Investigation of correlation between shear wave elastography and lymphangiogenesis in invasive breast cancer and diagnosis of axillary lymph node metastasis.

BACKGROUND: Accurate evaluation of axillary lymph node metastasis (LNM) in breast cancer is very important. A large number of hyperplastic and dilated lymphangiogenesis cases can usually be found in the pericancerous tissue of breast cancer to promote the occurrence of tumor metastasis.Shear wave elastography (SWE) can be used as an important means for evaluating pericancerous stiffness. We determined the stiffness of the pericancerous by SWE to diagnose LNM and lymphangiogenesis in invasive breast cancer (IBC). METHODS: Patients with clinical T1-T2 stage IBC who received surgical treatment in our hospital from June 2020 to December 2020 were retrospectively enrolled. A total of 299 patients were eventually included in the preliminary study, which included an investigation of clinicopathological features, ultrasonic characteristics, and SWE parameters. Multivariable logistic regression analysis was used to establish diagnostic model and evaluated its diagnostic performance of LNM. The correlation among SWE values, collagen volume fraction (CVF), and microlymphatic density (MLD) in primary breast cancer lesions was analyzed in another 97 patients. RESULTS: The logistic regression model is Logit(P)=-1.878 + 0.992*LVI-2.010*posterior feature enhancement + 1.230*posterior feature shadowing + 0.102*posterior feature combined pattern + 0.009*Emax. The optimum cutoff value of the logistic regression model was 0.365, and the AUC (95% CI) was 0.697 (0.636-0.758); the sensitivity (70.7 vs. 54.3), positive predictive value (PPV) (54.0 vs. 50.8), negative predictive value (NPV) (76.9 vs. 69.7), and accuracy (65.2 vs. 61.9) were all higher than Emax. There was no correlation between the SWE parameters and MLD in primary breast cancer lesions. CONCLUSIONS: The logistic regression model can help us to determine LNM, thus providing more imaging basis for the selection of preoperative treatment. The SWE parameter of the primary breast cancer lesion cannot reflect the peritumoral lymphangiogenesis, and we still need to find a new ultrasonic imaging method.

Ginsenoside Rg3 overcomes tamoxifen resistance through inhibiting glycolysis in breast cancer cells.

Tamoxifen (TAM) resistance poses a significant clinical challenge in human breast cancer and exhibits high heterogeneity among different patients. Rg3, an original ginsenoside known to inhibit tumor growth, has shown potential for enhancing TAM sensitivity in breast cancer cells. However, the specific role and underlying mechanisms of Rg3 in this context remain unclear. Aerobic glycolysis, a metabolic process, has been implicated in chemotherapeutic resistance. In this study, we demonstrate that elevated glycolysis plays a central role in TAM resistance and can be effectively targeted and overcome by Rg3. Mechanistically, we observed upregulation of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3), a key mediator of glycolysis, in TAM-resistant MCF-7/TamR and T-47D/TamR cells. Crucially, PFKFB3 is indispensable for the synergistic effect of TAM and Rg3 combination therapy, which suppresses cell proliferation and glycolysis in MCF-7/TamR and T-47D/TamR cells, both in vitro and in vivo. Moreover, overexpression of PFKFB3 in MCF-7 cells mimicked the TAM resistance phenotype. Importantly, combination treatment significantly reduced TAM-resistant MCF-7 cell proliferation in an in vivo model. In conclusion, this study highlights the contribution of Rg3 in enhancing the therapeutic efficacy of TAM in breast cancer, and suggests that targeting TAM-resistant PFKFB3 overexpression may represent a promising strategy to improve the response to combination therapy in breast cancer.

Prediction of Axillary Lymph Node Metastasis in Invasive Breast Cancer by Sound Touch Elastography.

