Clonal dominance defines metastatic dissemination in pancreatic cancer.

Tumors represent ecosystems where subclones compete during tumor growth. While extensively investigated, a comprehensive picture of the interplay of clonal lineages during dissemination is still lacking. Using patient-derived pancreatic cancer cells, we created orthotopically implanted clonal replica tumors to trace clonal dynamics of unperturbed tumor expansion and dissemination. This model revealed the multifaceted nature of tumor growth, with rapid changes in clonal fitness leading to continuous reshuffling of tumor architecture and alternating clonal dominance as a distinct feature of cancer growth. Regarding dissemination, a large fraction of tumor lineages could be found at secondary sites each having distinctive organ growth patterns as well as numerous undescribed behaviors such as abortive colonization. Paired analysis of primary and secondary sites revealed fitness as major contributor to dissemination. From the analysis of pro- and nonmetastatic isogenic subclones, we identified a transcriptomic signature able to identify metastatic cells in human tumors and predict patients' survival.

Trans-lesion synthesis and mismatch repair pathway crosstalk defines chemoresistance and hypermutation mechanisms in glioblastoma.

Almost all Glioblastoma (GBM) are either intrinsically resistant to the chemotherapeutical drug temozolomide (TMZ) or acquire therapy-induced mutations that cause chemoresistance and recurrence. The genome maintenance mechanisms responsible for GBM chemoresistance and hypermutation are unknown. We show that the E3 ubiquitin ligase RAD18 (a proximal regulator of TLS) is activated in a Mismatch repair (MMR)-dependent manner in TMZ-treated GBM cells, promoting post-replicative gap-filling and survival. An unbiased CRISPR screen provides an aerial map of RAD18-interacting DNA damage response (DDR) pathways deployed by GBM to tolerate TMZ genotoxicity. Analysis of mutation signatures from TMZ-treated GBM reveals a role for RAD18 in error-free bypass of O6mG (the most toxic TMZ-induced lesion), and error-prone bypass of other TMZ-induced lesions. Our analyses of recurrent GBM patient samples establishes a correlation between low RAD18 expression and hypermutation. Taken together we define molecular underpinnings for the hallmark tumorigenic phenotypes of TMZ-treated GBM.

Hydrophilic Peptide and Glycopeptide as Immobilized Sorbents for Glycosylation Analysis.

Hydrophilic interaction liquid chromatography (HILIC) is widely used for glycopeptide enrichment in shot-gun glycoproteomics to enhance the glycopeptide signal and minimize the ionization competition of peptides. In this work, we have developed a novel hydrophilic material (glycoHILIC) based on glycopeptides and peptides to provide hydrophilic properties. GlycoHILIC was synthesized by oxidizing cotton and then reacting the resulting aldehyde with the N-terminus of the glycopeptide or peptide by reductive amination. Due to the large amount of hydrophilic carbohydrates and hydrophilic amino acids contained in glycopeptides, glycoHILIC showed significantly better enrichment of glycopeptides than cotton itself. Our results demonstrate that glycoHILIC has high selectivity, a low detection limit, and good stability. Over 257 unique N-linked glycosylation sites in 1477 intact N-glycopeptides from 146 glycoproteins were identified from 1 µL of human serum using glycoHILIC. Serum analysis of pancreatic cancer patients found that 38 N-glycopeptides among 21 glycoproteins changed significantly, of which 7 N-glycopeptides increased and 31 N-glycopeptides decreased. These results demonstrate that glycoHILIC can be used for glycopeptide enrichment and analysis.

Mass spectrometry analysis of intact protein N-glycosylation signatures of cells and sera in pancreatic adenocarcinomas. / èƒ°è ºç™Œç»†èƒžå’Œè¡€æ¸ å®Œæ•´è›‹ç™½è´¨N-糖基化特征的质谱分析.

