hnRNPA1 couples nuclear export and translation of specific mRNAs downstream of FGF-2/S6K2 signalling

The increased cap-independent translation of anti-apoptotic proteins is involved in the development of drug resistance in lung cancer but signalling events regulating this are poorly understood. Fibroblast growth factor 2 (FGF-2) signalling-induced S6 kinase 2 (S6K2) activation is necessary, but the downstream mediator(s) coupling this kinase to the translational response is unknown. Here, we show that S6K2 binds and phosphorylates hnRNPA1 on novel Ser4/6 sites, increasing its association with BCL-XL and XIAP mRNAs to promote their nuclear export. In the cytoplasm, phosphoS4/6-hnRNPA1 dissociates from these mRNAs de-repressing their IRES-mediated translation. This correlates with the phosphorylation-dependent association of hnRNPA1 with 14-3-3 leading to hnRNPA1 sumoylation on K183 and its re-import into the nucleus. A non-phosphorylatible, S4/6A mutant prevented these processes, hindering the pro-survival activity of FGF-2/S6K2 signalling. Interestingly, immunohistochemical staining of lung and breast cancer tissue samples demonstrated that increased S6K2 expression correlates with decreased cytoplasmic hnRNPA1 and increased BCL-XL expression. In short, phosphorylation on novel N-term sites of hnRNPA1 promotes translation of anti-apoptotic proteins and is indispensable for the pro-survival effects of FGF-2

Analysis of Lymphocyfic Infiltration in Uveal Melanoma

Among 27 uveal melanomas, five were found to contain tumor infiltrating lymphocytes (TILs). Four had high levels of lymphocytes, and the fifth had comparatively low levels but adequate numbers for comprehensive analysis. The TILs were analyzed by flow cytometry to determine the relative proportions of lymphocyte subsets and markers of lymphocyte activation. The results show the predominance of T-suppressor/cytotoxic lymphocytes and insignificant levels of B-cells present in the infiltrate. The T-suppressor/cytotoxic cells were generally activated to a higher degree than the T-helper cells when assayed for levels of the histocompatibility antigen, HLA-DR. T-helper cells expressed more interleukin (IL-2) receptor (Tac) than T-suppressor/cytotoxic cells. Invest Ophthalmol Vis Sci 31: [2106][2107][2108][2109][2110]1990 Uveal melanomas, like their cutaneous counterpart, are considered to be relatively susceptible to immunologic influences because of reports of spontaneous regression, 1 -2 of the development of vitiligo and halo nevi, 5 Such intraocular transplantation can be prevented by prior adoptive transfer of humoral or cellular immunity to the recipient. " 14 Some investigators report the predominance of the T-cytotoxic/suppressor cell subpopulation; 1213 others found a majority of T-helper/inducer cells. 10 " 14 There is, however, no convincing evidence that a more favorable prognosis is associated with lymphocytic infiltration in uveal melanomas, 715 and it remains uncertain whether tumor infiltrating lymphocytes (TILs) play a significant role in tumor immunity. The availability of precisely defined monoclonal-antibody markers and flow-cytometric techniques makes it From the *Department of Biochemistry and the fTennent Institute of Ophthalmology, University of Glasgow, Glasgow, Scotland. Reprint requests: Fiona H. Durie, Department of Biochemistry, University of Glasgow, Glasgow G12 8QQ, UK. possible to do a detailed objective analysis of a large number of lymphocytes in any tumour. We characterized the phenotype of the TILs and studied the expression of the histocompatibility antigen, HLA-DR, and interleukin (IL-2) receptor (Tac), which are markers of activation, on the surface of T-helper and T-suppressor/cytotoxic lymphocytes in ocular melanoma. Materials and Methods Preparation of Cells Tissue was obtained from five choroidal melanomas which were removed either by local resection or by enucleation. Slices were cut from the apical part of the fresh specimen after preliminary examination, and these were transferred to RPMI-1640 medium (Gibco, Grand Island, NY). Spilled cells were teased out using a sterile needle. The cells were then harvested, washed by centrifugation at 1000 rpm for 15 min, counted, and adjusted to 1X10 6 cells/ml. The remaining cell suspensions of TILs and tumor cells were cryopreserved in medium containing 90% fetal calf serum and 10% dimethylsulfoxide and stored in liquid nitrogen until use. Four of the five patients consented to venipuncture, and peripheral blood lymphocytes were separated on a discontinuous ficoll density gradient. 1617 Preparation of Samples Cell suspensions of TILs and tumor cells were washed and resuspended in 1 ml of phosphate-buffered saline. Cells (50 ii\) were incubated on ice with appropriate fluorescein isothiocyanate (FITC)-conjugated leu series monoclonal antibody and phycoerythrin (PE) conjugated antibody (Becton-Dickinson, 2106 Downloaded from iovs.arvojournals.org on 06/28/201

