Inhibition of Proliferation and Induction of Apoptosis in Prostatic Carcinoma DU145 Cells by Polysaccharides from Yunnan Rosa roxburghii Tratt.

OBJECTIVE: This study aimed to investigate methodologies for the extraction and purification of polysaccharides from Rosa roxburghii Tratt fruits and their impact on various cellular processes in prostate cancer DU145 cells, including survival rate, migration, invasion, cell cycle, and apoptosis. RESULTS: Compared to the control group, the polysaccharide exhibited a significant reduction in the viability, migration, and invasion rates of DU145 cells in a time- and dose-dependent manner within the polysaccharide-treated groups. Additionally, it effectively arrested the cell cycle of DU145 cells at the G0/G1 phase by downregulating the expressions of CDK-4, CDK-6, and Cyclin D1. Furthermore, it induced apoptosis by upregulating the expressions of Caspase 3, Caspase 8, Caspase 9, and BAX. METHODS: Polysaccharides were extracted from Rosa roxburghii Tratt sourced from Yunnan, China. Extraction and decolorization methods were optimized using response surface methodology, based on a single-factor experiment. Polysaccharide purification was carried out using DEAE-52 cellulose and Sephadex G-100 column chromatography. The optimal dosage of R. roxburghii Tratt polysaccharide affecting DU145 cells was determined using the CCK-8 assay. Cell migration and invasion were assessed using transwell and scratch assays. Flow cytometry was employed to analyze the effects on the cell cycle and apoptosis. Western blotting and Quantitative real-time PCR were utilized to examine protein and mRNA expressions in DU145 cells, respectively. CONCLUSIONS: Rosa roxburghii Tratt polysaccharides, consisting of D-mannose, L-rhamnose, N-acetyl-D-glucosamine, D-galacturonic acid, D-glucose, D-galactcose, D-xylose, L-arabinose, and L-fucose, possess the ability to hinder DU145 cell proliferation, migration, and invasion while inducing apoptosis through the modulation of relevant protein and gene expressions.

Cardio-Oncologic Knowledge of Nurses in the Oncology Service: A Multi-Center Survey in China.

Purpose: Elucidation of the cardio-oncologic knowledge among the oncology nurses of tertiary hospitals in Shanxi Province to provide better insights and directions for management by nursing managers. Background: China's National Health and Wellness Commission issued the Action Plan for Further Improving Nursing Services in June 2023, which requires nurses to provide patients with physical and mental holistic nursing services, such as medical care, condition observation, assistance with treatment, and health guidance. Most oncology patients are treated with chemotherapy, but the modality can cause greater harm to patients, especially cardiotoxicity. How to provide precise care for chemotherapy patients is a problem for nursing managers. Methods: In order to investigate the level of cardio-oncologic knowledge among the oncology nurses of tertiary care hospitals in Shanxi Province, China, a questionnaire was created based on the relevant literature and the provided instructions on cardio-oncology. The chi-squared test was performed for multiple comparisons of the level of knowledge of disease observation, health guidance, and implementation of treatment. Spearman correlation analysis was performed to analyze the correlation between the levels of cardio-oncologic knowledge and general information of hospitals and nurses. Results: Cardio-oncology awareness among the oncology nurses was 0.1%-44.7%, the awareness rate of single dimension was 0 to 3.9%, and overall awareness rate was 0. A partially significant difference was revealed in the two-by-two comparisons of the awareness rates of the three dimensions of disease observation, health guidance, and implementation of treatment (P < 0.05). A correlation was observed between the cardio-oncologic knowledge and some of the hospital and the nurses' general information data (P < 0.05). Conclusion: Oncology nurses exhibited a low rate of awareness related to cardio-oncology. Hospitals could establish oncology nursing teams to train the oncology nurses to promote their cardio-oncologic knowledge and ensure the quality of daily care provided by these nurses.

TMEM215 Prevents Endothelial Cell Apoptosis in Vessel Regression by Blunting BIK-Regulated ER-to-Mitochondrial Ca Influx.

