4 research outputs found
Olokizumab, a monoclonal antibody against interleukin-6, in combination with methotrexate in patients with rheumatoid arthritis inadequately controlled by tumour necrosis factor inhibitor therapy: efficacy and safety results of a randomised controlled phase III study
No full textObjectives To assess the efficacy and safety of olokizumab (OKZ), a monoclonal antibody against the interleukin-6 (IL-6) cytokine, versus placebo (PBO) in patients with prior inadequate response to tumour necrosis factor inhibitors (TNFi-IRs).Methods In this 24-week multicentre, placebo-controlled, double-blind study, the patients were randomised in a 2:2:1 ratio to receive subcutaneously administered OKZ 64 mg once every 2 weeks (q2w), OKZ 64 mg once every 4 weeks (q4w) or PBO plus methotrexate. At week 16, the patients on PBO were randomised to receive either OKZ regime. The primary endpoint was the proportion of patients achieving an American College of Rheumatology 20% (ACR20) response at week 12. Disease Activity Score 28-joint count C-reactive protein (DAS28 (CRP)) <3.2 at week 12 was the major secondary efficacy endpoint. Safety and immunogenicity were assessed.Results In 368 patients randomised, ACR20 response rates were 60.9% in OKZ q2w, 59.6% in OKZ q4w and 40.6% in PBO (p<0.01 for both comparisons). Achievement of DAS28 (CRP) <3.2 was significantly different, favouring the OKZ arms. Improvements in efficacy and patientreported outcomes were maintained throughout 24 weeks and were noted after week 16 in patients who switched from PBO.Dose-related treatment-emergent serious adverse events were 7% in OKZ q2w, 3.2% in OKZ q4w and none in the PBO group.Conclusions Direct inhibition of IL-6 with OKZ resulted in significant improvements in the signs and symptoms of rheumatoid arthritis compared with PBO in TNFi-IR patients with a similar safety profile as observed for monoclonal antibodies to the IL-6 receptor
Олокизумаб, моноклональное антитело к интерлейкину 6, в комбинации с метотрексатом у пациентов с ревматоидным артритом и неадекватным контролем заболевания на фоне терапии ингибиторами фактора некроза опухоли: результаты оценки эффективности и безопасности в рандомизированном контролируемом исследовании III фазы
No full textObjectives To assess the efficacy and safety of olokizumab (OKZ), a monoclonal antibody against the interleukin-6 (IL-6) cytokine, versus placebo (PBO) in patients with prior inadequate response to tumour necrosis factor inhibitors (TNFi-IRs).Methods In this 24-week multicentre, placebo-controlled, double-blind study, the patients were randomised in a 2:2:1 ratio to receive subcutaneously administered OKZ 64 mg once every 2 weeks (q2w), OKZ 64 mg once every 4 weeks (q4w) or PBO plus methotrexate. At week 16, the patients on PBO were randomised to receive either OKZ regime. The primary endpoint was the proportion of patients achieving an American College of Rheumatology 20% (ACR20) response at week 12. Disease Activity Score 28-joint count C-reactive protein (DAS28 (CRP)) <3.2 at week 12 was the major secondary efficacy endpoint. Safety and immunogenicity were assessed.Results In 368 patients randomised, ACR20 response rates were 60.9% in OKZ q2w, 59.6% in OKZ q4w and 40.6% in PBO (p<0.01 for both comparisons). Achievement of DAS28 (CRP) <3.2 was significantly different, favouring the OKZ arms. Improvements in efficacy and patientreported outcomes were maintained throughout 24 weeks and were noted after week 16 in patients who switched from PBO.Dose-related treatment-emergent serious adverse events were 7% in OKZ q2w, 3.2% in OKZ q4w and none in the PBO group.Conclusions Direct inhibition of IL-6 with OKZ resulted in significant improvements in the signs and symptoms of rheumatoid arthritis compared with PBO in TNFi-IR patients with a similar safety profile as observed for monoclonal antibodies to the IL-6 receptor.