The implications of model-informed drug discovery and development for tuberculosis

Despite promising advances in the field and highly effective first-line treatment, an estimated 9.6 million people are still infected with tuberculosis (TB). Innovative methods are required to effectively transition the growing number of compounds into novel combination regimens. However, progression of compounds into patients occurs despite the lack of clear understanding of the pharmacokinetic–pharmacodynamic (PK/PD) relations. The PreDiCT-TB consortium was established in response to the existing gaps in TB drug development. The aim of the consortium is to develop new preclinical tools in concert with an in silico model-based approach, grounded in PKPD principles. Here, we highlight the potential impact of such an integrated framework on various stages in TB drug development and on the dose rationale for drug combinations

The use of multiplex platforms for absolute and relative protein quantification of clinical material

Abstract When introducing multiplex platforms to measure protein content in precious clinical material there is an increased risk of cross reactivity, loss of sensitivity as well as accuracy. In this paper, four multiplex platforms and one singleplex platform were compared by running pre- and post-treatment plasma samples from CML patients. We found a variation of absolute protein concentrations between platforms. For some of the analytes and platforms, relative differences between pre- and post-treatment samples correlated. We conclude that absolute concentrations measured by different platforms should be compared with caution and comparing relative differences could be more accurate.Peer reviewe

Inter-nanocarrier and nanocarrier-to-cell transfer assays demonstrate the risk of an immediate unloading of dye from labeled lipid nanocapsules

Release studies constitute a fundamental part of the nanovector characterization. However, it can be difficult to correctly assess the release of lipophilic compounds from lipid nanocarriers using conventional assays. Previously, we proposed a method including an extraction with oil to measure the loading stability of lipophilic dyes in lipid nanocapsules (LNCs). The method indicated a rapid release of Nile Red from LNCs, while the loading of lipophilic carbocyanine dyes remained stable. This method, although interesting for a rapid screening of the fluorescence labeling stability of nanocarriers, is far from what happens in vivo, where lipid acceptor phases are nanostructured. Here, lipophilic dye loading stability has been assessed, by monitoring dye transfer from LNCs toward stable colloidal lipid nanocompartments, i.e. non-loaded LNCs, using new methodology based on size exclusion chromatography (SEC) and Förster Resonance Energy Transfer (FRET). Dye transfer between LNCs and THP-1 cells (as model for circulating cells) has also been studied by FACS. The assays reveal an almost instantaneous transfer of Nile Red between LNCs, from LNCs to THP-1 cells, between THP-1 cells, and a reversal transfer from THP-1 cells to LNCs. On the contrary, there was no detectable transfer of the lipophilic carbocyanine dyes. Dye release was also analyzed using dialyses, which only revealed a very slow release of Nile Red from LNCs, demonstrating the weakness of membrane based assays for investigations of the lipophilic compound loading stability in lipid nanocarriers. These results highlight the importance of using relevant release assays, and the potential risk of an immediate unloading of lipophilic fluorescent dyes from lipid nanocarriers, in the presence of a lipid acceptor nanocompartment. Some misinterpretations of cellular trafficking and in vivo biodistribution of fluorescent nanoparticles should be avoided

Phosphorylated Nucleolin Interacts with Translationally Controlled Tumor Protein during Mitosis and with Oct4 during Interphase in ES Cells

BACKGROUND: Reprogramming of somatic cells for derivation of either embryonic stem (ES) cells, by somatic cell nuclear transfer (SCNT), or ES-like cells, by induced pluripotent stem (iPS) cell procedure, provides potential routes toward non-immunogenic cell replacement therapies. Nucleolar proteins serve as markers for activation of embryonic genes, whose expression is crucial for successful reprogramming. Although Nucleolin (Ncl) is one of the most abundant nucleolar proteins, its interaction partners in ES cells have remained unidentified. METHODOLOGY: Here we explored novel Ncl-interacting proteins using in situ proximity ligation assay (PLA), colocalization and immunoprecipitation (IP) in ES cells. PRINCIPAL FINDINGS: We found that phosphorylated Ncl (Ncl-P) interacted with translationally controlled tumor protein (Tpt1) in murine ES cells. The Ncl-P/Tpt1 complex peaked during mitosis and was reduced upon retinoic acid induced differentiation, signifying a role in cell proliferation. In addition, we showed that Ncl-P interacted with the transcription factor Oct4 during interphase in human as well as murine ES cells, indicating of a role in transcription. The Ncl-P/Oct4 complex peaked during early stages of spontaneous human ES cell differentiation and may thus be involved in the initial differentiation event(s) of mammalian development. CONCLUSIONS: Here we described two novel protein-protein interactions in ES cells, which give us further insight into the complex network of interacting proteins in pluripotent cells

Autologous endothelialisation by the stromal vascular fraction on laminin-bioconjugated nanocellulose-alginate scaffolds

Establishing a vascular network in biofabricated tissue grafts is essential for ensuring graft survival. Such networks are dependent on the ability of the scaffold material to facilitate endothelial cell adhesion; however, the clinical translation potential of tissue-engineered scaffolds is hindered by the lack of available autologous sources of vascular cells. Here, we present a novel approach to achieving autologous endothelialisation in nanocellulose-based scaffolds by using adipose tissue-derived vascular cells on nanocellulose-based scaffolds. We used sodium periodate-mediated bioconjugation to covalently bind laminin to the scaffold surface and isolated the stromal vascular fraction and endothelial progenitor cells (EPCs; CD31+CD45−) from human lipoaspirate. Additionally, we assessed the adhesive capacity of scaffold bioconjugation in vitro using both adipose tissue-derived cell populations and human umbilical vein endothelial cells. The results showed that the bioconjugated scaffold exhibited remarkably higher cell viability and scaffold surface coverage by adhesion regardless of cell type, whereas control groups comprising cells on non-bioconjugated scaffolds exhibited minimal cell adhesion across all cell types. Furthermore, on culture day 3, EPCs seeded on laminin-bioconjugated scaffolds showed positive immunofluorescence staining for the endothelial markers CD31 and CD34, suggesting that the scaffolds promoted progenitor differentiation into mature endothelial cells. These findings present a possible strategy for generating autologous vasculature and thereby increase the clinical relevance of 3D-bioprinted nanocellulose-based constructs

Nanovectors for Neurotherapeutic Delivery Part II: Polymeric Nanoparticles

Despite major advances in intracranial surgery and delivery of drugs to the brain, treatment of neurological diseases remains one of the great medical challenges of our days. The complexity of the organ makes surgical procedures complicated, and conventional systemic delivery of drugs to the brain is hampered by low drug selectivity and low drug partitioning over the blood-brain barrier. Due to the high social and economic impacts related to diseases of the central nervous system, development of new improved treatments of brain related disorders is of significant value, both for the patient and for the society. Nanomedicine is a rapidly growing field in the development of novel therapies for treatments of brain pathologies. The scientific progress in nanotechnology has resulted in several new innovative nano-assemblies, with promising medical potentials. Therapeutic benefits related to the use of nanovectors includes, reduced chemical and enzymatic degradation of drugs, increased uptake over biological barriers, improved selectivity by surface modification using targeting ligands, and reduced toxic side effects in non-target tissue. This review discusses various applications of polymeric nanoparticles as nanovectors in treatment of neuronal diseases, specifically illustrated for Alzheimer’s and Parkinson’s diseases and Glioblastoma