Endovascular treatment of complex abdominal aortic aneurysms by combining different types of endoprostheses.

PURPOSE: To describe an off-the-shelf method for the treatment of abdominal aortic aneurysms with hostile (large, >30 mm) neck and/or small (<20 mm) aortic bifurcation. CASE REPORT: We describe five patients with large aortic necks and/or small aortic bifurcations, which were treated by combining an AFX endoprosthesis with a Valiant Captiva endograft, and additional proximal endoanchors when deemed necessary. Initial technical success was 100%. Follow-up ranged from 228 to 875 days. One patient suffered a type 1A and 1B endoleak at 446 days follow-up, which were successfully treated by endovascular means. CONCLUSION: Combining the AFX and Valiant Captiva endografts is an off-the-shelf solution for treatment of large diameter aortic necks and small aortic bifurcations in patients deemed unfit for open repair or declined for fenestrated endografts. Longer follow-up is required to assess the long-term safety with special focus on aortic neck dilation.

An internal iliac artery aneurysm causing sudden buttock ischemia and nerve root compression.

Acute buttock ischemia can be a consequence of aneurysmatic disease and has a dramatic presentation. This case report describes an otherwise healthy patient with a simultaneous onset of buttock ischemia combined with sciatic nerve compression caused by a small distal internal iliac artery aneurysm. Coiling of the aneurysm prevented thromboembolism recurrence but was only partially successful in reducing the symptoms of nerve compression. Given the serious consequences, prophylactic treatment independent of aneurysm diameter can be considered.

Hypoxia after liver surgery imposes an aggressive cancer stem cell phenotype on residual tumor cells.

OBJECTIVE: To assess the contribution of hypoxia and bone marrow-derived cells to aggressive outgrowth of micrometastases after liver surgery. BACKGROUND: Liver surgery generates a microenvironment that fosters aggressive tumor recurrence. These areas are characterized by chronic hypoxia and influx of bone marrow-derived cells. METHODS: The contribution of hematopoietic cell types was studied in mice lacking specific components of the immune system and in irradiated mice lacking all bone marrow-derived cells. Tumor cells were derived from colorectal cancer patients and from a metastatic tumor cell line. Hypoxia-induced changes in stem cell and differentiation marker expression, clone-forming potential, and metastatic capacity were assessed. The effect of vascular clamping on cancer stem cell (CSC) characteristics was performed in mice bearing patient-derived liver metastases. RESULTS: Immune cells and bone marrow-derived cells were not required for aggressive outgrowth of micrometastases in livers treated with surgery. Rather, hypoxia was sufficient to promote invasion and accelerate metastatic outgrowth. This was associated with a rapid loss of differentiation markers and increased expression of CSC markers and clone-forming capacity. Likewise, metastases residing in ischemia-reperfusion-injured liver lobes acquired CSC characteristics. Despite their renowned general resistance to chemotherapy, clone-forming CSCs were readily killed by the hypoxia-activated prodrug tirapazamine. CONCLUSIONS: Surgery-generated hypoxia in the liver causes rapid dedifferentiation of tumor cells into immature CSCs with high clone- and metastasis-forming capacity. The results help explain the phenomenon of aggressive local tumor recurrence after liver surgery and offer a potential strategy to kill aggressive CSCs by hypoxia-activated prodrugs.

Circulating CD95-ligand as a potential prognostic marker for recurrence in patients with synchronous colorectal liver metastases.

AIM: To assess whether circulating soluble CD95 ligand (sCD95L) levels are associated with recurrence-free survival (RFS) in patients with synchronous colorectal liver metastases. PATIENTS AND METHODS: Blood samples were obtained from 62 patients with synchronous colorectal liver metastases before and after liver surgery. Serum sCD95L levels were determined using enzyme-linked immunosorbent assay (ELISA). Cox regression analysis was performed to determine the correlation between sCD95L levels and RFS and overall survival (OS). RESULTS: Median follow-up was 33 months. High pre-operative sCD95L levels were associated with poor RFS and OS in univariable (p=0.019 and p=0.020) and multivariable analyses (p=0.020 and p=0.003). CONCLUSION: Preoperatives CD95L is a potential prognostic factor for RFS and OS of patients undergoing surgery for synchronous colorectal liver metastases. Low preoperatives CD95L levels may help identify a subgroup of patients with synchronous liver metastases that are likely to benefit from liver surgery.

