Search

Nanotherapy has emerged as an improved anticancer therapeutic strategy to circumvent the harmful side effects of chemotherapy. It has been proven to be beneficial to offer multiple advantages, including their capacity to carry different therapeutic agents, longer circulation time and increased therapeutic index with reduced toxicity. Over time, nanotherapy evolved in terms of their designing strategies like geometry, size, composition or chemistry to circumvent the biological barriers. Multifunctional nanoscale materials are widely used as molecular transporter for delivering therapeutics and imaging agents. Nanomedicine involving multi-component chemotherapeutic drug-based combination therapy has been found to be an improved promising approach to increase the efficacy of cancer treatment. Next-generation nanomedicine has also utilized and combined immunotherapy to increase its therapeutic efficacy. It helps in targeting tumor immune response sparing the healthy systemic immune function. In this review, we have summarized the progress of nanotechnology in terms of nanoparticle designing and targeting cancer. We have also discussed its further applications in combination therapy and cancer immunotherapy. Integrating patient-specific proteomics and biomarker based information and harnessing clinically safe nanotechnology, the development of precision nanomedicine could revolutionize the effective cancer therapy.

RNA interference (RNAi) is a primitive evolutionary mechanism developed to escape incorporation of foreign genetic material. siRNA has been instrumental in achieving the therapeutic potential of RNAi by theoretically silencing any gene of interest in a reversible and sequence-specific manner. Extrinsically administered siRNA generally needs a delivery vehicle to span across different physiological barriers and load into the RISC complex in the cytoplasm in its functional form to show its efficacy. This review discusses the designing principles and examples of different classes of delivery vehicles that have proved to be efficient in RNAi therapeutics. We also briefly discuss the role of RNAi therapeutics in genetic and rare diseases, epigenetic modifications, immunomodulation and combination modality to inch closer in creating a personalized therapy for metastatic cancer. At the end, we present, strategies and look into the opportunities to develop efficient delivery vehicles for RNAi which can be translated into clinics.

Engineering bioinspired peptide-based molecular medicine is an emerging paradigm for the management of traumatic coagulopathies and inherent bleeding disorder. A hemostat-based strategy in managing uncontrolled bleeding is limited due to the lack of adequate efficacy and clinical noncompliance. In this study, we report an engineered adhesive peptide-based hybrid regenerative medicine, sealant 5, which is designed integrating the structural and functional features of fibrin and mussel foot-pad protein. AFM studies have revealed that sealant 5 (55.8 ± 6.8 nN adhesive force) has higher adhesive force than fibrin (46.4 ± 7.3 nN adhesive force). SEM data confirms that sealant 5 retains its network-like morphology both at 37 and 60 °C, inferring its thermal stability. Both sealant 5 and fibrin exhibit biodegradability in the presence of trypsin, and sealant 5 also showed biocompatibility in the presence of fibroblast cells. Engineered sealant 5 efficiently promotes hemostasis with enhanced adhesiveness and less blood-loss than fibrin. In vivo data suggests that in heparinized conditions, sealant 5 ceases bleeding at 212.3 ± 15.1 s, whereas fibrin halts bleeding at 294.3 ± 21.4 s and blood-loss is ∼4-fold less in sealant 5 than in fibrin. In a heparinized system, sealant 5 facilitates faster blood-clotting than fibrin (∼82 s faster) and RADA-16, a reported peptide-based sealant (∼113 s faster). Additionally, in the case of sealant 5, the process of clotting mimicry-like fibrin is independent of the body's own coagulation system. Sealant 5 efficiently halts bleeding for both external and internal wounds, even for a heparinized system overcoming the bacterial infection. ELISA data and PMBC cell proliferation data support the non-immunogenic feature of sealant 5. Though fibrin and sealant 5 have exhibited comparable efficacy in suture-free wound closure, in vivo H&E staining images have revealed infiltration of very few immune cells as well as the presence of abundant collagen formation in the case of sealant 5-treated wound. Such nature-inspired non-immunogenic sealants offer exciting possibilities for the treatment of uncontrolled bleeding vis-a-vis wound closure.

