17 research outputs found
Controlling the Growth of the Skin Commensal Staphylococcus epidermidis Using d-Alanine Auxotrophy.
Using live microbes as therapeutic candidates is a strategy that has gained traction across multiple therapeutic areas. In the skin, commensal microorganisms play a crucial role in maintaining skin barrier function, homeostasis, and cutaneous immunity. Alterations of the homeostatic skin microbiome are associated with a number of skin diseases. Here, we present the design of an engineered commensal organism, Staphylococcus epidermidis, for use as a live biotherapeutic product (LBP) candidate for skin diseases. The development of novel bacterial strains whose growth can be controlled without the use of antibiotics or genetic elements conferring antibiotic resistance enables modulation of therapeutic exposure and improves safety. We therefore constructed an auxotrophic strain of S. epidermidis that requires exogenously supplied d-alanine. The S. epidermidis NRRL B-4268 Δalr1 Δalr2 Δdat strain (SEΔΔΔ) contains deletions of three biosynthetic genes: two alanine racemase genes, alr1 and alr2 (SE1674 and SE1079), and the d-alanine aminotransferase gene, dat (SE1423). These three deletions restricted growth in d-alanine-deficient medium, pooled human blood, and skin. In the presence of d-alanine, SEΔΔΔ colonized and increased expression of human β-defensin 2 in cultured human skin models in vitro. SEΔΔΔ showed a low propensity to revert to d-alanine prototrophy and did not form biofilms on plastic in vitro. These studies support the potential safety and utility of SEΔΔΔ as a live biotherapeutic strain whose growth can be controlled by d-alanine.IMPORTANCE The skin microbiome is rich in opportunities for novel therapeutics for skin diseases, and synthetic biology offers the advantage of providing novel functionality or therapeutic benefit to live biotherapeutic products. The development of novel bacterial strains whose growth can be controlled without the use of antibiotics or genetic elements conferring antibiotic resistance enables modulation of therapeutic exposure and improves safety. This study presents the design and in vitro evidence of a skin commensal whose growth can be controlled through d-alanine. The basis of this strain will support future clinical studies of this strain in humans
The clustering of galaxies in the SDSS-III Baryon Oscillation Spectroscopic Survey: Analysis of potential systematics
We analyze the density field of galaxies observed by the Sloan Digital Sky
Survey (SDSS)-III Baryon Oscillation Spectroscopic Survey (BOSS) included in
the SDSS Data Release Nine (DR9). DR9 includes spectroscopic redshifts for over
400,000 galaxies spread over a footprint of 3,275 deg^2. We identify,
characterize, and mitigate the impact of sources of systematic uncertainty on
large-scale clustering measurements, both for angular moments of the
redshift-space correlation function and the spherically averaged power
spectrum, P(k), in order to ensure that robust cosmological constraints will be
obtained from these data. A correlation between the projected density of stars
and the higher redshift (0.43 < z < 0.7) galaxy sample (the `CMASS' sample) due
to imaging systematics imparts a systematic error that is larger than the
statistical error of the clustering measurements at scales s > 120h^-1Mpc or k
< 0.01hMpc^-1. We find that these errors can be ameliorated by weighting
galaxies based on their surface brightness and the local stellar density. We
use mock galaxy catalogs that simulate the CMASS selection function to
determine that randomly selecting galaxy redshifts in order to simulate the
radial selection function of a random sample imparts the least systematic error
on correlation function measurements and that this systematic error is
negligible for the spherically averaged correlation function. The methods we
recommend for the calculation of clustering measurements using the CMASS sample
are adopted in companion papers that locate the position of the baryon acoustic
oscillation feature (Anderson et al. 2012), constrain cosmological models using
the full shape of the correlation function (Sanchez et al. 2012), and measure
the rate of structure growth (Reid et al. 2012). (abridged)Comment: Matches version accepted by MNRAS. Clarifications and references have
been added. See companion papers that share the "The clustering of galaxies
in the SDSS-III Baryon Oscillation Spectroscopic Survey:" titl
Single-cell multiplexed cytokine profiling of CD19 CAR-T cells reveals a diverse landscape of polyfunctional antigen-specific response
Background:
It remains challenging to characterize the functional attributes of chimeric antigen receptor (CAR)-engineered T cell product targeting CD19 related to potency and immunotoxicity ex vivo, despite promising in vivo efficacy in patients with B cell malignancies.
Methods:
We employed a single-cell, 16-plex cytokine microfluidics device and new analysis techniques to evaluate the functional profile of CD19 CAR-T cells upon antigen-specific stimulation. CAR-T cells were manufactured from human PBMCs transfected with the lentivirus encoding the CD19-BB-z transgene and expanded with anti-CD3/anti-CD28 coated beads. The enriched CAR-T cells were stimulated with anti-CAR or control IgG beads, stained with anti-CD4 RPE and anti-CD8 Alexa Fluor 647 antibodies, and incubated for 16 h in a single-cell barcode chip (SCBC). Each SCBC contains ~12,000 microchambers, covered with a glass slide that was pre-patterned with a complete copy of a 16-plex antibody array. Protein secretions from single CAR-T cells were captured and subsequently analyzed using proprietary software and new visualization methods.
