Safety assessment and dose selection for first-in-human clinical trials with immunomodulatory monoclonal antibodies.

Modulating immune responses with monoclonal antibodies (mAbs) that target immune molecules has become a promising therapeutic strategy and is under investigation for the treatment of cancer and (auto)-immune diseases. A major hurdle to the development and early clinical investigation of many immunomodulatory mAbs is the inherent risk of adverse immune-mediated drug reactions in humans, such as cytokine storms, autoimmunity, and immunosuppression. Dose selection for first-in-human (FIH) clinical trials involving immunomodulatory mAbs, and mAbs in general, is based on specifically designed preclinical safety studies, primarily in nonhuman primates (NHPs), and on mechanistic ex vivo investigations. Dose selection in such trials is challenging for a number of reasons related to safety. In this context, safety-relevant differences between NHP and human immune systems, species selection/qualification and preclinical study design considerations, the receptor occupancy model and its calculation, the minimal anticipated biological effect level (MABEL) and its use in the selection of a safe starting dose in humans, microdosing and the impact of immunogenicity on safety assessment of mAbs, and safety-relevant formulation properties of therapeutic mAbs are critically reviewed. In addition, the current regulatory requirements are presented and discussed to demonstrate how the TeGenero TGN1412 case is leading to increased regulatory scrutiny regarding dose selection for FIH clinical trials.

Apolipoprotein E3Basel: new insights into a highly conserved protein region.

BACKGROUND: Apolipoprotein E is important for the receptor-mediated uptake of triglyceride-rich lipoproteins. Mutations in the gene encoding apolipoprotein E may cause a reduced uptake of these lipoproteins. Particular apolipoprotein E mutations have been also found to be associated with nephrologic, neurologic, and even ophthalmologic diseases. Hence, a continuously expanding role in biology is being attributed to this protein. DESIGN: Randomly selected volunteers from of a large Swiss cohort were genotyped for the common apolipoprotein E isoforms (apolipoprotein E2, apolipoprotein E3, apolipoprotein E4). RESULTS: In one of the volunteers, a novel C-to-T mutation causing an alanine-to-valine substitution (A106V, designated apolipoprotein E3Basel) was discovered. Alanine at residue 106 is highly conserved between mammalian species and is located in the immediate vicinity of the 112C/R polymorphism (apolipoprotein E4). Recombinant apolipoprotein E3Basel, expressed in the baculovirus system, displayed no detectable reduction in its low density lipoprotein (LDL) receptor- and heparin-binding activities. Despite normal binding functions, apolipoprotein E3Basel might cause modifications in the lipoprotein pattern. In the index case, plasma triglycerides were elevated and in two further apolipoprotein E3Basel-carriers, cholesterol, phospholipid, apolipoprotein CIII levels, LDL-cholesterol/apoB-100- and VLDL-triglyceride/VLDL-cholesterol-ratios were higher compared with apolipoprotein E3Basel-noncarriers when pair-matched for age and gender. One of the four apolipoprotein E3Basel-carriers from the index family had a personal history of Alzheimer's disease. CONCLUSIONS: Alanine at amino acid position 106 is highly conserved but not crucial in the receptor-mediated uptake of lipoprotein particles. Nevertheless, amino acid position 106 might be involved in the apolipoprotein E-dependent regulation of the lipoprotein lipase that hydrolyzes triglycerides and in the development of Alzheimer's disease.

Identification of mutations in the gene encoding sterol regulatory element binding protein (SREBP)-2 in hypercholesterolaemic subjects.