Developing non-immunogenic therapeutic biomolecules for facilitating blood clotting followed by wound healing via therapeutic angiogenesis, still remains a formidable challenge. Excessive blood loss of accident victims and battalions cause a huge number of deaths worldwide. Patients with inherited bleeding disorders face acute complications during injury and post-surgery. Biologically-inspired peptide-based hemostat can act as a potential therapeutic for handling coagulopathy. Additionally, non-healing wounds for patients having ischemic diseases can cause severe clinical complications. Advancement in stabilized growth-factor-based proangiogenic therapy may offer effective possibilities for the treatment of ischemic pathology. This review will discuss nature-inspired biocompatible stabilized peptide- and protein-based molecular medicines to serve unmet medical challenges for handling traumatic coagulopathy and impaired wound healing.

Traumatic coagulopathy due to severe external injury and internal hemorrhage is life-threatening to accident victims and soldiers on the battlefield, causing considerable number of deaths worldwide. Patients with inherited bleeding disorders (such as haemophilia, von Willebrand disease, inherited qualitative platelet defects, and afibrinogenemia) also contribute to the vast number of deaths due to abnormal bleeding, and these patients are difficult to handle during surgery. Platelets and different plasma proteins play an essential role in blood coagulation and in the maintenance of the body's hemostatic balance. The improper function or deficiency of these factors cause abnormal bleeding. To address such bleeding disorders, external clotting agents (such as extracellular protein-inspired natural and synthetic peptide-based sealants and peptide-functionalized polymer/liposome-based sealants) have been developed by different groups of researchers. The primary focus of this review is to provide molecular insights into the existing biologically inspired peptide-based sealants, highlighting the advantages and limitations of such reported designed sealants to handle blood clotting, and also provide insights into the design of improved next-generation surgical sealants.

Cytosolic delivery of functional siRNA remains the major challenge to develop siRNA-based therapeutics. Designing clinically safe and effective siRNA transporter to deliver functional siRNA across the plasma and endosomal membrane remains a key hurdle. With the aim of improving endosomal release, we have designed cyclic and linear peptide-based transporters having an Arg-DHis-Arg template. Computational studies show that the Arg-DHis-Arg template is also stabilized by the Arg-His side-chain hydrogen bonding interaction at physiological pH, which dissociates at lower pH. The overall atomistic interactions were examined by molecular dynamics simulations, which indicate that the extent of peptide_siRNA assembly formation depends greatly on physicochemical properties of the peptides. Our designed peptides having the Arg-DHis-Arg template and two lipidic moieties facilitate high yield of intracellular delivery of siRNA. Additionally, unsaturated lipid, linoleic acid moieties were introduced to promote fusogenicity and facilitate endosomal release and cytosolic delivery. Interestingly, such protease-resistant peptides provide serum stability to siRNA and exhibit high efficacy of erk1 and erk2 gene silencing in the triple negative breast cancer (TNBC) cell line. The peptide having two linoleyl moieties demonstrated comparable efficacy with commercial transfection reagent HiPerFect, as evidenced by the erk1 and erk2 gene knockdown experiment. Additionally, our study shows that ERK1/2 silencing siRNA and doxorubicin-loaded gramicidin-mediated combination therapy is more effective than siRNA-mediated gene silencing-based monotherapy for TNBC treatment.

Peptides have emerged as powerful platform technologies to engineer effective therapies against cancer. Peptides play significant role as clinically important molecules in developing next-generation cancer therapeutics by targeting signalling pathways, cell cycle, tumour suppressor protein, transcription factor, ion channels, several G-protein-coupled receptors and matrix metalloproteinases MMP 2 and MMP 9. Peptide-based cancer vaccines also hold huge promise in cancer treatment. Peptide-based biomarkers have immense potential to facilitate the early detection of cancer and thus increase the lifespan of cancer patients. Additionally, peptides also act as safe clinically suitable molecular medicine transporters for cancer treatment. Cell-penetrating cationic peptides, lipopeptides, amphipathic peptides and mini-proteins facilitate nucleic acid-based drug delivery and can be exploited for developing siRNA-based nanotherapeutics and combination therapy against cancer. Peptide-based medicines and molecular transporters play significant role in managing adaptive chemotherapeutic drug resistance and metastasis against cancer. Here we have discussed the current status and emerging opportunities of peptides as molecular medicines and molecular transporters.