Цель исследования – оценить эффективность и безопасность олокизумаба (ОКЗ), моноклонального антитела к интерлейкину (ИЛ) 6, по сравнению с плацебо у больных ревматоидным артритом (РА) с предшествующим неадекватным ответом на ингибиторы фактора некроза опухоли альфа (ФНО α ).Методы.В данном 24-недельном многоцентровом плацебо-контролируемом двойном слепом исследовании пациенты рандомизировались в соотношении 2:2:1 для проведения терапии ОКЗ подкожно в дозе 64 мг 1 раз в 2 нед; ОКЗ в дозе 64 мг 1 раз в 4 нед либо плацебо, в комбинации с метотрексатом. На неделе 16 пациентов, получавших плацебо, рандомизировали для проведения терапии ОКЗ в одном из двух режимов. Первичной конечной точкой являлась доля пациентов, у которых был достигнут ответ по ACR20 (20% улучшение согласно критериям ACR) на неделе 12. Важнейшей из вторичных конечных точек было достижение значения DAS28-CРБ <3,2 на неделе 12. Проводилась оценка безопасности и иммуногенности.Результаты. У 368 рандомизированных пациентов частота ответа по ACR20 составила 60,9% в группе ОКЗ 1 раз в 2 нед, 59,6% в группе ОКЗ 1 раз в 4 нед и 40,6% в группе плацебо (p<0,01 для обоих сравнений). Между группами отмечались значимые различия по частоте достижения DAS28-CРБ <3,2 в пользу групп ОКЗ. Достигнутое улучшение сохранялось на протяжении всех 24 нед, а у пациентов, которым плацебо заменялось на ОКЗ, улучшение выявлялось после недели 16. Частота связанных с терапией серьезных нежелательных явлений составила 7% в группе ОКЗ 1 раз в 2 нед и 3,2% в группе ОКЗ 1 раз в 4 нед, в то время как в группе плацебо их не было.Заключение. Прямое ингибирование ИЛ6 ОКЗ привело к значительному уменьшению выраженности проявлений РА по сравнению с плацебо у пациентов с неадекватным ответом на ингибиторы ФНО α , при этом профиль безопасности был схож с таковым при назначении моноклональных антител к рецептору ИЛ6
Development of a secreted cell-permeable NF-κB inhibitor to control inflammation
No full textPhDRheumatoid arthritis (RA) is an autoimmune chronic inflammatory disease, of
unknown aetiology. Several disease-modulating approaches have been developed in
the past years, however these are expensive, usually accompanied by unwanted
side-effects and 30% of the patients fail to respond. The transcription factor NF-kB is
a key factor in the development and perpetuation of the disease, as it regulates a
number of inflammatory genes. The activity of certain signalling pathways can be
modulated by delivering into cells inhibitors coupled to Protein Transduction Domains
(PTDs). The aim of this study was to develop a secretable PTD-fusion NF-κΒ inhibitor
that is produced and secreted by genetically engineered mammalian cells in sufficient
amounts to subsequently transduce and regulate NF-κΒ activity in neighbouring cells.
Such methodology could be useful for the management of RA by transplantation of
engineered cells or directly using gene delivery into the synovial joints.
In this study, PTD-fusion proteins were fused to the IL-2 secretion signal and
their ability to be secreted from mammalian cells was explored. Secretable forms of
TAT-IgG2A and TAT-eGFP were generated as control PTD-fusion proteins, and the
TAT-srIκΒα (super repressor IκBα, a NF-κΒ inhibitor) was generated as an NF-κΒ
PTD-fusion inhibitor. Western blotting analysis of supernatants from transiently
transfected 293T cells revealed that TAT-IgG2A, TAT-eGFP and TAT-srIκΒα are
secreted with variable efficiencies. When concentrated, PTD proteins were able to
transduce mammalian cells, as demonstrated with Jurkat cells by confocal
microscopy and western blotting analysis.