BACKGROUND: In developmental and pathological tissues, nascent vessel networks generated by angiogenesis require further pruning/regression to delete nonfunctional endothelial cells (ECs) by apoptosis and migration. Mechanisms underlying EC apoptosis during vessel pruning remain elusive. TMEM215 (transmembrane protein 215) is an endoplasmic reticulum-located, 2-pass transmembrane protein. We have previously demonstrated that TMEM215 knockdown in ECs leads to cell death, but its physiological function and mechanism are unclear. METHODS: We characterized the role and mechanism of TMEM215 in EC apoptosis using human umbilical vein endothelial cells by identifying its interacting proteins with immunoprecipitation-mass spectrometry. The physiological function of TMEM215 in ECs was assessed by establishing a conditional knockout mouse strain. The role of TMEM215 in pathological angiogenesis was evaluated by tumor and choroidal neovascularization models. We also tried to evaluate its translational value by delivering a Tmem215 small interfering RNA (siRNA) using nanoparticles in vivo. RESULTS: TMEM215 knockdown in ECs induced apoptotic cell death. We identified the chaperone BiP as a binding partner of TMEM215, and TMEM215 forms a complex with and facilitates the interaction of BiP (binding immunoglobin protein) with the BH (BCL-2 [B-cell lymphoma 2] homology) 3-only proapoptotic protein BIK (BCL-2 interacting killer). TMEM215 knockdown triggered apoptosis in a BIK-dependent way and was abrogated by BCL-2. Notably, TMEM215 knockdown increased the number and diminished the distance of mitochondria-associated endoplasmic reticulum membranes and increased mitochondrial calcium influx. Inhibiting mitochondrial calcium influx by blocking the IP3R (inositol 1,4,5-trisphosphate receptor) or MCU (mitochondrial calcium uniporter) abrogated TMEM215 knockdown-induced apoptosis. TMEM215 expression in ECs was induced by physiological laminar shear stress via EZH2 downregulation. In EC-specific Tmem215 knockout mice, induced Tmem215 depletion impaired the regression of retinal vasculature characterized by reduced vessel density, increased empty basement membrane sleeves, and increased EC apoptosis. Moreover, EC-specific Tmem215 ablation inhibited tumor growth with disrupted vasculature. However, Tmem215 ablation in adult mice attenuated lung metastasis, consistent with reduced Vcam1 expression. Administration of nanoparticles carrying Tmem215 siRNA also inhibited tumor growth and choroidal neovascularization injury. CONCLUSIONS: TMEM215, which is induced by blood flow-derived shear stress via downregulating EZH2, protects ECs from BIK-triggered mitochondrial apoptosis mediated by calcium influx through mitochondria-associated ER membranes during vessel pruning, thus providing a novel target for antiangiogenic therapy.

Repression of rRNA gene transcription by endothelial SPEN deficiency normalizes tumor vasculature via nucleolar stress.

Human cancers induce a chaotic, dysfunctional vasculature that promotes tumor growth and blunts most current therapies; however, the mechanisms underlying the induction of a dysfunctional vasculature have been unclear. Here, we show that split end (SPEN), a transcription repressor, coordinates rRNA synthesis in endothelial cells (ECs) and is required for physiological and tumor angiogenesis. SPEN deficiency attenuated EC proliferation and blunted retinal angiogenesis, which was attributed to p53 activation. Furthermore, SPEN knockdown activated p53 by upregulating noncoding promoter RNA (pRNA), which represses rRNA transcription and triggers p53-mediated nucleolar stress. In human cancer biopsies, a low endothelial SPEN level correlated with extended overall survival. In mice, endothelial SPEN deficiency compromised rRNA expression and repressed tumor growth and metastasis by normalizing tumor vessels, and this was abrogated by p53 haploinsufficiency. rRNA gene transcription is driven by RNA polymerase I (RNPI). We found that CX-5461, an RNPI inhibitor, recapitulated the effect of Spen ablation on tumor vessel normalization and combining CX-5461 with cisplatin substantially improved the efficacy of treating tumors in mice. Together, these results demonstrate that SPEN is required for angiogenesis by repressing pRNA to enable rRNA gene transcription and ribosomal biogenesis and that RNPI represents a target for tumor vessel normalization therapy of cancer.

miR-342-5p promotes vascular smooth muscle cell phenotypic transition through a negative-feedback regulation of Notch signaling via targeting FOXO3.