Combined KRAS and TP53 mutation status is not predictive in CAPOX-treated metastatic colorectal cancer.

BACKGROUND: The response of colorectal tumours to chemotherapy is highly variable. Preclinical work has shown that the Kirsten ras (KRAS) oncogene sensitizes colorectal tumour cells to oxaliplatin and capecitabine in a wild-type tumour suppressor p53 (TP53)-dependent manner. Therefore, whether or not the combined mutation status of KRAS and TP53 could predict response to chemotherapy in metastatic colorectal cancer was tested. PATIENTS AND METHODS: A subgroup of patients from the CAIRO2 study (randomized phase III study on capecitabine, oxaliplatin, bevacizumab with or without cetuximab in first-line advanced colorectal cancer) that received capecitabine plus oxaliplatin (CAPOX) treatment in combination with bevacizumab was selected. The tumours were analyzed for KRAS and TP53 mutations by PCR/sequencing. The relationship between tumour response and genotype was analyzed. RESULTS: The following KRAS/TP53 genotypes were identified: KRASmut/TP53mut n=21, KRASmut/TP53wt n=20, KRASwt/TP53mut n=25, KRASwt/TP53wt n=15. No genotype was associated with a significantly better or worse progression-free or overall survival. CONCLUSION: The combined mutation status of KRAS and TP53 does not predict response to CAPOX in patients with metastasized colorectal cancer.

Synergistic killing of colorectal cancer cells by oxaliplatin and ABT-737.

BACKGROUND: Oxaliplatin is frequently used in the treatment of metastatic colorectal cancer (CRC). Our previous work shows that oxaliplatin induces the pro-apoptotic protein Noxa in CRC cells. The Bcl2-inhibitor ABT-737 is particularly effective in cells with high Noxa levels. Therefore, we tested whether oxaliplatin and ABT-737 display synergy in killing CRC cells. METHODS: A panel of CRC cell lines was treated with oxaliplatin and ABT-737, either alone or in combination. Apoptosis was measured by FACS analysis of sub-G1 DNA content and by Western blot analysis of caspase-3 processing. Noxa expression was suppressed by lentiviral RNA interference. RESULTS: Oxaliplatin and ABT-737 displayed a strong synergistic apoptotic response, which was dependent on wildtype TP53 and oncogenic KRAS. TP53 and KRAS were required for drug-induced Noxa expression and this was essential for tumor cell apoptosis. Oxaliplatin, but not ABT-737, induced p53 accumulation, but both drugs stimulated Noxa expression. Combination treatment of mice with subcutaneous tumor xenografts drastically reduced tumor volume, while single drug treatment had no effect. CONCLUSION: ABT-737 synergizes with oxaliplatin to kill colorectal cancer cells. This requires induction of Noxa by wildtype TP53 and oncogenic KRAS. Future studies should explore the anti-tumor efficacy of this drug combination in mouse models for spontaneous CRC development and in patient-derived tumor cell cultures and xenografts.

Intermittent ischaemia maintains function after ischaemia reperfusion in steatotic livers.

BACKGROUND: Ischaemic preconditioning (IPC) and intermittent ischaemia (INT) reduce liver injury after ischaemia reperfusion (IR). Steatotic livers are at a higher risk of IR injury, but the protection offered by IPC and INT is not well understood. The aim of the present study was to determine the effectiveness of IPC and INT in maintaining liver function in steatotic livers. MATERIAL AND METHODS: A model of segmental hepatic ischaemia (45 min) and reperfusion (60 min) was employed using lean and obese Zucker rats. Bile flow recovery was measured to assess dynamic liver function, hepatocyte fat content quantified and blood electrolytes, metabolites and bile calcium measured to assess liver and whole body physiology. Liver marker enzymes and light and electron microscopy were employed to assess hepatocyte injury. RESULTS: IPC was not effective in promoting bile flow recovery after IR in either lean or steatotic livers, whereas INT promoted good bile flow recovery in steatotic as well as lean livers. However, the bile flow recovery in steatotic livers was less than that in lean livers. In steatotic livers, ischaemia led to a rapid and substantial decrease in fat content. Steatotic livers were more susceptible to IR injury than lean livers, as indicated by increased blood ALT concentrations and major histological injury. CONCLUSION: INT is more effective than IPC in restoring liver function in the acute phase of IR in steatotic livers. In obese patients, INT may be useful in promoting better liver function after IR after liver resection.