Main chain chiral poly(methacrylate)s have been synthesized by controlled reversible addition–fragmentation chain transfer (RAFT) copolymerization of chiral monomer Boc- l -alanine methacryloyloxyethyl ester (Boc- l -Ala-HEMA, A) with Boc- d -alanine methacryloyloxyethyl ester (Boc- d -Ala-HEMA, B) and achiral monomer Boc-2-methylalanine-methacryloyloxyethyl ester (Boc-Aib-HEMA, C), and B with C by changing comonomer feed compositions. We systematically examined the role of l -alanine, d -alanine and 2-aminoisobutyric acid (Aib) as folding nuclei in our designed hybrid polymer-amino acid conjugates. Specific optical rotation and circular dichroism (CD) spectroscopy studies reveal that poly(Boc- l -Ala-HEMA) and poly(Boc- d -Ala-HEMA) formed entirely opposite handed helical structure stabilized by intramolecular hydrogen bonding among the pendant chromophores of the chiral monomers. Poly(Boc-Aib-HEMA) formed random coil structures conformation (unstructured conformation), whereas the resultant copolymers of C with A and B showed an unusual linear relationship of optical activity against comonomer composition, in sharp contrast with Green’s “majority” and “sergeant-and-soldier” rule. After Boc-group expulsion in acidic environment from the polymer side-chains, backbone retained its helicity. Variable temperature chiroptical properties investigation was also performed to examine the thermal stability of the homo and copolymer’s helical conformations. Additionally, pH induced conformational transformation was observed for the homo and copolymers.

Designing an effective peptide based molecular transporter for the intracellular delivery of hydrophilic therapeutic biomacromolecules remains a considerable challenge. Highly basic oligoarginine and lipidated arginine rich cell penetrating peptides have been reported in the literature as molecular transporters, which were extensively used for cellular internalization of significantly large biopharmaceuticals. However, oligoarginine based molecular transporters with l-arginine residues pose significant challenges due to proteolytic instability and limited stability of noncovalent peptide-cargo nanocomplexes. Exploiting the rational peptide designing strategy, we have engineered protease-resistant facial lipopeptide based molecular transporter having arginine-sarcosine-arginine moiety to minimize adjacent arginine-arginine pair repulsion. N-Methylated amino acid sarcosine was incorporated as a spacer between two adjacent arginine residues, which provides proteolytic stability to the designed peptide and minimizes intermolecular aggregation of peptides. Two stearyl moieties were incorporated to facilitate cellular internalization. Interestingly, our designed lipopeptide exhibits significantly enhanced cellular internalization with only six l-arginine residues compared to stearylated oligo-nona-arginine. Additionally, enhanced proteolytic stability of such class of molecular transporter enables increased cargo internalization, and we anticipate that our engineered multifunctional, proteolytically stable, nanostructured facial lipopeptide based molecular transporter can have major impact in advancing drug delivery technologies.

Designing biologically inspired nanoscale molecular assembly with desired functionality is a challenging endeavour. Here we report the designing of fibrin-inspired nanostructured peptide based sealants which facilitate remarkably fast entrapping of blood corpuscles (~28 seconds) in contrast to fibrin (~56 seconds). Our engineered sealants are stabilized by lysine-aspartate ionic interactions and also by Nε(γ-glutamyl) lysine isopeptide bond mediated covalent interaction. Each sealant is formed by two peptides having complementary charges to promote lysine-aspartate ionic interactions and designed isopeptide bond mediated interactions. Computational analysis reveals the isopeptide bond mediated energetically favourable peptide assemblies in sealants 1–3. Our designed sealants 2 and 3 mimic fibrin-mediated clot formation mechanism in presence of transglutaminase enzyme and blood corpuscles. These fibrin-inspired peptides assemble to form sealants having superior hemostatic activities than fibrin. Designed sealants feature mechanical properties, biocompatibility, biodegradability and high adhesive strength. Such nature-inspired robust sealants might be potentially translated into clinics for facilitating efficient blood clotting to handle traumatic coagulopathy and impaired blood clotting.

