Antibody engineering for Alzheimer's disease immunotherapies

Many neurodegenerative diseases are characterized by the abnormal accumulation of specific proteins in brain tissue. In Alzheimer's disease, these aggregates consist of amyloid plaques and neurofibrillary tangles (comprised of amyloid-beta [Aβ] and tau, respectively). As both Aβ and tau are neurotoxic, tend to correlate with cognitive decline, and induce inflammation, current models suggest that their aggregation triggers a cascade of downstream pathologies, drives neurodegeneration, and ultimately results in Alzheimer's disease (Bloom 2014; Laurent, Buée, and Blum 2018).

These protein aggregates, therefore, represent an attractive therapeutic target. A major push in clinical research has been in the development and testing of anti-Aβ and, more recently, anti-tau antibodies. While the field has experienced setbacks (Honig et al., 2018; Selkoe 2019), a growing number of antibodies have moved into clinical trials (reviewed in Cummings et al. 2018) and offer the hope that clearing these aggregates may arrest, and potentially reverse, the progression of this debilitating disease. To facilitate this work, we offer end-to-end antibody engineering solutions for simple, rapid, and high-throughput antibody generation and purification.

Highlighted products

SMARTer amplification of antibody variable regions

Our SMARTer RACE kits leverage our unique SMART (Switching Mechanism at the 5' end of RNA Template) technology to provide a true representation of the original heavy (H), kappa light chain  [L (κ)], or lambda light chain [L (λ)] mRNA transcripts from as little as 10 ng of total RNA (Figure 1). Unlike many other RACE-PCR methodologies, our SMARTer RACE technology captures the 5' end of transcripts and, coupled with the addition of the SMART sequence at the 5' end, provides full-length, uniform gene-body coverage. This optimized technology is robust, boasting low background and high amplification efficiency, and can be used even for samples contaminated with gDNA. Finally, to streamline your workflow, these kits include our proprietary In-Fusion Cloning technology (see below) to facilitate the rapid, highly accurate cloning of your RACE fragments.

Efficient, high-throughput generation of antibody constructs

Our In-Fusion HD Cloning Plus technology (Figure 2) enhances antibody discovery and engineering workflows by allowing the rapid generation and screening of functional clones. Screening for individual clones is usually one of the most time-consuming steps of antibody engineering workflows. However, unlike traditional ligation-based methods, In-Fusion Cloning is highly efficient, has an extremely low background, and boasts a >95% success rate for single-insert cloning. Our In-Fusion technology is also fast (15 minutes) and seamless, allowing the directional cloning of up to several PCR fragments into any destination vector in a single reaction. Due to these benefits, our In-Fusion technology has powered multiple high-throughput antibody cloning workflows (Chen et al. 2014; Meng et al. 2015; Spidel et al. 2016; Rudkin et al. 2018).

After antibodies have been generated from your expression construct, our next-generation Capturem purification technology enables rapid, high-quality, and high-throughput antibody purification within 5–15 minutes. Capturem columns and plates are resin-free and leverage a high-capacity nylon-membrane-based system (Figure 3) that contains immobilized Protein A or G (Figure 3), allowing direct antibody purification from complex starting samples (such as hybridoma supernatants and serum). These technologies are available in a variety of formats (minipreps, maxipreps, 24-well plates, and 96-well plates) and are ideal for rapid, high-throughput screening of your antibody clones.

References for AD and the development of Αβ and tau-targeted antibodies

Bloom, G. S. Amyloid-β and tau. JAMA Neurol. 71, 505 (2014).

Cummings, J., Lee, G., Ritter, A. & Zhong, K. Alzheimer's disease drug development pipeline: 2018. Alzheimer's Dement. (New York, N. Y.) 4, 195–214 (2018).

Honig, L. S. et al. Trial of solanezumab for mild dementia due to Alzheimer's disease. N. Engl. J. Med. 378, 321–330 (2018).

Laurent, C., Buée, L. & Blum, D. Tau and neuroinflammation: What impact for Alzheimer's disease and tauopathies? Biomed. J. 41, 21–33 (2018).

Selkoe, D. J. Alzheimer disease and aducanumab: adjusting our approach. Nat. Rev. Neurol. 15, 365–366 (2019).

References citing the use of In-Fusion HD Cloning for high-throughput antibody cloning

Chen, C.-G., Fabri, L. J., Wilson, M. J. & Panousis, C. One-step zero-background IgG reformatting of phage-displayed antibody fragments enabling rapid and high-throughput lead identification. Nucleic Acids Res. 42, e26–e26 (2014).

Meng, W. et al. Efficient generation of monoclonal antibodies from single rhesus macaque antibody secreting cells. MAbs 7, 707–18 (2015).

Rudkin, F. M. et al. Single human B cell-derived monoclonal anti-Candida antibodies enhance phagocytosis and protect against disseminated candidiasis. Nat. Commun. 9, 5288 (2018).

Spidel, J. L., Vaessen, B., Chan, Y. Y., Grasso, L. & Kline, J. B. Rapid high-throughput cloning and stable expression of antibodies in HEK293 cells. J. Immunol. Methods 439, 50–58 (2016).