AIM: Under various pathological conditions such as cancer, vascular smooth muscle cells (vSMCs) transit their contractile phenotype into phenotype(s) characterized by proliferation and secretion, a process called vSMC phenotypic transition (vSMC-PT). Notch signaling regulates vSMC development and vSMC-PT. This study aims to elucidate how the Notch signal is regulated. MAIN METHODS: Gene-modified mice with a SM22α-CreERT2 transgene were generated to activate/block Notch signaling in vSMCs. Primary vSMCs and MOVAS cells were cultured in vitro. RNA-seq, qRT-PCR and Western blotting were used to evaluated gene expression level. EdU incorporation, Transwell and collagen gel contraction assays were conducted to determine the proliferation, migration and contraction, respectively. KEY FINDINGS: Notch activation upregulated, while Notch blockade downregulated, miR-342-5p and its host gene Evl in vSMCs. However, miR-342-5p overexpression promoted vSMC-PT as shown by altered gene expression profile, increased migration and proliferation, and decreased contraction, while miR-342-5p blockade exhibited the opposite effects. Moreover, miR-342-5p overexpression significantly suppressed Notch signaling, and Notch activation partially abolished miR-342-5p-induced vSMC-PT. Mechanically, miR-342-5p directly targeted FOXO3, and FOXO3 overexpression rescued miR-342-5p-induced Notch repression and vSMC-PT. In a simulated tumor microenvironment, miR-342-5p was upregulated by tumor cell-derived conditional medium (TCM), and miR-342-5p blockade abrogated TCM-induced vSMC-PT. Meanwhile, conditional medium from miR-342-5p-overexpressing vSMCs significantly enhanced tumor cell proliferation, while miR-342-5p blockade had the opposite effects. Consistently, in a co-inoculation tumor model, miR-342-5p blockade in vSMCs significantly delayed tumor growth. SIGNIFICANCE: miR-342-5p promotes vSMC-PT through a negative-feedback regulation of Notch signaling via downregulating FOXO3, which could be a potential target for cancer therapy.

Resistance to anti-HER2 therapy associated with the TSC2 nonsynonymous variant c.4349 C > G (p.Pro1450Arg) is reversed by CDK4/6 inhibitor in HER2-positive breast cancer.

HER2-positive breast cancer patients carrying the germline TSC2 nonsynonymous variant c.4349 C > G (p.Pro1450Arg) are resistant to anti-HER2 therapy. Multi-predictor in silico analysis reveals that this variant is deleterious. We explore the potential mechanism of this TSC2 variant and investigate methods for overcoming anti-HER2 resistance. TSC2 c.4349 C > G reverses the inhibitory effect on mTOR and downstream signaling by increasing TSC2 phosphorylation at Thr1462 and confers significant lapatinib resistance in vitro and in vivo. The combination of lapatinib and the CDK4/6 inhibitor palbociclib inhibits cyclin D1/CDK4/Rb alternative pathway and TSC2 phosphorylation, thereby partially attenuating mTOR activity and inducing TSC2-mutant cell blockage at G1/G0. In in vitro and xenograft models, palbociclib+lapatinib shows higher anti-tumor activity than monotherapy and overcomes the resistance of the TSC2 c.4349 C > G-related variant to anti-HER2 therapy. We reveal a new mechanism of resistance to anti-HER2 therapy and provide a strategy to increase the efficiency of anti-HER2 therapy in HER2-positive breast cancer.

EFEMP2 increases the invasion ability of cervical cancer cells by promoting EMT via the Raf/MEK/ERK signaling pathway.

EFEMP2 has been reported as a candidate oncogene. To investigate the role of EFEMP2 in cervical cancer cell proliferation and invasion, the mRNA and protein expressions of EFEMP2 in 5 different cervical cancer cell lines were detected. And then the effects of up- or down-regulation of EFEMP2 expression on the biological behavior of cervical cancer cells were further investigated by transfection experiments and cell function assays in vitro and in vivo. The results revealed that EFEMP2 was highly expressed in highly invasive Ca Ski cells and lowly expressed in less invasive HT-3 cells. When EFEMP2 was knocked down, the proliferation and invasion ability of cervical cancer cells were also reduced, accompanied by the decreased expression of MMP-1, MMP-13, MMP-3, and MMP-10, meanwhile, the EMT process was blocked and the Raf/MEK/ERK signaling pathway was inhibited. On the contrary, the upregulation of EFEMP2 could promote the proliferation and invasion of cervical cancer cells by inducing EMT and activating the Raf/MEK/ERK pathway. In conclusion, EFEMP2 could increase the invasion ability of cervical cancer cells by upregulating the expression of MMP-1, MMP-13, MMP-3, and MMP-10 and inducing the EMT process through the Raf/MEK/ERK pathway. EFEMP2 played a promoting role in the development of cervical cancer and provided a potential therapeutic target for inhibiting the invasion and metastasis of cancer cells and improving the prognosis of cervical cancer patients.

