OBJECTIVES/GOALS: #NAME? METHODS/STUDY POPULATION: Cell culture & protein identification: human T cells were purified from healthy blood, then activated & cultured for 5d. CAR-T cells were collected from infusion bags of cancer patients undergoing CAR-T. Silver staining of naive & activated healthy T-cell lysates was compared; B-II spectrin was upregulated and confirmed by Western blot. Migration assays: naive & activated T-cells were imaged during migration on ICAM-1 and ICAM-1 + CXCL12 coated plates. T-cells were transfected with BII-spectrin cDNA & the chemokine dependence of migration was compared with controls. In-vivo studies: in a melanoma mouse model, BII-spectrin transfected or control T-cells were injected; tumors were followed with serial imaging. Human patient records were examined to correlate endogenous BII-spectrin levels and CAR-T response. RESULTS/ANTICIPATED RESULTS: Activated T-cells downregulate the cytoskeletal protein B-II spectrin compared to naive cells, leading to chemokine-independent migration in in vitro assays and off-target trafficking when CAR-T cells are given in vivo. Restoration of B-II spectrin levels via transfection restores chemokine-dependence of activated T-cells. In a mouse melanoma model, control mice injected with standard activated T-cells showed fewer cells in the tumor site and more cells in the off-target organs (spleen, lungs) when compared to mice injected with B-II spectrin transfected cells. Furthermore, among 3 human patients undergoing CAR-T therapy, those with higher endogenous B-II spectrin levels experienced fewer side-effects, measured by the neurotoxicity and cytokine release syndrome grades. DISCUSSION/SIGNIFICANCE: A major hurdle to widespread CAR-T therapy for cancer is significant, often fatal side-effects. Our work shows that the protein B-II spectrin is downregulated during CAR-T production, and that restoring B-II spectrin levels decreases side-effects while increasing tumor clearance--hopefully translating to better CAR-T regimens for the future.

Based on the data from the Next Generation Virgo cluster Survey (NGVS), we statistically study the photometric properties of globular clusters (GCs), ultra-compact dwarfs (UCDs) and dwarf nuclei in the Virgo core (M87) region. We found an obvious negative color (g - z) gradient in GC system associate with M87, i.e. GCs in the outer regions are bluer. However, such color gradient does not exist in UCD system, neither in dwarf nuclei system around M87. In addition, we found that many UCDs are surrounded by extended, low surface brightness envelopes. The dwarf nuclei and UCDs show different spatial distributions from GCs, with dwarf nuclei and UCDs (especially for the UCDs with visible envelopes) lying at larger distances to the Virgo center. These results support the view that UCDs (at least for a fraction of UCDs) are more tied to dwarf nuclei than to GCs.

Discovery of ultra-compact dwarfs (UCDs) in the past 15 years blurs the once thought clear division between classic globular clusters (GCs) and early-type galaxies. The intermediate nature of UCDs, which are larger and more massive than typical GCs but more compact than typical dwarf galaxies, has triggered hot debate on whether UCDs should be considered galactic in origin or merely the most extreme GCs. Previous studies of various scaling relations, stellar populations and internal dynamics did not give an unambiguous answer to the primary origin of UCDs. In this contribution, we present the first ever detailed study of global dynamics of 97 UCDs (rh ≳ 10 pc) associated with the central cD galaxy of the Virgo cluster, M87. We found that UCDs follow a different radial number density profile and different rotational properties from GCs. The orbital anisotropies of UCDs are tangentially-biased within ~ 40 kpc of M87 and become radially-biased with radius further out. In contrast, the blue GCs, which have similar median colors to our sample of UCDs, become more tangentially-biased at larger radii beyond ~ 40 kpc. Our analysis suggests that most UCDs in M87 are not consistent with being merely the most luminous and extended examples of otherwise normal GCs. The radially-biased orbital structure of UCDs at large radii is in general agreement with the scenario that most UCDs originated from the tidally threshed dwarf galaxies.

Molecular genetic research has provided some evidence for the association between depression and metabolic disorders. We sought to determine if molecular findings are reflected in twin analyses testing if common genetic and environmental risk factors contribute to the co-occurrence of diabetes and depression. Data to derive depression and diabetes were collected from 1,237 male-male twins who participated in the 2005 Vietnam Era Twin Study of Aging (VETSA). The 1,237 twins were comprised of 347 MZ pairs, 3 MZ singletons, 267 DZ pairs and 6 unpaired twins. Depression was defined as a score below 46 on the Short Form-36 mental component summary score. Diabetes was defined by self report, use of anti-diabetic medications and insulin. Twin models were fit to estimate the correlation of genetic and environmental contributions to depression and diabetes. Consistent with other studies these data support the association between depression and diabetes (OR = 1.7; 95%CI: 1.1–2.7). Genetic vulnerability accounted for 50% (95%CI: 32%–65%) of the variance in risk for depression and 69% (95%CI: 52%–81%) of the variance in risk for diabetes. The genetic correlation between depression and diabetes was r = 0.19 (95%CI: 0–0.46) and the non-shared environmental correlation was r = 0.09 (95% CI: 0–0.45). Overall there is little evidence that common genetic and environmental factors account for the co-occurrence of depression and diabetes in middle aged men. Further research in female twins and larger cohorts is warranted.