The relation between genotype and phenotype in cystic fibrosis - Analysis of the most common mutation (ΔF 508)

Background and methods. Both the clinical manifestations of cystic fibrosis and the genotypes of patients are heterogeneous but the associations between the two are not known. We therefore studied blood samples from 293 patients with cystic fibrosis for the presence of the most common disease-causing mutation (ΔF 508) on chromosome 7 and compared the results with the clinical manifestations of the disease. Results. The prevalence of the ΔF 508 allele in the cohort was 71 percent; 52 percent of the patients were homozygous for the mutation, 40 percent were heterozygous, and 8 percent had other, undefined mutations. The patients who were homozygous for the mutation had received a diagnosis of cystic fibrosis at an earlier age and had a greater frequency of pancreatic insufficiency; pancreatic insufficiency was present in 99 percent of the homozygous patients, but in 72 percent of the heterozygous patients and only 36 percent of the patients with other genotypes. The patients with pancreatic insufficiency in all three genotype groups had similar clinical characteristics, reflected by an early age at diagnosis, similar sweat chloride values at diagnosis, similar severity of pulmonary disease, and similar percentiles for weight. In contrast, the patients in the heterozygous-genotype and other-genotype groups who did not have pancreatic insufficiency were older and had milder disease. They had lower sweat chloride value at diagnosis, normal nutritional status, and better pulmonary function after adjustment for age. Conclusions. The variable clinical course in patients with cystic fibrosis can be attributed at least in part to specific genotypes at the locus of the cystic fibrosis gene.published_or_final_versio

Role of radiography, MRI and FDG-PET/CT in diagnosing, staging and therapeutical evaluation of patients with multiple myeloma

Multiple myeloma is a malignant B-cell neoplasm that involves the skeleton in approximately 80% of the patients. With an average age of 60 years and a 5-years survival of nearly 45% Brenner et al. (Blood 111:2516–2520, 35) the onset is to be classified as occurring still early in life while the disease can be very aggressive and debilitating. In the last decades, several new imaging techniques were introduced. The aim of this review is to compare the different techniques such as radiographic survey, multidetector computed tomography (MDCT), whole-body magnetic resonance imaging (WB-MRI), fluorodeoxyglucose positron emission tomography- (FDG-PET) with or without computed tomography (CT), and 99mTc-methoxyisobutylisonitrile (99mTc-MIBI) scintigraphy. We conclude that both FDG-PET in combination with low-dose CT and whole-body MRI are more sensitive than skeleton X-ray in screening and diagnosing multiple myeloma. WB-MRI allows assessment of bone marrow involvement but cannot detect bone destruction, which might result in overstaging. Moreover, WB-MRI is less suitable in assessing response to therapy than FDG-PET. The combination of PET with low-dose CT can replace the golden standard, conventional skeletal survey. In the clinical practise, this will result in upstaging, due to the higher sensitivity

Consensus recommendations for risk stratification in multiple myeloma: report of the International Myeloma Workshop Consensus Panel 2.