Novel Chemicals Derived from Tadalafil Exhibit PRMT5 Inhibition and Promising Activities against Breast Cancer.

Breast cancer seriously endangers women's health worldwide. Protein arginine methyltransferase 5 (PRMT5) is highly expressed in breast cancer and represents a potential druggable target for breast cancer treatment. However, because the currently available clinical PRMT5 inhibitors are relatively limited, there is an urgent need to develop new PRMT5 inhibitors. Our team previously found that the FDA-approved drug tadalafil can act as a PRMT5 inhibitor and enhance the sensitivity of breast cancer patients to doxorubicin treatment. To further improve the binding specificity of tadalafil to PRMT5, we chemically modified tadalafil, and designed three compounds, A, B, and C, based on the PRMT5 protein structure. These three compounds could bind to PRMT5 through different binding modes and inhibit histone arginine methylation. They arrested the proliferation and triggered the apoptosis of breast cancer cells in vitro and also promoted the antitumor effects of the chemotherapy drugs cisplatin, doxorubicin, and olaparib in combination regimens. Among them, compound A possessed the highest potency. Finally, the anti-breast cancer effects of PRMT5 inhibitor A and its ability to enhance chemosensitivity were further verified in a xenograft mouse model. These results indicate that the new PRMT5 inhibitors A, B, and C may be potential candidates for breast cancer treatment.

Cardiotoxicity of Epidermal Growth Factor Receptor 2-Targeted Drugs for Breast Cancer.

Breast cancer is the most common form of cancer in women and its incidence has been increasing over the years. Human epidermal growth factor receptor 2 (HER2 or ErbB2) overexpression is responsible for 20 to 25% of invasive breast cancers, and is associated with poor prognosis. HER2-targeted therapy has significantly improved overall survival rates in patients with HER2-positive breast cancer. However, despite the benefits of this therapy, its cardiotoxicity is a major concern, especially when HER2-targeted therapy is used in conjunction with anthracyclines. At present, the mechanism of this cardiotoxicity is not fully understood. It is thought that HER2-targeting drugs inhibit HER2/NRG 1 dimer formation, causing an increase in ROS in the mitochondria of cardiomyocytes and inhibiting the PI3K/Akt and Ras/MAPK pathways, resulting in cell apoptosis. Antioxidants, ACE inhibitors, angiotensin II receptor blockers, ß-blockers, statins and other drugs may have a cardioprotective effect when used with ErbB2-targeting drugs. NT-proBNP can be used to monitor trastuzumab-induced cardiotoxicity during HER2-targeted treatment and may serve as a biological marker for clinical prediction of cardiotoxicity. Measuring NT-proBNP is non-invasive, inexpensive and reproducible, therefore is worthy of the attention of clinicians. The aim of this review is to discuss the potential mechanisms, clinical features, diagnostic strategies, and intervention strategies related to cardiotoxicity of ErbB2-targeting drugs.

Notch activation promotes endothelial quiescence by repressing MYC expression via miR-218.

