Machine learning application identifies plasma markers for proteinuria in metastatic colorectal cancer patients treated with Bevacizumab.

BACKGROUND AND OBJECTIVES: Proteinuria is a common complication after the application of bevacizumab therapy in patients with metastatic colorectal cancer, and severe proteinuria can lead to discontinuation of the drug. There is a lack of sophisticated means to predict bevacizumab-induced proteinuria, so the present study aims to predict bevacizumab-induced proteinuria using peripheral venous blood samples. METHODS: A total of 122 subjects were enrolled and underwent pre-treatment plasma markers, and we followed them for six months with proteinuria as the endpoint event. We then analyzed the clinical features and plasma markers for grade ≥ 2 proteinuria occurrence using machine learning to construct a model with predictive utility. RESULTS: One hundred sixteen subjects were included in the statistical analysis. We found that high baseline systolic blood pressure, low baseline HGF, high baseline ET1, high baseline MMP2, and high baseline ACE1 were risk factors for the development of grade ≥ 2 proteinuria in patients with metastatic colorectal cancer who received bevacizumab. Then, we constructed a support vector machine model with a sensitivity of 0.889, a specificity of 0.918, a precision of 0.615, and an F1 score of 0.727. CONCLUSION: We constructed a machine learning model for predicting grade ≥ 2 bevacizumab-induced proteinuria, which may provide proteinuria risk assessment for applying bevacizumab in patients with metastatic colorectal cancer.

Efficacy of roxadustat in maintenance hemodialysis patients with erythropoietin-hyporesponsive anemia.

OBJECTIVE: To investigate the efficacy of roxadustat in hemodialysis patients with erythropoietin (EPO) hypo-responsive anemia. MATERIALS AND METHODS: This retrospective study included 55 hemodialysis patients with erythropoietin hypo-responsive anemia at the First Affiliated Hospital of Chongqing Medical University from January to December 2020. We observed their hemoglobin (Hb) changes, inflammatory factors, and adverse reactions before and after 12 weeks of roxadustat treatment. RESULTS: Among the 55 patients, the average age was 60.75 ± 13.96 years old, and the median dialysis age was 48 months. All patients were taken off EPO and switched to roxadustat during the follow-up period. Compared with baseline, patients' Hb was significantly increased (90.64 ± 20.01 g/L, 98.52 ± 15.89 g/L, 104.34 ± 19.15 g/L, and 107.02 ± 20.54 g/L, respectively) (p < 0.05). At 12 weeks of roxadustat treatment, 34 patients (61.82%) met the target Hb levels (100 - 130 g/L). The multivariate logistic analysis showed that Hb response positively correlated with the Hb level before roxadustat treatment (p = 0.046), while responding well to roxadustat negatively correlated with blood platelet-to-lymphocyte ratio (PLR) and duration of dialysis (p = 0.029 and p = 0.046) in patients with EPO hypo-responsive anemia. CONCLUSION: Roxadustat could effectively improve anemia; the PLR and dialysis age were independent predictors of roxadustat efficacy in dialysis patients with EPO hypo-responsive anemia.

Analysis of the evolution trends and influential factors of bankfull discharge in the Lower Yellow River.

After the Xiaolangdi Reservoir (XLDR) was put into operation, new water and sediment conditions have improved the silting condition of the Lower Yellow River (LYR), but against this background, the flow capacity of the LYR has become more complex. In this research, the measured water and sediment data of seven hydrological stations in the LYR from 1950 to 2020 were systematically collated. The evolution trend and development period of the bankfull discharge in this area were studied based on the wavelet analysis method, and the main factors influencing the evolution of the bankfull discharge were explored. The results indicate that the evolution process of bankfull discharge in the LYR has experienced two phases in the last 70 years. XLDR has been impounded since October 1999. Before XLDR operation, the bankfull discharge of the LYR has a main time scale of about 26 years. After XLDR operation began, it has a time scale of about 10 years. The bankfull discharge of the LYR shows two phases of evolution, and these phases are mainly influenced by the factors of water and sediment conditions. This research is needed for a deeper understanding of flow-bed and river discharge and sediment transport capacity under the water and sediment conditions in the LYR.

