Citations

Abstract Hyperactive K-RAS signaling is a major menace that drives aggressive tumor growth and metastasis in human cancer. Currently, there are no effective ways to treat human cancers that have oncogenic K-RAS mutations that confer drug resistance, aggressive tumor growth, systemic metastasis, and poor clinical outcome. Therefore, finding novel approaches and new drug targets to inhibit oncogenic K-RAS is an urgent goal and the major challenge in cancer therapy and treatment. Instead of targeting an upstream signaling module such as EGFR/HER2/KRAS/B-RAF/MEK/MAPK/AKT/mT0R, we targeted the most downstream signaling module in the EGFR/HER2/K-RAS pathway called the SIAH-dependent proteolytic machinery. SIAHs are the human homologs of seven-in-absentia (SINA), an evolutionarily conserved RING E3 ligase, an essential downstream signaling module, and a critical gatekeeper required for proper ERBB/RAS/RAF signal transduction. Guided by the insights and principles learned from Drosophila RAS signaling pathway, we conducted preclinical studies to dissect SIAH function in the context of ERBB/K-RAS-mediated tumorigenesis and metastasis in human cancer cells and tumor specimens. We found that: 1) SIAH is a novel biomarker for human cancer, i.e., SIAH specifically decorates proliferating tumor cells (not stromal or immune cells in the tumor microenvironment). SIAH expression is progressively and markedly upregulated as human cancer progresses to the advanced stages in human pancreatic, lung, breast, and prostate cancers; and 2) inhibiting SIAH function is highly effective to abolish tumorigenesis and metastasis in multiple of the most aggressive human cancer cell lines known in the literature that include pancreatic cancer cells (MiaPaCa and Panc-1), lung cancer cells (A549, H727, and UMC11), invasive breast cancer cells (MDA-MB-231, MDA-MB-468, and MDA-MB-435), colorectal cancer cells (SW620 and HCT116) and hormonal-refractory prostate cancer cells (PC-3 and DU-145) in our preclinical studies. These findings demonstrate that SIAH is an attractive and logical new therapeutic target for novel anti-K-RAS and anticancer therapy in human cancer. Through our work, SIAH has emerged as a new and effective drug target against oncogenic K-RAS activation in human cancer cells. Using anti-SIAH molecules to block K-RAS signaling in human cancer is an excellent example of science going from the bench (basic research in fruit flies) to the bedside (preclinical studies and ultimately clinical trials). As a highly evolutionarily conserved E3 ligase and the most downstream gatekeeper of EGFR/HER2/K-RAS/B-RAF signaling pathway, SIAH is uniquely and logically positioned to become a next-generation anti-K-RAS drug target. Our preclinical studies have demonstrated that SIAH-dependent proteolysis is an Achilles' heel for human cancer cells. Albeit at an early preclinical stage, knowledge gained from our study has great promise that has generated many excitements with immediate translational values. Anti-SIAH-based small molecule inhibitors may aid in expanding our arsenal of novel anticancer therapies. By attacking the oncogenic ERBB/K-RAS/B-RAF pathway using multipronged synergistic inhibitions at upstream (EGFR/HER2 membrane receptors), midstream (K-RAS/B-RAF/MEK/mTOR), and downstream (SIAH E3 ligase) signaling modules in parallel, we will be in a great position to halt the genesis, progression, and metastasis of the most aggressive and the deadliest forms of human cancers. If successful, we hope to translate our findings to the cancer clinics to treat many cancer patients with metastatic diseases in the near future. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr B14.