Pancreatic cancer is among the most malignant cancers, and thus early intervention is the key to better survival outcomes. However, no methods have been derived that can reliably identify early precursors of development into malignancy. Therefore, it is urgent to discover early molecular changes during pancreatic tumorigenesis. As aberrant glycosylation is closely associated with cancer progression, numerous efforts have been made to mine glycosylation changes as biomarkers for diagnosis; however, detailed glycoproteomic information, especially site-specific N-glycosylation changes in pancreatic cancer with and without drug treatment, needs to be further explored. Herein, we used comprehensive solid-phase chemoenzymatic glycoproteomics to analyze glycans, glycosites, and intact glycopeptides in pancreatic cancer cells and patient sera. The profiling of N-glycans in cancer cells revealed an increase in the secreted glycoproteins from the primary tumor of MIA PaCa-2 cells, whereas human sera, which contain many secreted glycoproteins, had significant changes of glycans at their specific glycosites. These results indicated the potential role for tumor-specific glycosylation as disease biomarkers. We also found that AMG-510, a small molecule inhibitor against Kirsten rat sarcoma viral oncogene homolog (KRAS) G12C mutation, profoundly reduced the glycosylation level in MIA PaCa-2 cells, suggesting that KRAS plays a role in the cellular glycosylation process, and thus glycosylation inhibition contributes to the anti-tumor effect of AMG-510.

Causal effects of osteoporosis on structural changes in specific brain regions: a Mendelian randomization study.

Observational studies have reported that osteoporosis is associated with cortical changes in the brain. However, the inherent limitations of observational studies pose challenges in eliminating confounding factors and establishing causal relationships. And previous observational studies have not reported changes in specific brain regions. By employing Mendelian randomization, we have been able to infer a causal relationship between osteoporosis and a reduction in the surficial area (SA) of the brain cortical. This effect is partially mediated by vascular calcification. We found that osteoporosis significantly decreased the SA of global brain cortical (ß = -1587.62 mm2, 95%CI: -2645.94 mm2 to -529.32 mm2, P = 0.003) as well as the paracentral gyrus without global weighted (ß = - 19.42 mm2, 95%CI: -28.90 mm2 to -9.95 mm2, P = 5.85 × 10-5). Furthermore, we estimated that 42.25% and 47.21% of the aforementioned effects are mediated through vascular calcification, respectively. Osteoporosis leads to a reduction in the SA of the brain cortical, suggesting the presence of the bone-brain axis. Vascular calcification plays a role in mediating this process to a certain extent. These findings establish a theoretical foundation for further investigations into the intricate interplay between bone, blood vessels, and the brain.

Trans-Lesion Synthesis and Mismatch Repair Pathway Crosstalk Defines Chemoresistance and Hypermutation Mechanisms in Glioblastoma.

Almost all Glioblastoma (GBM) are either intrinsically resistant to the chemotherapeutical drug temozolomide (TMZ) or acquire therapy-induced mutations that cause chemoresistance and recurrence. The genome maintenance mechanisms responsible for GBM chemoresistance and hypermutation are unknown. We show that the E3 ubiquitin ligase RAD18 (a proximal regulator of TLS) is activated in a Mismatch repair (MMR)-dependent manner in TMZ-treated GBM cells, promoting post-replicative gap-filling and survival. An unbiased CRISPR screen provides a new aerial map of RAD18-interacting DNA damage response (DDR) pathways deployed by GBM to tolerate TMZ genotoxicity. Analysis of mutation signatures from TMZ-treated GBM reveals a role for RAD18 in error-free bypass of O6mG (the most toxic TMZ-induced lesion), and error-prone bypass of other TMZ-induced lesions. Our analyses of recurrent GBM patient samples establishes a correlation between low RAD18 expression and hypermutation. Taken together we define novel molecular underpinnings for the hallmark tumorigenic phenotypes of TMZ-treated GBM.

Trans-Lesion Synthesis and Mismatch Repair Pathway Crosstalk Defines Chemoresistance and Hypermutation Mechanisms in Glioblastoma.