Breast-feeding is the predominant postnatal transmission route for HIV-1 infection in children. However, a majority of breast-fed infants do not become HIV-infected despite continuous exposure to the virus through their mothers' milk over many months. What protects some breast-fed infants from HIV-1 infection? HIV-1 entry across the infant's mucosal barrier is partially mediated through binding of the HIV-1 surface glycoprotein gp120 to dendritic cell-specific ICAM3-grabbing non-integrin (DC-SIGN) on human dendritic cells. Lewis antigen glycans, present in human milk, bind to DC-SIGN and inhibit HIV-1 transfer to CD4+T lymphocytes. Human milk contains a high amount of unbound, complex oligosaccharides (5–10 g/l) that carry one or more Lewis antigen glycans, and we hypothesized that they compete with gp120 for DC-SIGN binding. Here, we show in two independent assays that physiological concentrations of human milk oligosaccharides significantly reduce gp120 binding to DC-SIGN by more than 80 %. These results may provide an additional explanation for the inhibitory effects of human milk on HIV-1 mother-to-child-transmission. Identifying the specific milk oligosaccharides that interact with DC-SIGN may guide the development of glycan-based drugs that prevent transmission of HIV-1 and other pathogens that use DC-SIGN as an entry point. However, blocking DC-SIGN may be a two-edged sword.

Our main approach to the synthesis and study of hybrid organic/inorganic materials involves incorporating nano-size inorganic polyhedral oligomeric silsesquioxane (POSS) clusters into various polymeric resins. A typical POSS cluster is a discrete silicon and oxygen framework solubilized with organic groups and contains a single reactive site. This lone site of reactivity is used to covalently attach the inorganic macromers pendent to a polymer backbone without causing any crosslinking. This strategy permits the synthesis of melt processable, linear hybrid polymers containing pendent inorganic clusters, and allows us to study the effect these clusters have on chain motion, polymer properties and morphology. The synthesis of norbornenyl-based (POSS) macromers, their ring opening metathesis copolymerizations with varying amounts of norbornene, and analysis of the effect of the pendent POSS group is presented. Ring opening metathesis polymerization permits the easy synthesis of both random and diblock copolymers. Transmission electron microscopy (TEM) clearly images POSS-rich domains against the POSS-free regions. Major differences in polymer morphology are observed as the amount of inorganic POSS is varied, between random and diblock copolymers, as well as between polymers that differ only in the solubilizing cycloalkyl groups on the POSS cluster.

In this work, we present the synthesis and characterization of a new series of wholly-aromatic copolyesters derived from the condensation of various weight fractions of 4,4'-(o- phenylenedioxy)dibenzoic acid (OPDB) and substituted terephthalic acid (BTA) with 2-phenylhydroquinone (PHQ). The Higashi method, involving tosyl chloride and pyridine as solvent, was employed to yield polymer with significant molecular weight. These polymers are intended to enable accessible clearing transition and to control the balance of stiffness and toughness in melt-spun fibers systematically. We report the synthetic details along with characterization of quiescent phase behavior and morphology.

We have taken a unique approach to the synthesis and study of hybrid organic/inorganic materials. Our method involves synthesizing nano-size inorganic P1R7Si8O12 clusters which contain seven inert “R” groups for solubility and only one functional “P” group for polymerization. This strategy permits the synthesis of melt processable, linear hybrid polymers containing pendent inorganic clusters and allows us to study the effect these clusters have on chain motions and polymer properties. The synthesis of norbomenyl-based polyhedral oligomeric silsesquioxane (POSS) macromers, their ring opening metathesis copolymerizations with varying amounts of norbornene, and analysis of the effect of the pendent POSS group is presented. The mechanical relaxation behavior and microstructure of norbornyl-POSS hybrid copolymers have been examined for their dependencies on the mole fraction of POSS-norbornyl monomer, as well as for potential sensitivity to the seven inert “R” groups present in each POSS macromer. POSS copolymerization is observed to enhance the α-relaxation temperature, Tα, in proportion to the mole fraction of POSS-norbornyl comonomer. Interestingly, however, the magnitude of this dependence is larger for POSS-norbornyl comonomer possessing cyclohexyl groups (CyPOSS) than for cyclopentyl groups (CpPOSS). While POSS copolymerization yields only slight enhancement of the tensile storage modulus measured near room temperature, at temperatures lower than a strong mechanical relaxation (β-relaxation near T = -75 °C), there is a significant POSS-reinforcement of the storage modulus.

The effect of the average grading rate and of the number of incremental Ge alloy steps on the threading dislocation density has been studied in 30% Ge relaxed films formed by rapid thermal chemical vapor deposition (RTCVD) on Si substrates. Electron beam induced current (EBIC) images and transmission electron microscopy (TEM) show that threading defects fall in two categories: individual threading dislocations (dark spot defects), and organized clusters of these threads (pile-ups, or dark line defects). The overall surface defect density must include both categories to properly characterize the material. The lowest defect density, 4 × 105cm−2, was found in specimens grown at an average grading rate of 10% Ge per pm thickness.

Co(SiGe)x contacts have been formed on low defect density, relaxed Si0.7Ge0.3 layers by conventional self-aligned contact processing techniques. Test structures measuring the contact metal sheet resistance indicate that the resistivity is high for anneal temperatures from 450°C to 750°C. The lowest sheet resistivity was 100 Ω/□, about 10 times the resistivity of a comparable amount of CoSi2. Contact resistivities, measured by the transmission line method, were as low as 2 × 10−5 Ω cm2. There is a large discrepancy between contact resistivities measured by transmission line and 4 point Kelvin test structures that may be due to the fabricated contact sizes.