Therapeutic angiogenesis is an emerging paradigm for the management of ischemic pathologies. Proangiogenic Therapy is limited, however, by the current inability to deliver angiogenic factors in a sustained manner at the site of pathology. In this study, we investigated a unique nonglycosylated active fragment of hepatocyte growth factor/scatter factor, 1K1, which acts as a potent angiogenic agent in vitro and in a zebrafish embryo and a murine matrigel implant model. Furthermore, we demonstrate that nanoformulating 1K1 for sustained release temporally alters downstream signaling through the mitogen activated protein kinase pathway, and amplifies the angiogenic outcome. Merging protein engineering and nanotechnology offers exciting possibilities for the treatment of ischemic disease, and furthermore allows the selective targeting of downstream signaling pathways, which translates into discrete phenotypes.

The effects of inserting unsubstituted -amino acids into the strand segments of model -hairpin peptides was investigated by using four synthetic decapeptides, Boc-Leu-Val-Xxx-Val-D-Pro-Gly-Leu-Xxx-Val-Val-OMe: peptide 1 (Xxx=Gly), peptide 2 (Xxx=$\beta$Gly=$\beta$ahGly=homoglycine, -$\beta$glycine), peptide 3 (Xxx=$\gamma$Abu=-$\gamma$aminobutyric acid), peptide 4 (Xxx=$\delta$Ava=-$\delta$aminovaleric acid). $^1H\hspace{2mm}NMR$ studies (500 MHz, methanol) reveal several critical cross-strand NOEs, providing evidence for -$\beta$hairpin conformations in peptides 2-4. In peptide 3, the NMR results support the formation of the nucleating turn, however, evidence for cross-strand registry is not detected. Single-crystal X-ray diffraction studies of peptide 3 reveal a -$\beta$hairpin conformation for both molecules in the crystallographic asymmetric unit, stabilized by four cross-strand hydrogen bonds, with the $\gamma$Abu residues accommodated within the strands. The D-Pro-Gly segment in both molecules (A,B) adopts a type II -$\beta$turn conformation. The circular dichroism spectrum for peptide 3 is characterized by a negative CD band at 229 nm, whereas for peptides 2 and 4, the negative band is centered at 225 nm, suggesting a correlation between the orientation of the amide units in the strand segments and the observed CD pattern.

The incporation of beta-amino acid residues into the strand segments of designed beta-hairpin leads to the formation of polar sheets, since in the case of beta-peptide strands, all adjacent carbonyl groups point in one direction and the amide groups orient in the opposite direction. The conformational analysis of two designed peptide hairpins composed of alpha/beta-hybrid segments are described: Boc-Leu-betaPhe-Val-(D)-Pro-Gly-Leu-betaPhe-Val-OMe (1) and Boc-betaLeu-Phe-betaVal-D-Pro-Gly-betaLeu-Phe-betaVal-OMe (2). A 500-MHz 1H-NMR (nuclear magnetic resonance) analysis in methanol supports a significant population of hairpin conformations in both peptides. Diagnostic nuclear Overhauser effects (NOEs) are observed in both cases. X-ray diffraction studies on single crystals of peptide 1 reveal a beta-hairpin conformation in both the molecules, which constitute the crystallographic asymmetric unit. Three cross-strand hydrogen bonds and a nucleating type II' beta-turn at the D-Pro-Gly segment are observed in the two independent molecules. In peptide 1, the betaPhe residues at positions 2 and 7 occur at the nonhydrogen-bonding position, with the benzyl side chains pointing on opposite faces of the beta-sheet. The observed aromatic centroid-to-centroid distances are 8.92 A (molecule A) and 8.94 A (molecule B). In peptide 2, the aromatic rings must occupy facing positions in antiparallel strands, in the NMR-derived structure. Peptide 1 yields a normal "hairpin-like" CD spectrum in methanol with a minimum at 224 nm. The CD spectrum of peptide 2 reveals a negative band at 234 nm and a positive band at 221 nm, suggestive of an exciton split doublet. Modeling of the facing Phe side chains at the hydrogen-bonding position of a canonical beta-hairpin suggests that interring separation is approximately 4.78 A for the gauche+ gauche- (g+ g-) rotamer. A previously reported peptide beta-hairpin composed of only alpha-amino acids, Boc-Leu-Phe-Val-D-Pro-Gly-Leu-Phe-Val-OMe also exhibited an anomalous far-UV (ultraviolet) CD (circular dichroism) spectrum, which was interpreted in terms of interactions between facing aromatic chromophores, Phe 2 and Phe 7 (C. Zhao, P. L. Polavarapu, C. Das, and P. Balaram, Journal of the American Chemical Society, 2000, Vol 122, pp. 8228-8231).

