The mRNA-binding protein HuR promotes hypoxia-induced chemoresistance through posttranscriptional regulation of the proto-oncogene PIM1 in pancreatic cancer cells.

Previously, it has been shown that pancreatic ductal adenocarcinoma (PDA) tumors exhibit high levels of hypoxia, characterized by low oxygen pressure (pO2) and decreased O2 intracellular perfusion. Chronic hypoxia is strongly associated with resistance to cytotoxic chemotherapy and chemoradiation in an understudied phenomenon known as hypoxia-induced chemoresistance. The hypoxia-inducible, pro-oncogenic, serine-threonine kinase PIM1 (Proviral Integration site for Moloney murine leukemia virus 1) has emerged as a key regulator of hypoxia-induced chemoresistance in PDA and other cancers. Although its role in therapeutic resistance has been described previously, the molecular mechanism behind PIM1 overexpression in PDA is unknown. Here, we demonstrate that cis-acting AU-rich elements (ARE) present within a 38-base pair region of the PIM1 mRNA 3'-untranslated region mediate a regulatory interaction with the mRNA stability factor HuR (Hu antigen R) in the context of tumor hypoxia. Predominantly expressed in the nucleus in PDA cells, HuR translocates to the cytoplasm in response to hypoxic stress and stabilizes the PIM1 mRNA transcript, resulting in PIM1 protein overexpression. A reverse-phase protein array revealed that HuR-mediated regulation of PIM1 protects cells from hypoxic stress through phosphorylation and inactivation of the apoptotic effector BAD and activation of MEK1/2. Importantly, pharmacological inhibition of HuR by MS-444 inhibits HuR homodimerization and its cytoplasmic translocation, abrogates hypoxia-induced PIM1 overexpression and markedly enhances PDA cell sensitivity to oxaliplatin and 5-fluorouracil under physiologic low oxygen conditions. Taken together, these results support the notion that HuR has prosurvival properties in PDA cells by enabling them with growth advantages in stressful tumor microenvironment niches. Accordingly, these studies provide evidence that therapeutic disruption of HuR's regulation of PIM1 may be a key strategy in breaking an elusive chemotherapeutic resistance mechanism acquired by PDA cells that reside in hypoxic PDA microenvironments.

Acute aortic regurgitation due to spontaneous rupture of a fenestrated cusp: report in a 65-year-old man and review of seven additional cases.

A 65-year-old man with chronic hypertension developed dyspnea, a cough, and a new diastolic murmur. Two-dimensional echocardiography showed severe aortic regurgitation. No valvular vegetations were identified and blood cultures were negative. Surgical intervention was recommended, but the patient died of an acute intracranial hemorrhage two weeks later. At autopsy, the posterior aortic cusp was flail, due to rupture of the residual cord above two large fenestrations. There was no acute or healed endocarditis. To our knowledge, this is the eighth reported case of aortic valve incompetence due to spontaneous rupture of a fenestrated cusp. Patients ranged in age from 31-67 years (mean, 54), and 4 (50%) were older than 60 years. Seven (88%) of the 8 were men, and 4 (57%) of 7 had chronic hypertension. Analogously, in another four reported cases, aortic insufficiency developed following spontaneous rupture of the fenestrated raphe of an atypical congenitally bicuspid aortic valve. Noninfective and nontraumatic rupture of cord-like aortic valve structures may result in severe acute aortic regurgitation, particularly in men with chronic hypertension.

Death from pulmonary thromboembolism in severe obesity: lack of association with established genetic and clinical risk factors.

Several clinical and environmental conditions are causally related to sudden death from acute pulmonary thromboembolism (APT). Morbid obesity, despite its frequency and association with adverse health effects, is usually considered at most only an additive risk factor for APT. We reviewed protocols and histories from 7227 consecutive autopsies performed between 1985 and 1996 at the Mayo Clinic, including all deaths from APT where no clinical or environmental risk factor could be identified in the study. Body mass indices (BMI) were calculated and compared with those of age- and sex-matched controls who had died suddenly and naturally without evidence of APT. Resistance to activated protein C is the most common molecular clotting defect predisposing to APT, and it is caused by a point mutation in the factor V gene (R506Q). Genomic DNA was extracted from archival tissues of all cases and controls, and the R506Q status was determined by polymerase chain reaction amplification, restriction endonuclease digestion, and direct sequencing. APT was found as the immediate cause of death in 433 patients, with 36 (8%) having no previously established risk factors. Twenty-four of these persons (67%) were morbidly obese (BMI >30 kg/m2). compared with only five controls (14%, P<0.0001). Four patients in both groups, each with a BMI <30 kg/m2. had at least one allele positive for R506Q. Morbid obesity is an independent risk factor in cases of sudden death from APT after the exclusion of previously established clinical, environmental, and molecular risk factors.

The retinoblastoma tumor suppressor pathway modulates the invasiveness of ErbB2-positive breast cancer.