Efficacy and safety of neoadjuvant pertuzumab plus trastuzumab in combination with chemotherapy regimen in Chinese patients with HER2-positive early breast cancer: a real-world retrospective multi-center cohort study.

Background: Pertuzumab plus trastuzumab combined with chemotherapy has become a standard neoadjuvant therapy option for patients with high-risk human epidermal growth factor receptor 2 (HER2)-positive breast cancer (BC). There is still not enough evidence for the efficacy and safety of neoadjuvant pertuzumab and trastuzumab plus chemotherapy in HER2-positive BC patients in China, both in clinical trials and real-world settings. This study aimed to assess the efficacy and safety of neoadjuvant pertuzumab plus trastuzumab in combination with chemotherapy in Chinese patients with HER2-positive BC in real-world clinical application. Methods: We retrospectively collected the data from the electronic medical records of HER2-positive patients treated with neoadjuvant trastuzumab and pertuzumab plus chemotherapy from December 2018 to May 2021 at 21 hospitals located in Hunan Province, China, including age, American Joint Committee on Cancer (AJCC) stage, clinical tumor size, clinical lymph node status, pathological characteristics (before neoadjuvant systemic therapy), treatment approach, adverse events to neoadjuvant therapy, and achievement of pathological complete response (pCR). The primary endpoint was the total rate of pCR, and the secondary endpoints were the rate of pCR of each subgroup and the safety of dual anti-HER2 therapy. Results: A total of 188 patients met the inclusion criteria and were included in the analysis. Of the 188 patients, 119 (63.3%) were diagnosed at stage II and 64 (34.0%) at stage III; 163 (86.7%) were cT2-3; 149 patients (79.3%) were ≥ cN1; 84 patients (44.7%) were hormone receptor (HR)-positive. pCR was observed in 88 of 188 patients (46.8%). The pCR rate of HR-negative patients (54.8%) was higher (P=0.014) than that of HR-positive patients (36.9%). Patients with Ki-67 <15% achieved a higher (P=0.033) pCR rate (68.2%) than those with Ki-67 ≥15% (44.0%). Anemia was the most common adverse event (63.4%), and the most common grade 3-4 adverse event was nausea and vomiting (8.5%). Conclusions: Our study confirmed the benefit of neoadjuvant pertuzumab plus trastuzumab in combination with chemotherapy on pCR with a tolerable safety profile in routine clinical practice in Chinese patients with HER2-positive BC. HR-negativity and Ki-67 <15% were associated with pCR in these patients.

Ambrosin sesquiterpene lactone exerts selective and potent anticancer effects in drug-resistant human breast cancer cells (MDA-MB-231) through mitochondrial mediated apoptosis, ROS generation and targeting Akt/ß-Catenin signaling pathway.

PURPOSE: Breast cancer accounts for a significant proportion of cancer burden among women world over. Concerning breast cancer treatment, there are only few chemotherapeutic agents available, which also have serious side effects. The present study was thus designed to explore in vitro the antitumor effects of ambrosin sesquiterpene lactone against human drug-resistant breast cancer cells (MDA-MB-231). METHODS: WST-1 assay was used to determine cell viability. The fact that ambrosin induced apoptosis was studied through acridine orange (AO)/ethidium bromide (EB) staining using fluorescence microscopy as well as using flow cytometry in association with annexin-v/propidium iodide (PI) staining. Furthermore, western blot assay was used to study effects of ambrosin on apoptosis-related protein expressions including Bax and Bcl-2, as well as to study the effects on numerous caspases and Akt/ß-Catenin Signaling Pathway. The effects on reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) were evaluated by flow cytometry. RESULTS: The results showed that ambrosin with an IC50 value of 25 µM decreased the viability of the MDA-MB-231 cells. The cytotoxicity of ambrosin was also investigated on the MCF-12A normal breast cells which showed that it exerted very low toxic effects on these cells. Ambrosin also caused remarkable changes in the morphology and suppressed the colony forming potential of MDA-MB-231 cells. The AO/EB staining assay showed that ambrosin inhibits the viability of cancer cells via induction of apoptotic cell death which was associated with increase in Bax and reduction in Bcl-2 levels. The apoptotic cells increased from 3.5% in the controls to around 56% at 50 µM concentration in the MDA-MB-231 cells. It was also seen that ambrosin treatment to these cancer cells resulted in substantial suppression in MMP and remarkable rise in ROS in a dose-dependent manner. This molecule also significantly inhibited the Akt/ß-catenin signalling pathway by reducing the expressions of phosphorylated GSK-3ß and Akt. CONCLUSIONS: Taken all together, the results of our study indicate that ambrosin sesquiterpene may be developed as a promising anticancer agent in human breast cancer provided further in-depth studies are performed.

