In Vitro Synergistic Inhibition of HT-29 Proliferation and 2H-11 and HUVEC Tubulogenesis by Bacopaside I and II Is Associated with Ca2+ Flux and Loss of Plasma Membrane Integrity.

We previously showed how triterpene saponin bacopaside (bac) II, purified from the medicinal herb Bacopa monnieri, induced cell death in colorectal cancer cell lines and reduced endothelial cell migration and tube formation, and further demonstrated a synergistic effect of a combination of bac I and bac II on the inhibition of breast cancer cell line growth. Here, we assessed the effects of bac I and II on the colorectal cancer HT-29 cell line, and mouse (2H-11) and human umbilical vein endothelial cell (HUVEC) lines, measuring outcomes including cell viability, proliferation, migration, tube formation, apoptosis, cytosolic Ca2+ levels and plasma membrane integrity. Combined bac I and II, each applied at concentrations below IC50 values, caused a synergistic reduction of the viability and proliferation of HT-29 and endothelial cells, and impaired the migration of HT-29 and tube formation of endothelial cells. A significant enhancement of apoptosis was induced only in HUVEC, although an increase in cytosolic Ca2+ was detected in all three cell lines. Plasma membrane integrity was compromised in 2H-11 and HUVEC, as determined by an increase in propidium iodide staining, which was preceded by Ca2+ flux. These in vitro findings support further research into the mechanisms of action of the combined compounds for potential clinical use.

Bacopasides I and II Act in Synergy to Inhibit the Growth, Migration and Invasion of Breast Cancer Cell Lines.

Bacopaside (bac) I and II are triterpene saponins purified from the medicinal herb Bacopa monnieri. Previously, we showed that bac II reduced endothelial cell migration and tube formation and induced apoptosis in colorectal cancer cell lines. The aim of the current study was to examine the effects of treatment with combined doses of bac I and bac II using four cell lines representative of the breast cancer subtypes: triple negative (MDA-MB-231), estrogen receptor positive (T47D and MCF7) and human epidermal growth factor receptor 2 (HER2) positive (BT-474). Drug treatment outcome measures included cell viability, proliferation, cell cycle, apoptosis, migration, and invasion assays. Relationships were analysed by one- and two-way analysis of variance with Bonferroni post-hoc analysis. Combined doses of bac I and bac II, each below their half maximal inhibitory concentration (IC50), were synergistic and reduced the viability and proliferation of the four breast cancer cell lines. Cell loss occurred at the highest dose combinations and was associated with G2/M arrest and apoptosis. Migration in the scratch wound assay was significantly reduced at apoptosis-inducing combinations, but also at non-cytotoxic combinations, for MDA-MB-231 and T47D (p < 0.0001) and BT-474 (p = 0.0003). Non-cytotoxic combinations also significantly reduced spheroid invasion of MDA-MB-231 cells by up to 97% (p < 0.0001). Combining bac I and II below their IC50 reduced the viability, proliferation, and migration and invasiveness of breast cancer cell lines, suggesting synergy between bac I and II.

The Purified Extract from the Medicinal Plant Bacopa monnieri, Bacopaside II, Inhibits Growth of Colon Cancer Cells In Vitro by Inducing Cell Cycle Arrest and Apoptosis.

Aquaporin-1 (AQP1), a transmembrane pore-forming molecule, facilitates the rapid movement of water and small solutes across cell membranes. We have previously shown that bacopaside II, an extract from the medicinal herb Bacopa monnieri, blocks the AQP1 water channel and impairs migration of cells that express AQP1. The aim of this study was to further elucidate the anti-tumour potential of bacopaside II in colon cancer cells. Expression of AQP1 in HT-29, SW480, SW620 and HCT116 was determined by quantitative PCR and western immunoblot. Cells were treated with bacopaside II, and morphology, growth, autophagy, cell cycle and apoptosis assessed by time-lapse microscopy, crystal violet, acridine orange, propidium iodide (PI) and annexin V/PI staining respectively. AQP1 expression was significantly higher in HT-29 than SW480, SW620 and HCT116. Bacopaside II significantly reduced growth at ≥20 µM for HT-29 and ≥15 µM for SW480, SW620 and HCT116. Inhibition of HT-29 at 20 µM was primarily mediated by G0/G1 cell cycle arrest, and at 30 µM by G2/M arrest and apoptosis. Inhibition of SW480, SW620 and HCT116 at ≥15 µM was mediated by G2/M arrest and apoptosis. These results are the first to show that bacopaside II inhibits colon cancer cell growth by inducing cell cycle arrest and apoptosis.