The processes that control the progression of ductal carcinoma in situ (DCIS) to invasive breast cancer remain poorly understood. Epidermal growth factor receptor 2 (ErbB2) overexpression is common in DCIS, as is disruption of the retinoblastoma tumor suppressor (RB) pathway. Here, we examined the cooperative impact of ErbB2 and RB deregulation on facets of disease progression. Our studies demonstrate that RB deficiency altered the expression of key molecules needed for proper cellular organization and epithelial cell-cell adhesion as part of a program related to the epithelial-to-mesenchymal transition (EMT). An increase in the invasive potential of ErbB2-overexpressing cells was observed upon RB depletion. Further, stable knockdown of RB resulted in invasive lesions in orthotopic xenograft assays, compared with DCIS-like lesions developing from RB-proficient cells. Conversely, the invasive phenotype observed in ErbB2-positive cancer models was inhibited through CDK4/6 inhibition in an RB-dependent manner. Finally, in a cohort of DCIS cases, we show that, although elevated levels of ErbB2 are associated with increased risk of a subsequent DCIS recurrence, it is not associated with progression to invasive disease. In contrast, RB loss in ErbB2-positive DCIS cases was associated with increased risk for invasive breast cancer. Taken together, these data demonstrate a key role for the RB pathway in invasion associated with breast tumor progression, and shed light on the key molecular events that promote the progression of DCIS to invasive disease.

N-cadherin haploinsufficiency increases survival in a mouse model of pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDA) is often detected at a late stage, hence the identification of new therapies that have potential to block tumor progression is critical for this lethal disease. N-cadherin upregulation has been observed in many cancers including PDA, however, a causal role for this cell adhesion receptor in disease progression has yet to be defined. The concomitant expression of oncogenic Kras(G12D) and mutant p53 (Trp53(R172H)) in the murine pancreas results in metastatic PDA that recapitulates the cognate features of human pancreatic cancer providing an excellent animal model to identify genes required for tumor progression. Here we determine the consequences of genetically manipulating N-cadherin expression in a mouse model of PDA. Remarkably, mice with reduced N-cadherin expression (that is, Ncad(-/+)) survived 25% longer (177 vs 142 days, P<0.05) than animals expressing two wild-type N-cadherin (Cdh2) alleles. The survival benefit is likely due to a cumulative effect of N-cadherin's role in different aspects of tumorigenesis including tumor-cell survival, growth, migration and invasion. Interestingly, reduced hedgehog signaling may contribute to the better prognosis for the Ncad(-/+) mice. Moreover, the matrix metalloproteinase MMP-7, associated with poor prognosis in PDA, was reduced in Ncad(-/+) tumors. Finally, Ncad(-/+) tumor cells exhibited decreased FGF-stimulated ERK1/2 activation consistent with N-cadherin's ability to promote FGFR signaling. These data support a critical role for N-cadherin in PDA and its potential prognostic value. Additionally, this study provides in vivo genetic evidence that the cell-surface protein N-cadherin represents a promising therapeutic target for the treatment of pancreatic cancer.

RB has a critical role in mediating the in vivo checkpoint response, mitigating secondary DNA damage and suppressing liver tumorigenesis initiated by aflatoxin B1.

Hepatocellular carcinoma (HCC) is a significant worldwide health concern that is associated with discrete etiological events, encompassing viral infection, metabolic stress and genotoxic compounds. In particular, exposure to the genotoxic hepatocarcinogen aflatoxin B1 (AFB1) is a significant factor in the genesis of human liver cancer. Presumably, genetic events associated with HCC could influence the effect of environmental insults, yielding a predilection for tumor development. The retinoblastoma (RB) tumor suppressor pathway is functionally inactivated in HCC through discrete mechanisms; however, the role of RB in suppressing tumorigenesis in this disease is poorly understood. Therefore, we analysed how RB status affects the response to AFB1 in reference to acute exposures and tumor development reflective of chronic exposure. Liver-specific Rb deletion resulted in an aberrant proliferative response to AFB1. This cell-cycle induction was associated with increased levels of secondary genetic damage and failure in appropriate cell-cycle coupling. This effect of RB loss was unique to AFB1 and involved the induction of a non-canonical proliferative pathway, and was not merely reflective of the overall cell-cycle deregulation or aberrant regenerative responses. The acute responses to AFB1 exposure presaged aberrations in hepatocyte nuclear morphology and ploidy with RB loss. Correspondingly, RB-deficient livers showed significantly enhanced susceptibility to liver tumorigenesis initiated by AFB1. Combined, these studies show that RB has a critical role in mediating checkpoint responses in liver tissue to maintain genome integrity and in suppressing tumorigenesis.

Cyclin D1b protein expression in breast cancer is independent of cyclin D1a and associated with poor disease outcome.

Aberrant expression of cyclin D1 protein is a common feature of breast cancer. However, the CCND1 gene encodes two gene products, cyclin D1a and cyclin D1b, which have discrete mechanisms of regulation and impact on cell behavior. A polymorphism at nucleotide 870 in the CCND1 gene, rs603965, influences the relative production of the encoded proteins and can impart increased risk for tumor development. Here, the impact of both the G/A870 polymorphism and cyclin D1b protein production on breast cancer risk, disease phenotype and patient outcome was analysed. In a large multiethnic case-control study, the G/A870 polymorphism conferred no significant risk for breast cancer overall or by stage or estrogen receptor (ER) status. However, the cyclin D1b protein was found to be upregulated in breast cancer, independent of cyclin D1a levels, and exhibited heterogeneous levels in breast cancer specimens. High cyclin D1a expression inversely correlated with the Ki67 proliferation marker and was not associated with clinical outcome. In contrast, elevated cyclin D1b expression was independently associated with adverse outcomes, including recurrence, distant metastasis and decreased survival. Interestingly, cyclin D1b was particularly associated with poor outcome in the context of ER-negative breast cancer. Thus, specific cyclin D1 isoforms are associated with discrete forms of breast cancer and high cyclin D1b protein levels hold prognostic potential.