We designed β-strand peptides that stabilize integral membrane proteins (IMPs). β-strand peptides self-assemble in solution as filaments and become restructured upon association with IMPs; resulting IMP-β-strand peptide complexes resisted aggregation when diluted in detergent-free buffer and were visible as stable, single particles with low detergent background in electron micrographs. β-strand peptides enabled clear visualization of flexible conformations in the highly dynamic ATP-binding cassette (ABC) transporter MsbA.

Conformational studies on the synthetic 11-aa peptide t -butoxycarbonyl (Boc)-Val-Ala-Phe-α-aminoisobutyric acid (Aib)-( R )-β 3 -homovaline (βVal)-( S )-β 3 -homophenylalanine (βPhe)-Aib-Val-Ala-Phe-Aib-methyl ester (OMe) (peptide 1; βVal and βPhe are β amino acids generated by homologation of the corresponding l -residues) establish that insertion of two consecutive β residues into a polypeptide helix can be accomplished without significant structural distortion. Crystal-structure analysis reveals a continuous helical conformation encompassing the segment of residues 2–10 of peptide 1. At the site of insertion of the ββ segment, helical hydrogen-bonded rings are expanded. A C 15 hydrogen bond for the αββ segment and two C 14 hydrogen bonds for the ααβ or βαα segments have been characterized. The following conformational angles were determined from the crystal structure for the β residues: βVal-5 (φ = –126°, θ = 76°, and ψ = –124) and βPhe-6 (φ =–88°, θ = 80°, and ψ =–118). The N terminus of the peptide is partially unfolded in crystals. The 500-MHz 1 H-NMR studies establish a continuous helix over the entire length of the peptide in CDCl 3 solution, as evidenced by diagnostic nuclear Overhauser effects. The presence of seven intramolecular hydrogen bonds is also established by using solvent dependence of NH chemical shifts.

The incorporation of the -amino acid residues into specific positions in the strands and -turn segments of peptide hairpins is being systematically explored. The presence of an additional torsion variable about the C()C() bond () enhances the conformational repertoire in -residues. The conformational analysis of three designed peptide hairpins composed of /-hybrid segments is described: Boc-Leu-Val-Val-DPro-Phe- Leu-Val-Val-OMe (1), Boc-Leu-Val-Val-DPro-Gly-Leu-Val-Val-OMe (2), and Boc-Leu-Val-Phe-Val-DPro- Gly-Leu-Phe-Val-Val-OMe (3). 500-MHz 1H-NMR Analysis supports a preponderance of -hairpin conformation in solution for all three peptides, with critical cross-strand NOEs providing evidence for the proposed structures. The crystal structure of peptide 2 reveals a -hairpin conformation with two -residues occupying facing, non-H-bonded positions in antiparallel -strands. Notably, Val(3 ) adopts a gauche conformation about the C()C() bond (65) without disturbing cross-strand H-bonding. The crystal structure of 2, together with previously published crystal structures of peptides 3 and Boc-Phe-Phe-DPro-Gly- Phe-Phe-OMe, provide an opportunity to visualize the packing of peptide sheets with local −polar segments× formed as a consequence of reversal peptide-bond orientation. The available structural evidence for hairpins suggests that -residues can be accommodated into nucleating turn segments and into both the H-bonding and non-H-bonding positions on the strands.

Ischemic disease causes a large number of deaths and significant clinical problems worldwide. Therapeutic angiogenesis, strengthened by advances in growth-factor-based therapies, is a promising solution to ischemic pathologies. Major challenges in therapeutic angiogenesis are the lack of stability of native angiogenic proteins and also providing sustained delivery of biologically active proteins at the ischemic sites. This paper will discuss various protein engineering strategies to develop stabilized proangiogenic proteins and several biomaterial technologies used to amplify the angiogenic outcome by delivering biologically active growth factors in a sustained manner.

