PTEN deficiency induces an extrahepatic cholangitis-cholangiocarcinoma continuum via aurora kinase A in mice.

BACKGROUND & AIMS: The PTEN-AKT pathway is frequently altered in extrahepatic cholangiocarcinoma (eCCA). We aimed to evaluate the role of PTEN in the pathogenesis of eCCA and identify novel therapeutic targets for this disease. METHODS: The Pten gene was genetically deleted using the Cre-loxp system in biliary epithelial cells. The pathologies were evaluated both macroscopically and histologically. The characteristics were further analyzed by immunohistochemistry, reverse-transcription PCR, cell culture, and RNA sequencing. Some features were compared to those in human eCCA samples. Further mechanistic studies utilized the conditional knockout of Trp53 and Aurora kinase A (Aurka) genes. We also tested the effectiveness of an Aurka inhibitor. RESULTS: We observed that genetic deletion of the Pten gene in the extrahepatic biliary epithelium and peri-ductal glands initiated sclerosing cholangitis-like lesions in mice, resulting in enlarged and distorted extrahepatic bile ducts in mice as early as 1 month after birth. Histologically, these lesions exhibited increased epithelial proliferation, inflammatory cell infiltration, and fibrosis. With aging, the lesions progressed from low-grade dysplasia to invasive carcinoma. Trp53 inactivation further accelerated disease progression, potentially by downregulating senescence. Further mechanistic studies showed that both human and mouse eCCA showed high expression of AURKA. Notably, the genetic deletion of Aurka completely eliminated Pten deficiency-induced extrahepatic bile duct lesions. Furthermore, pharmacological inhibition of Aurka alleviated disease progression. CONCLUSIONS: Pten deficiency in extrahepatic cholangiocytes and peribiliary glands led to a cholangitis-to-cholangiocarcinoma continuum that was dependent on Aurka. These findings offer new insights into preventive and therapeutic interventions for extrahepatic CCA. IMPACT AND IMPLICATIONS: The aberrant PTEN-PI3K-AKT signaling pathway is commonly observed in human extrahepatic cholangiocarcinoma (eCCA), a disease with a poor prognosis. In our study, we developed a mouse model mimicking cholangitis to eCCA progression by conditionally deleting the Pten gene via Pdx1-Cre in epithelial cells and peribiliary glands of the extrahepatic biliary duct. The conditional Pten deletion in these cells led to cholangitis, which gradually advanced to dysplasia, ultimately resulting in eCCA. The loss of Pten heightened Akt signaling, cell proliferation, inflammation, fibrosis, DNA damage, epigenetic signaling, epithelial-mesenchymal transition, cell dysplasia, and cellular senescence. Genetic deletion or pharmacological inhibition of Aurka successfully halted disease progression. This model will be valuable for testing novel therapies and unraveling the mechanisms of eCCA tumorigenesis.

A Tale of Two Checkpoints: ATR Inhibition and PD-(L)1 Blockade.

Innate immunity and the DNA damage response (DDR) pathway are inextricably linked. Within the DDR, ataxia telangiectasia and Rad3-related (ATR) is a key kinase responsible for sensing replication stress and facilitating DNA repair through checkpoint activation, cell cycle arrest, and promotion of fork recovery. Recent studies have shed light on the immunomodulatory role of the ATR-CHK1 pathway in the tumor microenvironment and the specific effects of ATR inhibition in stimulating an innate immune response. With several potent and selective ATR inhibitors in developmental pipelines, the combination of dual ATR and PD-(L)1 blockade has attracted increasing interest in cancer therapy. In this review, we summarize the clinical and preclinical data supporting the combined inhibition of ATR and PD-(L)1, discuss the potential challenges surrounding this approach, and highlight biomarkers relevant for selected patients who are most likely to benefit from the blockade of these two checkpoints.

Dna2 nuclease deficiency results in large and complex DNA insertions at chromosomal breaks.