Expression of glucose-regulated protein 78 as prognostic biomarkers for triple-negative breast cancer.

INTRODUCTION: Glucose-regulated protein78(GRP78) is a stress - induced endoplasmic reticulum chaperone protein. it is closely related to the occurrence, development, proliferation, differentiation and drug resistance of breast cancer. However, the association and clinicopathological features between GRP78 and triple negative breast cancer (TNBC) remain to be studied. MATERIAL AND METHODS: Clinical and pathological characteristics and overall survival were analysed retrospectively in 179 surgically resected TNBC patients. GRP78 was detected by immunohistochemistry (IHC) using breast cancer tissue microarrays (TMAs), and the association between GRP78 levels and clinicopathological factors and prognosis was analyzed. Furthermore, GRP78 expression in human TNBC and NTNBC cell lines was detected by Western blot and qRT-PCR. After Si-GRP78 knocked-down GRP78 in MDA-MB-231 and BT549 cell lines, cell proliferation was detected using Cell Counting Kit-8 (CCK-8) and cell colony formation was detected by crystal violet staining, respectively. RESULTS: GRP78 was expressed in triple negative breast cancer (TNBC). GRP78 expression was significantly associated with invasive, distant metastasis and proliferation of TNBC (P<0.05). In addition, patients with positive GRP78 expression had shorter overall survival (OS) and disease-free survival (DFS). And the high expression of GRP78 was significantly associated with disease-free survival (DFS) in patients with TNBC (P<0.001). CONCLUSIONS: These findings improve our understanding of the expression pattern of GRP78 in TNBC and clarify the role of GRP78 as promising prognostic biomarkers for triple-negative breast cancer.

Inhibition of cancer cell growth by Tangeretin flavone in drug-resistant MDA-MB-231 human breast carcinoma cells is facilitated via targeting cell apoptosis, cell cycle phase distribution, cell invasion and activation of numerous Caspases.

PURPOSE: In this study, the anticancer effects of a natural flavonoid-Tangeretin, were examined against the drug-resistant MDA-MB-231 breast cancer (BC) cell line and the normal breast cell line Hs 841.T. METHODS: The MTT assay was employed for cell viability determination. Apoptosis was demonstrated by DAPI and Annexin V/propidium iodide (PI) staining. Flow cytometric analyses were performed to gain insights about cell cycle distribution. Western blot assay was used for protein expression determination. RESULTS: Tangeretin inhibited the growth of the drug-resistant MDA-MB-231 cells concentration-dependently and its IC50 was 9 µM, whereas the IC50 was >100 µM against the normal cells. The anti-proliferative effects were due to induction of apoptotic cell death. The apoptotic cell percentage was increased from 5.7% to around 69% as the concentration of Tangeretin was increased. Tangeretin also caused an increase in the Bax/Bcl-2 ratio and activation of the Caspase 3, 8 and 9. In addition, Tangeretin led to arrest of the cells at G2/M phase which was accompanied by depletion of cyclin B1 and D. Transwell assay showed that Tangeretin also reduced the invasion of the MDA-MB-231 cells. CONCLUSION: The findings of this study suggest that Tangeretin exerts potent anticancer effects on the MDA-MB-231 cells and may therefore prove a beneficial lead molecule in BC research.