Oncogenic KRAS desensitizes colorectal tumor cells to epidermal growth factor receptor inhibition and activation.

Epidermal growth factor receptor (EGFR)-targeting therapeutics have shown efficacy in the treatment of colorectal cancer patients. Clinical studies have revealed that activating mutations in the KRAS protooncogene predict resistance to EGFR-targeted therapy. However, the causality between mutant KRAS and resistance to EGFR inhibition has so far not been demonstrated. Here, we show that deletion of the oncogenic KRAS allele from colorectal tumor cells resensitizes those cells to EGFR inhibitors. Resensitization was accompanied by an acquired dependency on the EGFR for maintaining basal extracellular signal-regulated kinase (ERK) activity. Deletion of oncogenic KRAS not only resensitized tumor cells to EGFR inhibition but also promoted EGF-induced NRAS activation, ERK and AKT phosphorylation, and c-FOS transcription. The poor responsiveness of mutant KRAS tumor cells to EGFR inhibition and activation was accompanied by a reduced capacity of these cells to bind and internalize EGF and by a failure to retain EGFR at the plasma membrane. Of 16 human colorectal tumors with activating mutations in KRAS, 15 displayed loss of basolateral EGFR localization. Plasma membrane localization of the EGFR could be restored in vitro by suppressing receptor endocytosis through Rho kinase inhibition. This caused an EGFR-dependent increase in basal and EGF-stimulated ERK phosphorylation but failed to restore tumor cell sensitivity to EGFR inhibition. Our results demonstrate a causal role for oncogenic KRAS in desensitizing tumor cells not only to EGFR inhibitors but also to EGF itself.

Oncogenic K-ras activates p38 to maintain colorectal cancer cell proliferation during MEK inhibition.

BACKGROUND: Colon carcinomas frequently contain activating mutations in the K-ras proto-oncogene. K-ras itself is a poor drug target and drug development efforts have mostly focused on components of the classical Ras-activated MEK/ERK pathway. Here we have studied whether endogenous oncogenic K-ras affects the dependency of colorectal tumor cells on MEK/ERK signaling. METHODS: K-ras mutant colorectal tumor cell lines C26, HCT116 and L169 were used. K-ras or components of the MEK/ERK and p38 pathway were suppressed by RNA interference (RNAi). MEK was inhibited by U0126. p38 was inhibited by SB203850. RESULTS: MEK inhibition, or suppression of MEK1/2 or ERK1/2 by RNA interference, reduced the proliferation rate of all colorectal cancer cell lines. However, cell proliferation returned to normal after two weeks of chronic inhibition, despite the continued suppression of MEK or ERK. In contrast, K-ras-suppressed tumor cells entered an irreversible senescent-like state following ERK pathway inhibition. MEK inhibition or ERK1/2 suppression caused activation of p38alpha in a K-ras-dependent manner. Inhibition or suppression of p38alpha prevented the recovery of K-ras mutant tumor cells during prolonged MEK inhibition. CONCLUSION: Oncogenic K-ras activates p38alpha to maintain cell proliferation during MEK inhibition. MEK-targeting therapeutics can create an acquired tumor cell dependency on p38alpha.

Oncogenic K-Ras turns death receptors into metastasis-promoting receptors in human and mouse colorectal cancer cells.