The Aquaporin 1 Inhibitor Bacopaside II Reduces Endothelial Cell Migration and Tubulogenesis and Induces Apoptosis.

Expression of aquaporin-1 (AQP1) in endothelial cells is critical for their migration and angiogenesis in cancer. We tested the AQP1 inhibitor, bacopaside II, derived from medicinal plant Bacopa monnieri, on endothelial cell migration and tube-formation in vitro using mouse endothelial cell lines (2H11 and 3B11) and human umbilical vein endothelial cells (HUVEC). The effect of bacopaside II on viability, apoptosis, migration and tubulogenesis was assessed by a proliferation assay, annexin-V/propidium iodide flow cytometry, the scratch wound assay and endothelial tube-formation, respectively. Cell viability was reduced significantly for 2H11 at 15 µM (p = 0.037), 3B11 at 12.5 µM (p = 0.017) and HUVEC at 10 µM (p < 0.0001). At 15 µM, the reduced viability was accompanied by an increase in apoptosis of 38%, 50% and 32% for 2H11, 3B11 and HUVEC, respectively. Bacopaside II at ≥10 µM significantly reduced migration of 2H11 (p = 0.0002) and 3B11 (p = 0.034). HUVECs were most sensitive with a significant reduction at ≥7.5 µM (p = 0.037). Tube-formation was reduced with a 15 µM dose for all cell lines and 10 µM for 3B11 (p < 0.0001). These results suggest that bacopaside II is a potential anti-angiogenic agent.

Comparable survival outcome of metastatic colorectal cancer in Indigenous and non-Indigenous patients: Retrospective analysis of the South Australian metastatic colorectal cancer registry.

OBJECTIVE: This study aims to investigate disparities in demographics, disease characteristics, treatment and overall survival between South Australian (SA) Indigenous and non-Indigenous patients with metastatic colorectal cancer (mCRC). DESIGN: This employs a retrospective population study using the SA mCRC registry. SETTING: The SA mCRC registry identifies mCRC patients from hospital encounters, histopathology reports, medical oncology letters, clinician notification, attendances at multidisciplinary meetings and death audits by the SA Cancer Registry. PARTICIPANTS: A total of 2865 adult mCRC patients including 14 Indigenous patients were identified through the SA mCRC registry between February 2006 and August 2013. Patients were linked to the SA Cancer Registry to obtain Indigenous status. MAIN OUTCOME MEASURES: Demographic, disease and treatment characteristics were compared using Chi-squared test and t-test; while overall survival defined as time to any cause of death was analysed using Cox regression. RESULTS: No difference was observed for clinical characteristics, except for a higher proportion of Indigenous patients receiving chemotherapy (85.7% versus 58.5%; P = 0.04). The rate of liver surgery was similar across the two groups (21.0% versus 15.1%; P = 0.40). The median overall survivals were equivalent (11.9 months versus 15.1 months; hazard ratio = 1.00; 95% confidence interval for hazard ratio, 0.54-1.86). CONCLUSIONS: Clinical characteristics and survival outcomes were similar between Indigenous and non-Indigenous patients captured on the SA mCRC registry, and outcome of those who have an access to comprehensive cancer care appeared independent of Indigenous status and in line with large clinical trials. Underestimation of Indigenous cases due to their lower utilisation of cancer service could not be excluded and ultimately the accurate reporting of these patients is crucial.

Does the Chemotherapy Backbone Impact on the Efficacy of Targeted Agents in Metastatic Colorectal Cancer? A Systematic Review and Meta-Analysis of the Literature.

