Background APOE genotype is the foremost genetic factor modulating β-amyloid (Aβ) deposition and risk of sporadic Alzheimer’s disease (AD). Here we investigated how APOE genotype influences response to anti-Aβ immunotherapy.MethodsAPPSW/PS1dE9 (APP) transgenic mice with targeted replacement of the murine Apoe gene for human APOE alleles received 10D5 anti-Aβ or TY11-15 isotype control antibodies between the ages of 12 and 15 months.ResultsAnti-Aβ immunization decreased both the load of fibrillar plaques and the load of Aβ immunopositive plaques in mice of all APOE backgrounds. Although the relative reduction in parenchymal Aβ plaque load was comparable across all APOE genotypes, APP/ε4 mice showed the greatest reduction in the absolute Aβ plaque load values, given their highest baseline. The immunization stimulated phagocytic activation of microglia, which magnitude adjusted for the post-treatment plaque load was the greatest in APP/ε4 mice implying association between the ε4 allele and impaired Aβ phagocytosis. Perivascular hemosiderin deposits reflecting ensued microhemorrhages were associated with vascular Aβ (VAβ) and ubiquitously present in control mice of all APOE genotypes, although in APP/ε3 mice their incidence was the lowest. Anti-Aβ immunization significantly reduced VAβ burden but increased the number of hemosiderin deposits across all APOE genotypes with the strongest and the weakest effect in APP/ε2 and APP/ε3 mice, respectively.ConclusionsOur studies indicate that APOE genotype differentially modulates microglia activation and Aβ plaque load reduction during anti-Aβ immunotherapy. The APOE ε3 allele shows strong protective effect against immunotherapy associated microhemorrhages; while, conversely, the APOE ε2 allele increases risk thereof.Electronic supplementary materialThe online version of this article (doi:10.1186/s13024-017-0156-1) contains supplementary material, which is available to authorized users.

The cell cycle is a progression of 4 distinct phases (G1, S, G2, and M), with various cycle proteins being essential in regulating this process. We aimed to develop a radiolabeled cyclin-dependent kinase 4/6 (CDK4/6) inhibitor for breast cancer imaging. Our transfluorinated analog ( 18 F-CDKi) was evaluated and validated as a novel PET imaging agent to quantify CDK4/6 expression in estrogen receptor (ER)-positive human epidermal growth factor receptor 2 (HER 2 )-negative breast cancer. Methods: 18 F-CDKi was synthesized and assayed against CDK4/6 kinases. 18 F-CDKi was prepared with a 2-step automated synthetic strategy that yielded the final product with remarkable purity and molar activity. In vitro and in vivo biologic specificity was assessed in a MCF-7 cell line and in mice bearing MCF-7 breast tumors. Nonradioactive palbociclib was used as a blocking agent to investigate the binding specificity and selectivity of 18 F-CDKi. Results: 18 F-CDKi was obtained with an overall radiochemical uncorrected yield of 15% and radiochemical purity higher than 98%. The total time from the start of synthesis to the final injectable formulated tracer is 70 min. The retention time reported for 18 F-CDKi and 19 F-CDKi is 27.4 min as demonstrated by coinjection with 19 F-CDKi in a high-pressure liquid chromatograph. In vivo blood half-life (weighted, 7.03 min) and octanol/ water phase partition coefficient (1.91 ± 0.24) showed a mainly lipophilic behavior. 18 F-CDKi is stable in vitro and in vivo (.98% at 4 h after injection) and maintained its potent targeting affinity to CDK4/6. Cellular uptake experiments performed on the MCF-7 breast cancer cell line (ER-positive and HER 2 -negative) demonstrated specific uptake with a maximum intracellular concentration of about 65% as early as 10 min after incubation. The tracer uptake was reduced to less than 5% when cells were coincubated with a molar excess of palbociclib. In vivo imaging and ex vivo biodistribution of ER-positive, HER 2 -negative MCF-7 breast cancer models showed a specific uptake of approximately 4% injected dose/g of tumor (reduced to ∼0.3% with a 50-fold excess of cold palbociclib). A comprehensive biodistribution analysis also revealed a significantly lower activation of CDK4/6 in nontargeting organs. Conclusion: 18 F-CDKi represents the first 18 F PET CDK4/6 imaging agent and a promising imaging agent for ER-positive, HER 2negative breast cancer.