The TAT PTD domain was replaced to a more stable, furin cleavage-resistant
and less positively charged PTD domain, the TAT3 PTD domain, to ensure that PTDfusion
proteins will be secreted more efficiently. This change of the PTD domain did
not increase secretion levels of the srΙκBα. Subsequently, the Latent Associated
Peptide (LAP) of TGFβ, was fused to the TAT3-srIκΒα inhibitor, via a matrix
metalloproteinase (MMP) cleavage linker. This LAP-MMP-PTD-fusion NF-κΒ inhibitor
was again poorly secreted. In turn, the srIκΒα inhibitor was replaced with a small
synthetic NF-κΒ inhibitor, termed Nemo Binding Domain (NBD), in the form of LAP
MMP-TAT3-NBD NF-κΒ inhibitor. Western blotting analysis of supernatants from
transiently transfected 293T cells revealed that the LAP-MMP-TAT3-NBD was
efficiently secreted.
The ability of LAP-MMP-TAT3-NBD to inhibit NF-κΒ was tested in vitro with the
use of a cell-assay based on HeLa cells that are permanently transfected with the
luciferase gene driven by an NF-κB regulated promoter. In this assay, HeLa cells that
were treated with the secreted LAP-MMP-TAT3-NBD, showed reduced levels of
luciferase activity after IL-1β stimulation. Subsequently, using a replication-deficient
lentiviral vector, genetically engeneered DBA/1 fibroblasts (DTF) able to produce the
secreted LAP-MMP-TAT3-NBD were generated. The NF-κB inhibitory properties of
the secreted LAP-MMP-TAT3-NBD were tested in vivo in the Carrageenan-induced
paw oedema, Antigen Induced Arthritis and Air-Pouch acute inflammation models.
Paws of mice that were treated with engineered cells or lentivirus encoding LAPMMP-
TAT3-NBD demonstrated milder paw swelling, suggesting that LAP-MMPTAT3-
NBD had a protective role in the induction of inflammation. However, the LAPMMP-
TAT3-NBD did not demonstrate anti-inflammatory effects in the Air-Pouch
model.
In this study, I present a method to design PTD-fusion proteins that can be
efficiently secreted from mammalian cells and I demonstrate a novel gene therapy
approach for the local delivery of a therapeutic agent
The Detection and Role of Human Endogenous Retrovirus K (HML-2) In Rheumatoid Arthritis
No full textA thesis submitted in partial fulfilment of the requirements of the University of
Wolverhampton for the degree of Doctor of PhilosophyHuman endogenous retroviruses are the remnants of ancient retroviral infections present within our genome. These molecular fossils show similarities with present day exogenous retroviruses but act as typical Mendelian elements that are passed vertically between generations. Despite being repeatedly linked to a number of autoimmune diseases and disorders, no conclusive proof has been identified. Rheumatoid arthritis (RA) is one such disease which has been associated with an increase in HERV expression, compared to controls. In order to elucidate a clear role for HERVs in RA pathogenesis, autoantigens implicated in disease pathogenesis were scanned for sequence homology to retroviral genes. Such epitopes would induce antibodies cross reactive with host proteins, resulting in disease. Short peptides mimicking these regions were synthesised and the prevalence of anti-HERV antibodies was determined in RA patients and disease controls. Additionally, a novel real-time Polymerase Chain Reaction (PCR) assay was developed to accurately quantify levels of HERV-K (HML-2) gag expression, relative to normalised levels of housekeeping gene expression. Both serological and molecular assays showed significant increases in HERV-K (HML-2) activity in RA patients compared to disease controls with CD4+ lymphocytes harbouring the highest activity. The real-time assay was also used to determine whether factors within the synovium could modulate HERVs, resulting in their upregulation. Exogenous viral protein expression and pro-inflammatory cytokines were shown to exert a significant modulatory effect over HERV-K (HML-2) transcription. From this data, it is clear that RA patients have increased levels of HERV-K (HML-2) gag activity compared to controls. Despite this it is likely that factors within the synovium such as exogenous viral expression and pro-inflammatory cytokines also influence HERV-K (HML-2) transcription possibly contributing to a role of bystander activation, i.e. being influenced by external factors, rather than actively contributing to disease processes. The exact role of HERVs in RA pathology remains elusive; however this research proposes several mechanisms by which HERV-K (HML-2) may contribute to disease
