraZeneca, Merck KGaA, Artios, and NeoPhore and consultancy, SAB membership, or honoraria payments from FoRx, Syncona, Sun Pharma, Gerson Lehrman Group, Merck KGaA, Vertex, AstraZeneca, Tango, 3rd Rock, Ono Pharma, Artios, Abingworth, Tesselate, Dark Blue Therapeutics, Pontifax, Astex, NeoPhore, Glaxo Smith Kline, and Dawn Bioventures. C.J.L. has stock in Tango, Ovibio, Hysplex, and Tesselate. C.J.L. is named inventor on patents describing use of DNA repair inhibitors and stands to gain from their development and use. J.C. is an advisor for Exai Bio. F.Y.F. has served on SAB or received consulting fees from Astellas, Bayer, Celgene, Clovis Oncology, Janssen, Genentech Roche, Myovant, Roivant, Sanofi, and Blue Earth Diagnostics. F.Y.F. is also an SAB member for Artera, ClearNote Genomics, Serimmune, and BMS (Microenvironment Division). K.D. is an advisor for Kinoteck Therapeutics and has received financial support from Livzon Pharmaceutical Group. Patents for CDK12/13 degraders/inhibitors used here have been filed by the University of Michigan and Shanghai Institute of Organic Chemistry, with A.M.C., K.D., X.W., J.Y., Y. Chang, and J.C.T. as co-inventors., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Context: The relationship between the genomic profile and prognosis of advanced thyroid carcinoma requiring drug therapy has not been reported., Objective: To evaluate the treatment period and overall survival time for each genetic alteration in advanced thyroid carcinoma that requires drug therapy., Methods: We conducted a retrospective observational study using a national database in Japan, which included 552 cases of thyroid carcinoma out of 53 543 patients in the database., Results: The database included anaplastic thyroid carcinoma (23.6%), poorly differentiated thyroid carcinoma (10.0%), and differentiated thyroid carcinoma (66.4%). The most common genetic abnormalities were TERT promoter (66.3%), BRAF (56.7%), and TP53 (32.2%). The typical driver genes were BRAF V600E (55.0%), RAS (18.5%), RET fusion (4.7%), NTRK fusion (1.6%), and ALK fusion (0.4%). The most common regimen was lenvatinib, and the time to treatment failure was not different despite the presence of BRAF or RAS mutations. In differentiated thyroid carcinoma and poorly differentiated thyroid carcinoma, TP53 alterations independently predicted worse overall survival (hazard ratio = 2.205, 95% confidence interval: 1.135-4.283). In anaplastic thyroid carcinoma, no genetic alterations were associated with overall survival., Conclusion: Genetic abnormalities with treatment options were found in 62.7% of advanced thyroid carcinomas. TP53 abnormality was an independent poor prognostic factor for overall survival in differentiated thyroid carcinoma. The time to treatment failure for lenvatinib was not different based on genetic profile., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society.)

Purpose: Leiomyosarcomas (LMS) are clinically and molecularly heterogeneous, occurring mostly in sporadic but also syndromic settings. The role of pathogenic germline variants (PGV) as LMS drivers and their impact on outcomes remains uncertain., Experimental Design: We performed a comprehensive clinicopathologic and molecular analysis using a tumor-normal DNA next-generation sequencing assay (Memorial Sloan Kettering-Integrated Mutational Profiling of Actionable Cancer Targets) of germline-associated LMS compared with sporadic LMS., Results: Among 285 LMS [120 soft-tissue LMS (STLMS) and 165 uterine LMS (ULMS)] with germline testing, 78 (27%, 43 STLMS and 35 ULMS) cases harbored PGV, with 35/78 (45%) of PGV carriers showing biallelic inactivation of the corresponding gene in the tumor (26 STLMS and nine ULMS). The most frequent germline predispositions were TP53 (Li-Fraumeni syndrome; 17 patients, 16 in STLMS) and RB1 (retinoblastoma; 13 patients, 11 in STLMS). Germline TP53 and somatic RB1 alterations often co-occurred in the tumor andvice versa. Other biallelically inactivated PGV were enriched in DNA damage repair-related genes: CHEK2, MSH2, MSH6, RAD51D, BRCA2, and FANCA. Monoallelic PGV were mostly in ULMS and associated with co-occurring TP53 and RB1 somatic alterations. Patients with STLMS with biallelic but not monoallelic PGV were significantly younger than patients with sporadic STLMS (median ages 38 vs. 52 vs. 58 years). No differences in disease-specific or progression-free survival were observed in germline-associated versus sporadic LMS regardless of biallelic status., Conclusions: Although patients with ULMS had a relatively low proportion of PGV, a high percentage of patients with STLMS with PGV had tumor biallelic status, indicating that PGV drive tumorigenesis in these individuals. These findings have significant implications for genetic testing recommendations., (©2024 American Association for Cancer Research.)

The tumor suppressor p53 is a transcription factor involved in a variety of crucial cellular functions, including cell cycle arrest, DNA repair and apoptosis. Still, a growing number of studies indicate that p53 plays multiple roles in spermatogenesis, as well as in the occurrence and development of male infertility. The representative functions of p53 in spermatogenesis include the proliferation of spermatogonial stem cells (SSCs), spermatogonial differentiation, spontaneous apoptosis, and DNA damage repair. p53 is involved in various male infertility-related diseases. Innovative therapeutic strategies targeting p53 have emerged in recent years. This review focuses on the role of p53 in spermatogenesis and male infertility and analyses the possible underlying mechanism involved. All these conclusions may provide a new perspective on drug intervention targeting p53 for male infertility treatment., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Li, Huang, Luo, Sun, Guo, Yang, Zhang and Ni.)

