A deimmunised form of the ribotoxin, α-sarcin, lacking CD4+ T cell epitopes and its use as an immunotoxin warhead

Fungal ribotoxins that block protein synthesis can be useful warheads in the context of a targeted immunotoxin. α-Sarcin is a small (17 kDa) fungal ribonuclease produced by Aspergillus giganteus that functions by catalytically cleaving a single phosphodiester bond in the sarcin–ricin loop of the large ribosomal subunit, thus making the ribosome unrecognisable to elongation factors and leading to inhibition of protein synthesis. Peptide mapping using an ex vivo human T cell assay determined that α-sarcin contained two T cell epitopes; one in the N-terminal 20 amino acids and the other in the C-terminal 20 amino acids. Various mutations were tested individually within each epitope and then in combination to isolate deimmunised α-sarcin variants that had the desired properties of silencing T cell epitopes and retention of the ability to inhibit protein synthesis (equivalent to wild-type, WT α-sarcin). A deimmunised variant (D9T/Q142T) demonstrated a complete lack of T cell activation in in vitro whole protein human T cell assays using peripheral blood mononuclear cells from donors with diverse HLA allotypes. Generation of an immunotoxin by fusion of the D9T/Q142T variant to a single-chain Fv targeting Her2 demonstrated potent cell killing equivalent to a fusion protein comprising the WT α-sarcin. These results represent the first fungal ribotoxin to be deimmunised with the potential to construct a new generation of deimmunised immunotoxin therapeutics.

Efficiency of chitosan and their combination with bentonite as retention aids in papermaking

In a previous work (Nicu et al. 2013), the flocculation efficiency of three chitosans differing by molecular weight and charge density were evaluated for their potential use as wet end additives in papermaking. According to the promising results obtained, chitosan (single system) and its combination with bentonite (dual system) were evaluated as retention aids, and their efficiency was compared with poly(diallyl dimethyl ammonium chloride) (PDADMAC) and polyethylenimine (PEI). In single systems, chitosan was clearly more efficient in drainage rate than PDADMAC and PEI, especially those with the lowest molecular weights; however, retention is considerably lower. This drawback can be overcome by using dual systems with anionic bentonite microparticles, with the optimum ratio of polymer:bentonite being 1:4 (wt./wt.). In dual systems, the differences in retention were almost negligible, and the difference in drainage rate was even higher, together with better floc reversibility. The most efficient chitosan in single systems was Ch.MMW, while Ch.LMW was the most efficient in dual systems. The flocculation mechanism of chitosan was a combination of patch formation, charge neutralization, and partial bridge formation, and the predominant mechanism depended on the molecular weight and charge density of the chitosan.