The antihypertensive influence of fish oil is controversial, and the mechanisms remain unclear. Because the inverse relation between fish oil and hypertension appears to be partially dependent on the degree of hypertension, we tested the hypothesis that fish oil would elicit more dramatic reductions in mean arterial pressure (MAP) and muscle sympathetic nerve activity (MSNA) in prehypertensive (PHT) compared with normotensive (NT) subjects. Resting MAP, MSNA, and heart rate (HR) were examined before and after 8 wk of fish oil (9 g/day; 1.6 g eicosapentaenoic acid and 1.1 g docosahexaenoic acid) or placebo (olive oil; 9 g/day) in 38 NT (19 fish oil; 19 placebo) and 29 PHT (15 fish oil; 14 placebo) volunteers. Fish oil did not alter resting MAP, MSNA, or HR in either NT (80 ± 1 to 80 ± 1 mmHg; 11 ± 2 to 10 ± 1 bursts/min; 71 ± 2 to 71 ± 2 beats/min) or PHT (88 ± 2 to 87 ± 1 mmHg; 11 ± 2 to 10 ± 2 bursts/min; 73 ± 2 to 73 ± 2 beats/min) subjects. When NT and PHT groups were consolidated, analysis of covariance confirmed that pretreatment resting MAP was not associated with changes in MSNA after fish oil. In contrast, pretreatment resting HR was correlated with changes in MSNA ( r = 0.47; P = 0.007) and MAP ( r = 0.42; P < 0.007) after fish oil but not placebo. In conclusion, fish oil did not alter sympathetic neural control in NT or PHT subjects. However, our findings suggest that fish oil is associated with modest sympathoinhibition in individuals with higher resting heart rates, a finding that is consistent with a recent meta-analysis examining the relations among fish oil, HR, and the risk of cardiovascular disease.

The effect of exercise on the pulmonary circulation was studied in a group of 6 intact dogs and in a second group of 4 subjected to ligation and division of the left pulmonary artery. Observations were made with the dogs standing and running on a horizontal treadmill up to the maximal speed they could maintain for periods of at least 2 minutes, usually 12 or 14 km./hr. Continuous records were made of the difference in pressure between the main pulmonary artery (PA) and a pulmonary-artery wedge (PAW) position. Cardiac output was determined by the indicator-dilution method, with injection of indocyanine (cardiogreen) dye into the main pulmonary artery and continuous sampling from the root of the aorta. The mean transit time of indicator betweell the pulmonary and aortic valves was obtained from the dilution curves, a correction being made for the transit time through the sampling apparatus. From these measurements the blood volume of the lungs and left heart chambers was derived. In the intact dogs, cardiac output increased approximately threefold from rest to severe exercise. There was an increase of only 12 per cent to 50 per cent in PA minus PAW pressure. Thus a continued fall occurred in pulmonary vascular resistance. The blood volume of the lungs and left heart of 3 intact dogs was measured. In 2 it did not change through the range of activity, and in the third the increase with maximal exercise was 18 per cent. The dogs with ligated left pulmonary artery also showed a threefold increase in output with severe exercise. Since the entire output passed through a single lung, the effect on the pulmonary vessels was comparable to that of a sixfold increase in output in a normal dog. Despite such large flows, in 3 of 4 dogs the difference between PA and PAW pressure rose only 37 per cent to 50 per cent. The volume of blood in the lungs and left heart was unchanged between rest and 8 km./hr., and at higher speeds showed a 0 per cent to about 20 per cent increase.

Measurements of stroke volume were made in dogs during graded exercise on a horizontal treadmill up to the maximal exercise which they could maintain for 2 to 3 minutes. Eleven series of experiments were performed on 7 dogs. The cardiac output was measured by the indicator-dilution technic and the heart rate was measured from the aortic pressure tracing. At all grades of exercise, there was little or no change in stroke volume. Even with severe exercise, when the cardiac output had increased from 1.5 to 3.5 (average, 2.4) times the values obtained with the dog standing at rest, the stroke volume showed less than 5 per cent change over resting values in 5 experiments on 4 dogs, while in the remainder the increases were 8, 11, 11, 14, 15, and 19 per cent. Thus, the increase in cardiac output in the untrained exercising dog is almost directly proportional to the increase in heart rate. This is in contrast to other situations, such as the combination of exercise with infusion of catecholamines, where an increase in stroke volume may make an important contribution to the increase in cardiac output.

The indicator-dilution method was used in the dog in an attempt to define the proportion of the "central" blood volume contained in the lung vessels. When injections were made into the pulmonary artery with sampling from the femoral artery only half or even less of the calculated blood volume could be accounted for by the lungs. When a venous injection site was used, the proportion in the lungs was reduced to a third or less. Thus when peripheral sites are used for the inscription of indicator-dilution curves few deductions can be made about either the actual volume or even relative changes in the volume of blood in the lungs.