FBXO protein family plays an essential role in the ubiquitination process acting as E3 ligases, which may contribute to the progression of cancers. However, the molecular functions of FBXOs in hepatocellular carcinoma (HCC) remain incompletely understood. Here, we investigated the overlapping genes between the FBXOs and differentially expressed genes (DEGs) of HCC identified by utilizing The Cancer Genome Atlas (TCGA) dataset, then, a prognostic model with effective predictive capacity was constructed based on the uni-cox and LASSO regression analyses. To elucidate the underlying mechanism of the FBXO model genes, KEGG analysis was carried out. Drug metabolism-cytochrome P450 and retinol metabolism were revealed as the potential pathway, which Increased the credibility of subsequent drug prediction research. Meanwhile, patients divided by the prognostic model showed a different immune infiltrating status and we also found FBXO model genes may ubiquitinate P53, inducing TP53 more prone to mutations, thereby promoting the occurrence and development of tumors. Consistent with these findings, the result of immunohistochemistry (IHC) validated an elevated expression of these model genes in HCC tissues than in the adjacent tissues. The primary aim of this investigation is to formulate a prognostic model while exploring the underlying mechanisms associated with FBXO genes in HCC. These findings offer initial research perspectives on the involvement of FBXO genes in HCC and contribute to the discovery of dependable biomarkers for the management, prognostication, and early detection of HCC in patients., (© 2024. The Author(s).)

Aims: Although the cannabinoid CB1 receptor has been implicated in atherosclerosis, its cell-specific effects in this disease are not well understood. To address this, we generated a transgenic mouse model to study the role of myeloid CB1 signalling in atherosclerosis., Methods and Results: Here, we report that male mice with myeloid-specific Cnr1 deficiency on atherogenic background developed smaller lesions and necrotic cores than controls, while only minor genotype differences were observed in females. Male Cnr1-deficient mice showed reduced arterial monocyte recruitment and macrophage proliferation with less inflammatory phenotype. The sex-specific differences in proliferation were dependent on oestrogen receptor (ER)α-oestradiol signalling. Kinase activity profiling identified a CB1-dependent regulation of p53 and cyclin-dependent kinases. Transcriptomic profiling further revealed chromatin modifications, mRNA processing, and mitochondrial respiration among the key processes affected by CB1 signalling, which was supported by metabolic flux assays. Chronic administration of the peripherally restricted CB1 antagonist JD5037 inhibited plaque progression and macrophage proliferation, but only in male mice. Finally, CNR1 expression was detectable in human carotid endarterectomy plaques and inversely correlated with proliferation, oxidative metabolism, and inflammatory markers, suggesting a possible implication of CB1-dependent regulation in human pathophysiology., Conclusion: Impaired macrophage CB1 signalling is atheroprotective by limiting their arterial recruitment, proliferation, and inflammatory reprogramming in male mice. The importance of macrophage CB1 signalling appears to be sex-dependent., Competing Interests: Conflict of interest: none declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Spinal cord injury (SCI) is a debilitating central nervous system (CNS) disorder that leads to significant motor and sensory impairments. Given the limited regenerative capacity of adult mammalian neurons, this study presents an innovative strategy to enhance axonal regeneration and functional recovery by identifying a novel factor that markedly promotes axonal regeneration. Employing a zebrafish model with targeted single axon injury in Mauthner cells (M-cells) and utilizing the Tg (Tol056: EGFP) transgenic line for in vivo monitoring, we investigate the intrinsic mechanisms underlying axonal regeneration. This research specifically examines the role of amino acid transport, emphasizing the role of the solute carrier 1A4 amino acid transporter in axonal regeneration. Our findings demonstrate that Slc1a4 overexpression significantly enhances axonal regeneration in M-cells, whereas Slc1a4 deficiency impedes this process, which is concomitant with the downregulation of the P53/Gap43 signaling pathway. By elucidating the fundamental role of Slc1a4 in axonal regeneration and uncovering its underlying mechanisms, this study thus provides novel insights into therapeutic strategies for SCI.

Colorectal cancer CRC remains one of the leading causes of cancer-related deaths worldwide, with chronic intestinal inflammation identified as a major risk factor. Notably, the tumor suppressor TP53 undergoes mutation at higher rates and earlier stages during human inflammation-driven colon tumorigenesis than in sporadic cases. We investigated whether deleting Trp53 affects inflammation-induced tumor growth and the expression of Lgr5+ cancer stem cells in mice. We examined azoxymethane (AOM)/dextran sodium sulfate (DSS)-induced colon tumorigenesis in wild-type Trp53 (+/+) , heterozygous (+/-) , and knockout (-/-) mice. Trp53-/- mice showed increased sensitivity to DSS colitis and earlier accelerated tumorigenesis with 100% incidence. All groups could develop invasive tumors, but knockouts displayed the most aggressive features. Unlike wild-type CRC, knockouts selectively showed increased populations of Lgr5+ colon cancer stem-like cells. Trp53 loss also boosted laminin, possibly facilitating the disruption of the tumor border. This study highlights how Trp53 deletion promotes the perfect storm of inflammation and stemness, driving colon cancer progression. Trp53 deletion dramatically shortened AOM/DSS latency and improved tumor induction efficiency, offering an excellent inflammation-driven CRC model.