Therapeutic activity of DCC-2036, a novel tyrosine kinase inhibitor, against triple-negative breast cancer patient-derived xenografts by targeting AXL/MET.

Triple-negative breast cancer (TNBC) is insensitive to endocrine therapies and targeted therapies to human epidermal growth factor receptor-2 (HER2), estrogen receptor (ER) and progesterone receptor (PR). New targets and new targeted therapeutic drugs for TNBC are desperately needed. Our study confirmed that DCC-2036 inhibited the proliferation, invasion, migration and epithelial-mesenchymal transition (EMT) of TNBC cells as well as induced apoptosis. Moreover, the antiproliferative activity of DCC-2036 was more efficient than that of most clinical drugs. In addition, the combination of DCC-2036 and cisplatin or lapatinib had synergistic effects on TNBC cells. Mechanistically, DCC-2036 targeted AXL/MET, especially AXL, and regulated the downstream PI3K/Akt-NFκB signaling to exert its antitumor effect in TNBC. DCC-2036 also inhibited the growth and metastasis of xenografted MDA-MB-231 cells (AXL/MET-high TNBC cells) but not MDA-MB-468 cells (AXL-low TNBC cells) in NSG mice in vivo. Furthermore, DCC-2036 significantly inhibited tumor growth and invasion of AXL/MET-high TNBC PDX tumors but not AXL/MET-low TNBC PDX tumors. These results highlighted the roles of AXL/MET in cancer growth and metastasis and further verified that the critical targets of DCC-2036 are AXL and MET, especially AXL. In addition, there was no significant toxicity of DCC-2036 even at a high dosage. Therefore, DCC-2036 may be a potential compound to treat TNBC, especially for tumors with AXL/MET overexpression.

Role of TXNDC5 in tumorigenesis of colorectal cancer cells: In vivo and in vitro evidence.

Thioredoxin domain­containing 5 (TXNDC5) is reportedly overexpressed in colorectal cancer (CRC) and is therefore considered an oncogene. However, the role of TXNDC5 in CRC tumorigenesis remains unclear. The present study aimed to explore the role of TXNDC5 in CRC tumorigenesis in vitro and in vivo under hypoxic and normoxic conditions. Analyses of patient tissue samples revealed a positive association between the expression of hypoxia­inducible factor­1α (HIF­1α) or TXNDC5 and the TNM stage of CRC. In addition, a positive correlation between the expression levels of HIF­1α and TXNDC5 was observed in CRC tissues. Furthermore, culturing RKO and HCT­116 human CRC cell lines under hypoxic conditions significantly increased the expression levels of HIF­1α and TXNDC5, whereas knockdown of HIF­1α abolished the hypoxia­induced expression of TXNDC5. Knockdown of TXNDC5 significantly decreased cell proliferation and colony formation, and incre-ased apoptosis of both cell lines. Furthermore, knockdown of TXNDC5 markedly increased hypoxia­induced reactive oxygen species (ROS) generation, and the expression of hypoxia­induced endoplasmic reticulum stress (ER) markers (CCAAT­enhancer­binding protein homologous protein, glucose­regulated protein 78 and activating transcription factor 4) and apoptotic markers (B­cell lymphoma 2­associated X protein and cleaved caspase­8). In addition, the expression levels of TXNDC5 were significantly increased in tumor tissues compared with in adenoma and normal tissues in a mouse model of CRC tumorigenesis. In conclusion, the in vivo data demonstrated that TXNDC5 is overexpressed in CRC tissues, and this overexpression may be associated with unfavorable clinicopathological features. The in vitro data indicated that hypoxia may induce TXNDC5 expression via upregulating HIF­1α; this effect promoted CRC cell proliferation and survival under hypoxic conditions, likely via inhibiting hypoxia­induced ROS/ER stress signaling. These findings suggested that TXNDC5 functions as an important stress survival factor to maintain tumorigenesis of CRC cells under hypoxia by regulating hypoxia­induced ROS/ER stress signaling. The present study provided novel insights into the role of TXNDC5 in the tumorigenesis of CRC.

