Inhomogeneous Diastereomeric Composition of Mongersen Antisense Phosphorothioate Oligonucleotide Preparations and Related Pharmacological Activity Impairment.

Mongersen is a 21-mer antisense oligonucleotide designed to downregulate Mothers against decapentaplegic homolog 7 (SMAD7) expression to treat Crohn's disease. Mongersen was manufactured in numerous batches at different scales during several years of clinical development, which all appeared identical, using common physicochemical analytical techniques, while only phosphorous-31 nuclear magnetic resonance (31P-NMR) in solution showed marked differences. Close-up analysis of 27 mongersen batches revealed marked differences in SMAD7 downregulation in a cell-based assay. Principal component analysis of 31P-NMR profiles showed strong correlation with SMAD7 downregulation and, therefore, with pharmacological efficacy in vitro. Mongersen contains 20 phosphorothioate (PS) linkages, whose chirality (Rp/Sp) was not controlled during manufacturing. A different diastereomeric composition throughout batches would lead to superimposable analytical data, but to distinct 31P-NMR profiles, as indeed we found. We tentatively suggest that this may be the origin of different biological activity. As similar manifolds are expected for other PS-based oligonucleotides, the protocol described here provides a general method to identify PS chirality issues and a chemometric tool to score each preparation for this elusive feature.

Explaining biosimilars and how reverse engineering plays a critical role in their development.

INTRODUCTION: Biologicals are protein-based therapeutics consisting of larger and more complex structures than small molecule medicines. As the patents for originator biological therapeutics expire, biosimilar products are licensed for the same indications as their marketed reference biologics across different specialities. Owing to the complex nature of the manufacturing process for a biological therapy compared to conventional chemically synthetized medicines, the development of biosimilars is more complicated and costly than the manufacture of generic small molecules. AREAS COVERED: The manufacturing process of the originator biologic is in most cases largely unknown to biosimilar developers and therefore reverse engineering through extensive analysis of the originator is a fundamental and critical step for successful biosimilar development. In this review, the authors examine the abbreviated roadmap for biosimilar approval which must be underpinned by the same rigorous standards that apply to all biological medicines. They discuss various aspects of biosimilar manufacturing with a focus on reverse engineering. EXPERT OPINION: The biosimilar approval pathway places a greater emphasis on preclinical assessments in comparison to the development of originator biologics. Multiple comparative clinical studies add little to the confirmation of the efficacy of the molecule under study whilst adding considerably to the cost and time of bringing a biosimilar into clinical use. A successful demonstration of biosimilarity to the reference product is therefore essential at a structural and functional level but this could not be achieved without well-designed and quality-driven reverse engineering of the originator production process.

The unleashing of the immune system in COVID-19 and sepsis: the calm before the storm?

The novel coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is sorely testing health care systems and economies around the world and is rightly considered as the major health emergency in a century. Despite the course of the disease appearing to be mild in many cases, a significant proportion of symptomatic patients develop pneumonia requiring hospitalisation or progress to manifest respiratory complications leading to intensive care treatment. Potential interventions for SARS-CoV2-associated pneumonia are being tested, some of which holding promise, but as of today none of these has yet demonstrated outstanding efficacy in treating COVID-19. In this article, we discuss fresh perspectives and insights into the potential role of immune dysregulation in COVID-19 as well as similarities with systemic inflammatory response in sepsis and the rationale for exploring novel treatment options affecting host immune response.

Can we wean patients with inflammatory arthritis from biological therapies?

Biological therapies have represented a cornerstone in the treatment of immune-mediated inflammatory diseases. Their advent combined with implementation of a treat-to-target approach has meant that remission or low disease activity are now realistic targets for treatment achieved by a significant number of patients. However, biologicals are not risk free and their elevated costs continue to present an important economic burden to national healthcare services. "Can we wean patients with inflammatory arthritis from biological therapies?" Over the last decade this question has become increasingly important as to define the best management strategies in terms of efficacy, safety and economic outcomes. Not surprisingly this has generated an interesting debate as to whether reasons to taper biologics outweigh reasons not to taper and evidence in support of either of these schools of thought is persistently growing. AIM: In this article we reviewed the contents of the relevant session from the 2019 Controversies in Rheumatology and Autoimmunity meeting in Florence.

Adalimumab Biosimilars in Europe: An Overview of the Clinical Evidence.

Adalimumab, the first fully humanised monoclonal antibody against tumour necrosis factor alpha (TNF-α), has played a leading role in the revolution brought about by the introduction of biologics, and has received the widest range of indications among TNF-α inhibitors. Post-registration, observational and registry studies of real-life use have largely supported the outcomes seen in registrational clinical trials. With the recent loss of exclusivity for the originator medicinal product in Europe, a number of biosimilar adalimumab molecules have been licensed for use in the same indications as the originator molecule across rheumatology, dermatology, gastroenterology and ophthalmology. Clinicians in these areas first gained experience with biosimilar infliximab, followed by etanercept and rituximab. However, adalimumab is likely to present unique challenges given the numbers of patients treated and the range of biosimilar adalimumab products available. The biosimilar approval pathway has an emphasis on the pre-clinical analytic data in combination with clinical studies conducted to confirm therapeutic equivalence. To date, several adalimumab biosimilars have entered the EU market following successful marketing authorisation applications and recent expiration of originator patent protection. This overview covers the extent of use of adalimumab and summarises the regulatory process involved in the development of biosimilars as well as their use in clinical practice. The authors also discuss clinical data available so far on adalimumab biosimilars and their envisaged impact in the field of immune-mediated inflammatory diseases.