After angiogenesis-activated embryonic and early postnatal vascularization, endothelial cells (ECs) in most tissues enter a quiescent state necessary for proper tissue perfusion and EC functions. Notch signaling is essential for maintaining EC quiescence, but the mechanisms of action remain elusive. Here, we show that microRNA-218 (miR-218) is a downstream effector of Notch in quiescent ECs. Notch activation upregulated, while Notch blockade downregulated, miR-218 and its host gene Slit2, likely via transactivation of the Slit2 promoter. Overexpressing miR-218 in human umbilical vein ECs (HUVECs) significantly repressed cell proliferation and sprouting in vitro. Transcriptomics showed that miR-218 overexpression attenuated the MYC proto-oncogene, bHLH transcription factor (MYC, also known as c-myc) signature. MYC overexpression rescued miR-218-mediated proliferation and sprouting defects in HUVECs. MYC was repressed by miR-218 via multiple mechanisms, including reduction of MYC mRNA, repression of MYC translation by targeting heterogeneous nuclear ribonucleoprotein A1 (hnRNPA1), and promoting MYC degradation by targeting EYA3. Inhibition of miR-218 partially reversed Notch-induced repression of HUVEC proliferation and sprouting. In vivo, intravitreal injection of miR-218 reduced retinal EC proliferation accompanied by MYC repression, attenuated pathological choroidal neovascularization, and rescued retinal EC hyper-sprouting induced by Notch blockade. In summary, miR-218 mediates the effect of Notch activation of EC quiescence via MYC and is a potential treatment for angiogenesis-related diseases.

Computer-aided diagnostic system based on deep learning for classifying colposcopy images.

BACKGROUND: Colposcopy is widely used to detect cervical cancer, but developing countries lack the experienced colposcopists necessary for accurate diagnosis. Artificial intelligence (AI) is being widely used in computer-aided diagnosis (CAD) systems. In this study, we developed and validated a CAD model based on deep learning to classify cervical lesions on colposcopy images. METHODS: Patient data, including clinical information, colposcopy images, and pathological results, were collected from Qilu Hospital. The study included 15,276 images from 7,530 patients. We performed two tasks in this study: normal cervix (NC) vs. low grade squamous intraepithelial lesion or worse (LSIL+) and high-grade squamous intraepithelial lesion (HSIL)- vs. HSIL+. The residual neural network (ResNet) probability was calculated for each patient to reflect the probability of lesions through a ResNet model. Next, a combination model was constructed by incorporating the ResNet probability and clinical features. We divided the dataset into a training set, validation set, and testing set at a ratio of 7:1:2. Finally, we randomly selected 300 patients from the testing set and compared the results with the diagnosis of a senior colposcopist and a junior colposcopist. RESULTS: The model that combines ResNet and clinical features performs better than ResNet alone. In the classification of NC and LSIL+, the area under the receiver operating characteristic curve (AUC), accuracy, sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) were 0.953, 0.886, 0.932, 0.846, 0.838, and 0.936, respectively. In the classification of HSIL- and HSIL+, the AUC, accuracy, sensitivity, specificity, PPV, and NPV were 0.900, 0.807, 0.823, 0.800, 0.618, and 0.920, respectively. In the two classification tasks, the diagnostic performance of the model was determined to be comparable to that of the senior colposcopist and exhibited a stronger diagnostic performance than the junior colposcopist. CONCLUSIONS: The CAD system for cervical lesion diagnosis based on deep learning performs well in the classification of cervical lesions and can provide an objective diagnostic basis for colposcopists.

Efficacy of Pyrotinib in HER2-Overexpressing Salivary Duct Carcinoma With Lung Metastasis: A Case Report.

Background: Salivary duct carcinoma (SDC), an aggressive and rare malignancy with poor prognosis, is mostly associated with the overexpression of the androgen receptor (AR) and human epidermal growth factor receptor 2 (HER2). However, limited data are available for the targeting of both HER2 and AR in advanced/metastatic SDC. Case Presentation: A 62-year-old man with advanced SDC accompanied by lung and lymph node metastasis showed disease progression after two lines of chemotherapy and endocrine therapy. Metastatic lesions from the lung biopsy were obtained, and immunohistochemistry (IHC) indicated the overexpression of AR and HER2 (3+). The patient was administered pyrotinib (a pan-ErbB receptor tyrosine kinase inhibitor) and bicalutamide (an androgen receptor antagonist) as a third-line treatment. During the ten months of follow-up, a durable partial response was achieved with this combination. Conclusions: This is the first clinical study to report the successful application of pyrotinib in a patient with advanced SDC. We recommend that pyrotinib and bicalutamide be used as salvage therapy for AR and HER2-positive advanced metastases in SDC, given the favorable response and clinical benefit.

SM22α+ vascular mural cells are essential for vessel stability in tumors and undergo phenotype transition regulated by Notch signaling.

