# Generation of aptamers for the selective detection and neutralization of soluble lymphotoxin alpha

## TL;DR

Researchers developed DNA aptamers that specifically detect and neutralize a form of the inflammatory protein lymphotoxin alpha, offering a new tool for studying immune signaling.

## Contribution

The paper introduces DNA aptamers that selectively target homotrimeric lymphotoxin alpha without affecting its heterotrimeric form.

## Key findings

- Four aptamer candidates were generated that selectively detect LTα3 but not LTα1β2.
- Optimized aptamers LTa1 and LTa5 significantly reduced LTα3-TNFR1 engagement-related cytotoxicity in vitro.
- The aptamers show potential for future therapeutic use with minimal off-target effects.

## Abstract

Lymphotoxin alpha (LTα) is a potent inflammatory cytokine implicated in the pathophysiology of numerous human autoimmune and inflammatory diseases. Existing as a soluble homotrimer (LTα3) or membrane-bound heterotrimer (LTα1β2), the differential and distinct functions of lymphotoxin signaling have meant that selective targeting of the cytokine with traditional pharmacological agents has proven difficult. While monoclonal antibodies that can neutralize human LTα3in vivo do exist (e.g., pateclizumab), their efficacy and subsequent use within the clinic have been limited. This may be in part perhaps due to cross-reactivity between the homotrimeric and heterotrimeric forms, leading to LTα3-independent effects. Herein, we implement a counter-Systematic Evolution of Ligands by Exponential Enrichment (SELEX) protocol to enrich aptamers targeting LTα3 but not LTα1β2. Through a combination of in silico and in vitro tests, we also refine the aptamer sequences and test their ability to limit LT⍺3-TNFR1 engagement and subsequent cellular cytotoxicity in L929 fibroblasts. We highlight the generation of 4 aptamer candidates that can selectively detect LTα3 but not LTα1β2. Using rational design, we optimize the sequences and show that LTa1 and LTa5 can significantly reduce LTa3-TNFR1 engagement-associated cytotoxicity in vitro, highlighting their future therapeutic potential. These data highlight aptamers for the future investigation of lymphotoxin signaling, limiting off-target impacts on other LT⍺3-dependent mechanisms.

Stephens and colleagues report newly developed DNA aptamers that selectively bind and neutralize homotrimeric lymphotoxin alpha. Achieving structural specificity within a pleiotropic cytokine family, their work introduces powerful molecular tools to interrogate lymphotoxin biology and highlights aptamers as precision modulators of immune signaling.