CD28 is a coreceptor expressed on T lymphocytes. Signaling downstream of CD28 promotes multiple T cell functions such as proliferation, survival, and cytokine secretion. Adhesion to APCs is another function of T cells; however, little is known with regard to the role of CD28 in this process. Our previous studies have shown that CD28 inhibits T cell adhesion, but the underlying mechanism that mediates this process remains unknown. In the present study we discovered that signaling downstream of CD28 resulted in inhibition of Rap1 activity and decreased LFA-1–mediated adhesion. We showed that this was regulated by the recruitment of calcium-promoted Ras inactivator (CAPRI), a GTPase-activating protein, to the plasma membrane downstream of CD28 signaling. CAPRI trafficking to the plasma membrane was secondary to calcium influx and was mediated by its C2A and C2B domains. We conclude that CD28 inhibits Rap1-mediated adhesion by recruiting the protein CAPRI to the plasma membrane.

3737 Background: Hodgkin lymphoma (HL) is among the most curable lymphomas, however 20%-30% of patients relapse after initial chemotherapy, or have primary refractory disease. While 30–50% of these patients may be cured with second line chemotherapy and autologous stem cell transplant, patients who do not obtain a complete response (CR) prior to transplantation, or who relapse after second line therapy have few effective therapeutic options. Novel treatment strategies for these patients are needed. It has been previously shown that in patients with classical HL the malignant Hodgkin Reed-Sternberg cells (HRS) express a high level of the Interleukin-3 receptor (IL-3R). Therefore, we hypothesized that SL-401, a novel biologic conjugate consisting of IL-3 linked to diphtheria toxin, which targets IL-3R, may be an effective approach for selectively targeting and killing HRS cells. Methods: We first assessed the expression of the IL-3R α-chain (CD123) on two nodular sclerosing (HDLM-2 and L-428) and two mixed cellularity (KM:H2 and L-1236) HL cell lines by flow cytometry. Cells were washed with PBS and stained with CD123 PE (BD Pharmingen) as well as other antibodies, including CD15 FITC (BD Pharmingen), for 20 minutes at 4°C and again washed with PBS. Stained cells were acquired using the LSR II (BD) and data were analyzed using Flow Jo (Tree Star). We then tested the sensitivity of these cell lines to SL-401 using a CellTiter Glo in vitro cytotoxicity assay. A CD123 positive eyrthroleukemic cell line (TF-1/H-ras) with known sensitivity to SL-401 was used as a positive control. The cell lines were cultured in the presence or absence of SL-401 for 48 h and assessed for cell viability at concentrations ranging from 3×10−7 to 1.3 μg/ml. Results: We found high expression of CD123 on HDLM-2 and L-428 HL lines (99% and 89%, respectively) and low-to-moderate expression of the receptor on the L1236 and KM-H2 lines (19.5% and 1%, respectively). Based on CD123 expression, we assessed the sensitivity of these cell lines to SL-401. The L428 and HDLM2 cell lines, which exhibited a high expression of CD123, showed sensitivity to SL-401 relative to control starting at approximately 0.3 ng/ml. Cell viability was reduced to 64.8 ± 5% for L-428 and 68.2 ± 8% for HDLM2 when incubated at the highest concentration of drug (1 μg/ml (L428) and 0.7 μg/ml (HDLM2)). The low CD123 expressing cell lines L1236 and KM:H2 were less sensitive to SL-401 and did not exhibit a significant reduction in cell viability even at the highest concentration of SL-401 tested (1.3 ug/ml; 100 ± 2% and 88.4 ± 6%, respectively). Other lymphoid malignant cell lines with CD123 expression reported in the literature were also tested. Interestingly, the T Acute Lymphocytic Leukemia (T-ALL) cell lines DND41 and P12 showed marked sensitivity to SL-401, with a reduction in cell viability to 55.9 ± 4% and 47.9 ± 6%, respectively, in the presence of 1.3 μg/ml of drug. Conclusion: These results suggest that CD123 expression may vary