BACKGROUND: Malformation of blood vessels represents a hallmark of cancers, but the role and regulation of vascular mural cells (vMCs), including vascular smooth muscle cells (vSMCs) and pericytes, in tumors has not been fully understood. SM22α has been identified as a marker of vSMCs. This study aims at elucidating the function and regulation of SM22α+ mural cells (SM22-MCs) in tumor stroma. METHODS: Gene-modified mice with a SM22α-CreERT2 transgene were employed to deplete SM22-MCs or activate/block Notch signaling in these cells. vSMCs from mouse dorsal aorta (vSMCs-DA) were cultured in vitro. RNA-seq was used to compare gene expression profiles. qRT-PCR and western blotting were used to determine gene expression level. Immunofluorescence was used to observe morphological alterations in tumors. RESULTS: SM22-MCs are essential for stabilizing tumor vasculature. Notch signaling was downregulated in tumor-derived SM22-MCs and vSMCs-DA treated with cancer cell-derived conditioned medium. Notch activation in SM22-MCs normalized tumor vasculature and repressed tumor growth. On the other hand, Notch disruption aggravated abnormal tumor vasculature and promoted growth and metastasis. Gene expression profiling of vSMCs-DA showed that Notch activation enhances their contractile phenotype and suppresses their secretory phenotype, further attenuating the invasion and proliferation of tumor cells. In contrast, Notch blockade in vSMCs-DA mitigated their contractile phenotype while strengthened the secretory phenotype. CONCLUSION: SM22-MCs facilitate vessel stability in tumors, and they gain a secretory phenotype and promote tumor malignancy in the absence of Notch signaling.

The Prognostic Impact of Hormonal Receptor and HER-2 Expression Discordance in Metastatic Breast Cancer Patients.

BACKGROUND: Hormone receptor (HR) and human epidermal growth factor receptor (HER2) discordance between primary and metastatic breast cancer lesions is common. However, its impact on long-term survival remains unclear. We aimed to determine the prognostic value of this discordance in patients with metastaticf breast cancer (MBC). METHODS: A total of 270 patients with MBC who were underwent re-biopsy of progressive metastases at Zhejiang Cancer Hospital from January 1, 2012 to December 31, 2015 with patients consent and then review their primary tumors pathological findings. The HR and HER2 status in both primary and progressive metastatic lesions was determined by immunohistochemistry and/or fluorescence in situ hybridization. The discordance rates were correlated with the clinicopathologic characteristics, metastatic lesions, salvage treatment, and survival analysis in this population. RESULTS: A total of 142 (52.6%) MBC patients were diagnosed with discordant HR and HER2 status. Alterations in estrogen receptor (ER), progesterone receptor (PR), and HER2 status were observed in 20.70%, 37.78%, and 11.48% cases, respectively. Chemotherapy (P=0.0192) and endocrine therapy (P=0.048) significantly affected the conversion of HR status. Endocrine therapy was positively correlated with PR discordance (P=0.002), while ER discordance was associated with adjuvant chemotherapy (P=0.031). Survival analysis showed that ER status alterations between primary and metastatic lesions were associated with overall survival (P=0.002). The clinical prognosis was significantly worse with HR losses than with persistent HR positivity (P=0.023). In Cox multivariate analysis, the loss of HR expression and conversion to triple negative were independent prognostic indicators. CONCLUSION: Discordance in HR status between primary and metastatic lesions may impact the prognosis of MBC, and HR conversion has independent prognostic value.

Endothelial Notch activation promotes neutrophil transmigration via downregulating endomucin to aggravate hepatic ischemia/reperfusion injury.