A review article on biosimilar infliximab SB2 in the treatment of rheumatoid arthritis.

TNF inhibition has had a major impact as an approach for treating rheumatoid arthritis and a series of biologic agents directed against TNF have been developed for clinical use. Infliximab, a chimeric monoclonal antibody against soluble and membrane-bound TNF-α, was the biopharmaceutical to lead this 'biologics revolution'. However, with expiration of patent protection of the originator medicinal product, biosimilar versions of infliximab have been developed through biosimilarity studies and randomized controlled trials aiming to assess pharmacokinetic, pharmacodynamic and clinical equivalence to their originator (reference product) in patients with moderate-to-severe disease activity. This review summarizes the clinical development of SB2, a biosimilar of infliximab, in rheumatoid arthritis.

Necrotizing Panniculitis as an Uncommon Manifestation of Acute Pancreatitis.

Pancreatic panniculitis is a rare disorder affecting 2-3% of patients with pancreatic disease. The findings are characterized by tender, erythematous, subcutaneous nodules which may undergo spontaneous ulceration with discharge of brownish and viscous material derived from colliquative necrosis of adipocytes. The lesions are usually localized in the lower limbs, although they may also extend to the buttocks and also involve the trunk, upper limbs and scalp. They can precede overt pancreatic disease in 40% of cases. The typical histological features observed in these lesions are characterized by necrotic adipocytes with absent nuclei (better known as 'ghost cells') in the context of a predominantly lobular panniculitis. We describe the case of a 78-year-old cirrhotic woman admitted to our department with abdominal pain affecting the upper abdomen and a 3-day fever. On physical examination, multiple tender erythematous nodules, with irregular margins, were present on the pretibial regions of both lower legs, ranging in size from 0.8 to 1.5 cm. Pancreatic amylase and lipase were elevated and abdominal computed tomography revealed acute pancreatitis with oedema, focal gland enlargement of the pancreatic tail and perivisceral inflammation. Histological examination of the lesions was consistent with a diagnosis of necrotizing granulomatous panniculitis. LEARNING POINTS: Identification of the aetiological factors of tender erythematous nodules is challenging.Careful examination and history taking is essential for correct diagnosis and proper treatment.Pancreatic panniculitis should be included in the differential diagnosis as it can indicate developing acute pancreatitis.

Weekly low-dose methotrexate for reduction of Global Initiative for Asthma Step 5 treatment in severe refractory asthma: study protocol for a randomized controlled trial.

BACKGROUND: Patients with chronic severe asthma (CSA) have a crippling disease and current available treatments are not satisfactory. Thus, management of CSA remains a major unmet need. Although the evidence from existing randomized controlled trials fails to support a definite role for immunomodulatory drugs in these patients due to major methodologic drawbacks, findings with low-dose methotrexate (MTX) are encouraging. However, larger and well-designed clinical trials are required to establish the beneficial role of MTX in CSA, and for the detection of the key characteristics of those who are going to respond to this drug. METHODS/DESIGN: Patients will be recruited from the accessible asthmatic patients lists of tertiary referral centers. All patients will meet the stringent diagnostic criteria for CSA, including the requirement for the regular use of Global Initiative for Asthma (GINA) Global Strategy for Asthma Management and Prevention Step 5 medications (oral prednisone and/or omalizumab). The experimental design of the proposed study will take the form of a double-blind parallel-randomized placebo-controlled trial consisting of a total of eight visits, including run-in and run-out periods. Patients will be randomly allocated to receive either MTX or a matched placebo once a week as an add-on therapy to their existing medication after run-in. Physiological, laboratory and clinical assessments will be measured regularly throughout the study and compared with baseline assessments. DISCUSSION: We expect that MTX will reduce Step 5 medications dosage in patients with CSA without compromising the overall disease control. Improvement in several indicators of asthma severity and control will be also investigated. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02124226 (assigned 25 April 2014).

5-aminosalicylic acid enhances anchorage-independent colorectal cancer cell death.

Resistance to anoikis, the cell death triggered by the loss of anchorage to the substratum, is an essential prerequisite in the proliferation and diffusion of colorectal cancer (CRC) cells. We examined whether 5-aminosalicylic acid (5-ASA), a drug that seems to reduce the risk of colitis-associated CRC, enhances CRC cell anoikis. To this end, Colo205 cells were treated with 5-ASA in the presence or absence of inhibitors of caspases (zVAD-fmk) and reactive oxygen species (ROS). We demonstrate that 5-ASA enhances Colo205 cell death. Although 5-ASA induces dissipation of mitochondrial transmembrane potential and caspase-3 activation, zVAD-fmk does not completely prevent the 5-ASA-induced cell death. 5-ASA also enhances the synthesis of ROS. However, inhibitors of ROS reduce the fraction of 5-ASA-induced Colo205 cell death but do not confer protection. In contrast, the 5-ASA-mediated Colo205 cell death is preventable by Bcl-2 over-expression. These data suggest a mechanism by which 5-ASA interferes with colon carcinogenesis.