as a function of HL histology, and that sensitivity to SL-401 may correlate with CD123 expression. Based on these results, SL-401, which is currently being evaluated in clinical trials of patients with acute myeloid leukemia, myelodysplastic syndrome, and chronic myeloid leukemia, may be a potential treatment strategy in refractory HL, and warrants further exploration in T-ALL. Exploration of CD123 expression in primary and relapsed/refractory patients with lymphoid malignancies as well as in vivo studies with SL-401 in this setting is currently underway. Disclosures: Brooks: Stemline Therapeutics, Inc: Employment, equity options. Cirrito:Stemline Therapeutics Inc.: Employment, Equity Ownership, Patents & Royalties. Bergstein:Stemline Therapeutics Inc.: Employment, Equity Ownership, Patents & Royalties. O'Connor:Merck: Research Funding; Spectrum: Research Funding; Novartis: Research Funding; Celgene: Consultancy, Research Funding.

Background: Idiopathic normal pressure hydrocephalus (iNPH) is a treatable yet highly underdiagnosed disorder. So far, few epidemiological have been conducted, and little is known about the longterm outcome, risk of symptom progression and natural course. We examined long-term outcome among untreated individuals with suspected iNPH, using a representative, population-based cohort. Outcomes were mortality, dementia and progression of hydrocephalic symptoms. Methods:A population sample of 1235 persons aged 70 years or more were examined between 1986 and 2000 and followed until 2012. iNPH was diagnosed using criteria from international consensus guidelines. Previously shunted individuals were excluded from all analyses. We identified 53 persons (4.3 %) with hydrocephalic ventricular enlargement on CT. Of these, 24 fulfilled criteria for probable iNPH and 29 were classified as asymptomatic or having possible iNPH. Comparisons were made with individuals without hydrocephalic ventricular enlargement (n¼1180). Outcome data were obtained from clinical examinations, the Swedish National Inpatient Register and the Swedish Population Register. Results:Median follow-up time was 11.5 years (Maximum 25 years). Crude five-year mortality was 87.5% among those with probable iNPH, and 19.1% in those without iNPH (p<0.001). In an adjusted Cox regression model, the hazard ratio (HR) for death was 3.8 (95% CI: 2.5-6.0) for probable iNPH. Those with possible iNPH and asymptomatic hydrocephalic ventricular enlargement (n¼29) had increased risk of developing dementia, HR 2.8 (95% CI: 1.5-5.2). Only two individuals (3.8 %) with hydrocephalic ventricular enlargement remained asymptomatic during follow-up. Conclusions: Untreated iNPH is associated with substantial excess mortality and an overall poor prognosis. Most persons with imaging signs of iNPH develop dementia or signs of iNPH. Radiological findings consistent with iNPH are probably more important than previously supposed.Background: White matter hyperintensities (WMHs) are a common MRI finding in individuals over 65 years old. Although commonly associated with neurodegenerative diseases coursing with cognitive decline, the relationship between WMHs and cognition remains controversial. Previous studies have shown an association between WM disruption in the fornix and memory. Our study was designed to test the hypothesis that the relationship between WMHs and memory impairment depends on neuronal degeneration and, in addition, that this association is confined to WM tracts associated with memory function. Methods: The data used in this abstract were obtained from the "Pre-symptomatic Evaluation of Novel or Experimental Treatments for Alzheimer's Disease" (PREVENT-AD). The PREVENT-AD cohort (n¼241) is a first-degree family history positive sample of aging cognitively normal persons. We assessed a subsample (n¼61) who underwent CSF phosphorylated tau (neuronal degeneration) and memory score of the Repeatable Battery for the Assessment of Neuropsychological Status (Memory). In addition,...