Background: Mutations in the TP53 tumor suppressor gene are well-established drivers of colorectal cancer (CRC) development. However, data on the prevalence of TP53 variants and their association with consensus molecular subtype (CMS) classification in patients with CRC from Rwanda are currently lacking. This study addressed this knowledge gap by investigating TP53 mutation status concerning CMS classification in a CRC cohort from Rwanda., Methods: Formalin-fixed paraffin-embedded (FFPE) tissue blocks were obtained from 51 patients with CRC at the University Teaching Hospital of Kigali, Rwanda. Exons 4 to 11 and their flanking intron-exon boundaries in the TP53 gene were sequenced using Sanger sequencing to identify potential variants. The recently established immunohistochemistry-based classifier was employed to determine the CMS of each tumor., Results: Sequencing analysis of cancerous tissue DNA revealed TP53 pathogenic variants in 23 of 51 (45.1%) patients from Rwanda. These variants were predominantly missense types (18/23, 78.3%). The most frequent were c.455dup (p.P153Afs*28), c.524G > A (p.R175H), and c.733G > A (p.G245S), each identified in three tumors. Trinucleotide sequence context analysis of the 23 mutations (20 of which were single-base substitutions) revealed a predominance of the [C > N] pattern among single-base substitutions (SBSs) (18/20; 90.0%), with C[C > T]G being the most frequent mutation (5/18, 27.8%). Furthermore, pyrimidine bases (C and T) were preferentially found at the 5' flanking position of the mutated cytosine (13/18; 72.2%). Analysis of CMS subtypes revealed the following distribution: CMS1 (microsatellite instability-immune) (6/51, 11.8%), CMS2 (canonical) (28/51, 54.9%), CMS3 (metabolic) (9/51, 17.6%), and CMS4 (mesenchymal) (8/51, 15.7%). Interestingly, the majority of TP53 variants were in the CMS2 subgroup (14/23; 60.1%)., Conclusion: Our findings indicate a high frequency of TP53 variants in CRC patients from Rwanda. Importantly, these variants are enriched in the CMS2 subtype. This study, representing the second investigation into molecular alterations in patients with CRC from Rwanda and the first to explore TP53 mutations and CMS classification, provides valuable insights into the molecular landscape of CRC in this understudied population., (© 2024. The Author(s).)

The p53 mutation is the most common genetic mutation associated with human neoplasia. TP53 missense mutations, which frequently arise early in breast cancer, are present in over thirty percent of breast tumors. In breast cancer, p53 mutations are linked to a more aggressive course of the disease and worse overall survival rates. TP53 mutations are mostly seen in triple-negative breast cancer, a very diverse kind of the disease. The majority of TP53 mutations originate in the replacement of individual amino acids within the p53 protein's core domain, giving rise to a variety of variations referred to as "mutant p53s." In addition to gaining carcinogenic qualities through gain-of-function pathways, these mutants lose the typical tumor-suppressive features of p53 to variable degrees. The gain-of-function impact of stabilized mutant p53 causes tumor-specific dependency and resistance to therapy. P53 is a prospective target for cancer therapy because of its tumor-suppressive qualities and the numerous alterations that it experiences in tumors. Phenotypic abnormalities in breast cancer, notably poorly differentiated basal-like tumors are frequently linked to high-grade tumors. By comparing data from cell and animal models with clinical outcomes in breast cancer, this study investigates the molecular mechanisms that convert gene alterations into the pathogenic consequences of mutant p53's tumorigenic activity. The study delves into current and novel treatment approaches aimed at targeting p53 mutations, taking into account the similarities and differences in p53 regulatory mechanisms between mutant and wild-type forms, as well., (© 2024. The Author(s).)

Abstract: T-prolymphocytic leukemia (T-PLL) is a mature T-cell neoplasm associated with marked chemotherapy resistance and continued poor clinical outcomes. Current treatments, that is, the CD52-antibody alemtuzumab, offer transient responses, with relapses being almost inevitable without consolidating allogeneic transplantation. Recent more detailed concepts of T-PLL's pathobiology fostered the identification of actionable vulnerabilities: (1) altered epigenetics, (2) defective DNA damage responses, (3) aberrant cell-cycle regulation, and (4) deregulated prosurvival pathways, including T-cell receptor and JAK/STAT signaling. To further develop related preclinical therapeutic concepts, we studied inhibitors of histone deacetylases ([H]DACs), B-cell lymphoma 2 (BCL2), cyclin-dependent kinase (CDK), mouse double minute 2 (MDM2), and classical cytostatics, using (1) single-agent and combinatorial compound testing in 20 well-characterized and molecularly profiled primary T-PLL (validated by additional 42 cases) and (2) 2 independent murine models (syngeneic transplants and patient-derived xenografts). Overall, the most efficient/selective single agents and combinations (in vitro and in mice) included cladribine, romidepsin ([H]DAC), venetoclax (BCL2), and/or idasanutlin (MDM2). Cladribine sensitivity correlated with expression of its target RRM2. T-PLL cells revealed low overall apoptotic priming with heterogeneous dependencies on BCL2 proteins. In additional 38 T-cell leukemia/lymphoma lines, TP53 mutations were associated with resistance toward MDM2 inhibitors. P53 of T-PLL cells, predominantly in wild-type configuration, was amenable to MDM2 inhibition, which increased its MDM2-unbound fraction. This facilitated P53 activation and downstream signals (including enhanced accessibility of target-gene chromatin regions), in particular synergy with insults by cladribine. Our data emphasize the therapeutic potential of pharmacologic strategies to reinstate P53-mediated apoptotic responses. The identified efficacies and their synergies provide an informative background on compound and patient selection for trial designs in T-PLL., (© 2024 American Society of Hematology. Published by Elsevier Inc. All rights are reserved, including those for text and data mining, AI training, and similar technologies.)