Almost all Glioblastoma (GBM) are either intrinsically resistant to the chemotherapeutical drug temozolomide (TMZ) or acquire therapy-induced mutations that cause chemoresistance and recurrence. The genome maintenance mechanisms responsible for GBM chemoresistance and hypermutation are unknown. We show that the E3 ubiquitin ligase RAD18 (a proximal regulator of TLS) is activated in a Mismatch repair (MMR)-dependent manner in TMZ-treated GBM cells, promoting post-replicative gap-filling and survival. An unbiased CRISPR screen provides a new aerial map of RAD18-interacting DNA damage response (DDR) pathways deployed by GBM to tolerate TMZ genotoxicity. Analysis of mutation signatures from TMZ-treated GBM reveals a role for RAD18 in error-free bypass of O6mG (the most toxic TMZ-induced lesion), and error-prone bypass of other TMZ-induced lesions. Our analyses of recurrent GBM patient samples establishes a correlation between low RAD18 expression and hypermutation. Taken together we define novel molecular underpinnings for the hallmark tumorigenic phenotypes of TMZ-treated GBM.

Inflammatory arthritis and eye diseases: a Mendelian randomization study.

Objectives: The aim of this study was to determine causal associations between inflammatory arthritis and eye diseases (disorders of sclera, cornea, iris, and ciliary body [DSCIC] and disorders of choroid and retina [DCR]). Methods: Genome-wide association studies' summary data of rheumatoid arthritis (RA) from a large-scale meta-analysis were used to identify genetically predicted RA. UK Biobank source data predicted ankylosing spondylitis (AS), psoriatic arthritis (PsA), and juvenile idiopathic arthritis (JIA). Furthermore, data from the FinnGen Biobank were used to identify genetically predicted eye diseases. Two-sample Mendelian randomization analysis was used to assess the causal relationship between inflammatory arthritis and eye diseases in the European population. Inverse-variance weighting (IVW) was used as the primary method, while MR-Egger, weighted median, and MR-PRESSO outlier test were used to detect heterogeneity and pleiotropy. Results: Genetically determined RA was indeed observed to have a causal effect on DSCIC (odds ratio [OR] = 1.084, p = 2.353 × 10-10) and DCR (OR = 1.151, p = 1.584 × 10-19). AS was causally associated with DSCIC (OR = 1.068, p < 2.024 × 10-8). In addition, PsA was also found to have a causal association with an increased risk of 17.9% for the development of DSCIC (OR = 1.179, p = 0.003). On the flip side, DSCIC increased the risk of JIA (OR = 2.276, p = 0.003). Conclusion: Our study provided genetic evidence for the causal associations of RA, AS, and PsA with an increased risk of DSCIC, and a causal association between RA and DCR was also identified. In addition, DSCIC greatly increased the risk of JIA.

Mass Spectrometric Analysis of Urinary N-Glycosylation Changes in Patients with Parkinson's Disease.

Urine is thought to provide earlier and more sensitive molecular changes for biomarker discovery than blood. Numerous glycoproteins, peptides, and free glycans are present in urine through glomerular filtration of plasma, cell shedding, apoptosis, proteolytic cleavage, and exosome secretion. Urine biomarkers have enormous diagnostic potential, and the use of these biomarkers is a long-standing practice. The discovery of non-urological disease biomarkers from urine is also gaining attention due to its non-invasive sample collection and ease of analysis. Abnormal protein glycosylation in plasma or cerebrospinal fluid has been associated with Parkinson's disease, however, whether urine with Parkinson's disease has characteristic glycosylation remains to be explored. Here, we use mass spectrometry-based glycomics and glycoproteomics approaches to analyze urine samples for glycans, glycosites, and intact glycopeptides of urine samples. Reduced abundance of N-glycans was detected at the level of total glycans as well as specific glycosites of glycopeptides. The most abundant N-glycan in urine is S(6)1H5N4F1; S(6)2H5N4 and N4H4F1 are highly present in serum and urine, and 10 biantennary galactosylated N-glycans in the urine of PD patients were significantly decreased. The downregulation of sialylation may be due to the reduction of ST3GAL2. Site-specific N-glycosylation analysis revealed that AMBP, UMOD, and RNase1 have PD-specific N-glycosylation sites. GO and KEGG analysis revealed that N-glycosylation changes may provide clues to identify disease-specific glycosylation biomarkers in Parkinson's disease.

Novel Approach to Enriching Glycosylated RNAs: Specific Capture of GlycoRNAs via Solid-Phase Chemistry.

