Therapeutic Applications of Peptides against Zika Virus: A Review.

Zika Virus (ZIKV) belongs to the class of flavivirus that can be transmitted by Aedes mosquitoes. The number of Zika virus caused cases of acute infections, neurological disorders and congenital microcephaly are rapidly growing and therefore, in 2016, the World Health Organization declared a global "Public Health Emergency of International Concern". Anti-ZIKV therapeutic and vaccine development strategies are growing worldwide in recent years, however, no specific and safe treatment is available till date to save the human life. Currently, development of peptide therapeutics against ZIKV has attracted rising attention on account of their high safety concern and low development cost, in comparison to small therapeutic molecules and antibody-based anti-viral drugs. In present review, an overview of ZIKV inhibition by peptide-based inhibitors including E-protein derived peptides, antimicrobial peptides, frog skin peptides and probiotic peptides has been discussed. Peptides inhibitors have also been reported to act against NS5, NS2B-NS3 protease and proteasome in order to inhibit ZIKV infection. Recent advances in peptide-based therapeutics and vaccine have been reviewed and their future promise against ZIKV infections has been explored.

Sulfur-containing angiotensin-converting enzyme inhibitor 3-thienylalanine-ornithyl-proline activates endothelial function and expression of genes involved in Renin-Angiotensin system.

Experiments were performed to elucidate the mechanism of action of a 7-day oral administration of the sulfur-containing angiotensin-converting enzyme (ACE) inhibitor 3-thienylalanine-ornithyl-proline (TOP; 10 mg/kg/d) on endothelial dysfunction and oxidative stress compared with that of captopril (control; 40 mg/kg/d) in spontaneously hypertensive rats. The differential expression of messenger RNA by real-time reverse-transcriptase-polymerase chain reaction and protein by Western blot analysis was assessed for the markers nicotinamide adenine dinucleotide phosphate oxidase, p22phox, endothelial nitric oxide (NO) synthase, and AT1 receptor. Furthermore, TOP-induced vascular relaxation was also investigated using rat aortic rings in an organ bath. TOP significantly downregulated both messenger RNA and protein expressions of p22phox and AT1 receptor; the latter facilitates vasoconstriction through angiotensin II. In addition, TOP upregulated endothelial NO synthase, thus enhancing the production of NO. Vascular studies revealed that TOP caused endothelium-dependent vasorelaxation. In conclusion, unlike the free sulfur in captopril, the thiophene ring in TOP may act as a better scavenger of free radicals. Therefore, TOP exerted more significant antihypertensive effects than captopril, not only through angiotensin-converting enzyme inhibition but also through more effective antioxidation, because the inherent thiophene moiety resulted in the enhanced production of NO.

Rationally designed chimeric peptide of met-enkephalin and FMRFa-[D-Ala2,p-Cl-Phe4]YFa induce multiple opioid receptors mediated antinociception and up-regulate their expression.

The physiological role of NPFF/FMRFa family of peptides appears to be complex and exact mechanism of action of these peptides is not yet completely understood. In same line of scrutiny, another analog of YGGFMKKKFMRFamide (YFa), a chimeric peptide of met-enkephalin and FMRFamide, was rationally designed and synthesized which contain D-alanine and p-Cl-phenylalanine residues at 2nd and 4th positions, respectively i.e., Y-(D-Ala)-G-(p-Cl-Phe)-MKKKFMRFamide ([D-Ala(2), p-Cl-Phe(4)]YFa) in order to achieve improved bioavailability and blood brain barrier penetration. Therefore, present study investigates the possible antinociceptive effect of [D-Ala(2), p-Cl-Phe(4)]YFa on intra-peritoneal (i.p.) administration using tail-flick test in rats followed by its opioid receptor(s) specificity using mu, delta and kappa receptor antagonists. Further, its antinociceptive effect was examined during 6 days of chronic i.p. treatment and assessed effect of this treatment on differential expression of opioid receptors. [D-Ala(2), p-Cl-Phe(4)]YFa in comparison to parent peptide YFa, induce significantly higher dose dependent antinociception in rats which was mediated by all three opioid receptors (mu, delta and kappa). Importantly, it induced comparable antinociception in rats throughout the chronic i.p. treatment and significantly up-regulated the overall expression (mRNA and protein) of mu, delta and kappa opioid receptors. Therefore, pharmacological and molecular behavior of [D-Ala(2), p-Cl-Phe(4)]YFa demonstrate that incorporation of D-alanine and p-Cl-phenylalanine residues at appropriate positions in chimeric peptide leads to altered opioid receptor selectivity and enhanced antinociceptive potency, relative to parent peptide.