Deep mutational scanning enables high-throughput functional assessment of genetic variants. While phenotypic measurements from screening assays generally align with clinical outcomes, experimental noise may affect the accuracy of individual variant estimates. We developed the FUSE (functional substitution estimation) pipeline, which leverages measurements collectively within screening assays to improve the estimation of variant impacts. Drawing data from 115 published functional assays, FUSE assesses the mean functional effect per amino acid position and makes estimates for individual allelic variants. It enhances the correlation of variant functional effects from different assay platforms and increases the classification accuracy of missense variants in ClinVar across 29 genes (area under the receiver operating characteristic [ROC] curve [AUC] from 0.83 to 0.90). In UK Biobank patients with rare missense variants in BRCA1, LDLR, or TP53, FUSE improves the classification accuracy of associated phenotypes. FUSE can also impute variant effects for substitutions not experimentally screened. This approach improves accuracy and broadens the utility of data from functional screening., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

This study aimed to investigate the activation of error-prone DNA repair pathway in response to Helicobacter pylori infection. Relative changes in the expression levels of genes involved in the non-homologous end-joining pathway (NHEJ) in H. pylori-infected (Cases) and non-infected patients (Controls) with chronic gastritis were measured. A significant increase in the relative expression level of TP53, and significant decrease in the relative transcription of lncRNA LINP1 and XRCC5 were detected in the case group. The transcription of Lig4 and XRCC6 was increased in the case group, which was not statistically significant. The Spearman's Correlation Coefficient analysis showed a significant positive-correlation between the transcriptional levels of LINP1 and XRCC4/XRCC5/Lig4, and between XRCC5 and TP53/Lig4 both in the case and control groups. Moreover, a significant positive correlation between LinP1 and XRCC6 in the case, and a significant positive correlation between XRCC4 and Lig4, and a negative correlation between TP53 and LinP1/XRCC4/XRCC5 in the control group was detected. Although a relative difference was detected in transcriptional levels of the NHEJ gene mediators, downregulation of LinP1 in H. pylori-infected patients proposed the activation of a negative feedback loop, which may interfere with the NHEJ activity at the early stages of gastritis., (© 2024. The Author(s).)

Mitotic Arrest Deficient 1 (gene name MAD1L1), an essential component of the mitotic spindle assembly checkpoint, is frequently overexpressed in colon cancer, which correlates with poor disease-free survival. MAD1 upregulation induces two phenotypes associated with tumor promotion in tissue culture cells-low rates of chromosomal instability (CIN) and destabilization of the tumor suppressor p53. Using CRISPR/Cas9 gene editing, we generated a novel mouse model by inserting a doxycycline (dox)-inducible promoter and HA tag into the endogenous mouse Mad1l1 gene, enabling inducible expression of HA-MAD1 following exposure to dox in the presence of the reverse tet transactivator (rtTA). A modest 2-fold overexpression of MAD1 in murine colon resulted in decreased p53 expression and increased mitotic defects consistent with CIN. After exposure to the colon-specific inflammatory agent dextran sulfate sodium (DSS), 31% of mice developed colon lesions, including a mucinous adenocarcinoma, while none formed in control animals. Lesion incidence was particularly high in male mice, 57% of which developed at least one hyperplastic polyp, adenoma or adenocarcinoma in the colon. Notably, mice expressing HA-MAD1 also developed lesions in tissues in which DSS is not expected to induce inflammation. These findings demonstrate that MAD1 upregulation is sufficient to promote colon tumorigenesis in the context of inflammation in immune-competent mice., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Copeland et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

A cell's fate involves transitions among its various states, each defined by a distinct gene expression profile governed by the topology of gene regulatory networks, which are affected by 3D genome organization. Here, we develop thermodynamic models to determine the fate of a malignant cell as governed by the tumor suppressor p53 signaling network, taking into account long-range chromatin interactions in the mean-field approximation. The tumor suppressor p53 responds to stress by selectively triggering one of the potential transcription programs that influence many layers of cell signaling. These range from p53 phosphorylation to modulation of its DNA binding affinity, phase separation phenomena, and internal connectivity among cell fate genes. We use the minimum free energy of the system as a fundamental property of biological networks that influences the connection between the gene network topology and the state of the cell. We constructed models based on network topology and equilibrium thermodynamics. Our modeling shows that the binding of phosphorylated p53 to promoters of target genes can have properties of a first order phase transition. We apply our model to cancer cell lines ranging from breast cancer (MCF-7), colon cancer (HCT116), and leukemia (K562), with each one characterized by a specific network topology that determines the cell fate. Our results clarify the biological relevance of these mechanisms and suggest that they represent flexible network designs for switching between developmental decisions., (© 2024 Author(s). All article content, except where otherwise noted, is licensed under a Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).)

