[(WR)8WKβA]-Doxorubicin Conjugate: A Delivery System to Overcome Multi-Drug Resistance against Doxorubicin

Doxorubicin (Dox) is an anthracycline chemotherapeutic agent used to treat breast, leukemia, and lymphoma malignancies. However, cardiotoxicity and inherent acquired resistance are major drawbacks, limiting its clinical application. We have previously shown that cyclic peptide [WR]9 containing alternate tryptophan (W) and arginine (R) residues acts as an efficient molecular transporter. An amphiphilic cyclic peptide containing a lysine (K) residue and alternative W and R was conjugated through a free side chain amino group with Dox via a glutarate linker to afford [(WR)8WKβA]-Dox conjugate. Antiproliferative assays were performed in different cancer cell lines using the conjugate and the corresponding physical mixture of the peptide and Dox to evaluate the effectiveness of synthesized conjugate compared to the parent drug alone. [(WR)8WKβA]-Dox conjugate showed higher antiproliferative activity at 10 µM and 5 µM than Dox alone at 5 μM. The conjugate inhibited the cell viability of ovarian adenocarcinoma (SK-OV-3) by 59% and the triple-negative breast cancer cells MDA-MB-231 and MCF-7 by 71% and 77%, respectively, at a concentration of 5 μM after 72 h of incubation. In contrast, Dox inhibited the proliferation of SK-OV-3, MDA-MB-231, and MCF-7 by 35%, 63%, and 57%, respectively. Furthermore, [(WR)8WKβA]-Dox conjugate (5 µM) inhibited the cell viability of Dox-resistant cells (MES-SA/MX2) by 92%, while the viability of cells incubated with free Dox was only 15% at 5 μM. Confocal microscopy images confirmed the ability of both Dox conjugate and the physical mixture of the peptide with the drug to deliver Dox through an endocytosis-independent pathway, as the uptake was not inhibited in the presence of endocytosis inhibitors. The stability of Dox conjugate was observed at different time intervals using analytical HPLC when the conjugate was incubated with 25% human serum. Half-life (t1/2) for [(WR)8WKβA]-Dox conjugate was (∼6 h), and more than 80% of the conjugate was degraded at 12 h. The release of free Dox was assessed intracellularly using the CCRF-CEM cell line. The experiment demonstrated that approximately 100% of free Dox was released from the conjugate intracellularly within 72 h. These data confirm the ability of the cyclic cell-penetrating peptide containing tryptophan and arginine residues as an efficient tool for delivery of Dox and for overcoming resistance to it.

Ribosomally derived lipopeptides containing distinct fatty acyl moieties

Lipopeptides represent a large group of microbial natural products that include important antibacterial and antifungal drugs and some of the most-powerful known biosurfactants. The vast majority of lipopeptides comprise cyclic peptide backbones N-terminally equipped with various fatty acyl moieties. The known compounds of this type are biosynthesized by nonribosomal peptide synthetases, giant enzyme complexes that assemble their products in a non–gene-encoded manner. Here, we report the genome-guided discovery of ribosomally derived, fatty-acylated lipopeptides, termed selidamides. Heterologous reconstitution of three pathways, two from cyanobacteria and one from an arctic, ocean-derived alphaproteobacterium, allowed structural characterization of the probable natural products and suggest that selidamides are widespread over various bacterial phyla. The identified representatives feature cyclic peptide moieties and fatty acyl units attached to (hydroxy)ornithine or lysine side chains by maturases of the GCN5-related N-acetyltransferase superfamily. In contrast to nonribosomal lipopeptides that are usually produced as congener mixtures, the three selidamides are selectively fatty acylated with C10, C12, or C16 fatty acids, respectively. These results highlight the ability of ribosomal pathways to emulate products with diverse, nonribosomal-like features and add to the biocatalytic toolbox for peptide drug improvement and targeted discovery.

A new amino acid for improving permeability and solubility in macrocyclic peptides through side chain-to-backbone hydrogen bonding.

Despite the notoriously poor membrane permeability of peptides in general, many cyclic peptide natural products show high passive membrane permeability and potently inhibit a variety of “undruggable” intracellular targets. A major impediment to designing cyclic peptides with good permeability is the high desolvation energy associated with the peptide backbone amide NH groups. Strategies for mitigating the deleterious effect of the backbone NH group on permeability include N-methylation, steric occlusion, and the formation of intramolecular hydrogen bonds with backbone carbonyl oxygens, while there have been relatively few studies on the use of polar side chains to sequester backbone NH groups. We investigated the ability of N,N-pyrrolidinyl glutamine (Pye), whose side chain contains a powerful hydrogen bond accepting C=O amide group but no hydrogen bond donors, to sequester exposed backbone NH groups in a series of cyclic hexapeptide diastereomers. Analyses of partition coefficients, lipophilic permeability efficiencies (LPE), artificial and cell-based permeability assays revealed that specific Leu-to-Pye substitutions conferred dramatic improvements in aqueous solubility and permeability in a scaffold- and position-dependent manner. Introduction of the Pye residue thus offers a complementary tool, alongside traditional approaches, for improving membrane permeability and solubility in cyclic peptides.