Effect of chronic intra-peritoneally administered chimeric peptide of met-enkephalin and FMRFa-[D-Ala2]YFa-on antinociception and opioid receptor regulation.

The physiological role of NPFF/FMRFa family of peptides is complex and exact mechanism of action of these peptides is not yet completely understood. In same line of scrutiny, previously we reported an enzymatically stable chimeric analog of YGGFMKKKFMRFamide (YFa) i.e., [D-Ala(2)]YAGFMKKKFMRFamide ([D-Ala(2)]YFa) which have a role in antinociception and modulatory effect on opioid analgesia. In continuation, presently we investigated using tail-flick test whether [D-Ala(2)]YFa on systemic administration induced any antinociception in rats and if so then which specific opioid receptor(s) mu, delta or kappa mediated it. Further, the antinociceptive effect of [D-Ala(2)]YFa on 6 days chronic intra-peritoneal (i.p.) treatment in rats was examined and finally, effect of this chronic treatment on the differential expression of opioid receptors was assessed. [D-Ala(2)]YFa on i.p. administration induced dose dependent antinociception which was mainly mediated by delta (DOR) and partially by mu (MOR) and kappa (KOR) opioid receptors. Moreover, its antinociceptive effect remained comparable throughout the chronic treatment even during insufficient availability of DOR1. Importantly, during this treatment the mRNA expression of all three opioid receptors (MOR1, KOR1 and DOR1) was increased as assessed by real-time RTPCR though subsequent western blot analysis revealed a selective increase in the protein level of DOR1, only. Thus, pharmacological behavior of [d-Ala(2)]YFa suggests that competency of an opioid agonist to bind with multiple opioid receptors may enhance its potency to induce tolerance free analgesia.

Endogenous peptide: Met-enkephalin-Arg-Phe, differently regulate expression of opioid receptors on chronic treatment.

Endogenous peptide, Met-enkephalin-Arg-Phe (Tyr-Gly-Gly-Phe-Met-Arg-Phe; MERF) induces effects like antinociception, inhibit contraction of guinea pig ileum, mouse vas deferens and anti-tussive action. However, results regarding its functional efficiency and selectivity are controversial. Therefore, present study was undertaken to investigate whether MERF on systemic (intra-peritoneal, i.p.) route of administration induce any antinociception or not; to scrutinize the effect of 6 days chronic i.p. treatment of MERF on expression of mu (MOR1), delta (DOR1) and kappa (KOR1) opioid receptors; and finally, the antinociceptive effect of two synthetic peptides, MERFamide and (D-Ala(2))-MERFamide was compared with MERF on intracerebroventricular administration in order to understand the role of FMRF moiety in analgesic effect of MERF. Pharmacological results revealed that only 68.4 and 91.2 micromol/kg dose induce significant antinociception among various doses. Further, on 6 days chronic treatment, MERF induced significant antinociception in comparison to saline. Differential expression of MOR1 and KOR1 showed continuous up-regulation throughout the treatment whereas DOR1 showed down-regulation in initial 3 days followed by subsequently up-regulation during the latter observable period. Moreover, variation in opioid receptors expression had not affected the MERF antinociception. In conclusion, present study discursively demonstrates that MERF during chronic treatment interacts with all three opioid receptors (mu, delta and kappa) in rats and differently regulates their expression. Further, the interaction was such that the induction was mainly observed at molecular/expression level and not at pharmacological level to affect antinociception.

Effect of varying chain length between P(1) and P(1') position of tripeptidomimics on activity of angiotensin-converting enzyme inhibitors.