Serum miR-20a is a promising biomarker for gastric cancer.

Gastric cancer (GC) is a common type of cancer, particularly in China. Numerous studies have demonstrated that circulating microRNAs (miRNAs) have potential applications as noninvasive biomarkers for cancer diagnosis and prognosis. Microarray-based serum miRNA profiling was performed on the serum of 12 paired pre- and post-operative GC patients to screen differentially expressed serum miRNAs. Twelve different serum miRNAs between pre- and post-operative GC patients were identified. Those miRNAs were verified by real-time quantitative polymerase chain reaction in 110 paired pre- and post-operative serum samples from 55 GC patients. miR-20a was confirmed and demonstrated potential as a GC-associated biomarker. Furthermore, the levels of serum miR-20a were significantly different between GC, nasopharyngeal cancer, colorectal carcinoma, breast cancer and non-cancerous controls. In addition, it was found that serum miR-20a levels correlated with age, tumor stage, differentiated degree and lymph node metastasis in GC. Survival analysis indicated that GC patients with elevated levels of serum miR-20a had poor survival. Thus, serum miR-20a may serve as a molecular marker for diagnosis, evaluating therapeutic efficacy and prognosis, as well as monitoring recurrence in GC patients.

Heterogeneous nuclear ribonucleoprotein K is overexpressed and associated with poor prognosis in gastric cancer.

Heterogeneous nuclear ribonucleoprotein K (hnRNP K) is one of the major pre-mRNA-binding proteins, that is involved in translational modifications. In our previous studies, we found that hnRNP K is associated with human gastric cancer. The protein levels of hnRNP K were detected in cell lines and tissue microarrays. The correlation between hnRNP K expression and patient survival rate was evaluated by Kaplan-Meier survival analysis. In addition, we also detected hnRNP K expression in preoperative and postoperative serum samples from patients with gastric cancer, and serum samples from healthy volunteers. We found that hnRNP K was overexpressed in the gastric cancer cell lines. The levels of hnRNP K were significantly elevated in the gastric cancer tissues compared with that noted in the tumor-adjacent gastric mucosal and normal gastric mucosal sampes, and hnRNP K expression was found to correlate with tumor stage and lymph node metastasis. However, the level of serum hnRNP K did not differ significantly between gastric cancer patients and healthy volunteers. We also found that patients whose tumors showed elevated expression of hnRNP K had poor survival. The present study suggests that hnRNP K is a promising tissue biomarker for diagnosing gastric cancer and is a prognostic indicator for patients with gastric cancer.

Hypoxia regulates stemness of breast cancer MDA-MB-231 cells.

Human breast cancers include cancer stem cell populations as well as non-tumorigenic cancer cells. Breast cancer stem cells possess self-renewal capability and thus are the root cause of recurrence and metastasis of malignant tumors. Hypoxia is a fundamental pathological feature of solid tumor tissues and exerts a wide range of effects on the biological behavior of cancer cells. However, there is little information on the role of hypoxia in modulating the stemness of breast cancer cells. In the present study, we cultured MDA-MB-231 cells in a hypoxic gas mixture to simulate the hypoxic environment in tissues and to determine how hypoxia conditions could affect the cell proliferation, apoptosis, cytotoxicity, and colony-forming ability. Expression of the stem cell phenotype CD24(-)CD44(+)ESA(+) was analyzed to assess the effects of hypoxia on stemness transformation in MDA-MB-231 cells. Our results found that the cell toxicity of MDA-MB-231 cells was not affected by hypoxia. Hypoxia could slightly inhibit the growth of MDA-MB-231 cells, but the inhibitory effect is not significant when compared with normoxic control. Moreover, hypoxia significantly blocked the apoptosis in MDA-MB-231 cells (P < 0.05). The proportion of CD24(-)CD44(+)ESA(+) cells in MDA-MB-231 cells was increased greatly after they were treated with hypoxia, and cell colony-formation rate of MDA-MB-231 cells also increased significantly in hypoxia-treated cells. These results encourage the exploration of hypoxia as a mechanism which might not be underestimated in chemo-resistant breast cancer treatment.