A panel of members of the 2009 International Myeloma Workshop developed guidelines for risk stratification in multiple myeloma. The purpose of risk stratification is not to decide time of therapy but to prognosticate. There is general consensus that risk stratification is applicable to newly diagnosed patients; however, some genetic abnormalities characteristic of poor outcome at diagnosis may suggest poor outcome if only detected at the time of relapse. Thus, in good-risk patients, it is necessary to evaluate for high-risk features at relapse. Although detection of any cytogenetic abnormality is considered to suggest higher-risk disease, the specific abnormalities considered as poor risk are cytogenetically detected chromosomal 13 or 13q deletion, t(4; 14) and del17p, and detection by fluorescence in situ hybridization of t(4; 14), t(14; 16), and del17p. Detection of 13q deletion by fluorescence in situ hybridization only, in absence of other abnormalities, is not considered a high-risk feature. High serum beta(2)-microglobulin level and International Staging System stages II and III, incorporating high beta(2)-microglobulin and low albumin, are considered to predict higher risk disease. There was a consensus that the high-risk features will change in the future, with introduction of other new agents or possibly new combinations. (Blood. 2011; 117(18): 4696-4700

Mobilization in myeloma revisited: IMWG consensus perspectives on stem cell collection following initial therapy with thalidomide-, lenalidomide-, or bortezomib-containing regimens

The past decade has witnessed a paradigm shift in the initial treatment of multiple myeloma with the introduction of novel agents such as thalidomide, lenalidomide, and bortezomib, leading to improved outcomes. High-dose therapy and autologous stem cell transplantation remains an important therapeutic option for patients with multiple myeloma eligible for the procedure. Before the advent of the novel agents, patients underwent stem cell collection prior to significant alkylating agent exposure, given its potential deleterious effect on stem cell collection. With increasing use of the novel agents in the upfront setting, several reports have emerged raising concerns about their impact on the ability to collect stem cells. An expert panel of the International Myeloma Working Group (IMWG) was convened to examine the implications of these therapies on stem collection in patients with myeloma and to develop recommendations for addressing these issues. Here we summarize the currently available data and present our perspective on the problem and potential options to overcome this problem. Specifically, we recommend early mobilization of stem cells, preferably within the first 4 cycles of initial therapy, in patients treated with novel agents and encourage participation in clinical trials evaluating novel approaches to stem cell mobilization. (Blood. 2009;114:1729-1735

Clinical predictors of long-term survival in newly diagnosed transplant eligible multiple myeloma - an IMWG Research Project

Purpose: multiple myeloma is considered an incurable hematologic cancer but a subset of patients can achieve long-term remissions and survival. The present study examines the clinical features of long-term survival as it correlates to depth of disease response. Patients & Methods: this was a multi-institutional, international, retrospective analysis of high-dose melphalan-autologous stem cell transplant (HDM-ASCT) eligible MM patients included in clinical trials. Clinical variable and survival data were collected from 7291 MM patients from Czech Republic, France, Germany, Italy, Korea, Spain, the Nordic Myeloma Study Group and the United States. Kaplan–Meier curves were used to assess progression-free survival (PFS) and overall survival (OS). Relative survival (RS) and statistical cure fractions (CF) were computed for all patients with available data. Results: achieving CR at 1 year was associated with superior PFS (median PFS 3.3 years vs. 2.6 years, p < 0.0001) as well as OS (median OS 8.5 years vs. 6.3 years, p < 0.0001). Clinical variables at diagnosis associated with 5-year survival and 10-year survival were compared with those associated with 2-year death. In multivariate analysis, age over 65 years (OR 1.87, p = 0.002), IgA Isotype (OR 1.53, p = 0.004), low albumin < 3.5 g/dL (OR = 1.36, p = 0.023), elevated beta 2 microglobulin ≥ 3.5 mg/dL (OR 1.86, p < 0.001), serum creatinine levels ≥ 2 mg/dL (OR 1.77, p = 0.005), hemoglobin levels < 10 g/dL (OR 1.55, p = 0.003), and platelet count < 150k/μL (OR 2.26, p < 0.001) appeared to be negatively associated with 10-year survival. The relative survival for the cohort was ~0.9, and the statistical cure fraction was 14.3%. Conclusions: these data identify CR as an important predictor of long-term survival for HDM-ASCT eligible MM patients. They also identify clinical variables reflective of higher disease burden as poor prognostic markers for long-term survival