Background —Unstable angina (UA) is associated with systemic inflammation and with expansion of interferon-γ–producing T lymphocytes. The cause of T-cell activation and the precise role of activated T cells in plaque instability are not understood. Methods and Results —Peripheral blood T cells from 34 patients with stable angina and 34 patients with UA were compared for the distribution of functional T-cell subsets by flow cytometric analysis. Clonality within the T-cell compartment was identified by T-cell receptor spectrotyping and subsequent sequencing. Tissue-infiltrating T cells were examined in extracts from coronary arteries containing stable or unstable plaque. The subset of CD4 + CD28 null T cells was expanded in patients with UA and infrequent in patients with stable angina (median frequencies: 10.8% versus 1.5%, P <0.001). CD4 + CD28 null T cells included a large monoclonal population, with 59 clonotypes isolated from 20 UA patients. T-cell clonotypes from different UA patients used antigen receptors with similar sequences. T-cell receptor sequences derived from monoclonal T-cell populations were detected in the culprit but not in the nonculprit lesion of a patient with fatal myocardial infarction. Conclusions —UA is associated with the emergence of monoclonal T-cell populations, analogous to monoclonal gammopathy of unknown significance. Shared T-cell receptor sequences in clonotypes of different patients implicate chronic stimulation by a common antigen, for example, persistent infection. The unstable plaque but not the stable plaque is invaded by clonally expanded T cells, suggesting a direct involvement of these lymphocytes in plaque disruption.

Abstract Background: Patients with lung cancer with mutations in EGF receptor (EGFR) tyrosine kinase have improved prognosis when treated with EGFR inhibitors. We hypothesized that EGFR mutations may be related to residential radon or passive tobacco smoke. Methods: This hypothesis was investigated by analyzing EGFR mutations in 70 lung tumors from a population of never and long-term former female smokers from Missouri with detailed exposure assessments. The relationship with passive smoking was also examined in never-smoking female lung cancer cases from the Mayo clinic. Results: Overall, the frequency of EGFR mutation was 41% [95% confidence interval (CI), 32%–49%]. Neither radon nor passive-smoking exposure was consistently associated with EGFR mutations in lung tumors. Conclusions: The results suggest that EGFR mutations are common in female, never-smoking lung cancer cases from the United States, and EGFR mutations are unlikely due to exposure to radon or passive smoking. Cancer Epidemiol Biomarkers Prev; 21(6); 988–92. ©2012 AACR.

Object Glomus tumors are rare skull base neoplasms that frequently involve critical cerebrovascular structures and lower cranial nerves. Complete resection is often difficult and may increase cranial nerve deficits. Stereotactic radiosurgery has gained an increasing role in the management of glomus tumors. The authors of this study examine the outcomes after radiosurgery in a large, multicenter patient population. Methods Under the auspices of the North American Gamma Knife Consortium, 8 Gamma Knife surgery centers that treat glomus tumors combined their outcome data retrospectively. One hundred thirty-four patient procedures were included in the study (134 procedures in 132 patients, with each procedure being analyzed separately). Prior resection was performed in 51 patients, and prior fractionated external beam radiotherapy was performed in 6 patients. The patients' median age at the time of radiosurgery was 59 years. Forty percent had pulsatile tinnitus at the time of radiosurgery. The median dose to the tumor margin was 15 Gy. The median duration of follow-up was 50.5 months (range 5–220 months). Results Overall tumor control was achieved in 93% of patients at last follow-up; actuarial tumor control was 88% at 5 years postradiosurgery. Absence of trigeminal nerve dysfunction at the time of radiosurgery (p = 0.001) and higher number of isocenters (p = 0.005) were statistically associated with tumor progression–free tumor survival. Patients demonstrating new or progressive cranial nerve deficits were also likely to demonstrate tumor progression (p = 0.002). Pulsatile tinnitus improved in 49% of patients who reported it at presentation. New or progressive cranial nerve deficits were noted in 15% of patients; improvement in preexisting cranial nerve deficits was observed in 11% of patients. No patient died as a result of tumor progression. Conclusions Gamma Knife surgery was a well-tolerated management strategy that provided a high rate of long-term glomus tumor control. Symptomatic tinnitus improved in almost one-half of the patients. Overall neurological status and cranial nerve function were preserved or improved in the vast majority of patients after radiosurgery.