Ribonuclease (RNA) modifications can alter cellular function and lead to differential immune responses by acting as discriminators between RNAs from different phyla. RNA glycosylation has recently been observed at the cell surface, and its dysregulation in disease may change RNA functions. However, determining which RNA substrates can be glycosylated remains to be explored. Here, we develop a solid-phase chemoenzymatic method (SPCgRNA) for targeting glycosylated RNAs, by which glycosylated RNA substrates can be specifically recognized. We found the differential N-glycosylation of small RNAs in hTERT-HPNE and MIA PaCa-2 cancer cells using SPCgRNA. RNA-Seq showed that the changes in glyco-miRNAs prepared from SPCgRNA were consistent with those of traditional methods. The KEGG signaling pathway analysis revealed that differential miRNA glycosylation can affect tumor cell proliferation and survival. Further studies found that NGI-1 significantly inhibited the proliferation, migration, and circulation of MIA PaCa-2 and promoted cell apoptosis. In addition, ß-1,4-galactosyltransferase 1 (B4GALT1) not only affected the expression level of glycosylated miRNAs hsa-miR-21-5p but also promoted cell apoptosis and inhibited the cell cycle possibly through the p53 signaling pathway, while B4GALT1 and p53 were also affected following the hsa-miR-21-5p increase. These results suggest that B4GALT1 may catalyze miRNAs glycosylation, which further promotes cancer cell progression.

Comparison of the femoral condylar ellipse line and the surgical epicondylar axis: 3D measurement by MRI scans in healthy Chinese people.

OBJECTIVE: The sagittal shapes of the femoral condyles were thought to consist of circles. However, the line connecting the centers of circles was not consistent with the surgical epicondylar axis (SEA) which was commonly used in surgery. Recently, ellipses have been proposed as an alternative method to represent the sagittal femoral condylar shape. Does the condylar ellipse line (CEL) coincide with the SEA in 3D MRI reconstruction analysis? METHODS: From May to August 2021, a total of 80 healthy subjects were scanned by MRI on the right knee in this retrospective study. The ellipses on the most distal slices of the medial and lateral condyles were determined. A line connecting the centers of the medial and lateral ellipses was the CEL. A line connecting the deepest point of the medial sulcus and the most prominent point of the lateral epicondyle was the SEA. Angular measurement of the SEA and the CEL relative to the posterior condylar line (PCL) and the distal condylar line (DCL) was performed on an axial and coronal view of the 3D model, respectively. Measurements were compared between males and females by using the independent-samples t-test. Pearson correlation was used to analyze the relationship between SEA-PCL and CEL-PCL, SEA-DCL, and CEL-DCL. RESULTS: On the axial view, the mean SEA-CEL was 0.35° ± 0.96°. SEA-PCL (2.91° ± 1.40°) had a high correlation with CEL-PCL (3.27° ± 1.11°) (r = 0.731, p < 0.001). On the coronal view, the mean coronal SEA-CEL was 1.35° ± 1.13°. SEA-DCL (1.35° ± 1.13°) had a low correlation with CEL-DCL (0.18° ± 0.84°) (r = 0.319, p = 0.007). On the sagittal view, the outlet points of the CEL on the medial and lateral epicondyles were anatomically located in the anteroinferior direction to the SEA. CONCLUSIONS: CEL traversed the medial and lateral epicondyles, which has a mean deviation of 0.35° with SEA on axial view and a mean deviation of 0.18° with DCL on coronal view. This study suggested that the ellipse approach is an improved scheme for representing the femoral condylar shape.

PHD-finger family genes in wheat (Triticum aestivum L.): Evolutionary conservatism, functional diversification, and active expression in abiotic stress.