IMPORTANCE: The EGFR inhibitors (EGFR-I) cetuximab and panitumumab and the angiogenesis inhibitors (AIs) bevacizumab and aflibercept have demonstrated varying efficacy in mCRC. OBJECTIVE: To document the overall impact of specific chemotherapy regimens on the efficacy of targeted agents in treating patients with mCRC. DATA SOURCES: MEDLINE, EMBASE and Cochrane databases were searched to 2014, supplemented by hand-searching ASCO/ESMO conference abstracts. STUDY SELECTION: Published RCTs of patients with histologically confirmed mCRC were included if they investigated either 1) chemotherapy with or without a biological agent or 2) different chemotherapy regimens with the same biological agent. EGFR-I trials were restricted to KRAS exon 2 wild-type (WT) populations. DATA EXTRACTION AND SYNTHESIS: Data were independently abstracted by two authors and trial quality assessed according to Cochrane criteria. The primary outcome was overall survival with secondary endpoints progression free survival (PFS), overall response rate (ORR) and toxicity. RESULTS: EGFR-I added to irinotecan-based chemotherapy modestly improved OS with HR 0.90 (95% CI 0.81-1.00, p = 0.04), but more so PFS with HR 0.77 (95% CI 0.69-0.86, p<0.00001). No benefit was evident for EGFR-I added to oxaliplatin-based chemotherapy (OS HR 0.97 (95% CI 0.87-1.09) and PFS HR 0.92 (95% CI 0.83-1.02)). Significant oxaliplatin-irinotecan subgroup interactions were present for PFS with I2 = 82%, p = 0.02. Further analyses of oxaliplatin+EGFR-I trials showed greater efficacy with infusional 5FU regimens (PFS HR 0.82, 95% CI 0.72-0.94) compared to capecitabine (HR 1.09; 95% CI 0.91-1.30) and bolus 5FU (HR 1.07; 95% CI 0.79-1.45); subgroup interaction was present with I2 = 72%, p = 0.03. The oxaliplatin-irinotecan interaction was not evident for infusional 5FU regimens. For AIs, OS benefit was observed with both oxaliplatin-based (HR 0.83) and irinotecan-based (HR 0.77) regimens without significant subgroup interactions. Oxaliplatin+AI trials showed no subgroup interactions by type of FP, whilst an interaction was present for irinotecan+AI trials although aflibercept was only used with infusional FP (I2 = 89.7%, p = 0.002). CONCLUSION AND RELEVANCE: The addition of EGFR-I to irinotecan-based chemotherapy has consistent efficacy, regardless of FP regimen, whereas EGFR-I and oxaliplatin-based regimens were most active with infusional 5FU. No such differential activity was observed with the varying chemotherapy schedules when combined with AIs.

Rechallenge with oxaliplatin and fluoropyrimidine for metastatic colorectal carcinoma after prior therapy.

OBJECTIVES: Patients with advanced colorectal cancer (CRC) who have received oxaliplatin, 5-fluoropyrimidine, and irinotecan chemotherapy (with or without bevacizumab) and antiepidermal growth factor receptor therapy (if KRAS is wild type) have no further standard treatment options. Although repeating a prior chemotherapy [in particular, oxaliplatin and fluoropyrimidine (FOX)] is an option, there is very little evidence in the literature for this approach; thus, we reviewed our registry to assess the frequency and outcome of rechallenging with FOX. METHODS: Patients who had been rechallenged with FOX were identified from the South Australian metastatic CRC database. Patient characteristics were analyzed, and tumor response was retrospectively assessed using Response Evaluation Criteria in Solid Tumors criteria. RESULTS: Twenty patients were eligible for inclusion in this analysis. The number of prior lines of therapy received for metastatic CRC was 4 lines for 2 patients, 3 lines for 6 patients, 2 lines for 7 patients, and 1 line for 3 patients, with 3 patients having received oxaliplatin as adjuvant therapy. Four patients had received bevacizumab previously, 7 patients had undergone antiepidermal growth factor receptor treatment, and 4 patients had undergone liver resection earlier. Response rate was 18%, and 47% had stable disease. The median progression-free survival was 3.7 months, median overall survival was 7.8 months, and 1-year survival was 37%. CONCLUSIONS: In this selected population, there is evidence of modest activity of rechallenge with FOX chemotherapy, although radiologic response is uncommon.