Regulation of cytosolic Ca2+ concentration ([Ca2+]cyt) in airway smooth muscle (ASM) is a key aspect of airway contractility and can be modulated by inflammation. Mitochondria have tremendous potential for buffering [Ca2+]cyt, helping prevent Ca2+ overload, and modulating other intracellular events. Here, compartmentalization of mitochondria to different cellular regions may subserve different roles. In the present study, we examined the role of Ca2+ buffering by mitochondria and mitochondrial Ca2+ transport mechanisms in the regulation of [Ca2+]cyt in enzymatically dissociated human ASM cells upon exposure to the proinflammatory cytokines TNF-α and IL-13. Cells were loaded simultaneously with fluo-3 AM and rhod-2 AM, and [Ca2+]cyt and mitochondrial Ca2+ concentration ([Ca2+]mito) were measured, respectively, using real-time two-color fluorescence microscopy in both the perinuclear and distal, perimembranous regions of cells. Histamine induced a rapid increase in both [Ca2+]cyt and [Ca2+]mito, with a significant delay in the mitochondrial response. Inhibition of the mitochondrial Na+/Ca2+ exchanger (1 μM CGP-37157) increased [Ca2+]mito responses in perinuclear mitochondria but not distal mitochondria. Inhibition of the mitochondrial uniporter (1 μM Ru360) decreased [Ca2+]mito responses in perinuclear and distal mitochondria. CGP-37157 and Ru360 significantly enhanced histamine-induced [Ca2+]cyt. TNF-α and IL-13 both increased [Ca2+]cyt, which was associated with decreased [Ca2+]mito in the case of TNF-α but not IL-13. The effects of TNF-α on both [Ca2+]cyt and [Ca2+]mito were affected by CGP-37157 but not by Ru360. Overall, these data demonstrate that in human ASM cells, mitochondria buffer [Ca2+]cyt after agonist stimulation and its enhancement by inflammation. The differential regulation of [Ca2+]mito in different parts of ASM cells may serve to locally regulate Ca2+ fluxes from intracellular sources versus the plasma membrane as well as respond to differential energy demands at these sites. We propose that such differential mitochondrial regulation, and its disruption, may play a role in airway hyperreactivity in diseases such as asthma, where [Ca2+]cyt is increased.

Abstract Purpose: miRNA plays an important role in human disease and cancer. We seek to investigate the expression status, clinical relevance, and functional role of miRNA in non–small cell lung cancer. Experimental Design: We conducted miRNA expression profiling in matched lung adenocarcinoma and uninvolved lung using 56 pairs of fresh-frozen (FF) and 47 pairs of formalin-fixed, paraffin-embedded (FFPE) samples from never smokers. The most differentially expressed miRNA genes were evaluated by Cox analysis and log-rank test. Among the best candidate, miR-708 was further examined for differential expression in two independent cohorts. Functional significance of miR-708 expression in lung cancer was examined by identifying its candidate mRNA target and through manipulating its expression levels in cultured cells. Results: Among the 20 miRNAs most differentially expressed between tested tumor and normal samples, high expression level of miR-708 in the tumors was most strongly associated with an increased risk of death after adjustments for all clinically significant factors including age, sex, and tumor stage (FF cohort: HR, 1.90; 95% CI, 1.08–3.35; P = 0.025 and FFPE cohort: HR, 1.93; 95% CI, 1.02–3.63; P = 0.042). The transcript for TMEM88 gene has a miR-708 binding site in its 3′ UTR and was significantly reduced in tumors high of miR-708. Forced miR-708 expression reduced TMEM88 transcript levels and increased the rate of cell proliferation, invasion, and migration in culture. Conclusions: miRNA-708 acts as an oncogene contributing to tumor growth and disease progression by directly downregulating TMEM88, a negative regulator of the Wnt signaling pathway in lung cancer. Clin Cancer Res; 18(13); 3658–67. ©2012 AACR.