BACKGROUND & AIMS: Death receptors expressed on tumor cells can prevent metastasis formation by inducing apoptosis, but they also can promote migration and invasion. The determinants of death receptor signaling output are poorly defined. Here we investigated the role of oncogenic K-Ras in determining death receptor function and metastatic potential. METHODS: Isogenic human and mouse colorectal cancer cell lines differing only in the presence or absence of the K-Ras oncogene were tested in apoptosis and invasion assays using CD95 ligand and tumor necrois factor-related apoptosis-inducing ligand (TRAIL) as stimuli. Metastatic potential was assessed by intrasplenic injections of green fluorescent protein- or luciferase-expressing tumor cells, followed by intravital fluorescence microscopy or bioluminescence imaging, and confocal microscopy and immunohistochemistry. Ras-effector pathway control of CD95 output was assessed by an RNA-interference and inhibitor-based approach. RESULTS: CD95 ligand and TRAIL stimulated invasion of colorectal tumor cells and liver metastases in a K-Ras-dependent fashion. Loss of mutant K-Ras switched CD95 and TRAIL receptors back into apoptosis mode and abrogated metastatic potential. Raf1 was essential for the switch in CD95 function, for tumor cell survival in the liver, and for K-Ras-driven formation of liver metastases. K-Ras and Raf1 suppressed Rho kinase (ROCK)/LIM kinase-mediated phosphorylation of the actin-severing protein cofilin. Overexpression of ROCK or LIM kinase allowed CD95L to induce apoptosis in K-Ras-proficient cells and prevented metastasis formation, whereas their suppression protected K-Ras-deficient cells against apoptosis. CONCLUSIONS: Oncogenic K-Ras and its effector Raf1 convert death receptors into invasion-inducing receptors by suppressing the ROCK/LIM kinase pathway, and this is essential for K-Ras/Raf1-driven metastasis formation.

Accelerated perinecrotic outgrowth of colorectal liver metastases following radiofrequency ablation is a hypoxia-driven phenomenon.

OBJECTIVE: The aim of this study was to assess how thermal ablation of colorectal liver metastases affects the outgrowth of micrometastases in the transition zone (TZ) between ablated tissue and the unaffected reference zone (RZ) in 2 different murine models. BACKGROUND: Thermal destruction therapies of nonresectable colorectal liver metastases, including radiofrequency ablation (RFA), can provide tumor clearance, but local recurrences are common. METHODS: Three days after intrasplenic injection of C26 colon carcinoma cells, RFA was applied to the left liver lobe. Perinecrotic microcirculation, tissue hypoxia, hypoxia inducible factor (HIF)-1alpha and HIF-2alpha, and the outgrowth of micrometastases both in the TZ and in the RZ were evaluated over time. RESULTS: In 2 different animal models, the outgrowth of micrometastases in the TZ following RFA was stimulated approximately 4-fold compared to tumor growth in the RZ. Accelerated tumor growth in the TZ was associated with microcirculatory disturbances, prolonged hypoxia, and stabilization of HIF-1alpha and HIF-2alpha in the tumor cells. In addition, RFA induced the formation of new hepatic vessels that sprouted from existing sinusoids and grew into the generated necrotic lesion. Surprisingly, the accelerated tumor growth was not associated with these vessels. Treatment with 17DMAG prevented HIF-1alpha and HIF-2alpha stabilization and selectively reduced tumor growth in the TZ by approximately 40% without affecting tumor growth in sham-operated mice or in the RZ of RFA-treated mice. PTK787/ZK-222584, a nonselective Vascular Endothelial Growth Factor (VEGF)-receptor inhibitor, reduced RFA-stimulated tumor growth and tumor growth in the RZ to a similar extent. CONCLUSIONS: We conclude that RFA stimulates the outgrowth of tumor cells at the lesion periphery. Angiogenesis is not the driving force behind RFA-stimulated tumor growth, but other hypoxia/HIF-activated pathways are likely to be important.

Ischemic preconditioning and intermittent ischemia preserve bile flow in a rat model of ischemia/reperfusion injury.

Ischemia and reperfusion (IR) injury of the liver is associated with impaired bile secretion, but the effects of ischemic preconditioning (IPC) and intermittent ischemia (INT) on bile flow are unknown. A rat model of segmental (60%-70%) hepatic ischemia and reperfusion was employed to test the effects of IPC and INT on bile flow. Continuous clamping for 45 min (CC) substantially reduced bile flow, and this did not recover after 60 min of reperfusion. IPC and INT caused a significant recovery of bile flow. The elevation in plasma liver marker enzymes induced by CC was not reduced by IPC and INT. Light microscopy showed mild hepatocyte damage in all groups. In the CC group, the amount of F-actin localized around the bile canaliculi in the ischemic lobes was less than that in the nonischemic lobes, but this difference was not observed in the IPC and INT groups. It is concluded that IPC and INT substantially alleviate the decrease in bile flow induced by ischemia. Bile flow may be useful in the assessment of IR injury.