Plant homeodomain (PHD) transcription factors (TFs) are a class of proteins with conserved Cys4-His-Cys3 domains that play important roles in plant growth and development and in response to abiotic stresses. Although characterization of PHDs has been performed in plants, little is known about their function in wheat (Triticum aestivum L.), especially under stress conditions. In the present study, 244 TaPHDs were identified in wheat using comparative genomics. We renamed them TaPHD1-244 based on their chromosomal distribution, and almost all PHD proteins were predicted to be located in the nucleus. According to the unrooted neighbor-joining phylogenetic tree, gene structure, and motif analyses, PHD genes were divided into four clades. A total of 149 TaPHD genes were assigned to arise from duplication events. Furthermore, 230 gene pairs came from wheat itself, and 119, 186, 168, 7, 2, and 6 gene pairs came from six other species (Hordeum vulgareto, Zea mays, Oryza sativa, Arabidopsis thaliana, Brassica rapa, and Gossypium raimondii, respectively). A total of 548 interacting protein branches were identified to be involved in the protein interaction network. Tissue-specific expression pattern analysis showed that TaPHDs were highly expressed in the stigma and ovary during flowering, suggesting that the TaPHD gene plays an active role in the reproductive growth of wheat. In addition, the qRT-PCR results further confirmed that these TaPHD genes are involved in the abiotic stress response of wheat. In conclusion, our study provides a theoretical basis for deciphering the molecular functions of TaPHDs, particularly in response to abiotic stress.

Recent development in hydrophilic interaction liquid chromatography stationary materials for glycopeptide analysis.

Protein glycosylation is one of the most important post-translational modifications, and aberrant glycosylation is associated with the occurrence and development of diseases. Deciphering abnormal glycosylation changes can identify disease-specific signatures to facilitate the discovery of potential disease biomarkers. However, glycosylation analysis is challenging due to the diversity of glycans, heterogeneity of glycosites, and poor electrospray ionization of mass spectrometry. To overcome these obstacles, glycosylation is often elucidated using enriched glycopeptides by removing highly abundant non-glycopeptides. Hydrophilic interaction liquid chromatography (HILIC) is widely used for glycopeptide enrichment due to its excellent selectivity and specificity to hydrophilic glycans and compatibility with mass spectrometry. However, the development of HILIC has lagged far behind hydrophobic interaction chromatography, so efforts to further improve the performance of HILIC are beneficial for glycosylation analysis. This review discusses recent developments in HILIC materials and their advanced applications. Based on the physiochemical properties of glycopeptides, the use of amino acids or peptides as stationary phases showed improved enrichment and separation of glycopeptides. We can envision that the use of glycopeptides as stationary phases would definitely further improve the selectivity and specificity of HILIC for glycosylation analysis.

Contribution of Alate and Apterous Morphs to Demographic Characteristics, and Stable Stage Distribution of Acyrthosiphon pisum (Hemiptera: Aphididae) on Four Different Alfalfa Varieties.

Life table data of the green pea aphid, Acyrthosiphon pisum (Harris) reared on four different resistant alfalfa varieties, i.e., 5S43, TG4 CW044026 (abbreviated as TG4), TG7 CW2883 (abbreviated as TG7), and Aurora were analyzed using the age-stage, two-sex life table. A higher proportion of alate adults were observed on 5S43, TG7, and Aurora; while a higher proportion of apterous adults occurred on TG4. The contributions of alate aphids to the finite rate of increase (λ), intrinsic rate of increase (r), and net reproductive rate (R0) were higher than apterous aphids on 5S43, TG7, and Aurora, while apterous aphids contributed more to λ, r, and R0 on TG4. The highest population parameters were observed on TG4 (r = 0.208 d-1, λ = 1.231 d-1, and R0 = 18.8 offspring/individual), while the lowest values were on TG7 (r = 0.129 d-1, λ = 1.138 d-1, and R0 = 9.9 offspring/individual). Because the age-stage, two-sex life table is capable of describing the stage differentiation, it enables the calculation of the stable stage distribution (SSD). A higher proportion of adult A. pisum was observed in SSD than in fourth instar nymphs. Population simulation showed the stage structure will approach SSD. Because the R0 and the mean generation time (T) values do not reflect the population growth rate, their use as population fitness parameters should be avoided. These findings can be utilized in helping to select resistant alfalfa varieties to effectively manage the pea aphid.

Pro-angiognetic and pro-osteogenic effects of human umbilical cord mesenchymal stem cell-derived exosomal miR-21-5p in osteonecrosis of the femoral head.