Insertions of mobile elements1-4, mitochondrial DNA5 and fragments of nuclear chromosomes6 at DNA double-strand breaks (DSBs) threaten genome integrity and are common in cancer7-9. Insertions of chromosome fragments at V(D)J recombination loci can stimulate antibody diversification10. The origin of insertions of chromosomal fragments and the mechanisms that prevent such insertions remain unknown. Here we reveal a yeast mutant, lacking evolutionarily conserved Dna2 nuclease, that shows frequent insertions of sequences between approximately 0.1 and 1.5 kb in length into DSBs, with many insertions involving multiple joined DNA fragments. Sequencing of around 500 DNA inserts reveals that they originate from Ty retrotransposons (8%), ribosomal DNA (rDNA) (15%) and from throughout the genome, with preference for fragile regions such as origins of replication, R-loops, centromeres, telomeres or replication fork barriers. Inserted fragments are not lost from their original loci and therefore represent duplications. These duplications depend on nonhomologous end-joining (NHEJ) and Pol4. We propose a model in which alternative processing of DNA structures arising in Dna2-deficient cells can result in the release of DNA fragments and their capture at DSBs. Similar DNA insertions at DSBs are expected to occur in any cells with linear extrachromosomal DNA fragments.

Percutaneous External Fixation of Redistributed Orbital Fat in Transcutaneous Lower Eyelid Blepharoplasty.

BACKGROUND: While percutaneous external fixation has gained popularity in transconjunctival blepharoplasty due to its simplicity and minimal incision, internal fixation remains the standard approach in transcutaneous blepharoplasty. OBJECTIVES: This study aims to assess the safety and effectiveness of percutaneous external fixation specifically in transcutaneous fat repositioning blepharoplasty. METHODS: From May 2022 to December 2023, a consecutive cohort of 97 patients underwent this surgical technique. Building upon the principles of transconjunctival fat repositioning, the procedure involved a minimal incision targeting the muscle layer, a conservative release of the tear trough ligament, and the repositioning of fat secured with percutaneous external fixation. Surgical outcomes were evaluated using the Barton grading system and FACE-Q scales for patient-reported outcomes. RESULTS: Following a mean follow-up of 7.2 months (ranging from 6 to 17 months), a resolution of tear trough deformities and eyelid bags was achieved in over 92% of cases. Notably, significant improvements (P < 0.05) were observed in lower eyelid aesthetics as measured by the FACE-Q scale, with a concomitant enhancement in patients' social confidence (P < 0.05). Overall, patients expressed satisfaction with their decision to undergo the procedure (71.0±19.1). Complications necessitating revision surgery included isolated cases of hematoma evacuation, fat injection, excision of excess fat, and skin excision. CONCLUSIONS: External fixation in transcutaneous blepharoplasty is technically less complex and minimally invasive, effectively securing transposed fat pedicles safely, presenting a viable alternative to conventional fixation techniques.

Clinical efficacy of blood derivatives on wound healing: A systematic review and network meta-analysis.

This study aims to evaluate the clinical effects of different blood derivatives on wound healing using network meta-analysis. PubMed, Embase, OVID, Web of Science, SCOPUS and Cochrane Central were searched to obtain studies about blood derivatives on wound healing until October 2023. R 4.2.0 and Stata 15.0 softwares were used for data analysis. Forty-four studies comprising 5164 patients were included. The results of network meta-analysis showed that the healing area from high to low was GF + ORCCB, ORCCB, GF, PRF, Unnas paste dressing, APG, PRP injection, PRP, PRP + thrombin gel, PPP, HPL, CT. The healing time from low to high was PRP + thrombin gel, GF, PRP, PC + K, PC, APG, PRF, CT, Silver sulfadiazine ointment. The number of patients cured from high to low was APG, PRP injection, PRP, Aurix, PRF, Leucopatch, HPL, Antimicrobial Ointment Dressing, CT, 60 µg/cm2 repifermin, 120 µg/cm2 repifermin, AFG, PPP. The order of analgesic effect from high to low was AFG, Aminogam gel, PRF, PRP, Oxidised oil, APG, GF, CT. The order of the number of wound infection cases from low to high is APG, 20 µg/cm2 repifermin, 60 µg/cm2 repifermin, PRP, LeucoPatch, CT, PPP, Antiseptic ointment dressing. Healing area: GF + ORCCB had the best effect; Healing time: PRP + thrombin gel took the shortest time. The number of cured patients and the reduction of wound infection: APG has the best effect. Analgesic effect: AFG has the best effect. More studies with large sample sizes are needed to confirm the above findings.

