Chimeric antigen receptor T-cell therapy in glioblastoma: charging the T cells to fight.

Glioblastoma multiforme (GBM) is the most common malignant brain cancer that invades normal brain tissue and impedes surgical eradication, resulting in early local recurrence and high mortality. In addition, most therapeutic agents lack permeability across the blood brain barrier (BBB), further reducing the efficacy of chemotherapy. Thus, effective treatment against GBM requires tumor specific targets and efficient intracranial drug delivery. With the most recent advances in immunotherapy, genetically engineered T cells with chimeric antigen receptors (CARs) are becoming a promising approach for treating cancer. By transducing T lymphocytes with CAR constructs containing a tumor-associated antigen (TAA) recognition domain linked to the constant regions of a signaling T cell receptor, CAR T cells may recognize a predefined TAA with high specificity in a non-MHC restricted manner, and is independent of antigen processing. Active T cells can travel across the BBB, providing additional advantage for drug delivery and tumor targeting. Here we review the CAR design and technical innovations, the major targets that are in pre-clinical and clinical development with a focus on GBM, and multiple strategies developed to improve CAR T cell efficacy.

Neutrophil Granule Proteins Inhibit Amyloid Beta Aggregation and Neurotoxicity.

BACKGROUND: A role for neutrophils in the pathogenesis of Alzheimer's disease (AD) is emerging. We previously showed that the neutrophil granule proteins cationic antimicrobial protein of 37 kDa (CAP37), cathepsin G (CG), and neutrophil elastase (NE) directly bind the amyloid-beta peptide Aß1-42, a central player in AD pathogenesis. CAP37, CG, and NE are serine proteases that can cleave Aß1-42 at different sites and with different catalytic activities. OBJECTIVE: In this study, we compared the effects of these three proteins on Aß1-42 fibrillation and neurotoxicity. METHODS: Using mass spectrometry and in vitro aggregation assay, we found that NE and CG efficiently cleave Aß1-42. This cleavage correlates well with the inhibition of Aß1-42 aggregation into fibrils. In contrast, CAP37 did not efficiently cleave Aß1-42, but was still able to inhibit its fibrillation, most likely through a quenching effect. Inhibition of Aß1-42 aggregation by NE and CG neutralized its toxicity measured in cultured neurons. In contrast, inhibition of Aß1-42 aggregation by CAP37 did not inhibit its neurotoxicity. RESULTS: We found that a peptide derived from CAP37 could mimic the quenching and inhibition of Aß1-42 aggregation effects of the full-length protein. Additionally, this peptide was able to inhibit the neurotoxicity of the most toxic Aß1-42 aggregate, an effect that was not found with the full-length CAP37. CONCLUSION: These results shed light on the mechanisms of action of neutrophil granule proteins with regard to inhibition of Aß1-42 aggregation and neurotoxicity and open up a possible strategy for the discovery of new disease-modifying drugs for AD.

Antimicrobial Peptides Derived from the Immune Defense Protein CAP37 Inhibit TLR4 Activation by S100A9.

Purpose: Corneal abrasion is a common eye injury, and its resolution can be seriously complicated by bacterial infection. We showed that topical application of the cationic antimicrobial protein of 37 kDa (CAP37) promotes corneal re-epithelialization in mice, and peptides derived from CAP37 can recapitulate the antibacterial and wound-healing effects of the full-length protein. The current study was designed to identify the molecular mechanisms mediating the wound-healing effect of CAP37 and derived bioactive peptides. Methods: We used a TriCEPS-based, ligand-receptor glycocapture method to identify the binding partners of CAP37 on live human corneal epithelial cells using the hTCEpi cell line. We used an ELISA method to confirm binding with identified partners and test the binding with CAP37-derived peptides. We used a reporter cell line to measure activation of the identified membrane receptor by CAP37 and derived peptides. Results: We pulled down S100 calcium-binding protein A9 (S100A9) as a binding partner of CAP37 and found that CAP37 and four derived peptides encompassing two regions of CAP37 bind S100A9 with high affinities. We found that CAP37 and the S100A9-binding peptides could also directly interact with the Toll-like receptor 4 (TLR4), a known receptor for S100A9. CAP37 and one peptide partially activated TLR4. The other three peptides did not activate TLR4. Finally, we found that CAP37 and all four peptides could inhibit the activation of TLR4 by S100A9. Conclusions: This study identifies a mechanism of action for CAP37 and derived antimicrobial peptides that may restrain inflammatory responses to corneal injury and favor corneal re-epithelialization.

Overexpression of HGF/MET axis along with p53 inhibition induces de novo glioma formation in mice.

BACKGROUND: Aberrant MET receptor tyrosine kinase (RTK) activation leads to invasive tumor growth in different types of cancer. Overexpression of MET and its ligand hepatocyte growth factor (HGF) occurs more frequently in glioblastoma (GBM) than in low-grade gliomas. Although we have shown previously that HGF-autocrine activation predicts sensitivity to MET tyrosine kinase inhibitors (TKIs) in GBM, whether it initiates tumorigenesis remains elusive. METHODS: Using a well-established Sleeping Beauty (SB) transposon strategy, we injected human HGF and MET cDNA together with a short hairpin siRNA against Trp53 (SB-hHgf.Met.ShP53) into the lateral ventricle of neonatal mice to induce spontaneous glioma initiation and characterized the tumors with H&E and immunohistochemistry analysis. Glioma sphere cells also were isolated for measuring the sensitivity to specific MET TKIs. RESULTS: Mixed injection of SB-hHgf.Met.ShP53 plasmids induced de novo glioma formation with invasive tumor growth accompanied by HGF and MET overexpression. While glioma stem cells (GSCs) are considered as the tumor-initiating cells in GBM, both SB-hHgf.Met.ShP53 tumor sections and glioma spheres harvested from these tumors expressed GSC markers nestin, GFAP, and Sox 2. Moreover, specific MET TKIs significantly inhibited tumor spheres' proliferation and MET/MAPK/AKT signaling. CONCLUSIONS: Overexpression of the HGF/MET axis along with p53 attenuation may transform neural stem cells into GSCs, resulting in GBM formation in mice. These tumors are primarily driven by the MET RTK pathway activation and are sensitive to MET TKIs. The SB-hHgf.Met.ShP53 spontaneous mouse glioma model provides a useful tool for studying GBM tumor biology and MET-targeting therapeutics.