A Nonlinear Peptide Topology for Synthetic Virions.

a nonlinear de novo peptide topology for the assembly of synthetic virions is reported. The topology is a backbone cyclized amino-acid sequence in which polar l- and hydrophobic d-amino acid residues of the same-type alternate. This arrangement introduces pseudo C4 symmetries of side chains within the same cyclopeptide ring, allowing for the lateral propagation of cyclopeptides into networks with a [3/6, 4]-fold rotational symmetry closing into virus-like shells. A combination of computational and experimental approaches was used to establish that the topology forms morphologically uniform, nonaggregating and nontoxic nanoscale shells. These effectively encapsulate genetic cargo and promote its intracellular delivery and a target genetic response. The design introduces a nanotechnology inspired solution for engineering virus-like systems thereby expanding traditional molecular biology approaches used to create artificial biology to chemical space.

DNA synthesis technologies to close the gene writing gap.

Synthetic DNA is of increasing demand across many sectors of research and commercial activities. Engineering biology, therapy, data storage and nanotechnology are set for rapid developments if DNA can be provided at scale and low cost. Stimulated by successes in next generation sequencing and gene editing technologies, DNA synthesis is already a burgeoning industry. However, the synthesis of >200 bp sequences remains unaffordable. To overcome these limitations and start writing DNA as effectively as it is read, alternative technologies have been developed including molecular assembly and cloning methods, template-independent enzymatic synthesis, microarray and rolling circle amplification techniques. Here, we review the progress in developing and commercializing these technologies, which are exemplified by innovations from leading companies. We discuss pros and cons of each technology, the need for oversight and regulatory policies for DNA synthesis as a whole and give an overview of DNA synthesis business models.

Folding-Mediated DNA Delivery by α-Helical Amphipathic Peptides.

The renaissance gene therapy experiences these days requires specialist biomaterials and a systemic understanding of major factors influencing their ability to deliver genetic material. Peptide transfection systems represent a major class of such biomaterials. Several peptidic reagents have been commercialized to date. However, a comparative assessment of peptide sequences alone without auxiliary support or excipients against a common determinant for their ability to complex and deliver DNA has been lacking. This study cross-compares commercial and experimental transfection reagents from the same family of helical amphiphiles. Factors defining the efficacy of DNA delivery including cell uptake and gene expression are assessed along with cytotoxicity and DNA complexation. The results show that despite differences in sequence composition, length, and origin, peptide reagents of the same structural family exhibit similar characteristics and limitations with common variability trends. The cross-comparison revealed that functional DNA delivery is independent of the peptide sequence used but is mediated by the ability of the reagents to co-fold with DNA. Peptide folding proved to be the common determinant for DNA complexation and delivery by peptidic transfection reagents.

Designer protein pseudo-capsids targeting intracellular bacteria.

The emergence of multidrug-resistant bacteria stimulates the search for antimicrobial materials capable of addressing challenges conventional antibiotics fail to address. The ability to target intracellular bacteria remains one of the most fundamental tasks for contemporary antimicrobial treatments. Here we report engineered protein pseudo-capsids targeting bacteria internalised in macrophages. Using a combination of live-cell imaging and single-cell electron microscopy analysis we show that these materials effectively disrupt the bacteria without affecting the host cells. The study offers a disruptive antimicrobial strategy demonstrating potential for developing principally more challenging mechanisms for bacteria to overcome.

A KDM4A-PAF1-mediated epigenomic network is essential for acute myeloid leukemia cell self-renewal and survival.

Epigenomic dysregulation is a common pathological feature in human hematological malignancies. H3K9me3 emerges as an important epigenomic marker in acute myeloid leukemia (AML). Its associated methyltransferases, such as SETDB1, suppress AML leukemogenesis, whilst H3K9me3 demethylases KDM4C is required for mixed-lineage leukemia rearranged AML. However, the specific role and molecular mechanism of action of another member of the KDM4 family, KDM4A has not previously been clearly defined. In this study, we delineated and functionally validated the epigenomic network regulated by KDM4A. We show that selective loss of KDM4A is sufficient to induce apoptosis in a broad spectrum of human AML cells. This detrimental phenotype results from a global accumulation of H3K9me3 and H3K27me3 at KDM4A targeted genomic loci thereby causing downregulation of a KDM4A-PAF1 controlled transcriptional program essential for leukemogenesis, distinct from that of KDM4C. From this regulatory network, we further extracted a KDM4A-9 gene signature enriched with leukemia stem cell activity; the KDM4A-9 score alone or in combination with the known LSC17 score, effectively stratifies high-risk AML patients. Together, these results establish the essential and unique role of KDM4A for AML self-renewal and survival, supporting further investigation of KDM4A and its targets as a potential therapeutic vulnerability in AML.

The Emerging Role of H3K9me3 as a Potential Therapeutic Target in Acute Myeloid Leukemia.

Growing evidence has demonstrated that epigenetic dysregulation is a common pathological feature in human cancer cells. Global alterations in the epigenetic landscape are prevalent in malignant cells across different solid tumors including, prostate cancer, non-small-cell lung cancer, renal cell carcinoma, and in haemopoietic malignancy. In particular, DNA hypomethylation and histone hypoacetylation have been observed in acute myeloid leukemia (AML) patient blasts, with histone methylation being an emerging area of study. Histone 3 lysine 9 trimethylation (H3K9me3) is a post-translational modification known to be involved in the regulation of a broad range of biological processes, including the formation of transcriptionally silent heterochromatin. Following the observation of its aberrant methylation status in hematological malignancy and several other cancer phenotypes, recent studies have associated H3K9me3 levels with patient outcome and highlighted key molecular mechanisms linking H3K9me3 profile with AML etiology in a number of large-scale meta-analysis. Consequently, the development and application of small molecule inhibitors which target the histone methyltransferases or demethylase enzymes known to participate in the oncogenic regulation of H3K9me3 in AML represents an advancing area of ongoing study. Here, we provide a comprehensive review on how this particular epigenetic mark is regulated within cells and its emerging role as a potential therapeutic target in AML, along with an update on the current research into advancing the generation of more potent and selective inhibitors against known H3K9 methyltransferases and demethylases.

Solid-Phase Synthesis of Oxetane Modified Peptides.

Solid-phase peptide synthesis (SPPS) is used to create peptidomimetics in which one of the backbone amide C═O bonds is replaced by a four-membered oxetane ring. The oxetane containing dipeptide building blocks are made in three steps in solution, then integrated into peptide chains by conventional Fmoc SPPS. This methodology is used to make a range of peptides in high purity including backbone modified derivatives of the nonapeptide bradykinin and Met- and Leu-enkephalin.