[Network Meta-analysis of Chinese patent medicines in treatment of chronic pulmonary heart disease].

Network Meta-analysis was performed to compare the efficacy and safety of Chinese patent medicines in treating chronic pulmonary heart disease. CNKI, VIP, Wanfang, SinoMed, PubMed, Web of Science, EMbase, and Cochrane Library were searched for randomized controlled trial(RCT) of treating chronic pulmonary heart disease with Chinese patent medicines with the time interval from inception to December 2023. The Cochrane risk-of-bias tool was used for quality assessment of the included articles. RevMan 5.4 and Stata 17.0 were employed to establish the risk of bias map and perform the network Meta-analysis, respectively. Ultimately, a total of 95 RCTs involving 8 787 cases and 11 different Chinese patent medicines were included. Network Meta-analysis yielded the following results based on the surface under the cumulative ranking curve(SUCRA).(1)In terms of cardiac function improves clinical total effective rate, SUCRA the top three were Wenxin Granules + conventional western medicine, Tongxinluo Capsules + conventional western medicine, and Qishen Yiqi Dropping Pills + conventional western medicine.(2)For improving forced expiratory volume in the first se-cond(FEV1), SUCRA the top three were Danting Feixin Granules + conventional western medicine, Tongxinluo Capsules + conventional western medicine, and Bufei Huoxue Capsules + conventional western medicine.(3)Regarding increasing the FEV1/forced vital capacity(FVC%) value, SUCRA the top three were Qili Qiangxin Capsules + conventional western medicine, Shexiang Baoxin Pills + conventional western medicine, and Qishen Yiqi Dropping Pills + conventional western medicine.(4)In terms of increasing the partial pressure of oxygen(PaO_2), SUCRA the top three were Qili Qiangxin Capsules + conventional western medicine, Qishen Yiqi Dropping Pills + conventional western medicine, and Shexiang Baoxin Pills + conventional western medicine.(5)In terms of reducing the partial pressure of carbon dioxide(PaCO_2), SUCRA the top three were Tongxinluo Capsules + conventional western medicine, Qishen Yiqi Dropping Pills + conventional western medicine, and Shexiang Baoxin Pills + conventional western medicine.(6)In terms of increasing left ventricular ejection fraction(LVEF), SUCRA the top three were Bufei Huoxue Capsules + conventional western medicine, Qishen Yiqi Dropping Pills + conventional western medicine, and Shexiang Baoxin Pills + conventional western medicine.(7)In terms of decreasing brain natriu-retic peptide(BNP), SUCRA the top three were Compound Danshen Dropping Pills + conventional western medicine, Qili Qiangxin Capsules + conventional western medicine, and Tongxinluo Capsules + conventional western medicine.(8)In terms of improving the hematocrit level, SUCRA the top three were Qishen Yiqi Dropping Pills + conventional western medicine, Compound Danshen Dropping Pills + conventional western medicine, and Tongxinluo Capsules + conventional western medicine. In terms of safety, 26 RCTs reported adverse reactions, which primarily involved the circulatory and digestive systems. The combination of Chinese patent medicines with conventional western medicine has demonstrated enhanced therapeutic effects on chronic pulmonary heart disease. However, due to the varying quality and sample sizes of included studies and the absence of direct comparisons between Chinese patent medicines, the conclusions should be further validated by multicenter studies with larger sample sizes and higher methodological rigor.

[Network Meta-analysis of Chinese patent medicines in treatment of coronary heart disease complicated with heart failure].

