Novel neutralizing mAb offers potential treatment for cerebral ischaemic injury

Researchers from a joint team at the University of Hong Kong (HKU) and Chinese Academy of Sciences have developed a novel neutralizing monoclonal antibody (mAb; 6H2) that targets circulating adipocyte fatty acid–binding protein (A-FABP), offering potential in alleviating cerebral ischaemic injury.

“Our research will lay a foundation for further advancement in ischaemic stroke treatment,” said Professor Ruby Lai-Chong Hoo of the Department of Pharmacology and Pharmacy, HKU. Patent applications for 6H2 have been filed in China, Europe and US.

Circulating A-FABP can exacerbate cerebral ischaemia injury by disrupting the blood-brain barrier (BBB) through inducing expression of matrix metallopeptidase–9 (MMP-9) via activating the c-Jun N-terminal kinase (JNK)/c-Jun pathway. Although a small-molecule inhibitor of A-FABP demonstrated effectiveness in treating ischaemic stroke, its off-target effects on the heart limit further clinical development. [Sci Rep 2017;7:40657; Eur Heart J 2020;41:3169-3180; PLoS One 2012;7:e44570; Naunyn Schmiedebergs Arch Pharmacol 2011;384:255-263]

In the current study, the researchers generated four murine mAbs against human A-FABP using hybridoma technology. Among these mAbs, only 6H2 exhibited high affinity and specificity to A-FABP expressed in humans and mice, while showing no evidence of cross reactivity with human or mouse epidermal FABP and heart FABP. [Br J Pharmacol 2023;181:1238-1255]

Additionally, 6H2 demonstrated strong neutralizing activity against A-FABP, suppressing both the p-JNK/JNK ratio and MMP-9 expression activated by A-FABP in peritoneal macrophages isolated from C57BL/6N mice, compared with negative control of mouse immunoglobulin G (IgG).

The efficacy of 6H2 for cerebral ischaemia was subsequently assessed with the mouse model subjected to middle cerebral artery occlusion (MCAO) or sham operation. Mice received intravenous 6H2 (1.8 mg/kg or 3.6 mg/kg) or IgG control (1.8 mg/kg) 1 hour after surgery, followed by every 3 days in the 7-day postsurgery period, until reperfusion was achieved.

At 24 hours after MCAO, mice in the 6H2 1.8 mg/kg group showed significant reduction in infract volume vs the control group (12 percent vs 40 percent; p<0.005), with a similar effect observed in the 6H2 3.6 mg/kg group. Additionally, ischaemic oedema and levels of proinflammatory cytokines in the brain (monocyte chemoattractant protein–1, tumour necrosis factor–α and interleukin [IL]-1β, and IL-6) significantly decreased in the 6H2 1.8 mg vs control group (p<0.05 for each). Over the 7-day postsurgery period, the 6H2 group achieved improved neurological deficits and survival vs the IgG control group (p<0.05 for both). No significant differences were observed among experiment groups in the sham operation setting.

The improved outcomes in 6H2-treated mice may be due to its protective effect on the BBB. After MCAO, a single dose of 6H2 1.8 mg/kg showed decreased ischaemia-induced BBB leakage, as evidenced by a significant reduction in leakage of Evans blue dye vs IgG control (p<0.05). Ischaemia-induced degradation of tight junction proteins (zonula occluden-1 and occludin) were also significantly reduced with 6H2 vs IgG control (p<0.05 for both).

Safety assessments revealed no significant changes in systemic metabolic profile, glucose tolerance and serum lipid levels in healthy mice receiving 6H2 1.8 mg/kg every 3 days for 21 days vs those receiving IgG control.

“6H2 can [potentially] be an effective and safe therapeutic approach for improving ischaemic stroke outcomes,” the researchers suggested. “We are currently investigating 6H2’s mode of action and its humanization. Further research on its clinical use is warranted.”