Objective: To assess the distribution of molecular classes and their impact on the risk of recurrence in endometrial cancer patients with lymph node metastasis at the time of primary surgery., Methods: Endometrial cancer patients with lymph node micrometastasis or macrometastasis (International Federation of Gynecology and Obstetrics (FIGO) 2009 stage IIIC) after surgical staging at five referral centers worldwide from October 2013 to September 2022 who underwent molecular classification were identified. Endometrial cancers were categorized into four molecular classes: POLE mutated, mismatch repair deficient, p53 abnormal, and no specific molecular profile. Survival analyses using Kaplan-Meier and Cox models (univariate and multivariate) were conducted to evaluate the relationship between molecular class and 5-year recurrence free survival., Results: 131 patients were included: 55 (42.0%) no specific molecular profile, 46 (35.1%) mismatch repair deficient, 1 (0.8%) POLE mutated, and 29 (22.1%) p53 abnormal. During a 5 year follow-up period, 50 (38.2%) patients experienced a recurrence with a median time of 1.2 years (interquartile range (IQR) 0.5-1.8). Median follow-up for the remaining 81 patients was 3.1 years (IQR 1.3-4.5). Survival analysis revealed a significant difference in recurrence-free survival between no specific molecular profile, mismatch repair deficient, and p53 abnormal classes (log rank p<0.01). In a model adjusted for type of lymph node metastasis and tumor grade, the molecular class did not retain significance (p=0.13), while in a model adjusted for type of lymph node metastasis and adjuvant therapy, the molecular class retained significance (p<0.01)., Conclusion: Among patients with stage IIIC endometrial cancer, POLE mutated tumors exhibited an extremely low prevalence, with no specific molecular profile emerging as the largest molecular subgroup. Despite the significant difference in recurrence-free survival between molecular classes, conventional histopathologic parameters retained crucial prognostic value. Our findings highlight the necessity of integrating molecular classes with pathological characteristics, rather than considering them in isolation as crucial prognostic factors in stage IIIC endometrial cancer., Competing Interests: Competing interests: None declared., (© IGCS and ESGO 2024. No commercial re-use. See rights and permissions. Published by BMJ.)

Objective: Circulating tumour DNA (ctDNA) is a promising non-invasive biomarker in cancer. We aim to assess the dynamic of ctDNA in patients with hepatocellular carcinoma (HCC)., Design: We analysed 772 plasmas from 173 patients with HCC collected at the time of diagnosis or treatment (n=502), 24 hours after locoregional treatment (n=154) and during follow-up (n=116). For controls, 56 plasmas from patients with chronic liver disease without HCC were analysed. All samples were analysed for cell free DNA (cfDNA) concentration, and for mutations in TERT promoter, CTNNB1 , TP53 , PIK3CA and NFE2L2 by sequencing and droplet-based digital PCR. Results were compared with 232 corresponding tumour samples., Results: In patients with active HCC, 40.2% of the ctDNA was mutated vs 14.6% in patients with inactive HCC and 1.8% in controls (p<0.001). In active HCC, we identified 27.5% of mutations in TERT promoter, 21.3% in TP53 , 13.1% in CTNNB1 , 0.4% in PIK3CA and 0.2% in NFE2L2, most of the times similar to those identified in the corresponding tumour. CtDNA mutation rate increased with advanced tumour stages (p<0.001). In 103 patients treated by percutaneous ablation, the presence and number of mutations in the ctDNA before treatment were associated with higher risk of death (p=0.001) and recurrence (p<0.001). Interestingly, cfDNA concentration and detectable mutations increased 24 hours after a locoregional treatment. Among 356 plasmas collected in 53 patients treated by systemic treatments, we detected mutations at baseline in 60.4% of the cases. In patients treated by atezolizumab-bevacizumab, persistence of mutation in ctDNA was associated with radiological progression (63.6% vs 36.4% for disappearance, p=0.019). In two patients progressing under systemic treatments, we detected the occurrence of mutations in CTNNB1 in the plasma that was subclonal in the tumour for one patient and not detectable in the tumour for the other one., Conclusion: ctDNA offers dynamic information reflecting tumour biology. It represents a non-invasive tool useful to guide HCC clinical management., Competing Interests: Competing interests: PN has received honoraria from and/or consults for AstraZeneca, Bayer, Bristol-Myers Squibb, Eisai, Gilead, Ipsen and Roche. He received research grants from AstraZeneca, AbbVie, Bristol–Myers Squibb and Eisai. NGC received travel and congress fees, consulting fees or honoraria for lectures, presentations, speakers’ bureaus from Abbvie, Bayer, Gilead, Ipsen, Intercept and Roche. J-CN received research funding from Bayer and Ipsen. CC, SI, GC, MZ, TZH, SR, BN, KH, SS, OS, VT, PL-P and JZ-R have nothing to disclose., (© Author(s) (or their employer(s)) 2024. No commercial re-use. See rights and permissions. Published by BMJ.)