The aims of this study were to investigate the value of sound touch elastography (STE) in predicting axillary lymph node metastasis (ALNM) in patients with invasive breast cancer (IBC) and to explore whether lysyl oxidase (LOX) is correlated with increasing stiffness and promotion of metastasis in IBC. A total of 142 lesions in 142 patients were assessed by STE. The STE values of IBCs in the two groups were compared and the best cutoff values for diagnosing ALNM determined. Immunohistochemistry was used to detect LOX expression. Collagen fiber and elastic fiber content was determined by Masson and Weigert elastic fiber staining. Correlation analyses were performed to identify the associations of the data. The optimal cutoff values of Emax (maximum stiffness value of the tumor) and Smax (maximum stiffness value of the shell) for predicting ALNM of IBC were 94.58 and 148.78 kPa. Immunohistochemistry and Masson and Weigert elastic fiber staining were performed on 67 samples. LOX expression and collagen volume fraction were significantly higher in the ALNM+ group than in the ALNM- group (p = 0.04 and 0.03), except for elastic fiber content (p = 0.628). Moreover, Emax, Smax and LOX expression were positively correlated with collagen volume fraction (r = 0.624, 0.512, and 0.533, respectively). Emax and Smax were found to be predictors for ALNM of IBC. STE could serve as a non-invasive method for assessing lymph node status before surgery. Overexpression of LOX and increased collagen fiber contributed to the increased stiffness in the lesions and metastases of IBC.

miR-4478 sensitizes ovarian cancer cells to irradiation by inhibiting Fus and attenuating autophagy.

Ovarian cancer (OC) is a type of cancer with high prevalence and shocking mortality in women around the world. Radioresistance is a major reason for OC relapse. Mounting studies have shown the significant function of dysregulated microRNAs (miRNAs) in cancer progression and the cellular response to irradiation. The present study inquired about the function and mechanism of microRNA (miR)-4478 in regulating radiosensitivity of OC cells. Results showed that miR-4478 was downregulated in OC, and a low miR-4478 level indicated a disappointing prognosis for OC patients. Besides, in OC cells exposed to irradiation, the expression of miR-4478 decreased over time. Functionally, the upregulation of miR-4478 retarded OC cell proliferation and sensitized OC cells to irradiation. Mechanistically, miR-4478 targeted and inhibited fused in sarcoma (Fus). Additionally, Fus was upregulated in OC and its expression further elevated in OC cells under irradiation. Furthermore, miR-4478 targeted Fus to inhibit autophagy, therefore sensitizing OC cells to irradiation. Collectively, our study uncovered miR-4478 as a novel radiosensitizer by targeting Fus in OC cells, which may shed a new light on developing targets for treating patients with OC, particularly those with radioresistance.

Dynamics of angiogenesis and cellularity in rabbit VX2 tumors using contrast-enhanced magnetic resonance imaging and diffusion-weighted imaging.

A number of studies have demonstrated that dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) may be used to evaluate microvessel density (MVD), and may quantitatively reflect tumor angiogenesis. To investigate the dynamics, including angiogenesis and tumor cellularity, of rabbit VX2 tumors during the 4 weeks following tumor implantation, the present study used DCE-MRI combined with diffusion-weighted imaging (DWI) to scan the tumors at 3 days, and then at 1, 2, 3 and 4-week intervals, following tumor implantation. The dynamics, volume transfer coefficient (Ktrans) and apparent diffusion coefficient (ADC) of the tumor parenchyma were analyzed. Furthermore, the associations between Ktrans and MVD at 4 weeks after tumor implantation were analyzed. Tumor Ktrans was positively correlated with MVD at 4 weeks (r=0.674, P<0.001). Following tumor implantation, the tumor Ktrans level rose for 2 weeks and then began to decline, reaching its lowest point at 4 weeks (P<0.001). ADC values at 1 week were higher than at 3 days, but declined thereafter (P<0.001). Tumor necrosis appeared by 1 week after tumor implantation. The necrosis degree of tumor was gradually increased from the occurrence of necrosis within the 4-week time span of the present study (1 vs. 2 weeks, P=0.008; 2 vs. 3 weeks, P<0.001; 3 vs. 4 weeks, P<0.001). The present study identified that tumor angiogenesis is a dynamic process that serves a function in tumor growth, and that DCE-MRI may reflect tumor parenchymal MVD and be useful in evaluating angiogenesis.

