Preclinical Evaluation of Recombinant T Cell Receptor Ligand RTL1000 as a Therapeutic Agent in Ischemic Stroke

Abstract

Recombinant T-cell Receptor Ligand 1000 (RTL1000), a partial human major histocompatibility complex (MHC) molecule coupled to a human myelin peptide, reduces infarct size after experimental stroke in HLA-DRB1*1502 transgenic (DR2-Tg) mice. In this study, we characterized the therapeutic time window of opportunity for RTL1000; we explored the efficacy of single dose of RTL1000 administration and determined if RTL1000 affordslong-term neurobehavioral functional improvement after ischemic stroke. Male DR2-Tg mice underwent 60 min of intraluminal reversible middle cerebral artery occlusion (MCAO). RTL1000 or vehicle was injected 4, 6 or 8 h after MCAO, followed by 3 daily injections. In single dose study, one-time injection of RTL1000 was applied 4 h after MCAO. Cortical, striatal and hemispheric infarct sizes were measured 24 h or 96 h after stroke. Behavioral testing, including neuroscore evaluation, open field, paw preference and novel object recognition was performed up to 28 days after stroke. Our data showed RTL1000 significantly reduced infarct size 96 h after MCAO when first injection was given 4 and 6, but not 8 h after the onset of stroke. A single dose of 400 µg or 100 µg RTL1000 also significantly reduced infarct size 24 h after MCAO. Behavioral testing showed RTL1000 treatment used 4 h after MCAO improved long-term cognitive outcome 28 days after stroke. Taken together, RTL1000 protects against acute injury if applied within a 6-h time window and improves long-term functional recovery after experimental stroke in DR2-Tg mice.

DRα1-MOG-35-55 treatment reduces lesion volumes and improves neurological deficits after traumatic brain injury

“…Recombinant T-cell receptor (TCR) ligands (RTL) are molecular constructs comprised of covalently linked α1 and β1 domains of MHC II molecules with an attached antigenic peptide (Pan et al 2014; Zhu et al 2015; Zhu et al 2014). Unlike four-domain MHC II molecules that induce T cell activation, RTLs are partial agonists of TCR and can deviate autoreactive T cell responses towards an anti-inflammatory phenotype (Burrows et al 2001; Wang et al 2003).…”

Chronic methamphetamine exposure prior to middle cerebral artery occlusion increases infarct volume and worsens cognitive injury in Male mice

“…tMCAO was induced in male C57BL/6J mice for 1 hour by reversible right MCAO under isoflurane anesthesia followed by 96 hours of reperfusion as described previously (Banerjee et al, 2013). The 96 hour time point was selected based on our previous work (Zhu et al, 2015), indicating that it is a valid time point for assessment of infarct volume by 2 ,3,5-Triphenyltetrazolium Chloride (TTC) staining post-ischemia. The investigator performing the tMCAO surgery was blinded to treatment group.…”

Recombinant T-cell receptor ligand RTL1000 limits inflammation and decreases infarct size after experimental ischemic stroke in middle-aged mice

“…The dose and injection protocol of RTL1000 was established in our previous studies (Subramanian et al, 2009; Zhu et al, 2014A; Zhu et al, 2014B). …”

“…These findings indicate that RTL551 may not work in patients with stroke considering species-differences (murine vs. human) in antigens and MHC II molecules. To determine if a RTL strategy would work against human stroke, we determined the efficacy of humanized RTL1000, which contains a human MHC moiety (HLA-DR2) covalently linked to a human myelin peptide (hMOG-35-55) in experimental stroke in humanized DR2-Tg mice which expresses human TCR(Subramanian et al, 2009; Zhu et al, 2014A). We found that RTL1000 indeed protects against ischemic injury in young male DR2-Tg mice.…”

“…We found that RTL1000 indeed protects against ischemic injury in young male DR2-Tg mice. Behavioral testing showed that RTL1000 improves long-term cognitive function 28 days after stroke (Zhu et al, 2014A). A similar effect has also been demonstrated in young female DR2-Tg mice (Pan et al, 2014).…”