Results:
We demonstrate a new method for single-cell profiling of CD19 CAR-T pre-infusion products prepared from 4 healthy donors. CAR-T single cells exhibited a marked heterogeneity of cytokine secretions and polyfunctional (2+ cytokine) subsets specific to anti-CAR bead stimulation. The breadth of responses includes anti-tumor effector (Granzyme B, IFN-γ, MIP-1α, TNF-α), stimulatory (GM-CSF, IL-2, IL-8), regulatory (IL-4, IL-13, IL-22), and inflammatory (IL-6, IL-17A) functions. Furthermore, we developed two new bioinformatics tools for more effective polyfunctional subset visualization and comparison between donors.
Conclusions:
Single-cell, multiplexed, proteomic profiling of CD19 CAR-T product reveals a diverse landscape of immune effector response of CD19 CAR-T cells to antigen-specific challenge, providing a new platform for capturing CAR-T product data for correlative analysis. Additionally, such high dimensional data requires new visualization methods to further define precise polyfunctional response differences in these products. The presented biomarker capture and analysis system provides a more sensitive and comprehensive functional assessment of CAR-T pre-infusion products and may provide insights into the safety and efficacy of CAR-T cell therapy
The clustering of galaxies at z~0.5 in the SDSS-III Data Release 9 BOSS-CMASS sample: a test for the LCDM cosmology
We present results on the clustering of 282,068 galaxies in the Baryon
Oscillation Spectroscopic Survey (BOSS) sample of massive galaxies with
redshifts 0.4<z<0.7 which is part of the Sloan Digital Sky Survey III project.
Our results cover a large range of scales from ~0.5 to ~90 Mpc/h. We compare
these estimates with the expectations of the flat LCDM cosmological model with
parameters compatible with WMAP7 data. We use the MultiDark cosmological
simulation together with a simple halo abundance matching technique, to
estimate galaxy correlation functions, power spectra, abundance of subhaloes
and galaxy biases. We find that the LCDM model gives a reasonable description
to the observed correlation functions at z~0.5, which is a remarkably good
agreement considering that the model, once matched to the observed abundance of
BOSS galaxies, does not have any free parameters. However, we find a deviation
(>~10%) in the correlation functions for scales less than ~1 Mpc/h and ~10-40
Mpc/h. A more realistic abundance matching model and better statistics from
upcoming observations are needed to clarify the situation. We also estimate
that about 12% of the "galaxies" in the abundance-matched sample are satellites
inhabiting central haloes with mass M>~1e14 M_sun/h. Using the MultiDark
simulation we also study the real space halo bias b(r) of the matched catalogue
finding that b=2.00+/-0.07 at large scales, consistent with the one obtained
using the measured BOSS projected correlation function. Furthermore, the linear
large-scale bias depends on the number density n of the abundance-matched
sample as b=-0.048-(0.594+/-0.02)*log(n/(h/Mpc)^3). Extrapolating these results
to BAO scales we measure a scale-dependent damping of the acoustic signal
produced by non-linear evolution that leads to ~2-4% dips at ~3 sigma level for
wavenumbers k>~0.1 h/Mpc in the linear large-scale bias.Comment: Replaced to match published version. Typos corrected; 25 pages, 17
figures, 9 tables. To appear in MNRAS. Correlation functions (projected and
redshift-space) and correlation matrices of CMASS presented in Appendix B.
Correlation and covariance data for the combined CMASS sample can be
downloaded from http://www.sdss3.org/science/boss_publications.ph
The clustering of galaxies in the SDSS-III Baryon Oscillation Spectroscopic Survey: measurements of the growth of structure and expansion rate at z=0.57 from anisotropic clustering
We analyze the anisotropic clustering of massive galaxies from the Sloan
Digital Sky Survey III Baryon Oscillation Spectroscopic Survey (BOSS) Data
Release 9 (DR9) sample, which consists of 264,283 galaxies in the redshift
range 0.43 < z < 0.7 spanning 3,275 square degrees. Both peculiar velocities
and errors in the assumed redshift-distance relation ("Alcock-Paczynski
effect") generate correlations between clustering amplitude and orientation
with respect to the line-of-sight. Together with the sharp baryon acoustic
oscillation (BAO) standard ruler, our measurements of the broadband shape of
the monopole and quadrupole correlation functions simultaneously constrain the
comoving angular diameter distance (2190 +/- 61 Mpc) to z=0.57, the Hubble
expansion rate at z=0.57 (92.4 +/- 4.5 km/s/Mpc), and the growth rate of
structure at that same redshift (d sigma8/d ln a = 0.43 +/- 0.069). Our
analysis provides the best current direct determination of both DA and H in
galaxy clustering data using this technique. If we further assume a LCDM
expansion history, our growth constraint tightens to d sigma8/d ln a = 0.415
+/- 0.034. In combination with the cosmic microwave background, our
measurements of DA, H, and growth all separately require dark energy at z >
0.57, and when combined imply \Omega_{\Lambda} = 0.74 +/- 0.016, independent of
the Universe's evolution at z<0.57. In our companion paper (Samushia et al.
prep), we explore further cosmological implications of these observations.Comment: 19 pages, 11 figures, submitted to MNRAS, comments welcom
The Baryon Oscillation Spectroscopic Survey of SDSS-III
The Baryon Oscillation Spectroscopic Survey (BOSS) is designed to measure the
scale of baryon acoustic oscillations (BAO) in the clustering of matter over a
larger volume than the combined efforts of all previous spectroscopic surveys
of large scale structure. BOSS uses 1.5 million luminous galaxies as faint as
i=19.9 over 10,000 square degrees to measure BAO to redshifts z<0.7.