Proteomic Profiling of Paclitaxel Treated Cells Identifies a Novel Mechanism of Drug Resistance Mediated by PDCD4.

Paclitaxel (PTX) is a widely used chemotherapeutic drug effective against numerous cancers. To elucidate cellular pathways targeted by PTX and identify novel mechanisms of PTX resistance, we used a SILAC based quantitative proteomic approach to evaluate global changes of cellular protein abundance in HeLa cells. We identified 347 proteins involved in a number of biological processes including spindle assembly, mitotic exit, and extracellular adhesion whose abundance changes upon PTX treatment. Notably, the tumor suppressor PDCD4 involved in translation suppression was down-regulated by PTX. We demonstrated that PDCD4 is a cell-cycle regulated protein and that changes in its abundance are sufficient to alter PTX sensitivity in multiple human cancer cell lines. Immunoprecipitation of PDCD4-RNA complexes and RT-PCR revealed that PDCD4 mediated PTX sensitivity acts through its interaction with mRNA of UBE2S, a ubiquitin K11 linkage conjugating enzyme critical for mitotic exit. Lastly, high levels of PDCD4 in lung cancer tissues are positively correlated with the longer overall survival time of the examined lung cancer patients with PTX involved adjuvant therapy. Therefore, our proteomic screen for paclitaxel targets not only provided novel insight into the cellular resistance to paclitaxel via the PDCD4-mitotic exit regulation axis, but also offered a predictive biomarker for paclitaxel-based personalized chemotherapy in the treatment of lung cancer.

An ShRNA Based Genetic Screen Identified Sesn2 as a Potential Tumor Suppressor in Lung Cancer via Suppression of Akt-mTOR-p70S6K Signaling.

BACKGROUND: Lung cancer is emerging rapidly as the leading death cause in Chinese cancer patients. The causal factors for Chinese lung cancer development remain largely unclear. Here we employed an shRNA library-based loss-of-function screen in a genome-wide and unbiased manner to interrogate potential tumor suppressor candidates in the immortalized human lung epithelial cell line BEAS-2B. METHODS/RESULTS: Soft agar assays were conducted for screening BEAS-2B cells infected with the retroviral shRNA library with the acquired feature of anchorage-independent growth, large (>0.5mm in diameter) and well-separated colonies were isolated for proliferation. PCRs were performed to amplify the integrated shRNA fragment from individual genomic DNA extracted from each colony, and each PCR product is submitted for DNA sequencing to reveal the integrated shRNA and its target gene. A total of 6 candidate transformation suppressors including INPP4B, Sesn2, TIAR, ACRC, Nup210, LMTK3 were identified. We validated Sesn2 as the candidate of lung cancer tumor suppressor. Knockdown of Sesn2 by an shRNA targeting 3' UTR of Sesn2 transcript potently stimulated the proliferation and malignant transformation of lung bronchial epithelial cell BEAS-2B via activation of Akt-mTOR-p70S6K signaling, whereas ectopic expression of Sens2 re-suppressed the malignant transformation elicited by the Sesn2 shRNA. Moreover, knockdown of Sesn2 in BEAS-2B cells promoted the BEAS-2B cell-transplanted xenograft tumor growth in nude mice. Lastly, DNA sequencing indicated mutations of Sesn2 gene are rare, the protein levels of Sesn2 of 77 Chinese lung cancer patients varies greatly compared to their adjacent normal tissues, and the low expression level of Sesn2 associates with the poor survival in these examined patients by Kaplan Meier analysis. CONCLUSIONS: Our shRNA-based screen has demonstrated Sesn2 is a potential tumor suppressor in lung epithelial cells. The expression level of Sesn2 may serve as a prognostic marker for Chinese lung cancer patients in the clinic.

In vitro stimulation of calcium overload and apoptosis by sonodynamic therapy combined with hematoporphyrin monomethyl ether in C6 glioma cells.