Preclinical and randomized phase I studies of plitidepsin in adults hospitalized with COVID-19

Plitidepsin, a marine-derived cyclic-peptide, inhibits SARS-CoV-2 replication at nanomolar concentrations by targeting the host protein eukaryotic translation elongation factor 1A. Here, we show that plitidepsin distributes preferentially to lung over plasma, with similar potency against across several SARS-CoV-2 variants in preclinical studies. Simultaneously, in this randomized, parallel, open-label, proof-of-concept study (NCT04382066) conducted in 10 Spanish hospitals between May and November 2020, 46 adult hospitalized patients with confirmed SARS-CoV-2 infection received either 1.5 mg (n = 15), 2.0 mg (n = 16), or 2.5 mg (n = 15) plitidepsin once daily for 3 d. The primary objective was safety; viral load kinetics, mortality, need for increased respiratory support, and dose selection were secondary end points. One patient withdrew consent before starting procedures; 45 initiated treatment; one withdrew because of hypersensitivity. Two Grade 3 treatment-related adverse events were observed (hypersensitivity and diarrhea). Treatment-related adverse events affecting more than 5% of patients were nausea (42.2%), vomiting (15.6%), and diarrhea (6.7%). Mean viral load reductions from baseline were 1.35, 2.35, 3.25, and 3.85 log10 at days 4, 7, 15, and 31. Nonmechanical invasive ventilation was required in 8 of 44 evaluable patients (16.0%); six patients required intensive care support (13.6%), and three patients (6.7%) died (COVID-19-related). Plitidepsin has a favorable safety profile in patients with COVID-19.

1D alignment of proteins and other nanoparticles by using reversible covalent bonds on cyclic peptide nanotubes

Self-assembling cyclic peptide nanotubes are supramolecular structures whose diameter and external surface properties are precisely controlled. In this communication we describe a general strategy to align different molecules on top...

A rare case of Interstitial lung disease complicated by overlapping syndrome with obstructive ventilatory dysfunction as the first symptom

Interstitial lung disease is often a serious complication of connective tissue disease（CTD）, representing the first cause of death in CTD. However, there are few reports of SSc-RA overlap-associated interstitial pneumonia. Respiratory dysfunction as the first clinical manifestation is even rarely reported. We herein described a case of a male patient who developed significant respiratory dysfunction as the principal clinical sign for the past 6 months, plus newly developed skin thickening in bilateral upper limbs and pain and swelling of multiple joints for the past 6 weeks. Extensive immunological screening showed high titer of antinuclear antibodies(ANA), rheumatoid factor(RF), anti-cyclic peptide containing citrulline (anti-CCP)，and positive anti-Scl-70 antibody. Chest high resolution computed tomography(HRCT) was performed and hence ILD was confirmed. Pulmonary function test (PFT) revealed obstructive ventilatory dysfunction rather than restrictive ventilatory dysfunction. So bronchodilation test was performed and asthma was considered. Finally, after a multidisciplinary team (MDT) discussion, the diagnosis of asthma and ILD associated with SSc-RA overlap was established. It is important to note that CTD associated pulmonary interstitial fibrosis must be considered as the differential diagnosis of any newly diagnosed interstitial pulmonary disease. In addition, if patient with dyspnea is highly suspicious of pulmonary interstitial fibrosis, bronchodilation test is still needed to exclude asthma in order to avoid misdiagnosis.

A host-directed macrocyclic peptide therapeutic for MDR gram negative bacterial infections

The emergence of infections by carbapenem resistant Enterobacteriaceae (CRE) pathogens has created an urgent public health threat, as carbapenems are among the drugs of last resort for infections caused by a growing fraction of multi-drug resistant (MDR) bacteria. There is global consensus that new preventive and therapeutic strategies are urgently needed to combat the growing problem of MDR bacterial infections. Here, we report on the efficacy of a novel macrocyclic peptide, minimized theta-defensin (MTD)-12813 in CRE sepsis. MTD12813 is a theta-defensin inspired cyclic peptide that is highly effective against CRE pathogens K. pneumoniae and E. coli in vivo. In mouse septicemia models, single dose administration of MTD12813 significantly enhanced survival by promoting rapid host-mediated bacterial clearance and by modulating pathologic cytokine responses, restoring immune homeostasis, and preventing lethal septic shock. The peptide lacks direct antibacterial activity in the presence of mouse serum or in peritoneal fluid, further evidence for its indirect antibacterial mode of action. MTD12813 is highly stable in biological matrices, resistant to bacterial proteases, and nontoxic to mice at dose levels 100 times the therapeutic dose level, properties which support further development of the peptide as a first in class anti-infective therapeutic.