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Methods for Analysis of Nanoparticle Immunosuppressive Properties.

Edward Cedrone¹, Timothy M Potter¹, Barry W Neun¹

¹Nanotechnology Characterization Laboratory, Cancer Research Technology Program, Frederick National Laboratory for Cancer Research, Frederick, MD, USA.

Methods in Molecular Biology (Clifton, N.J.)

|March 20, 2024

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This study compares in vivo T cell dependent antibody response (TDAR) and in vitro human lymphocyte activation (HuLA) assays for assessing drug-induced immunosuppression, particularly for nanoparticles. The findings support using the HuLA assay as a viable alternative to TDAR in early drug development.

Area of Science:

Pharmacology
Immunotoxicology
Nanomedicine

Background:

Adverse drug effects on the immune system, specifically immunosuppression, are a major concern, leading to significant drug withdrawals.
Traditional methods like the T cell dependent antibody response (TDAR) assay are time-consuming and require substantial sample quantities.
Novel drug candidates necessitate efficient and reliable methods for early-stage immunotoxicity assessment.

Purpose of the Study:

To describe and compare the experimental procedures for evaluating nanoparticle immunosuppressive properties using both TDAR and HuLA assays.
To discuss the in vitro-in vivo correlation between the TDAR and HuLA methods.
To present a case study using an iron oxide nanoparticle formulation (Feraheme).

Main Methods:

Keywords:

Antigen HuLa Immunosuppression Leukocyte proliferation Nanoparticles TDAR

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In vivo assessment using the T cell dependent antibody response (TDAR) assay with Keyhole Limpet Hemocyanin (KLH) antigen over 28 days.

In vitro assessment using the human lymphocyte activation (HuLA) assay with donor leukocytes, Fluzone, and nanoparticles, measuring antigen-specific lymphocyte proliferation via BrdU incorporation.

Enzyme-linked immunosorbent assay (ELISA) for measuring BrdU incorporation in the HuLA assay.

Main Results:

The study details the methodologies for both TDAR and HuLA assays in the context of nanoparticle immunotoxicity.
The in vitro-in vivo correlation between the two assays is discussed, highlighting the potential of the HuLA assay.
A case study involving Feraheme, an iron oxide nanoparticle, is presented to illustrate the application of these methods.

Conclusions:

The human lymphocyte activation (HuLA) assay offers a promising in vitro alternative to the traditional T cell dependent antibody response (TDAR) assay for early preclinical assessment of drug-induced immunosuppression.
The HuLA assay is suitable for evaluating the immunomodulatory effects of nanoparticles.
Further validation and correlation studies are essential for refining the predictive value of in vitro assays in immunotoxicology.