The present study investigated enhancement of apoptosis induction and the mechanisms underlying calcium overload on C6 glioma cells in vitro, stimulated by low-level ultrasound in combination with hematoporphyrin monomethyl ether (HMME). The optimum frequency of ultrasound was determined by 3-(4,5-dimethythiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The apoptotic rate, reactive oxygen species concentration and decreased mitochondrial membrane potential (MMP) were analyzed by flow cytometry. Morphological changes were detected by a transmission electron microscope, and the concentration of intracellular Ca2+, [Ca2+]i, was detected by a confocal laser scanning microscope. In addition, the release of cytochrome c (cyt-c) was measured by western blot analysis. The results revealed that an increased apoptotic effect was induced by sonodynamic therapy (SDT), and this was found to correlate with the overloaded [Ca2+]i, derived from the intra- and extracellular sources in the early apoptotic process. The results also revealed an increased level of ROS production, a decreased MMP and an increased release of cyt-c. The present study indicated that low-level ultrasound in combination with HMME improved the apoptotic effect in C6 glioma cells. The overloaded [Ca2+]i was involved in the mechanism by which apoptosis was stimulated and enhanced by SDT.

Associations between mammography and ultrasound imaging features and molecular characteristics of triple-negative breast cancer.

BACKGROUND: The triple-negative breast cancer (TNBC) is an aggressive cancer characterized by the absence of estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2). Preoperative mammography and ultrasound features of TNBC may potentially suggest characteristics of the disease and assist in treatment decisions. MATERIALS AND METHODS: The study covered 153 patients with TNBC from May 2011 to May 2012 who were confirmed by postoperative pathology results in our hospital. We compared the radiological findings among the patients and sought to determine the significant iconographic features. The biomarkers p53 and Ki-67 are regarded as significant factors in TNBC. They were therefore used to divide the TNBC into four groups for assessment of relationships with TNBC imaging features. RESULTS: On mammography, most TNBCs exhibit obscure (44.3%) masses. On ultrasound, the majority of masses (95.4%) were predominantly indistinct (50.7%), irregular (76.0%) or featuring posterior echo enhancement/shadowing. Color Doppler flow imaging (CDFI) emphasized hypervascular (32.9%) masses. Differences in CDFI by ultrasound among the four groups were statistically significant (p=0.009). There were obvious differences in the percentages of spiculated margin (p=0.049) and intensive posterior echo (p=0.006) with spotty flow imaging by ultrasound between the Ki-67 (+) p53 (+) and other groups. CONCLUSIONS: A combination of mammography and ultrasound revealed the imaging characteristics of TNBC included an obscure mass with less attenuated posterior echoes and some vascularity. A worse prognosis was associated with spiculated margin and intensive posterior echoes with spotty flow imaging.

Apoptotic effect of sonodynamic therapy mediated by hematoporphyrin monomethyl ether on C6 glioma cells in vitro.

PURPOSE: Sonodynamic therapy (SDT) is a new method for treating cancer by inducing cell necrosis and/or apoptosis. However, the molecular mechanisms of cell apoptosis after SDT treatment remain unclear. In this study we investigated the apoptotic effect and mechanisms of SDT mediated by hematoporphyrin monomethyl ether (HMME) on C6 glioma cells in vitro. METHODS: The time of ultrasound irradiation was optimized by MTT. The apoptotic rate, generation of ROS (reactive oxygen species), and mitochondrial membrane potential (MMP) were detected by flow cytometry, and expressions of apoptosis proteins were measured by Western blot after SDT treatment. RESULTS: The results displayed that treatment of C6 glioma cells with SDT resulted in the occurrence of apoptosis, which was associated with the production of ROS and loss of MMP. The results also showed that protein expression of Bax, caspase-9, and caspase-3 significantly increased, protein expression of Bcl-2 and Fas-L significantly decreased, and Fas protein expression was unchanged after SDT treatment. CONCLUSIONS: Our results suggested that the mitochondrial signal pathway may play a pivotal role in the apoptosis induced by SDT in C6 glioma cells.