Ischemic preconditioning and intermittent ischemia preserve bile flow in a rat model of ischemia reperfusion injury.

Ischemia and reperfusion (IR) injury of the liver is associated with impaired bile secretion, but the effects of ischemic preconditioning (IPC) and intermittent ischemia (INT) on bile flow are unknown. A rat model of segmental (60%-70%) hepatic ischemia and reperfusion was employed to test the effects of IPC and INT on bile flow. Continuous clamping for 45 min (CC) substantially reduced bile flow, and this did not recover after 60 min of reperfusion. IPC and INT caused a significant recovery of bile flow. The elevation in plasma liver marker enzymes induced by CC was not reduced by IPC and INT. Light microscopy showed mild hepatocyte damage in all groups. In the CC group, the amount of F-actin localized around the bile canaliculi in the ischemic lobes was less than that in the nonischemic lobes, but this difference was not observed in the IPC and INT groups. It is concluded that IPC and INT substantially alleviate the decrease in bile flow induced by ischemia. Bile flow may be useful in the assessment of IR injury.

Hepatic ischemia-reperfusion injury: roles of Ca2+ and other intracellular mediators of impaired bile flow and hepatocyte damage.

Liver resection and liver transplantation have been successful in the treatment of liver tumors and end-stage liver disease. This success has led to an expansion in the pool of patients potentially treatable by liver surgery and, in the case of transplantation, to a shortage of liver donors. At present, there are significant numbers of potential candidates for liver resection and liver donation who have fatty livers, are aged, or have livers damaged by chemotherapy. All of these are at high risk for ischemic reperfusion (IR) injury. The aims of this review are to assess current knowledge of the clinical effectiveness of ischemic preconditioning and intermittent ischemia in reducing IR damage in liver surgery; to evaluate the use of bile flow as a sensitive indicator of IR liver damage; and to analyze the molecular mechanisms, especially intracellular Ca2+, involved in IR injury and ischemic preconditioning. It is concluded that bile flow is a sensitive indicator of IR injury. Together with reactive oxygen species (ROS) and other extracellular and intracellular signaling molecules, intracellular Ca2+ in hepatocytes plays a key role in the normal regulation of bile flow and in IR-induced injury and cell death. Ischemic preconditioning is an effective strategy to reduce IR injury but there is considerable scope for improvement, especially in patients with fatty and aged livers. The development of effective new strategies to reduce IR injury will depend on improved understanding of the molecular mechanisms involved, especially by gaining a better perspective of the relative importance of the various intrahepatocyte signaling pathways involved.

Effects of subtotal colectomy on bacterial translocation during experimental acute pancreatitis.

OBJECTIVES: The colon is considered a major source of bacteria causing infection of pancreatic necrosis in acute pancreatitis (AP). Subtotal colectomy before AP in rats reduces mortality, but its role in affecting small bowel flora, bacterial translocation, and infection of pancreatic necrosis is unknown. Our aim was to study these phenomena in rats with AP. METHODS: Fifty rats, allocated in 4 groups, underwent 2 laparotomies: group 1, sham laparotomy and saline biliopancreatic duct infusion; group 2, subtotal colectomy and saline infusion; group 3, sham laparotomy and AP (ductal infusion of glycodeoxycholic acid and intravenous cerulein); group 4, subtotal colectomy and AP. Seventy-two hours later, samples were collected for microbiological analysis. RESULTS: Subtotal colectomy caused small bowel bacterial overgrowth with gram-positive cocci (group 1 versus group 2, duodenum: P = 0.030, ileum: P = 0.029). Bacterial counts of gram-negative rods/anaerobes in the duodenum and ileum and pancreatic bacterial counts of rats with colectomy and AP were significantly higher than in rats with AP only (group 3 versus group 4, duodenum: P = 0.040, ileum: P = 0.029, pancreas: P = 0.017). Duodenal bacterial overgrowth and pancreatic infection correlate significantly (r = 0.45, P = 0.004). CONCLUSIONS: Subtotal colectomy induces small bowel bacterial overgrowth, which is associated with increased bacterial translocation to the pancreas.