Human cells maintain their cholesterol homeostasis by regulated cleavage of membrane bound transcription factors, so-called sterol regulatory element binding proteins (SREBPs). If cells are deprived of cholesterol, SREBPs are cleaved by two proteolytic steps. The NH2-terminal domain of the SREBPs is released from the membranes of the endoplasmic reticulum and transported into the nucleus, where it binds to specific nucleotide sequences in the promoters of the low density lipoprotein receptor gene and of key genes involved in cholesterol and triglyceride homeostasis. Given the central role of SREBPs in the regulation of cholesterol metabolism, we investigated whether subjects with inherited forms of high plasma cholesterol carry specific sequence variations in SREBP-2 that might be involved in the development of hypercholesterolaemia. Exons 5 to 10, encoding the DNA binding and the regulatory domains of SREBP-2, were screened for sequence variations in a cohort of 70 hypercholesterolaemic subjects. Two missense mutations (V623M, R645Q) in the regulatory domain, one single nucleotide polymorphism (R371K) in the DNA binding domain, and one translationally silent mutation (P433P) were identified in SREBP-2. However, none of the mutations found in the regulatory domain could be detected in 167 subjects of a random control sample. A potential causative mechanism of these mutations for high plasma cholesterol concentrations is discussed. In summary, this is the first report of mutations in the human SREBP-2 gene to suggest that these and/or other mutations in this key regulator of cholesterol metabolism are associated with hypercholesterolaemia.

A single-nucleotide polymorphism in the sterol-regulatory element-binding protein 1c gene is predictive of HIV-related hyperlipoproteinaemia.

A single-nucleotide polymorphism (3'322C/G) was identified in the gene encoding a key cholesterol/triglyceride regulator, sterol-regulatory element-binding protein 1c (SREBP-1c). Although it did not alter the amino acid sequence, SREBP-1c-3'322C/G was predictive of highly active antiretroviral therapy-related hyperlipoproteinaemia. Increases in cholesterol were less frequently associated with homozygous SREBP-1c-3'322G (genotype 22) than with heterozygous/homozygous SREBP-1c-3'322C (genotypes 11/12) and correlated with leptin and insulin increases, particularly in genotype 11/12 carriers. A functional mutation linked to SREBP-1c-3'322C/G or messenger RNA conformation differences may explain our findings.

RepGene: a spreadsheet template for the management of reporter gene assays.

One of the most powerful techniques in molecular biology is the controlled expression of specific proteins by transfection of eukaryotic cells. This method has become feasible and highly sensitive and, thus, suitable for high-throughput reporter gene assays in basic and applied research. Moreover, the limiting factors are neither the transfection efficiency nor the functional analysis, but rather the ability to manage complex experimental protocols when multiple genes are co-transfected and/or when the effects of several chemical compounds are investigated within the same experiment. Here, we describe an easy-to-use and highly flexible spreadsheet template intended to rationalize and expedite the organization and data management of multi-step reporter gene assays. The objectives of this spreadsheet template are the design of the transfection protocol, the coordination of the administration of test compounds, and the graphical presentation and statistical analysis of the results.

Molecular Biocomputing Suite: a word processor add-in for the analysis and manipulation of nucleic acid and protein sequence data.

In all fields of molecular biology, researchers are increasingly challenged by experiments planned and evaluated on the basis of nucleic acid and protein sequence data generally retrieved from public databases. Despite the wide spectrum of available Web-based software tools for sequence analysis, the routine use of these tools has disadvantages, particularly because of the elaborate and heterogeneous ways of data input, output, and storage. Here we present a Visual Basic-encoded Microsoft Word Add-In, the Molecular BioComputing Suite (MBCS), available at the BioTechniques Software Library (www.BioTechniques.com). The MBCS software aims to manage and expedite a wide range of sequence analyses and manipulations using an integrated text editor environment including menu-guided commands. Its independence of sequence formats enables MBCS to be used as a pivotal application between other software tools for sequence analysis, manipulation, annotation, and editing.

[Recurrence or persistence of an osteoid osteoma: a case report]. / Redicive ou persistance d'un ostéome ostéoïde: une observation.

Recurrence of an osteoid osteoma treated by complete excision is thought to be very rare. Persistence of the lesion and reappearance of clinical and radiological signs have a more frequent occurrence, and are due to inadequate resection. Our case appears to be a true recurrence. It required no less than four operations first of these was probably not extensive enough. The second consisted of curettage of the osteoid osteoma after it had been exposed by abrasion of its cortical bone covering. The third and fourth resections were carried out under bone scan guidance and were controlled by postoperative radiography and computerised tomography. The uncertain aetiology of osteoid osteoma is one factor in the mysteriousness of this serial recurrence of what was apparently an ordinary osteoid osteoma. Such recurrence might be explained by an unknown persisting pathological environment.