The distal region of the uterine (Fallopian) tube is commonly associated with high-grade serous carcinoma (HGSC), the predominant and most aggressive form of ovarian or extra-uterine cancer. Specific cell states and lineage dynamics of the adult tubal epithelium (TE) remain insufficiently understood, hindering efforts to determine the cell of origin for HGSC. Here, we report a comprehensive census of cell types and states of the mouse uterine tube. We show that distal TE cells expressing the stem/progenitor cell marker Slc1a3 can differentiate into both secretory (Ovgp1+) and ciliated (Fam183b+) cells. Inactivation of Trp53 and Rb1, whose pathways are commonly altered in HGSC, leads to elimination of targeted Slc1a3+ cells by apoptosis, thereby preventing their malignant transformation. In contrast, pre-ciliated cells (Krt5+, Prom1+, Trp73+) remain cancer-prone and give rise to serous tubal intraepithelial carcinomas and overt HGSC. These findings identify transitional pre-ciliated cells as a cancer-prone cell state and point to pre-ciliation mechanisms as diagnostic and therapeutic targets., (© 2024. The Author(s).)

Ethnopharmacological Relevance: Adinandra nitida Merr. ex Li leaves serve as a herbal tea and hold a significant role in traditional Chinese medicine, being applied to assist in tumor treatment. Flavonoids present the primary bioactive constituents in Adinandra nitida Merr. ex Li leaves., Aim of the Study: To explore the potential of total flavonoids from Adinandra nitida Merr. ex Li Leaves (TFAN) in inhibiting non-small cell lung cancer (NSCLC) and further elucidate the underlying mechanisms., Materials and Methods: Human NSCLC cell lines and normal lung cell line were employed to assess the impact of TFAN (0-160 μg/mL for 24, 28 and 72 h) on cell proliferation in vitro. Immunofluorescence (IF) staining gauged p53 expression changes in NSCLC cells under TFAN present condition (150 μg/mL for 24 h). In vivo study utilized NSCLC cell derived xenograft tumors in nude mice, administering TFAN orally (200 and 400 mg/kg) for 14 days. Immunohistochemistry assessed Cleaved Caspase 3 expression change in A549 xenograft tumors treated with TFAN (400 mg/kg for 14 days). RNA-seq and KEGG analysis identified gene expression changes and enriched processes in A549 xenograft tumors treated with TFAN. CM-H 2 DCFDA and metabolomics assessed ROS level and GSH/GSSG pool changes in A549 cells under TFAN present condition. Cell viability assay and IF staining assessed A549 cell proliferation and p53 expression changes under H 2 O 2 -induced oxidative stress (0-40 μM for 24 h) and TFAN present conditions. GSEA and N-Acetyl-L-cysteine (NAC) rescue (0-1 μM for 24 h) analyzed the impact of TFAN on GSH de novo synthesis. NADPH/NADP + pool measurement and NADPH rescue (0-10 μM for 24 h) analyzed the impact of TFAN on GSH salvage synthesis. GC-FID and HPLC-MS were utilized to detect ethanol and ethyl acetate residues, and to characterize the chemical constituents in TFAN, respectively. The total flavonoid content of TFAN was determined using a 330 nm wavelength., Results: TFAN significantly inhibited A549 cells (wild-type p53) but not NCI-H1299 cells (p53-deficient), NCI-H596 cells (p53-mutant) or BEAS-2B in vitro. IF staining validated p53 genotype for the cell lines and revealed an increase in p53 expression in A549 cells after TFAN treatment. In vivo, TFAN selectively inhibited A549 xenograft tumor growth without discernible toxicity, inducing apoptosis evidenced by Cleaved Caspase 3 upregulation. RNA-seq and KEGG analysis suggested ROS biosynthesis was involved in TFAN-induced p53 activation in A549 cells. Elevated ROS level in TFAN-treated A549 cells were observed. Moreover, TFAN sensitized A549 cells to H 2 O 2 -induced oxidative stress, with higher p53 expression. Additionally, A549 cells compensated with GSH de novo synthesis under TFAN present condition, confirmed by GSEA and NAC rescue experiment. TFAN disrupted NADPH homeostasis to impair GSH salvage biosynthesis, supported by NADPH/NADP + change and NADPH rescue experiment. The chemical constituents of TFAN, with acceptable limits for ethanol and ethyl acetate residues and a total flavonoid content of 68.87%, included Catechin, Epicatechin, Quercitroside, Camellianin A, and Apigenin., Conclusion: The disruption of NADPH homeostasis by TFAN triggers ROS-dependent p53 activation that leads to apoptotic cell death, ultimately suppressing NSCLC growth. These findings offer potential therapeutic implications of Adinandra nitida Merr. ex Li leaves in combating NSCLC., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The six most common missense mutations in the DNA binding domain of p53 are known as "hot spots" and include two of the most frequently occurring p53 mutations (p53-R175H and p53-R273H). p53 stability and function are regulated by various post-translational modifications such as phosphorylation, acetylation, sumoylation, methylation, and interactions with other proteins including plakoglobin. Previously, using various carcinoma cell lines we showed that plakoglobin interacted with wild-type and several endogenous p53 mutants (e.g., R280K, R273H, S241F, S215R, R175H) and restored their tumor suppressor activities in vitro. Since mutant p53 function is both mutant-specific and cell context-dependent, we sought herein, to determine if plakoglobin tumor suppressive effects on exogenously expressed p53-R273H and p53-R175H mutants are similarly maintained under the same genetic background using the p53-null and plakoglobin-deficient H1299 cell line. Functional assays were performed to assess colony formation, migration, and invasion while immunoblotting and qPCR were used to examine the subcellular distribution and expression of specific proteins and genes that are typically regulated by or regulate p53 function and are altered in mutant p53-expressing cell lines and tumors. We show that though, plakoglobin interacted with both p53-R273H and p53-R175H mutants, it had a differential effect on the transcription and subcellular distribution of their gene targets and their overall oncogenic properties in vitro. Notably, we found that plakoglobin's tumor suppressive effects were significantly stronger in p53-R175H expressing cells compared to p53-R273H cells. Together, our results indicate that exploring plakoglobin interactions with p53-R175H may be useful for the development of cancer therapeutics focused on the restoration of p53 function., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Lo et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