The use of cisplatin, a first line chemotherapy for most cancers, is dose-limited due to nephrotoxicity. While this toxicity can be addressed through nanotechnology, previous attempts at engineering cisplatin nanoparticles have been limited by the impact on the potency of cisplatin. Here we report the rational engineering of a novel cisplatin nanoparticle by harnessing a novel polyethylene glycol-functionalized poly-isobutylene-maleic acid (PEG-PIMA) copolymer, which can complex with cis-platinum (II) through a monocarboxylato and a coordinate bond. We show that this complex self-assembles into a nanoparticle, and exhibits an IC(50) = 0.77 ± 0.11 µM comparable to that of free cisplatin (IC(50) = 0.44 ± 0.09 µM). The nanoparticles are internalized into the endolysosomal compartment of cancer cells, and release cisplatin in a pH-dependent manner. Furthermore, the nanoparticles exhibit significantly improved antitumor efficacy in a 4T1 breast cancer model in vivo, with limited nephrotoxicity, which can be explained by preferential biodistribution in the tumor with reduced kidney concentrations. Our results suggest that the PEG-PIMA-cisplatin nanoparticle can emerge as an attractive solution to the challenges in cisplatin chemotherapy.

Therapeutic angiogenesis is an emerging concept that can potentially be harnessed for management of ischemic pathologies. Hepatocyte growth factor/scatter factor (HGF/SF) promotes angiogenesis but ...

Half a century has passed since the hydrogen-bonded secondary structures of polypeptides and proteins were first recognized. An extraordinary wealth of conformational information is now available on peptides and proteins, which are formed of alpha-amino acid residues. More recently, the discovery of well-folded structures in oligopeptides containing beta-amino acids has focused a great deal of current interest on the conformational properties of peptides constructed from higher homologues (omega) of alpha-amino acids. This review examines the nature of intramolecularly hydrogen-bonded conformations of hybrid peptides formed by amino acid residues, with a varying number of backbone atoms. The beta-turn, a ubiquitous structural feature formed by two residue (alphaalpha) segments in proteins and peptides, is stabilized by a 10-atom (C10) intramolecular 4--1 hydrogen bond. Hybrid turns may be classified by comparison with their alphaalpha counterparts. The available crystallographic information on hydrogen-bonded hybrid turns is surveyed in this review. Several recent examples demonstrate that individual omega-amino acid residues and hybrid dipeptide segments may be incorporated into the regular structures of alpha-peptides. Examples of both peptide helices and hairpins are presented. The present review explores the relationships between folded conformations in hybrid sequences and their counterparts in all alpha-residue sequences. The use of stereochemically constrained omega-residues promises to expand the range of peptide design strategies to include omega-amino acids. This approach is exemplified by well-folded structures like the C12 (alphagamma) and C14 (gammagamma) helices formed in short peptides containing multiply substituted gamma-residues. The achiral gamma-residue gabapentin is a readily accessible building block in the design of peptides containing gamma-amino acids. The construction of globular polypeptide structures using diverse hybrid sequences appears to be a realistic possibility.

The crystal structures of four protected beta-amino acid residues, Boc-(S)-beta3-HAla-NHMe (1); Boc-(R)-beta3-HVal-NHMe (2); Boc-(S)-beta3-HPhe-NHMe (3); Boc-(S)-beta3-HPro-OH (6) and two beta-dipeptides, Boc-(R)-beta3-HVal-(R)-beta3-HVal-OMe (4); Boc-(R)-beta3-HVal-(S)-beta3-HVal-OMe (5) have been determined. Gauche conformations about the C(beta)-C(alpha) bonds (theta approximately +/-60 degrees) are observed for the beta3-HPhe residues in and all four beta3-HVal residues in the dipeptides and . Trans conformations (theta is approximately 180 degrees) are observed for beta3-HAla residues in both independent molecules in and for the beta3-HVal and beta3-HPro residues in and , respectively. In the cases of compounds , molecules associate in the crystals via intermolecular backbone hydrogen bonds leading to the formation of sheets. The polar strands formed by beta3-residues aggregate in both parallel (1,3,5) and antiparallel (2,4 fashion. Sheet formation accommodates both the trans and gauche conformations about the C(beta)-C(alpha) bonds.