The efficacy and safety of different Chinese patent medicines in the treatment of coronary heart disease complicated with heart failure were evaluated by network Meta-analysis. The randomized controlled trial(RCT) of Chinese patent medicines for coronary heart disease complicated with heart failure was retrieved from CNKI, Wanfang, VIP, SinoMed, PubMed, Web of Science, EMbase, and Cochrane Library with the time interval from inception to July 5, 2023. The quality of the included RCT was evaluated by the Cochrane's risk of bias assessment tool, and a network Meta-analysis was performed in Stata 16.0. Finally, a total of 82 RCTs were included, involving 9 298 patients and 11 Chinese patent medicines. Network Meta-analysis yielded the following results based on the surface under the cumulative ranking curve(SUCRA).(1)In terms of improving the clinical response rate, the top three interventions were Qishen Yiqi Dripping Pills + conventional western medicine, Zhenyuan Capsules + conventional western medicine, and Tongxinluo Capsules + conventional western medicine.(2) In terms of increasing left ventricular ejection fraction(LVEF), the top three interventions were Shexiang Baoxin Pills + conventional western medicine, Compound Danshen Dripping Pills + conventional western medicine, and Tongxinluo Capsules + conventional western medicine.(3) In terms of reducing left ventricular end-diastolic diameter(LVEDD), the top three interventions were Shexiang Tongxin Dripping Pills + conventional western medicine, Tongxinluo Capsules + conventional western medicine, and Shexiang Baoxin Pills + conventional western medicine.(4) In terms of reducing N-terminal pro-brain natriuretic peptide(NT-proBNP), the top three interventions were Shexiang Baoxin Pills + conventional western medicine, Qi-shen Yiqi Dripping Pills + conventional western medicine, and Compound Danshen Dripping Pills + conventional western medicine.(5) In terms of reducing hyper-sensitive C-reactive protein(hs-CRP), the top three interventions were Naoxintong Capsules + conventional western medicine, Shexiang Baoxin Pills + conventional western medicine, and Compound Danshen Dripping Pills + conventional western medicine.(6) In terms of increasing the distance of the six-minute walking trail(6MWT), the top three interventions were Zhen-yuan Capsules + conventional western medicine, Qili Qiangxin Capsules + conventional western medicine, and Qishen Yiqi Dripping Pills + conventional western medicine. The results showed that Chinese patent medicines combined with conventional western medicine can effectively improve the clinical response rate, LVEF, and 6MWT and reduce LVEDD, NT-proBNP, and hs-CRP. However, due to the overall low quality of the articles included and the few articles of some Chinese patent medicines, direct comparison between diffe-rent Chinese patent medicines remains to be carried out and the results need to be further verified.

Epithelial SOX9 drives progression and metastases of gastric adenocarcinoma by promoting immunosuppressive tumour microenvironment.

OBJECTIVE: Many cancers engage embryonic genes for rapid growth and evading the immune system. SOX9 has been upregulated in many tumours, yet the role of SOX9 in mediating immunosuppressive tumour microenvironment is unclear. Here, we aim to dissect the role of SOX9-mediated cancer stemness attributes and immunosuppressive microenvironment in advanced gastric adenocarcinoma (GAC) for novel therapeutic discoveries. METHODS: Bulk RNAseq/scRNA-seq, patient-derived cells/models and extensive functional studies were used to identify the expression and functions of SOX9 and its target genes in vitro and in vivo. Immune responses were studied in PBMCs or CD45+ immune cells cocultured with tumour cells with SOX9high or knockout and the KP-Luc2 syngeneic models were used for efficacy of combinations. RESULTS: SOX9 is one of the most upregulated SOX genes in GAC and highly expressed in primary and metastatic tissues and associated with poor prognosis. Depletion of SOX9 in patient-derived GAC cells significantly decreased cancer stemness attributes, tumour formation and metastases and consistently increased CD8+ T cell responses when cocultured with PBMCs/CD45+ cells from GAC patients. RNA sequencing identified the leukaemia inhibitory factor (LIF) as the top secreted molecule regulated by SOX9 in tumour cells and was enriched in malignant ascites and mediated SOX9-induced M2 macrophage repolarisation and inhibited T cell function. CONCLUSION: Epithelial SOX9 is critical in suppressing CD8+ T cell responses and modified macrophage function in GAC through the paracrine LIF factor. Cotargeting LIF/LIFR and CSF1R has great potential in targeting SOX9-mediated cancer stemness, T cell immunosuppression and metastases suggesting the novel combination therapy against advanced GAC.

Phosphorylation-Dependent Enhancement of Rad53 Kinase Activity through the INO80 Chromatin Remodeling Complex.