Inflammatory leukocytes infiltration is orchestrated by mechanisms involving chemokines, selectins, addressins and other adhesion molecules derived from endothelial cells (ECs), but how they respond to inflammatory cues and coordinate leukocyte transmigration remain elusive. In this study, using hepatic ischemia/reperfusion injury (HIRI) as a model, we identified that endothelial Notch activation was rapidly and dynamically induced in liver sinusoidal endothelial cells (LSECs) in acute inflammation. In mice with EC-specific Notch activation (NICeCA), HIRI induced exacerbated liver damage. Consistently, endothelial Notch activation enhanced neutrophil infiltration and tumor necrosis factor (TNF)-α expression in HIRI. Transcriptome analysis and further qRT-PCR as well as immunofluorescence indicated that endomucin (EMCN), a negative regulator of leukocyte adhesion, was downregulated in LSECs from NICeCA mice. EMCN was downregulated during HIRI in wild-type mice and in vitro cultured ECs insulted by hypoxia/re-oxygenation injury. Notch activation in ECs led to increased neutrophil adhesion and transendothelial migration, which was abrogated by EMCN overexpression in vitro. In mice deficient of RBPj, the integrative transcription factor of canonical Notch signaling, although overwhelming sinusoidal malformation aggravated HIRI, the expression of EMCN was upregulated; and pharmaceutical Notch blockade in vitro also upregulated EMCN and inhibited transendothelial migration of neutrophils. The Notch activation-exaggerated HIRI was compromised by blocking LFA-1, which mediated leukocyte adherence by associating with EMCN. Therefore, endothelial Notch signaling controls neutrophil transmigration via EMCN to modulate acute inflammation in HIRI.

An insertion mutation of ERBB2 enhances breast cancer cell growth and confers resistance to lapatinib through AKT signaling pathway.

In clinical practice, some breast cancer (BC) patients carry a rare ERBB2 in-frame insertion (p. Pro780_Tyr781insGlySerPro) and are resistant to anti-ERBB2 therapy. To explore the potential procarcinogenic role of this ERBB2 mutation, we conducted the present study using BC cells overexpressing wild-type (WT) ERBB2 or P780-Y781 ERBB2 [mutated (MT)]. MDA-MB-231 and MCF-7 cells were transfected with the following plasmids using a lentivirus system: negative control (ERBB2-NC), WT ERBB2 overexpression (ERBB2-WT), and P780-Y781 ERBB2 overexpression (ERBB2-MT). P780-Y781 ERBB2 conferred significant resistance to lapatinib, as assessed by cell viability and colony counts. Analysis of the cell cycle showed that the P780-Y781 ERBB2 group showed an elevated proportion of cells in S, G2, and M phases compared with WT ERBB2 when exposed to lapatinib. Following lapatinib treatment, phosphorylated AKT (p-AKT) was strongly upregulated in the P780-Y781 ERBB2 group. Among ERBB2+ patients, the P780-Y781 ERBB2 group showed increased levels of p-AKT. Furthermore, the AKT inhibitor perifosine effectively suppressed lapatinib resistance, as indicated by the lapatinib inhibition curve and results of the colony formation assay, and decreased AKT phosphorylation. Altogether, we discovered a procarcinogenic mutation of ERBB2 that enhances BC cell growth through AKT signaling and causes resistance to lapatinib. Patients with this in-frame insertion mutation of ERBB2 should be recommended other therapeutic strategies apart from ERBB2 tyrosine kinase inhibitors, in particular lapatinib.

Monitoring treatment efficacy and resistance in breast cancer patients via circulating tumor DNA genomic profiling.

BACKGROUND: One of the major challenges in managing invasive breast cancer (BC) is the lack of reliable biomarkers to track response. Circulating tumor DNA (ctDNA) from liquid biopsy, as a candidate biomarker, provides a valuable assessment of BC patients. In this retrospective study, we evaluated the utility of ctDNA to reflect the efficacy of treatment and to monitor resistance mechanisms. METHODS: Targeted next-generation sequencing (NGS) of 416 cancer-relevant genes was performed on 41 plasma biopsy samples of 19 HER2+ and 12 HER2- BC patients. Longitudinal ctDNA samples were analyzed in three BC patients over the treatment course for detecting acquired mutations. RESULTS: In HER2+ BC patients, ERBB2 somatic copy numbers in ctDNA samples were significantly higher in patients progressed on HER2-targeted therapy than those who were still responding to the treatment. Recurrent acquired mutations were detected in genes including ERBB2, TP53, EGFR, NF1, and SETD2, which may contribute to trastuzumab resistance. In longitudinal analyses, the observed mutation allele frequencies were tracked closely in concordance with treatment responses. A novel ERBB2 p.(Leu869Arg) mutation was acquired in one patient upon resistant to trastuzumab therapy, which was further validated as an oncogenic mutation in vitro and contributed to resistance. In HER2- BC patients with chemotherapy resistance, genetic alterations on TP53, PIK3CA, and DNA damage repair genes were frequently observed. CONCLUSIONS: In summary, ctDNA monitoring, particularly longitudinal analyses, provides valuable insights into the assessment of targeted therapy efficacy and gene alterations underlying trastuzumab resistance and chemotherapy resistance in HER2+ and HER2- BC patients, respectively.