Mesenchymal stem cell (MSC)-derived exosomes (Exos) enhanced new bone formation, coupled with positive effects on osteogenesis and angiogenesis. This study aims to define the role of microRNA (miR)-21-5p delivered by human umbilical MSC-derived Exos (hucMSC-Exos) in the osteonecrosis of the femoral head (ONFH). We first validated that miR-21-5p expression was downregulated in the cartilage tissues of ONFH patients. Besides, hucMSCs delivered miR-21-5p to hFOB1.19 cells and human umbilical vein endothelial cells (HUVECs) through the secreted Exos. Loss- and gain-of-function approaches were performed to clarify the effects of Exo-miR-21-5p, SOX5, and EZH2 on HUVEC angiogenesis and hFOB1.19 cell osteogenesis. It was established that Exo-miR-21-5p augments HUVEC angiogenesis and hFOB1.19 cell osteogenesis in vitro, as reflected by elevated alkaline phosphatase (ALP) activity and calcium deposition, and increased the expression of osteogenesis-related markers OCN, Runx2 and Collagen I. Mechanistically, miR-21-5p targeted SOX5 and negatively regulated its expression, while SOX5 subsequently promoted the transcription of EZH2. Ectopically expressed SOX5 or EZH2 could counterweigh the effect of Exo-miR-21-5p. Further, hucMSC-Exos containing miR-21-5p repressed the expression of SOX5 and EZH2 and augmented angiogenesis and osteogenesis in vivo. Altogether, our study uncovered the role of miR-21-5p shuttled by hucMSC-Exos, in promoting angiogenesis and osteogenesis, which may be a potential therapeutic target for ONFH.

The link between aberrant splicing and B cell infiltration in breast cancer hormone therapy.

Increasing evidence has suggested alternative splicing (AS) is an essential mechanism for tumor initiation and progression. However, the functions of aberrant splicing have not been well understood in hormone therapy-treated breast cancer (HTBC) patients yet. By mining the SpliceSeq data of TCGA, we characterized 1388 survival-associated AS events (SASEs) that are markedly related to the overall survival (OS) of HTBC patients. Subsequently, a SASE prognostic signature was deduced via the least absolute shrinkage and selection operator (LASSO) and multivariate Cox analysis, and the HTBC patients were stratified into high- and low-risk groups based on the signature. The differentially expressed genes between the high- and low-risk patients were enriched with immune response genes. Furthermore, we revealed that B cell infiltration was significantly low in the high-risk patients with single-sample gene set enrichment analysis (ssGSEA), suggesting a link between AS abnormality and B cell infiltration in tumor immune microenvironment (TIME). Next, a similar prognostic signature based on splicing factor expression was deduced using Molecular Taxonomy of Breast Cancer (METABRIC) HTBC patients. When this prognostic signature was applied to stratify HTBC patients in the METABRIC and TCGA cohorts, the high-risk subgroups both had significantly low B cell infiltration in TIME. CIBERSORT analysis further demonstrated plasma cells infiltrated less in TIME. Also, both the AS (P < 0.0001) and SF (P = 0.009) prognostic signatures performed well in predicting OS of HTBC patients. In summary, our study has demonstrated that AS abnormality is a potential cause of B cell infiltration alteration, which is a critical factor for cancer death risks in HTBC patients and might be exploited therapeutically in the future.

Efficacy of combining erbium:YAG and fractional CO2 laser for the treatment of facial scarring

