Lipid mediated oral tolerance

Project Summary: Autoimmune conditions, allergies and immunogenicity against therapeutic proteins are
caused by unwanted immune responses against proteins. Our long-term goal is to rationally develop an
immunotherapy platform using protein delivery strategies to treat these clinical conditions. We observed that an
oral pre-administration of antigens loaded in lyso-phosphatidylserine (lysoPS), but not in double chain PS, based
nanoparticles effectively reduced unwanted immune response against several protein antigens, forming the
basis for an innovative immunotherapy. The overall objective of this application is to elucidate the cellular and
molecular mechanisms of oral delivery and tolerance mediated by lysoPS based nanoparticles. The rationale for
the proposed research is that once we gain a mechanistic understanding of the tolerogenic property of lysoPS,
it would result in an innovative immunotherapy approach and clinical strategies to prevent and reverse
immunogenicity of therapeutic proteins and to treat/cure autoimmune conditions and allergies. We also found
that there are differences in biophysical characteristics such as PS exposure of the nanoparticles comprised of
lyso and double chain PS. Based on these observations, our working hypothesis is that lysoPS mediated
tolerance is due to its distinct structural/biophysical characteristics that influences its access to gut associated
lymphoid tissue, receptor mediated cellular uptake/signaling and underlying molecular and cellular mechanism
of oral tolerance. Three specific aims (SA) are proposed to test our hypothesis and accomplish our objective
using a multidisciplinary approach. In SA 1, we will investigate the impact of biophysical properties of lysoPS
based lipidic particles and its impact on M-cell mediated particulate uptake. Biophysical characteristics such as
PS exposure, clustering, and lamellar properties will be investigated using orthogonal techniques. We will adopt
in vitro, ex vivo and in vivo studies using M-cell deficient mice to investigate the impact of M-cell engagement of
PS based particles containing model antigen Ovalbumin OVA. In SA 2, we will investigate the impact of PS
exposure on receptor binding and intracellular signaling. Based on our observation that functional blocking
antibody against Tim-4 receptor reversed PS mediated tolerance, we will investigate the role of this receptor
binding in tolerance using advanced cell-biological and imaging techniques and also using knock-out mice
models. In SA 3, we will investigate the mechanism of lysoPS mediated oral tolerance towards OVA, in particular,
the effects on gut resident dendritic cells, generation of regulatory T-cells and B-cell response. The generation
of Tregs will be investigated using OT-II transgenic mice and by adoptive transfer approaches. We will investigate
the impact of Tregs on B-cells by investigating the impact of lysoPS on the expression of IL-10, TGFb and
neuritin. Overall, our combined approach investigating the biophysical, cell biological, innate and adaptive
mechanisms of lysoPS tolerance will lay the foundation for a comprehensive clinical approach to reduce
immunogenicity of therapeutic proteins as well as prevent and reverse autoimmunity and allergies.

Grant Number: 4R01AI169296-04
NIH Institute/Center: NIH
Principal Investigator: SATHY BALU-IYER