ATP-dependent chromatin remodeling complexes such as INO80 have been implicated in checkpoint regulation in response to DNA damage. However, how chromatin remodeling complexes regulate DNA damage checkpoints remain unclear. Here, we identified a mechanism of regulating checkpoint effector kinase Rad53 through a direct interaction with the INO80 chromatin remodeling complex. Rad53 is a key checkpoint kinase downstream of Mec1. Mec1/Tel1 phosphorylates the Ies4 subunit of the INO80 complex in response to DNA damage. We find that the phosphorylated Ies4 binds to the N-terminal FHA domain of Rad53. In vitro, INO80 can activate Rad53 kinase activity in an Ies4-phosphorylation-dependent manner in the absence of known activators such as Rad9. In vivo, Ies4 and Rad9 function synergistically to activate Rad53. These findings establish a direct connection between ATP-dependent chromatin remodeling complexes and checkpoint regulation.

Regulation of Mec1 kinase activity by the SWI/SNF chromatin remodeling complex.

ATP-dependent chromatin remodeling complexes alter chromatin structure through interactions with chromatin substrates such as DNA, histones, and nucleosomes. However, whether chromatin remodeling complexes have the ability to regulate nonchromatin substrates remains unclear. Saccharomyces cerevisiae checkpoint kinase Mec1 (ATR in mammals) is an essential master regulator of genomic integrity. Here we found that the SWI/SNF chromatin remodeling complex is capable of regulating Mec1 kinase activity. In vivo, Mec1 activity is reduced by the deletion of Snf2, the core ATPase subunit of the SWI/SNF complex. SWI/SNF interacts with Mec1, and cross-linking studies revealed that the Snf2 ATPase is the main interaction partner for Mec1. In vitro, SWI/SNF can activate Mec1 kinase activity in the absence of chromatin or known activators such as Dpb11. The subunit requirement of SWI/SNF-mediated Mec1 regulation differs from that of SWI/SNF-mediated chromatin remodeling. Functionally, SWI/SNF-mediated Mec1 regulation specifically occurs in S phase of the cell cycle. Together, these findings identify a novel regulator of Mec1 kinase activity and suggest that ATP-dependent chromatin remodeling complexes can regulate nonchromatin substrates such as a checkpoint kinase.

Intense d-p Hybridization in Nb3 O15 Tripolymer Induced the Largest Second Harmonic Generation Response and Birefringence in Germanates.

We herein report two asymmetric germanate crystals, KNbGe3 O9 and K3 Nb3 Ge2 O13 , with different structures and optical properties derived from divergent polymerized forms of GeO4 and NbO6 groups. Remarkably, K3 Nb3 Ge2 O13 achieved a rare combination of the strongest second harmonic generation (SHG) response of 17.5×KDP @ 1064 nm and the largest birefringence of 0.196 @ 546 nm in germanates. It features unique [Nb3 O12 ]∞ tubular chains constructed by circular Nb3 O15 tripolymers. Theoretical calculations reveal that the d-p interactions in the Nb3 O15 group are responsible for outstanding optical properties. This work emphasizes the significance of the polymerizable functional units in obtaining high-performance nonlinear optical (NLO) crystals.

Loss of ARID1A activates mTOR signaling and SOX9 in gastric adenocarcinoma-rationale for targeting ARID1A deficiency.

BACKGROUND: Gastric adenocarcinoma (GAC) is a lethal disease with limited therapeutic options. Genetic alterations in chromatin remodelling gene AT-rich interactive domain 1A (ARID1A) and mTOR pathway activation occur frequently in GAC. Targeting the mechanistic target of rapamycin (mTOR) pathway in unselected patients has failed to show survival benefit. A deeper understanding of GAC might identify a subset that can benefit from mTOR inhibition. METHODS: Genomic alterations in ARID1A were analysed in GAC. Mouse gastric epithelial cells from CK19-Cre-Arid1Afl/fl and wild-type mice were used to determine the activation of oncogenic genes due to loss of Arid1A. Functional studies were performed to determine the significance of loss of ARID1A and the sensitivity of ARID1A-deficient cancer cells to mTOR inhibition in GAC. RESULTS: More than 30% of GAC cases had alterations (mutations or deletions) of ARID1A and ARID1A expression was negatively associated with phosphorylation of S6 and SOX9 in GAC tissues and patient-derived xenografts (PDXs). Activation of mTOR signalling (increased pS6) and SOX9 nuclear expression were strongly increased in Arid1A-/- mouse gastric tissues which could be curtailed by RAD001, an mTOR inhibitor. Knockdown of ARID1A in GAC cell lines increased pS6 and nuclear SOX9 and increased sensitivity to an mTOR inhibitor which was further amplified by its combination with fluorouracil both in vitro and in vivo in PDXs. CONCLUSIONS: The loss of ARID1A activates pS6 and SOX9 in GAC, which can be effectively targeted by an mTOR inhibitor. Therefore, our studies suggest a new therapeutic strategy of clinically targeting the mTOR pathway in patients with GAC with ARID1A deficiency.

