Clinical validation of the in silico prediction of immunogenicity of a human recombinant therapeutic protein

Antibodies elicited by protein therapeutics can cause serious side effects in humans. We studied immunogenicity of a recombinant fusion protein (FPX) consisting of two identical, biologically active, peptides attached to human Fc fragment. EpiMatrix, an in silico epitope-mapping tool, predicted promiscuous T-cell epitope(s) within the 14-amino-acid carboxy-terminal region of the peptide portion of FPX. On administration of FPX in 76 healthy human subjects, 37% developed antibodies after a single injection. A memory T-cell response against the above carboxy-terminus of the peptide was observed in antibody-positive but not in antibody-negative subjects. Promiscuity of the predicted T-cell epitope(s) was confirmed by representation of all common HLA alleles in antibody-positive subjects. As predicted by EpiMatrix, HLA haplotype DRB1*0701/1501 was associated with the highest T-cell and antibody response. In conclusion, in silico prediction can be successfully used to identify Class II restricted T-cell epitopes within therapeutic proteins and predict immunogenicity thereof in humans

Assessment of immunogenicity of romiplostim in clinical studies with ITP subjects

Romiplostim is an Fc-peptide fusion protein that activates intracellular transcriptional pathways via the thrombopoietin (TPO) receptor leading to increased platelet production. Romiplostim has been engineered to have no amino acid sequence homology to endogenous TPO. Recombinant protein therapeutics can be at a risk of development of an antibody response that can impact efficacy and safety. Hence, a strategy to detect potential antibody formation to the drug and to related endogenous molecules can be useful. The immunogenicity assessment strategy involved both the detection and characterization of binding and neutralizing antibodies. The method for detection was based on a surface plasmon resonance biosensor platform using the Biacore 3000. Samples that tested positive for binding antibodies in the Biacore immunoassay were then tested in a neutralization assay. Serum samples from 225 subjects with immune thrombocytopenic purpura (ITP) dosed with romiplostim and 45 ITP subjects dosed with placebo were tested for romiplostim and TPO antibodies. Prior to romiplostim treatment, 17 subjects (7%) tested romiplostim antibody positive and 12 subjects (5%) tested TPO antibody positive for pre-existing binding antibodies. After romiplostim exposure, 11% of the subjects exhibited binding antibodies against romiplostim and 5% of the subjects with ITP showed binding antibodies against TPO. The antibodies against romiplostim did not cross-react with TPO and vice versa. No cases of anti-TPO neutralizing antibodies were detected in romiplostim-treated subjects. The incidence of anti-romiplostim neutralizing antibodies to romiplostim was 0.4% (one subject); this subject tested negative at the time of follow-up 4 months later. No impact on platelet profiles were apparent in subjects that had antibodies to romiplostim to date. In summary, administration of romiplostim in ITP subjects resulted in the development of a binding antibody response against romiplostim and TPO ligand. One subject developed a neutralizing antibody response to romiplostim that impacted the platelet counts of this subject. No neutralizing antibodies to endogenous TPO were observed

Translatability of findings from cynomolgus monkey to human suggests a mechanistic role for IL-21 in promoting immunogenicity to an anti-PD-1/IL-21 mutein fusion protein

AMG 256 is a bi-specific, heteroimmunoglobulin molecule with an anti-PD-1 antibody domain and a single IL-21 mutein domain on the C-terminus. Nonclinical studies in cynomolgus monkeys revealed that AMG 256 administration led to the development of immunogenicity-mediated responses and indicated that the IL-21 mutein domain of AMG 256 could enhance the anti-drug antibody response directed toward the monoclonal antibody domain. Anti-AMG 256 IgE were also observed in cynomolgus monkeys. A first-in-human (FIH) study in patients with advanced solid tumors was designed with these risks in mind. AMG 256 elicited ADA in 28 of 33 subjects (84.8%). However, ADA responses were only robust and exposure-impacting at the 2 lowest doses. At mid to high doses, ADA responses remained low magnitude and all subjects maintained exposure, despite most subjects developing ADA. Limited drug-specific IgE were also observed during the FIH study. ADA responses were not associated with any type of adverse event. The AMG 256 program represents a unique case where nonclinical studies informed on the risk of immunogenicity in humans, due to the IL-21-driven nature of the response

Recreation area in the city - the revitalization of the Lagoon Bagry in Krakow

Tereny rekreacyjne odgrywają kluczową rolę w dużych miastach, dzięki możliwości wykorzystania ich do aktywnego wypoczynku na łonie natury. W artykule autor chce ukazać potencjał terenu Zalewu Bagry w Krakowie poprzez jego dokładną analizę architektoniczno - krajobrazową a także projekt koncepcyjny całego założenia.Recreational areas play a key role in the big cities, thanks to their use for an active recreation in the bosom of nature. In this article the author wants to show the potential of land in Krakow Bagry Lagoon through its thorough analysis of architectural - landscape and conceptual design of the complex