Recent literature suggests an increasing incidence of colorectal carcinoma in young patients. We performed a histologic, molecular, and immunophenotypic analysis of patients with sporadic early-onset (r40 years of age) colorectal carcinoma seen at our institution from the years 2000-2010 and compared these tumors to a cohort of consecutively resected colorectal carcinomas seen in patients 440 years of age. A total of 1160 primary colorectal adenocarcinomas were surgically resected for the years 2000 through 2010. Of these, 75 (6%) were diagnoses in patients r40 years of age of which 13 (17%) demonstrated abnormalities in DNA mismatch repair, 4 (5%) were in patients with known germline genetic disorders (two patients with familial adenomatous polyposis, one patient with juvenile polyposis, and one patient with Li-Fraumeni syndrome), and three patients (4%) had long-standing chronic inflammatory bowel disease. The sporadic early-onset colorectal carcinoma group comprised a total of 55 patients (55/1160, 5%) and were compared with a control group comprising 73 consecutively resected colorectal carcinomas with proficient DNA mismatch repair in patients 440 years of age. For the early-onset colorectal carcinoma group, most cases (33/55, 60%) were diagnosed between the age of 35 and 40 years of age. Compared with the control group, the early-onset colorectal carcinoma group was significantly different with respect to tumor location (Po0.007) with 80% (44/55 cases) identified in either the sigmoid colon (24/55, 44%) or rectum (20/55, 36%). Morphologically, early-onset colorectal carcinomas more frequently displayed adverse histologic features compared with the control colorectal carcinoma group such as signet ring cell differentiation (7/55, 13% vs 1/73, 1%, P ¼ 0.021), perineural invasion (16/55, 29% vs 8/73, 11%, P ¼ 0.009) and venous invasion (12/55, 22% vs 4/73, 6%, P ¼ 0.006). A precursor adenomatous lesion was less frequently identified in the early-onset colorectal carcinoma group compared with the control group (19/55, 35% vs 39/73, 53%, P ¼ 0.034). Of the early-onset colorectal carcinomas, only 2/45 cases (4%) demonstrated KRAS mutations compared with 11/73 (15%) of the control group colorectal adenocarcinomas harboring KRAS mutations, although this difference did not reach statistical significance (P ¼ 0.13). BRAF V600E mutations were not identified in the early-onset colorectal carcinoma group. No difference was identified between the two

Background: UV-induced damage can induce apoptosis or trigger DNA repair mechanisms. Minor DNA damage is thought to halt the cell cycle to allow effective repair, while more severe damage can induce an apoptotic program. Of the two major types of UV-induced DNA lesions, it has been reported that repair of CPD, but not 6-4PP, abrogates mutation. To address whether the two major forms of UV-induced DNA damage, can induce differential biological effects, NERdeficient cells containing either CPD photolyase or 6-4 PP photolyase were exposed to UV and examined for alterations in cell cycle and apoptosis. In addition, pTpT, a molecular mimic of CPD was tested in vitro and in vivo for the ability to induce cell death and cell cycle alterations.

Vascular endothelial growth factor (VEGF) exerts crucial functions during pathological angiogenesis and normal physiology. We observed increased hematocrit (60-75%) after high-grade inhibition of VEGF by diverse methods, including adenoviral expression of soluble VEGF receptor (VEGFR) ectodomains, recombinant VEGF Trap protein and the VEGFR2-selective antibody DC101. Increased production of red blood cells (erythrocytosis) occurred in both mouse and primate models, and was associated with near-complete neutralization of VEGF corneal micropocket angiogenesis. High-grade inhibition of VEGF induced hepatic synthesis of erythropoietin (Epo, encoded by Epo) >40-fold through a HIF-1alpha-independent mechanism, in parallel with suppression of renal Epo mRNA. Studies using hepatocyte-specific deletion of the Vegfa gene and hepatocyte-endothelial cell cocultures indicated that blockade of VEGF induced hepatic Epo by interfering with homeostatic VEGFR2-dependent paracrine signaling involving interactions between hepatocytes and endothelial cells. These data indicate that VEGF is a previously unsuspected negative regulator of hepatic Epo synthesis and erythropoiesis and suggest that levels of Epo and erythrocytosis could represent noninvasive surrogate markers for stringent blockade of VEGF in vivo.

A defect in RelB, a member of the Rel/nuclear factor (NF)-κB family of transcription factors, affects antigen presenting cells and the formation of lymphoid organs, but its role in T lymphocyte differentiation is not well characterized. Here, we show that RelB deficiency in mice leads to a selective decrease of NKT cells. RelB must be expressed in an irradiation-resistant host cell that can be CD1d negative, indicating that the RelB expressing cell does not contribute directly to the positive selection of CD1d-dependent NKT cells. Like RelB-deficient mice, aly/aly mice with a mutation for the NF-κB–inducing kinase (NIK), have reduced NKT cell numbers. An analysis of NK1.1 and CD44 expression on NKT cells in the thymus of aly/aly mice reveals a late block in development. In vitro, we show that NIK is necessary for RelB activation upon triggering of surface receptors. This link between NIK and RelB was further demonstrated in vivo by analyzing RelB+/− × aly/+ compound heterozygous mice. After stimulation with α-GalCer, an antigen recognized by NKT cells, these compound heterozygotes had reduced responses compared with either RelB+/− or aly/+ mice. These data illustrate the complex interplay between hemopoietic and nonhemopoietic cell types for the development of NKT cells, and they demonstrate the unique requirement of NKT cells for a signaling pathway mediated by NIK activation of RelB in a thymic stromal cell.