Transmembrane protein 215 promotes angiogenesis by maintaining endothelial cell survival.

Sprouting angiogenesis is a major form of neovascularization of tissues suffering from hypoxia and other related stress. Endothelial cells (ECs) undergo proliferation, differentiation, programmed death, and migration during angiogenic sprouting, but the underlying molecular mechanisms regulating ECs in angiogenesis have been incompletely elucidated. Here we report that the transmembrane protein 215 (TMEM215) is involved in angiogenesis by regulating EC survival. The murine TMEM215 gene, which possesses two transcriptional starting sites as determined by 5'-rapid amplification of complementary DNA (cDNA) ends (RACE), encodes a two-pass TMEM. The TMEM215 transcripts were detected in ECs in addition to other tissues by quantitative reverse transcription-polymerase chain reaction. Immunofluorescence showed that TMEM215 was expressed in the vasculature in retina, liver, and tumor, and colocalized with EC markers. We show that knockdown of TMEM215 in ECs induced strong cell death of ECs in vitro without affecting cell proliferation and migration, suggesting that TMEM215 was required for EC survival. Downregulation of TMEM215 expression compromised lumen formation and sprouting capacities of ECs in vitro. Moreover, intravitreous injection of TMEM215 small interfering RNA resulted in delayed and abnormal development of retinal vasculature with poor perfusion. These results identified TMEM215 as a novel molecule involved in angiogenesis by regulating the survival of ECs.

B7-H3 is related to tumor progression in ovarian cancer.

B7-H3, a co-stimulatory molecule, has been found expressed in ovarian cancer, but its role and mechanism is not clear. In this study, we further verified the expression of B7-H3 in ovarian carcinoma and normal epithelial ovarian tissues. Three ovarian cancer cell lines, A2780, SKOV3 and HO8910 were selected to explore the effects of B7-H3 on proliferation, apoptosis, migration and invasion. We found that B7-H3 was mainly located in the cytoplasm of ovarian cancer cells as determined by immunofluorescence staining. The ability of cell invasion, migration, proliferation decreased after silencing B7-H3 whereas the apoptosis increased, which was related to the upregulation of Bax, caspase-8, cleaved caspase-8 and the downregulation of Bcl-2, Bcl-xl, matrix metalloproteinase-2 (MMP2) by western blotting. In addition, B7-H3 enhanced the H08910 cell capacities in invasion, migration and proliferation. Expression of the phosphorylation signal transducer and activator of transcription 3 (pStat3) molecules and their upstream molecules phosphorylation Janus kinase 2 (pJak2) were significantly increased. In order to investigate whether B7-H3 plays a role in this pathway, we treated the overexpressed HO8910 cells with AG490 (inhibitors of Jak2). Our findings revealed that B7-H3 affect ovarian cancer progression through the Jak2/Stat3 pathway, indicating that B7-H3 has the potential to be a useful prognostic marker.

The Notch ligand delta-like 3 promotes tumor growth and inhibits Notch signaling in lung cancer cells in mice.

Although it has been suggested that Dll3, one of the Notch ligands, promotes the proliferation and inhibits the apoptosis of cancer cells, the role of Dll3 in cancers remains unclear. In this study, we found that in the murine Lewis lung carcinoma (LLC) cells, the level of Dll3 mRNA changed upon tumor microenvironment (TME) stimulation, namely, decreased under hypoxia or stimulated with tumor necrosis factor (TNF)-α. Dll3 was also expressed at higher level in human lung carcinoma tissues than in the para-carcinoma tissues. Overexpression of Dll3 in LLC cells promoted cell proliferation and reduced apoptosis in vitro, and enhanced tumor growth when inoculated in vivo in mice. The Dll3-mediated proliferation could be due to increased Akt phosphorylation in LLC cells, because an Akt inhibitor counteracted Dll3-induced proliferation. Moreover, Dll3 overexpression promoted PI3K/Akt signaling through inhibiting Notch signaling.