Enantio- and diastereoselective boron conjugate addition to α-alkyl α,ß-unsaturated esters.

We developed a copper-catalyzed enantio- and diastereoselective boron conjugate addition to α-alkyl α,ß-unsaturated esters under base-free conditions. The approach showed excellent enantioselectivities (87-99% ee) and moderate to good conversions (51-99%), albeit with moderate diastereoselectivities (1 : 1-17 : 1 dr). The synthetic utility of this protocol was demonstrated.

[Medication rule of traditional Chinese medicine for heart failure after myocardial infarction: based on data mining].

This study aims to explore the medication rule of traditional Chinese medicine(TCM) for heart failure after myocardial infarction via data mining. To be specific, articles on the treatment of the disease with Chinese medicine were retrieved from CNKI, Wanfang, VIP, and SinoMed and related information was collected. A database was created with Microsoft Excel 2019, and SPSS Clementine 12.0 and IBM SPSS Statistics 23.0 were applied for association rules analysis, cluster analysis, and factor analysis. Finally, a total of 81 TCM prescriptions were screened out, involving 91 medicinals with cumulative use frequency of 740. The main syndromes were Qi deficiency and blood stasis, Yang Qi deficiency and blood stasis together with retained morbid fluid, deficiency of both Qi and Yin and blood stasis. The medicinals with high-frequency were Astragali Radix, Salviae Miltiorrhizae Radix et Rhizoma, Ginseng Radix et Rhizoma, Poria, and Aconiti Lateralis Radix Praeparata. The effects of the medicinals were tonifying deficiency, activating blood and resolving stasis, and promoting urination and draining dampness. The association rules analysis yielded "Astragali Radix-Salviae Miltiorrhizae Radix et Rhizoma" "Astragali Radix-Aconiti Lateralis Radix Praeparata-Salviae Miltiorrhizae Radix et Rhizoma" "Aconiti Lateralis Radix Praeparata-Ginseng Radix et Rhizoma-Salviae Miltiorrhizae Radix et Rhizoma-Astragali Radix" combinations. Cluster analysis yielded 6 basic formulas for heart failure after myocardial infarction. Factor analysis extracted a total of 8 common factors. Heart failure after myocardial infarction is characterized by the syndrome of deficiency in nature and excess in superficiality. The core pathogenesis is "deficiency" "stasis" "retained morbid fluid", particularly "deficiency". This disease is closely related to the heart, lung, and spleen. The basic treatment principle is replenishing Qi and activating blood, and warming Yang, excreting water, and nourishing yin should also be emphasized. The common basic prescriptions, such as Siwu Decoction, Shengmai Powder, Xuefu Zhuyu Decoction, Linggui Zhugan Decoction, and Shenfu Decoction, have been discovered. This study provided data for clinical medication and drug development for heart failure after myocardial infarction.

YAP1 mediates gastric adenocarcinoma peritoneal metastases that are attenuated by YAP1 inhibition.

OBJECTIVE: Peritoneal carcinomatosis (PC; malignant ascites or implants) occurs in approximately 45% of advanced gastric adenocarcinoma (GAC) patients and associated with a poor survival. The molecular events leading to PC are unknown. The yes-associated protein 1 (YAP1) oncogene has emerged in many tumour types, but its clinical significance in PC is unclear. Here, we investigated the role of YAP1 in PC and its potential as a therapeutic target. METHODS: Patient-derived PC cells, patient-derived xenograft (PDX) and patient-derived orthotopic (PDO) models were used to study the function of YAP1 in vitro and in vivo. Immunofluorescence and immunohistochemical staining, RNA sequencing (RNA-Seq) and single-cell RNA-Seq (sc-RNA-Seq) were used to elucidate the expression of YAP1 and PC cell heterogeneity. LentiCRISPR/Cas9 knockout of YAP1 and a YAP1 inhibitor were used to dissect its role in PC metastases. RESULTS: YAP1 was highly upregulated in PC tumour cells, conferred cancer stem cell (CSC) properties and appeared to be a metastatic driver. Dual staining of YAP1/EpCAM and sc-RNA-Seq revealed that PC tumour cells were highly heterogeneous, YAP1high PC cells had CSC-like properties and easily formed PDX/PDO tumours but also formed PC in mice, while genetic knockout YAP1 significantly slowed tumour growth and eliminated PC in PDO model. Additionally, pharmacologic inhibition of YAP1 specifically reduced CSC-like properties and suppressed tumour growth in YAP1high PC cells especially in combination with cytotoxics in vivo PDX model. CONCLUSIONS: YAP1 is essential for PC that is attenuated by YAP1 inhibition. Our data provide a strong rationale to target YAP1 in clinic for GAC patients with PC.