Observations of neutral hydrogen in the Lyman alpha forest in more than 150,000
quasar spectra (g<22) will constrain BAO over the redshift range 2.15<z<3.5.
Early results from BOSS include the first detection of the large-scale
three-dimensional clustering of the Lyman alpha forest and a strong detection
from the Data Release 9 data set of the BAO in the clustering of massive
galaxies at an effective redshift z = 0.57. We project that BOSS will yield
measurements of the angular diameter distance D_A to an accuracy of 1.0% at
redshifts z=0.3 and z=0.57 and measurements of H(z) to 1.8% and 1.7% at the
same redshifts. Forecasts for Lyman alpha forest constraints predict a
measurement of an overall dilation factor that scales the highly degenerate
D_A(z) and H^{-1}(z) parameters to an accuracy of 1.9% at z~2.5 when the survey
is complete. Here, we provide an overview of the selection of spectroscopic
targets, planning of observations, and analysis of data and data quality of
BOSS.Comment: 49 pages, 16 figures, accepted by A
The Clustering of Galaxies in SDSS-III DR9 Baryon Oscillation Spectroscopic Survey: Constraints on Primordial Non-Gaussianity
We analyze the density field of 264,283 galaxies observed by the Sloan
Digital Sky Survey (SDSS)-III Baryon Oscillation Spectroscopic Survey (BOSS)
and included in the SDSS data release nine (DR9). In total, the SDSS DR9 BOSS
data includes spectroscopic redshifts for over 400,000 galaxies spread over a
footprint of more than 3,000 deg^2. We measure the power spectrum of these
galaxies with redshifts 0.43 < z < 0.7 in order to constrain the amount of
local non-Gaussianity, f_NL,local, in the primordial density field, paying
particular attention to the impact of systematic uncertainties. The BOSS galaxy
density field is systematically affected by the local stellar density and this
influences the ability to accurately measure f_NL,local. In the absence of any
correction, we find (erroneously) that the probability that f_NL,local is
greater than zero, P(f_NL,local >0), is 99.5%. After quantifying and correcting
for the systematic bias and including the added uncertainty, we find -45 <
f_NL,local 0) = 91.0%. A more
conservative approach assumes that we have only learned the k-dependence of the
systematic bias and allows any amplitude for the systematic correction; we find
that the systematic effect is not fully degenerate with that of f_NL,local, and
we determine that -82 < f_NL,local < 178 (at 95% confidence) and P(f_NL,local
>0) = 68%. This analysis demonstrates the importance of accounting for the
impact of Galactic foregrounds on f_NL,local measurements. We outline the
methods that account for these systematic biases and uncertainties. We expect
our methods to yield robust constraints on f_NL,local for both our own and
future large-scale-structure investigations.Comment: Matches version to be published in MNRAS. While in press, we found an
error that caused all of our fNL values to be off by a factor of h^2. Our
conclusions (and nearly 100% of the text) are unchanged because they were all
in reference to the probability of fNL > 0 and the relative effect of
systematics on the recovered constraint
The Baryon Oscillation Spectroscopic Survey of SDSS-III
The Baryon Oscillation Spectroscopic Survey (BOSS) is designed to measure the scale of baryon acoustic oscillations (BAO) in the clustering of matter over a larger volume than the combined efforts of all previous spectroscopic surveys of large-scale structure. BOSS uses 1.5 million luminous galaxies as faint as i = 19.9 over 10,000 deg(2) to measure BAO to redshifts z < 0.7. Observations of neutral hydrogen in the Ly alpha forest in more than 150,000 quasar spectra (g < 22) will constrain BAO over the redshift range 2.15 < z < 3.5. Early results from BOSS include the first detection of the large-scale three-dimensional clustering of the Ly alpha forest and a strong detection from the Data Release 9 data set of the BAO in the clustering of massive galaxies at an effective redshift z = 0.57. We project that BOSS will yield measurements of the angular diameter distance d(A) to an accuracy of 1.0% at redshifts z = 0.3 and z = 0.57 and measurements of H(z) to 1.8% and 1.7% at the same redshifts. Forecasts for Ly alpha forest constraints predict a measurement of an overall dilation factor that scales the highly degenerate D-A(z) and H-1(z) parameters to an accuracy of 1.9% at z similar to 2.5 when the survey is complete. Here, we provide an overview of the selection of spectroscopic targets, planning of observations, and analysis of data and data quality of BOSS