P53 is a transcription factor that controls a variety of genes, primarily involved in cell cycle and other processes related to cell survival and death. We have isolated peptides targeting P53 (protein and domains) using the "phage display" technique. Interestingly, adding ZnCl2 at 5-10 mM in panning solutions helped to recover more plaque-forming units at least at round one of the screening. Subtractive docking analyses were designed by using a pool of common redundant peptides known as parasites. This rationale helped us differentiate between possibly specific and non-specific bindings. We found notable differences in docking characteristics between different sets of peptides either related to different targets or related to zinc-conditions. The set of zinc-related peptides shows advantageous docking profiles: sharper binding for some positions and distinct exclusive bound residues, including the relevant R248 and R273. Zinc would have modulating/helping role in the targeting of protein P53 by phage displayed peptides in addition to an enhancement action on bacterial infection., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Ben Abid et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Multiple pathogenic single-nucleotide polymorphisms (SNPs) have been identified as contributing factors in the aggravation of cancer prognosis and emergence of drug resistance in various cancers. Here, we targeted mutated EGFR and TP53 oncogenes harboring single-nucleotide missense mutations (EGFR-T790M and TP53-R273H) that are associated with gefitinib resistance. Co-delivery of adenine base editor (ABE) and EGFR- and TP53-SNP specific single-guide RNA via adenovirus (Ad) resulted in precise correction of the oncogenic mutations with high accuracy and efficiency in vitro and in vivo. Importantly, compared with a control group treated only with gefitinib, an EGFR inhibitor, co-treatment with Ad/ABE targeting SNPs in TP53 and EGFR in combination with gefitinib increased drug sensitivity and suppressed abnormal tumor growth more efficiently. Taken together, these results indicate that ABE-mediated correction of dual oncogenic SNPs can be an effective strategy for the treatment of drug-resistant cancers., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

The FAM83 ( Fam ily with sequence similarity 83 ) family is highly conserved in vertebrates, but little is known of the functions of these proteins beyond their association with oncogenesis. Of the family, FAM83F is of particular interest because it is the only membrane-targeted FAM83 protein. When overexpressed, FAM83F activates the canonical Wnt signalling pathway and binds to and stabilizes p53; it therefore interacts with two pathways often dysregulated in disease. Insights into gene function can often be gained by studying the roles they play during development, and here we report the generation of fam83f knock-out (KO) zebrafish, which we have used to study the role of Fam83f in vivo . We show that endogenous fam83f is most strongly expressed in the hatching gland of developing zebrafish embryos, and that fam83f KO embryos hatch earlier than their wild-type (WT) counterparts, despite developing at a comparable rate. We also demonstrate that fam83f KO embryos are more sensitive to ionizing radiation than WT embryos-an unexpected finding, bearing in mind the previously reported ability of FAM83F to stabilize p53. Transcriptomic analysis shows that loss of fam83f leads to downregulation of phosphatidylinositol-3-phosphate (PI(3)P) binding proteins and impairment of cellular degradation pathways, particularly autophagy, a crucial component of the DNA damage response. Finally, we show that Fam83f protein is itself targeted to the lysosome when overexpressed in HEK293T cells, and that this localization is dependent upon a C' terminal signal sequence. The zebrafish lines we have generated suggest that Fam83f plays an important role in autophagic/lysosomal processes, resulting in dysregulated hatching and increased sensitivity to genotoxic stress in vivo .