Integrated genomic profiling and modelling for risk stratification in patients with advanced oesophagogastric adenocarcinoma.

OBJECTIVE: Prognosis of patients with advanced oesophagogastric adenocarcinoma (mEGAC) is poor and molecular determinants of shorter or longer overall survivors are lacking. Our objective was to identify molecular features and develop a prognostic model by profiling the genomic features of patients with mEGAC with widely varying outcomes. DESIGN: We profiled 40 untreated mEGACs (20 shorter survivors <13 months and 20 longer survivors >36 months) with whole-exome sequencing (WES) and RNA sequencing and performed an integrated analysis of exome, transcriptome, immune profile and pathological phenotypes to identify the molecular determinants, developing an integrated model for prognosis and comparison with The Cancer Genome Atlas (TCGA) cohorts. RESULTS: KMT2C alterations were exclusively observed in shorter survivors together with high level of intratumour heterogeneity and complex clonal architectures, whereas the APOBEC mutational signatures were significantly enriched in longer survivors. Notably, the loss of heterozygosity in chromosome 4 (Chr4) was associated with shorter survival and 'cold' immune phenotype characterised by decreased B, CD8, natural killer cells and interferon-gamma responses. Unsupervised transcriptomic clustering revealed a shorter survivor subtype with distinct expression features (eg, upregulated druggable targets JAK2, MAP3K13 and MECOM). An integrated model was then built based on clinical variables and the identified molecular determinants, which significantly segregated shorter and longer survivors. All the above features and the integrated model have been validated independently in multiple TCGA cohorts. CONCLUSION: This study discovered novel molecular features prognosticating overall survival in patients with mEGAC and identified potential novel targets in shorter survivors.

Combining UBR5 and CD163+ tumor-associated macrophages better predicts prognosis of clear cell renal cell carcinoma patients.

PURPOSE: Identification of reliable postoperative indicators for accurately evaluating prognosis of clear cell renal cell carcinoma (ccRCC) patients remains an important clinical issue. This study determined the prognostic value of UBR5 expression in ccRCC patients by combining with CD163+ tumor-associated macrophages (TAMs) and the established clinical parameters. METHODS: The expression of UBR5 was analyzed in ccRCC patients from TCGA databases. A total of 310 ccRCC patients were randomly divided into the training and validation cohorts at a 3:2 or 1:1 ratio, and immunohistochemistry (IHC) and statistical analyses were performed to examine the prognostic value of UBR5 and CD163+ TAMs. RESULTS: UBR5 expression was commonly downregulated in human ccRCC specimens, which was associated with TNM stage, SSIGN, WHO/ISUP Grading and poor prognosis of ccRCC patients. In addition, UBR5 expression was negatively correlated with CD163 expression (a TAM marker) in ccRCC tissues, and combining expressions of UBR5 and CD163 better predicted worse overall survival and progression-free survival of ccRCC patients. Even after multivariable adjustment, UBR5, CD163, TNM stage and SSIGN appeared to be independent risk factors. By time-dependent c-index analysis, the integration of intratumoral UBR5 and CD163 achieved higher c-index value than UBR5, CD163, TNM stage or SSIGN alone in predicting ccRCC patients' prognosis. Moreover, the incorporation of both UBR5 and CD163 into the clinical indicators TNM stage or SSIGN exhibited highest c-index value. CONCLUSIONS: Integrating intratumoral UBR5 and CD163+ TAMs with the current clinical parameters achieves better accuracy in predicting ccRCC patients' postoperative prognosis.