One of the efficient modes of treatments of chronic hypertension and cardiovascular disorders has been to restrain the formation of angiotensin-II by inhibiting the action of angiotensin-converting enzyme (ACE) on angiotensin-I. The efforts continue towards achieving superior molecules or drugs with improved affinities, better bioavailability and thus to achieve long duration of action with minimum side effects. Previously, we reported a library of tripeptidomimics of Ornithyl-Proline (Orn-Pro) conjugated with various unnatural amino acids and carboxylic acid derived heterocyclics based on the SAR studies of existing ACE inhibitors. Their synthesis and screening for possible inhibitors of angiotensin-converting enzyme (ACE) revealed that increase in the backbone chain length by one carbon atom results in a sudden decrease in their activity. Therefore, in the present study heterocycles with different chain length were introduced to interact with the hydrophobic S(2) sub-site of ACE and screened for their in vitro ACE inhibition activity. Further, their binding interaction with C-domain of somatic ACE was also determined. Docking and consequent LUDI scores showed good correlation with binding of these molecules in the active site of ACE. Results suggest that heterocycles with C(3) chain length are more appropriate for the effective binding of the tripeptidomimics within the active site of ACE.

Lack of tolerance and morphine-induced cross-tolerance to the analgesia of chimeric peptide of Met-enkephalin and FMRFa.

Chimeric peptide of Met-enkephalin and FMRFa (YGGFMKKKFMRFa-YFa), a kappa-opioid receptor specific peptide, did not induce tolerance and cross-tolerance effects to its analgesic action on day 5 after pretreatment with either YFa or morphine for 4 days. However, pretreatment with YFa for 4 days led to the development of cross-tolerance to the analgesic effects of morphine and also 4 days of pretreatment of morphine resulted in the expression of tolerance to its own analgesic effects. Similar expression of tolerance and cross-tolerance were also observed when YFa was compared with the kappa receptor agonist peptide dynorphin A(1-13) [DynA(1-13)]. Cross-tolerance effects between YFa and DynA(1-13) analgesia were also not observed on day 5. Interestingly, when YFa and DynA(1-13) were tested for their analgesic effects for 5 days, reduction in analgesia on day 3 was observed in case of DynA(1-13) whereas YFa maintained its analgesia for 5 days. Thus, chimeric peptide YFa may serve as a useful probe to understand pain modulation and expression of tolerance and cross-tolerance behavior with other opioids.

YFa, a chimeric opioid peptide, induces kappa-specific antinociception with no tolerance development during 6 days of chronic treatment.

Our previous study showed that YGGFMKKKFMRFamide (YFa), a chimeric peptide of Met-enkephalin, and Phe-Met-Arg-Phe-NH2 induced naloxone-reversible antinociception and attenuated the development of tolerance to morphine analgesia. In continuation, the present study investigated which specific opioid receptors-mu, delta or kappa-mediate the observed YFa antinociception pharmacologically using specific antagonists and whether chronic administration of YFa at 26.01 micromol/kg per day induces tolerance and its effect on the expression of mu and kappa opioid receptors from day 4 to day 6, with endomorphine-1 (EM-1) and saline taken as positive and negative controls, respectively. Quantitative differential expression analysis was carried out by real-time reverse-transcriptase polymerase chain reaction, and the corresponding changes in protein levels were assessed by Western blot. A pharmacological investigation revealed that nor-binaltorphimine, a specific kappa opioid receptor-1 (KOR1) antagonist, completely antagonized the antinociception induced by 39.01 micromol/kg of YFa. Importantly, its chronic intraperitoneal administration did not result in significant tolerance over 6 days, whereas EM-1 induced significant tolerance after day 4. Differential expression analysis revealed that EM-1 caused up-regulation of mu opioid receptor-1 on day 4, followed by down-regulation on later days. Interestingly, YFa treatment caused a decrease on day 4, followed by an increase in the expression of KOR1 from day 5 onward. In conclusion, YFa induces kappa-specific antinociception, with no development of tolerance during 6 days of chronic treatment, which further articulates new directions for improved designing of peptide-based analgesics that may be devoid of adverse effects like tolerance.