Exogenous administration of the histone deacetylation 4 (HDAC4) protein can effectively delay osteoarthritis (OA) progression. However, HDAC4 is unstable and easily degrades into N-terminal (HDAC4-NT) and C-terminal fragments, and the HDAC4-NT can exert biological effects, but little is known about its role in chondrocytes and cartilage. Thus, the roles of HDAC4-NT fragments (1-289aa, 1-326aa and 1-669aa) in chondrocytes and cartilage were evaluated via real-time cell analysis (RTCA), safranin O staining, Sirius Red staining and nanoindentation. Molecular mechanisms were profiled via whole-transcriptome sequencing (RNA-seq) and verified in vitro and in vivo by a live cell real-time monitoring system, flow cytometry, western blotting and immunohistochemistry. The results showed that 1-669aa induced chondrocyte death and cartilage injury significantly, and the differentially expressed genes (DEGs) were enriched mainly in the apoptotic term and p53 signalling pathway. The validation experiments showed that 1-669aa induced chondrocyte apoptosis via the endoplasmic reticulum stress (ERS) pathway, and up-regulated p53 expression was essential for this process. Thus, we concluded that the HDAC4-NT fragment 1-669aa induces chondrocyte apoptosis via the p53-dependent ERS pathway, suggesting that in addition to overexpressing HDAC4, preventing it from degradation may be a new strategy for the treatment of OA., (© 2024 The Author(s). Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Purpose: Pleuropulmonary blastoma (PPB) is an infrequently encountered childhood malignant intrathoracic neoplasm associated with unfavorable clinical behavior. Since a well-characterized preclinical model is essential for developing competent agents for PPB, we aim to establish and characterize the world's first cell line of PPB, and attempt to perform the cytotoxicity assay on the PPB cell line., Experimental Design: The index case is a 2-year-old female who developed a right thoracic tumor that was surgically removed and treated with multi-agent chemotherapy. The patient is free from recurrence, although it was 9 years after the diagnosis when she developed a thyroid tumor. We performed in vitro cultivation of the isolated neoplastic cells from the tumor, cytogenetic findings and molecular analysis, and tetrazolium colorimetric assay., Result: The histology was consistent with PPB. Serial passage of cultivation produced a continuously growing cell line, KCMC-PPB-1. Conventional cytogenetic analysis of the established cell line revealed complex numerical and structural chromosomal abnormalities, including add(17)(p11). Mutation analysis on the cultured cells revealed amino-acid substitution mutation on exon 4 of TP53 (NM&#95;001276760.3:c.212&#95;213delTG; NP&#95;001263689.1:p.Leu72ArgfsTer37) and compound heterozygous mutations of DICER1 (NM&#95;177438.3:c. 4910C>A; NP&#95;803187.1:Ser1637* and NM&#95;177438.3:c. 5114A>T; NP&#95;803187.1:Glu1705Val). The cultivated cells demonstrated vulnerability to bortezomib on cytotoxicity assay., Conclusion: Our KCMC-PPB-1 is the first genuine, molecularly characterized PPB cell line. The cell line is transplantable to nu/nu mice; therefore, it is suitable for a preclinical model for new drug development. The cytotoxicity assay demonstrated that bortezomib is active in the current PPB model., (© 2024 Wiley Periodicals LLC.)

Introduction: Blastoid variant-mantle cell lymphoma (BV-MCL) represents an aggressive subset of patients with no established standard of care treatment approach., Methods: We performed a retrospective analysis of the clinical characteristics and outcomes of 17 de novo BV-MCL patients treated at our institution between May 2009 and September 2023. In addition, we reviewed the literature supporting 2nd/3rd generation Bruton's Tyrosine-kinase (BTKi)-based therapies for upfront management., Results: The complete response rate to frontline and salvage therapy was 66.7% and 25%, respectively. Two-year overall survival (OS) was low at 62.5% (95% CI, 34.7%-81.1%). Variables associated with higher OS included receipt of consolidative HSCT (p = 0.017), TP53-wild type (p = 0.031), intermediate- versus high-risk Mantle Cell Lymphoma Prognostic Index score (p = 0.026), and achieving complete response with induction therapy (p = 0.027)., Discussion: Treatment outcomes with chemo-immunotherapy in BV-MCL patients are poor, especially among TP53-mutated patients. Recent findings describing positive outcomes with novel BTKi-based therapies are encouraging and should be considered in this high-risk population., (© 2024 The Author(s). Cancer Medicine published by John Wiley & Sons Ltd.)

Haploid larvae in non-mammalian vertebrates are lethal, with characteristic organ growth retardation collectively called 'haploid syndrome'. In contrast to mammals, whose haploid intolerance is attributed to imprinting misregulation, the cellular principle of haploidy-linked defects in non-mammalian vertebrates remains unknown. Here, we investigated cellular defects that disrupt the ontogeny of gynogenetic haploid zebrafish larvae. Unlike diploid control larvae, haploid larvae manifested unscheduled cell death at the organogenesis stage, attributed to haploidy-linked p53 upregulation. Moreover, we found that haploid larvae specifically suffered the gradual aggravation of mitotic spindle monopolarization during 1-3 days post-fertilization, causing spindle assembly checkpoint-mediated mitotic arrest throughout the entire body. High-resolution imaging revealed that this mitotic defect accompanied the haploidy-linked centrosome loss occurring concomitantly with the gradual decrease in larval cell size. Either resolution of mitotic arrest or depletion of p53 partially improved organ growth in haploid larvae. Based on these results, we propose that haploidy-linked mitotic defects and cell death are parts of critical cellular causes shared among vertebrates that limit the larval growth in the haploid state, contributing to an evolutionary constraint on allowable ploidy status in the vertebrate life cycle.

Therapy-related clonal hematopoiesis (t-CH) is defined as clonal hematopoiesis detected in individuals previously treated with chemotherapy and/or radiation therapy. With the increased use of genetic analysis in oncological care, the detection of t-CH among cancer patients is becoming increasingly common. t-CH arises through the selective bottleneck imposed by chemotherapies and potentially through direct mutagenesis from chemotherapies, resulting in a distinct mutational landscape enriched with mutations in DNA damage-response pathway genes such as TP53, PPM1D, and CHEK2. Emerging evidence sheds light on the mechanisms of t-CH development and potential strategies to mitigate its emergence. Due to its unique characteristics that predominantly affect cancer patients, t-CH has clinical implications distinct from those of CH in the general population. This Review discusses the potential mechanisms of t-CH development, its mutational landscape, mutant-drug relationships, and its clinical significance. We highlight the distinct nature of t-CH and call for intensified research in this field.