Identification of a nanobody specific to human pulmonary surfactant protein A

Abstract

Nanobody (Nb) is a promising vector for targeted drug delivery. This study aims to identify an Nb that can specifically target the lung by binding human pulmonary surfactant protein A (SP-A). Human lung frozen tissue sections were used for 3 rounds of biospanning of our previously constructed Nb library for rat SP-A to establish a sub-library of Nb, which specifically bound human lung tissues. Phage-ELISA was performed to screen the sub-library to identify Nb4, which specifically bound human SP-A. The binding affinity Kd of Nb4 to recombinant human SP-A was 7.48 × 10−7 M. Nb4 (19 kDa) was stable at 30 °C–37 °C and pH 7.0–7.6 and specifically bound the SP-A in human lung tissue homogenates, human lung A549 cells, and human lung tissues, whereas didn’t react with human liver L-02 cells, kidney 293T cells, and human tissues from organs other than the lung. Nb4 accumulated in the lung of nude mice 5 minutes after a tail vein injection of Nb4 and was excreted 3 hours. Short-term exposure (one month) to Nb4 didn’t cause apparent liver and kidney toxicity in rats, whereas 3-month exposure resulted in mild liver and kidney injuries. Nb4 may be a promising vector to specifically deliver drugs to the lung.

Development and assessment of the efficacy and safety of human lung-targeting liposomal methylprednisolone crosslinked with nanobody

“…Due to its abundant pulmonary expression but minimal expression in other tissues, SP-A is thought to be highly specific to the lung and thus can serve as an ideal target to the lung (Hamers-Casterman et al, 1993). Here, we used our previously developed nanobody to human lung SP-A (SPANb) (He et al, 2017), NSSLs as the drug delivery system, and MPS as the therapeutic drug to successfully prepare the human lung targeting GC drug MPS-NSSLs-SPANb (Figure 9). MPS-NSSLs-SPANb particles developed in the current study exhibit obvious advantages.…”

“…Alveolar surfactant protein A (SP-A) is expressed the most abundantly on type II alveolar epithelial cells whereas barely found in the extrapulmonary organs, and thus SP-A is believe to be the most ideal lung tissue target (Kuroki et al, 1998 ). In our previous study, we have successfully developed anti-rat SP-A Nbs (rSPANb) (Wang et al, 2015 ) and anti-human SP-ANbs (hSPANb) (He et al, 2017 ) and have found rSPANbs and hSPANbs target the rat lung and human lung tissues highly specifically. The current study aimed to use hSPANbs as the lung targeting molecule, the clinically commonly used GC hormone methylprednisolone (MPS), and nano-sterically stable liposome (NSSLs) as the drug delivery vector to first develop a GC hormone agent (MPS-NSSLs-SPANb) that specifically targets the human lung.…”

Nanobodies in the limelight: Multifunctional tools in the fight against viruses

“…Similar to immunohistochemistry, the permeabilization methods used in electron microscopy can affect the ultrastructure. The higher penetrability of nanobodies allows the use of milder permeabilization methods maintaining the ultrastructure [40,41]. In addition, and like in SRM, the use of nanobodies leads to a reduction in the linkage error and concomitant increase in image resolution.…”