We assessed the frequency and clinicopathologic significance of 19 genes currently identified as significantly mutated in myeloid neoplasms, RUNX1, ASXL1, TET2, CEBPA, IDH1, IDH2, DNMT3A, FLT3, NPM1, TP53, NRAS, EZH2, CBL, U2AF1, SF3B1, SRSF2, JAK2, CSF3R, and SETBP1, across 93 cases of acute myeloid leukemia (AML) using capture target enrichment and next-generation sequencing. Of these cases, 79% showed at least one nonsynonymous mutation, and cases of AML with recurrent genetic abnormalities showed a lower frequency of mutations versus AML with myelodysplasia-related changes (P<0.001). Mutational analysis further demonstrated that TP53 mutations are associated with complex karyotype AML, whereas ASXL1 and U2AF1 mutations are associated with AML with myelodysplasia-related changes. Furthermore, U2AF1 mutations were specifically associated with trilineage morphologic dysplasia. Univariate analysis demonstrated that U2AF1 and TP53 mutations are associated with absence of clinical remission, poor overall survival (OS), and poor disease-free survival (DFS; P<0.0001), whereas TET2 and ASXL1 mutations are associated with poor OS (P<0.03). In multivariate analysis, U2AF1 and TP53 mutations retained independent prognostic significance in OS and DFS, respectively. Our results demonstrate unique relationships between mutations in AML, clinicopathologic prognosis, subtype categorization, and morphologic dysplasia.

Phospholipid asymmetry in the red blood cell (RBC) lipid bilayer is well maintained during the life of the cell, with phosphatidylserine (PS) virtually exclusively located in the inner monolayer. Loss of phospholipid asymmetry, and consequently exposure of PS, is thought to play an important role in red cell pathology. The anemia in the human thalassemias is caused by a combination of ineffective erythropoiesis (intramedullary hemolysis) and a decreased survival of adult RBCs in the peripheral blood. This premature destruction of the thalassemic RBC could in part be due to a loss of phospholipid asymmetry, because cells that expose PS are recognized and removed by macrophages. In addition, PS exposure can play a role in the hypercoagulable state reported to exist in severe β-thalassemia intermedia. We describe PS exposure in RBCs of 56 comparably anemic patients with different genetic backgrounds of the α- or β-thalassemia phenotype. The use of fluorescently labeled annexin V allowed us to determine loss of phospholipid asymmetry in individual cells. Our data indicate that in a number of thalassemic patients, subpopulations of red cells circulate that expose PS on their outer surface. The number of such cells can vary dramatically from patient to patient, from as low as that found in normal controls (less than 0.2%) up to 20%. Analysis by fluorescent microscopy of β-thalassemic RBCs indicates that PS on the outer leaflet is distributed either over the entire membrane or localized in areas possibly related to regions rich in membrane-bound α-globin chains. We hypothesize that these membrane sites in which iron carrying globin chains accumulate and cause oxidative damage, could be important in the loss of membrane lipid organization. In conclusion, we report the presence of PS-exposing subpopulations of thalassemic RBC that are most likely physiologically important, because they could provide a surface for enhancing hemostasis as recently reported, and because such exposure may mediate the rapid removal of these RBCs from the circulation, thereby contributing to the anemia.

The importance of lymphotoxin (LT) βR (LTβR) as a regulator of lymphoid organogenesis is well established, but its role in host defense has yet to be fully defined. In this study, we report that mice deficient in LTβR signaling were highly susceptible to infection with murine CMV (MCMV) and early during infection exhibited a catastrophic loss of T and B lymphocytes, although the majority of lymphocytes were themselves not directly infected. Moreover, bone marrow chimeras revealed that lymphocyte survival required LTα expression by hemopoietic cells, independent of developmental defects in lymphoid tissue, whereas LTβR expression by both stromal and hemopoietic cells was needed to prevent apoptosis. The induction of IFN-β was also severely impaired in MCMV-infected LTα−/− mice, but immunotherapy with an agonist LTβR Ab restored IFN-β levels, prevented lymphocyte death, and enhanced the survival of these mice. IFN-αβR−/− mice were also found to exhibit profound lymphocyte death during MCMV infection, thus providing a potential mechanistic link between type 1 IFN induction and lymphocyte survival through a LTαβ-dependent pathway important for MCMV host defense.

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