Neuron navigator 2 overexpression indicates poor prognosis of colorectal cancer and promotes invasion through the SSH1L/cofilin-1 pathway.

BACKGROUND: Neuron navigator 2 (NAV2) encodes a member of the neuron navigator gene family, which plays a role in tumorigenesis and cell migration. However, the prognostic value of NAV2 expression in colorectal cancer (CRC) patients and the potential pathway through which NAV2 promotes migration and invasion in CRC cell lines is poorly understood. METHODS: The expression level of NAV2 was detected in CRC tissues from two different CRC cohorts by immunohistochemistry, qRT-PCR and Western blotting; the correlation between NAV2 expression and clinicopathological characters was analyzed, and the prognostic value of NAV2 expression was analyzed using a Cox regression model. CRC cell lines with NAV2 knocked out were used to validate the function and potential pathway used by NAV2 to promote CRC cell migration and invasion. RESULTS: The results showed that NAV2 was overexpressed in CRC tissues, and it was closely correlated with depth of invasion, and lymph and distant metastasis. Multivariate analysis indicated that high NAV2 expression was a poor prognostic indicator of recurrence-free survival and overall survival in CRC patients. Furthermore, Cox regression analysis revealed that high NAV2 expression integrated with high tumor budding grade was a powerful independent predictive factor of CRC clinical outcome. In vitro and in vivo assays demonstrated that knockdown of NAV2 led to reduced migration and invasion of cancer cells, and the process involved the regulation of F-actin polymerization through the SSH1L/cofilin-1 pathway. CONCLUSION: Based on these findings, NAV2 could serve as both a prognostic biomarker and a potential therapeutic target for patients with NAV2-positive CRC.

U6 is not a suitable endogenous control for the quantification of circulating microRNAs.

Recently, microRNAs have been detected in serum and plasma, and circulating microRNA (miRNA) profiles have now been associated with many diseases such as cancers and heart disease, as well as altered physiological states. Because of their stability and disease resistance, circulation miRNAs appear to be an ideal material for biomarkers of diseases and physiological states in blood. However, the lack of a suitable internal reference gene (internal reference miRNA) has hampered research and application of circulating miRNAs. Currently, U6 and miR-16 are the most common endogenous controls in the research of miRNAs in tissues and cells. We performed microarray-based serum miRNA profiling on the serum of 20 nasopharyngeal carcinoma patients and 20 controls to detect the expressions of U6 and miRNAs. Profiling was followed by real-time quantitative Polymerase Chain Reaction (qPCR) in 80 patients (20 each with gastric cancer, nasopharyngeal carcinoma, colorectal cancer, and breast cancer) and 30 non-cancerous controls. qPCR was also performed to detect miRNAs in serum with repeated freezing and thawing. The results of microarray showed that with the exception of U6, Ct values of miR-16, miR-24, miR-142-3p, miR-19b and miR-192 in serum samples of nasopharyngeal carcinoma were greater than control samples. The results of 110 cases showed large fluctuations in U6 expression. The difference between the greatest and the least levels of expression was 3.29 for delta Ct values, and 1.23 for miR-16. The expressions of U6, miR-16 and miR-24 in serum subjected to different freeze-thaw cycles showed that U6 expression gradually decreased after 1, 2, and 4 cycles of freezing and thawing, while the expression of miR-16 and miR-24 remained relatively stable. Collectively, our results suggested that U6 is unsuitable as an internal reference gene in the research of circulating miRNAs.