Integrating HECW1 expression into the clinical indicators exhibits high accuracy in assessing the prognosis of patients with clear cell renal cell carcinoma.

BACKGROUND: Although many intratumoral biomarkers have been reported to predict clear cell renal cell carcinoma (ccRCC) patient prognosis, combining intratumoral and clinical indicators could predict ccRCC prognosis more accurately than any of these markers alone. This study mainly examined the prognostic value of HECT, C2 and WW domain-containing E3 ubiquitin protein ligase 1 (HECW1) expression in ccRCC patients in combination with established clinical indicators. METHODS: The expression level of HECW1 was screened out by data-independent acquisition mass spectrometry (DIA-MS) and analyzed in ccRCC patients from the The Cancer Genome Atlas (TCGA) database and our cohort. A total of 300 ccRCC patients were stochastically divided into a training cohort and a validation cohort, and real-time PCR, immunohistochemistry (IHC) and statistical analyses were employed to examine the prognostic value of HECW1 in ccRCC patients. RESULTS: The expression level of HECW1 usually decreased in human ccRCC specimens relative to control specimens in TCGA (p < 0.001). DIA-MS, Real-time PCR, and IHC analyses also showed that the majority of ccRCCs harbored decreased HECW1 expression compared with that in normal adjacent tissues (p < 0.001). Additionally, HECW1 expression was reduced in ccRCC cell lines compared with the normal renal cell line HK-2 (p < 0.001). Moreover, lower HECW1 expression was found in ccRCC patients with a higher tumor node metastasis (TNM) stage, bone metastasis, or first-line targeted drug resistance (p < 0.001). Low HECW1 expression indicated higher TNM stage, SSIGN (Stage, Size, Grade, and Necrosis) score and WHO/ISUP grade and poor prognosis in ccRCC patients (p < 0.05). Even after multivariable adjustment, HECW1, TNM stage, and SSIGN score served as independent risk factors. The c-index analysis showed that integrating intratumoral HECW1 expression into TNM stage or SSIGN score resulted in a higher c-index value than these indicators alone for predicting ccRCC patient prognosis. CONCLUSION: HECW1 is a novel prognostic biomarker and therapeutic target in ccRCC, and integrating intratumoral HECW1 expression with established clinical indicators yields higher accuracy in assessing the postoperative prognosis of ccRCC patients.

ITCH nuclear translocation and H1.2 polyubiquitination negatively regulate the DNA damage response.

The downregulation of the DNA damage response (DDR) enables aggressive tumors to achieve uncontrolled proliferation against replication stress, but the mechanisms underlying this process in tumors are relatively complex. Here, we demonstrate a mechanism through which a distinct E3 ubiquitin ligase, ITCH, modulates DDR machinery in triple-negative breast cancer (TNBC). We found that expression of a nuclear form of ITCH was significantly increased in human TNBC cell lines and tumor specimens. Phosphorylation of ITCH at Ser257 by AKT led to the nuclear localization of ITCH and ubiquitination of H1.2. The ITCH-mediated polyubiquitination of H1.2 suppressed RNF8/RNF168-dependent formation of 53BP1 foci, which plays important roles in DDR. Consistent with these findings, impaired ITCH nuclear translocation and H1.2 polyubiquitination sensitized cells to replication stress and limited cell growth and migration. AKT activation of ITCH-H1.2 axis may confer TNBC cells with a DDR repression to counteract the replication stress and increase cancer cell survivorship and growth potential.

Regulatory Mechanisms of Retinal Photoreceptors Development at Single Cell Resolution.

Photoreceptors are critical components of the retina and play a role in the first step of the conversion of light to electric signals. With the discovery of the intrinsically photosensitive retinal ganglion cells, which regulate non-image-forming visual processes, our knowledge of the photosensitive cell family in the retina has deepened. Photoreceptor development is regulated by specific genes and proteins and involves a series of molecular processes including DNA transcription, post-transcriptional modification, protein translation, and post-translational modification. Single-cell sequencing is a promising technology for the study of photoreceptor development. This review presents an overview of the types of human photoreceptors, summarizes recent discoveries in the regulatory mechanisms underlying their development at single-cell resolution, and outlines the prospects in this field.