Background: Laser intervention is increasingly recognized as an effective approach to preventing scars, however, as lasers of different wavelengths reach different target tissues, an appropriate combination of laser treatment is required. Objectives: To evaluate the efficacy of combining erbium: yttrium aluminium garnet (Er:YAG) 2,940-nm laser and fractional carbon dioxide (CO2) 10,600-nm laser for the treatment of facial scarring. Materials & Methods: Medical records of 39 patients with facial scars were retrospectively reviewed. Treatments initiated within six months post-injury were defined as "early-intervention", whereas those initiated >six months post-injury were defined as "delayed-intervention". Patients' scores on the Vancouver Scar Scale (VSS) and Visual Analogue Scale (VAS) were collected and compared between different timepoints using the paired t-test. Improvement in pigmentation, height, vascularity and pliability was analysed between baseline and after final treatment. Results: Between March 2020 and March 2022, 39 patients underwent dual-laser therapy at our laser clinic. The average number of treatments was 4.69 1.54; 18 patients in the early-intervention and 21 patients in the delayedintervention group. Facial scars significantly improved after the first treatment in the delayed-intervention group (p < 0.001), however, there was no significant improvement until two sessions were completed in the early-intervention group (p < 0.001). In the delayed-intervention group, improvement in vascularity was insignificant (p = 0.083), however, improvement in height and flexibility was prominent (p < 0.001). In the early-intervention group, facial scars improved in all four dimensions of VSS (p < 0.05), with improvement of vascularity being the most significant. Side effects were self-limited, and complications were not noted. Conclusion: The combination of Er:YAG and fractional CO2 laser is a safe and effective therapeutic modality for facial scarring.

An Immune-Related Gene Prognostic Index for Triple-Negative Breast Cancer Integrates Multiple Aspects of Tumor-Immune Microenvironment.

Tumor-immune cell compositions and immune checkpoints comprehensively affect TNBC outcomes. With the significantly improved survival rate of TNBC patients treated with ICI therapies, a biomarker integrating multiple aspects of TIME may have prognostic value for improving the efficacy of ICI therapy. Immune-related hub genes were identified with weighted gene co-expression network analysis and differential gene expression assay using The Cancer Genome Atlas TNBC data set (n = 115). IRGPI was constructed with Cox regression analysis. Immune cell compositions and TIL status were analyzed with CIBERSORT and TIDE. The discovery was validated with the Molecular Taxonomy of Breast Cancer International Consortium data set (n = 196) and a patient cohort from our hospital. Tumor expression or serum concentrations of CCL5, CCL25, or PD-L1 were determined with immunohistochemistry or ELISA. The constructed IRGPI was composed of CCL5 and CCL25 genes and was negatively associated with the patient's survival. IRGPI also predicts the compositions of M0 and M2 macrophages, memory B cells, CD8+ T cells, activated memory CD4 T cells, and the exclusion and dysfunction of TILs, as well as PD-1 and PD-L1 expression of TNBC. IRGPI is a promising biomarker for predicting the prognosis and multiple immune characteristics of TNBC.

Identification of spliceosome components pivotal to breast cancer survival.

Cancer cells employ alternative splicing (AS) to acquire splicing isoforms favouring their survival. However, the causes of aberrant AS in breast cancer are poorly understood. In this study, the METABRIC (Molecular Taxonomy of Breast Cancer International Consortium) data were analysed with univariate feature selection. Of 122 analysed spliceosome components, U2SURP, PUF60, DDX41, HNRNPAB, EIF4A3, and PPIL3 were significantly associated with breast cancer survival. The top 4 four genes, U2SURP, PUF60, DDX41, and HNRNPAB, were chosen for further analyses. Their expression was significantly associated with cancer molecular subtype, tumour stage, tumour grade, overall survival (OS), and cancer-specific survival in the METABRIC data. These results were verifiable using other cohorts. The Cancer Genome Atlas data unveiled the elevated expression of PUF60, DDX41, and HNRNPAB in tumours compared with the normal tissue and confirmed the differential expression of the four genes among cancer molecular subtypes, as well as the associations of U2SURP, PUF60, and DDX41 expression with tumour stage. A meta-analysis data verified the associations of U2SURP, PUF60, and HNRNPAB expression with tumour grade, the associations of PUF60, DDX41, and HNRNPAB expression with OS and distant metastasis-free survival, and the associations of U2SURP and HNRNPAB expression with relapse-free survival. Experimentally, we demonstrated that inhibiting the expression of the four genes separately suppressed cell colony formation and slowed down cell growth considerably in breast cancer cells, but not in immortal breast epithelial cells. In conclusion, we have identified U2SURP, PUF60, DDX41, and HNRNPAB are spliceosome-related genes pivotal for breast cancer survival.