Amifostine analog, DRDE-30, alleviates radiation induced lung damage by attenuating inflammation and fibrosis.

BACKGROUND: Radiotherapy of thoracic neoplasms and accidental radiation exposure often results in pneumonitis and fibrosis of lungs. Here, we investigated the potential of amifostine analogs: DRDE-07, DRDE-30, and DRDE-35, in alleviating radiation-induced lung damage. METHODS: C57BL/6 mice were exposed to 13.5 Gy thoracic irradiation, 30 min after intraperitoneal administration of the analogs, and assessed for modulation of the pathological response at 12 and 24 weeks. KEY FINDINGS: DRDE-07, DRDE-30 and DRDE-35 increased the survival of irradiated mice from 20% to 30%, 80% and 70% respectively. Reduced parenchymal opacity (X-ray CT) in the lungs of DRDE-30 pre-treated mice corroborated well with the significant decrease in Ashcroft score (p < 0.01). Two-fold increase in SOD and catalase activities (p < 0.05), coupled with a 50% increase in GSH content and a 60% decrease in MDA content (p < 0.05) suggested restoration of the antioxidant defence system. A 20% to 40% decrease in radiation-induced apoptotic and mitotic death in the lung tissue (micronuclei: p < 0.01), resulted in attenuated lung and vascular permeability (FITC-Dextran leakage) by 50% (p < 0.01), and a commensurate reduction (~50%) in leukocyte infiltration in the injured tissue (p < 0.05). DRDE-30 abrogated the activation of pro-inflammatory NF-κB and p38/MAPK signaling cascades, suppressing the release of pro-inflammatory cytokines (IL-1ß: p < 0.05; TNF-α: p < 0.05; IL-6: p < 0.05) and up-regulation of CAMs on the endothelial cell surface. Reduction in hydroxyproline content (p < 0.01) and collagen suggested inhibition of lung fibrosis which was associated with attenuation of TGF-ß/Smad pathway-mediated-EMT. CONCLUSION: DRDE-30 could be a potential prophylactic agent against radiation-induced lung injury.

E-N-(2-acetyl-phenyl)-3-phenyl-acrylamide targets abrin and ricin toxicity: Hitting two toxins with one stone.

The efficacy of small molecule inhibitors (SMIs) against the enzymatic activity of Shiga toxin prompted the evaluation of their efficacy on related toxins viz. ricin and abrin. Ricin, like Shiga toxin, is listed as a category B bioweapon and belongs to the type II family of ribosome inactivating proteins (RIPs). Abrin though structurally and functionally similar to ricin, is considerably more toxic. In the present study, 35 compounds were evaluated in A549 cells in in vitro assays, of which 5 offered protection against abrin and 2 against ricin, with IC50 values ranging between 30.5-1379 µM and 300-341 µM, respectively. These findings are substantiated by fluorescence based thermal shift assay. Moreover, the binding of the promising compounds to the toxin components has been validated by Surface Plasmon Resonance assay and in vitro protein synthesis assay. In vivo studies reveal complete protection of mice with compound 4 E-N-(2-acetyl-phenyl)-3-phenyl-acrylamide against orally administered lethal doses of, both, abrin and ricin. The present study thus proposes the emergence of E-N-(2-acetyl-phenyl)-3-phenyl-acrylamide as a lead compound against RIPs.

A dual-channel optical chemical sensing system for selective detection of nerve agent simulant DFP.

This paper reports a novel optical chemical sensing system for selective detection of diisopropylfluorophosphate (DFP), a simulant of fluorine-containing nerve agents (Sarin and Soman). Contrary to the reported methods involving only single sensing probe, this sensing system is comprised of two molecular sensing probes (1 and 2) having intrinsic affinities for reactive subunits of DFP (electrophilic phosphorus and fluoride ion). On exposure to DFP, two molecular probes react in tandem with electrophilic phosphorus and fluoride ion (by-product of the initial phosphorylation reaction) to induce a unique modulation in the optical properties of the sensing system which leads to selective detection of DFP in solution as interferents like phosphorus-containing compounds, acids, and anions were unable to induce similar optical modulation due to lack of both electrophilic phosphorus and fluorine in the same molecule. Calibration curve between the amount of DFP added and the absorption intensity revealed the colorimetric detection limit of the system to be 4.50 µM which was further lowered to 2.22 µM by making use of a self-immolative fluoride sensing probe 5.

Amifostine Analog, DRDE-30, Attenuates Bleomycin-Induced Pulmonary Fibrosis in Mice.

Bleomycin (BLM) is an effective curative option in the management of several malignancies including pleural effusions; but pulmonary toxicity, comprising of pneumonitis and fibrosis, poses challenge in its use as a front-line chemotherapeutic. Although Amifostine has been found to protect lungs from the toxic effects of radiation and BLM, its application is limited due to associated toxicity and unfavorable route of administration. Therefore, there is a need for selective, potent, and safe anti-fibrotic drugs. The current study was undertaken to assess the protective effects of DRDE-30, an analog of Amifostine, on BLM-induced lung injury in C57BL/6 mice. Whole body micro- computed tomography (CT) was used to non-invasively observe tissue damage, while broncheo-alveolar lavage fluid (BALF) and lung tissues were assessed for oxidative damage, inflammation and fibrosis. Changes in the lung density revealed by micro-CT suggested protection against BLM-induced lung injury by DRDE-30, which correlated well with changes in lung morphology and histopathology. DRDE-30 significantly blunted BLM-induced oxidative stress, inflammation and fibrosis in the lungs evidenced by reduced oxidative damage, endothelial barrier dysfunction, Myeloperoxidase (MPO) activity, pro-inflammatory cytokine release and protection of tissue architecture, that could be linked to enhanced anti-oxidant defense system and suppression of redox-sensitive pro-inflammatory signaling cascades. DRDE-30 decreased the BLM-induced augmentation in BALF TGF-ß and lung hydroxyproline levels, as well as reduced the expression of the mesenchymal marker α-smooth muscle actin (α-SMA), suggesting the suppression of epithelial to mesenchymal transition (EMT) as one of its anti-fibrotic effects. The results demonstrate that the Amifostine analog, DRDE-30, ameliorates the oxidative injury and lung fibrosis induced by BLM and strengthen its potential use as an adjuvant in alleviating the side effects of BLM.

In Silico Discovery and Validation of Amide Based Small Molecule Targeting the Enzymatic Site of Shiga Toxin.

Shiga toxin (Stx), a category B biothreat agent, is a ribosome inactivating protein and toxic to human and animals. Here, we designed and synthesized small molecules that block the active site of the Stx A subunit. On the basis of binding energy, 20 molecules were selected for synthesis and evaluation. These molecules were primarily screened using fluorescence-based thermal shift assay and in vitro in Vero cells. Among 32 molecules (including 12 reported), six molecules offered protection with IC50 of 2.60-23.90 µM. 4-Nitro-N-[2-(2-phenylsulfanylethylamino)ethyl]benzamide hydrochloride is the most potent inhibitor with IC50 at 7.96 µM and selectivity index of 22.23 and is better than any known small molecule inhibitor of Stx. Preincubation with Stx offered full protection against Shiga toxin in mice. Surface plasmon resonance assay further confirmed that these molecules bind specifically to Stx A subunit. Further optimization is continued to identify a potential candidate which will be in vivo effective.

Mutagenicity and antimutagenicity studies of DRDE-07 and its analogs against sulfur mustard in the in vitro Ames Salmonella/microsome assay.

Sulfur mustard (bis(2-chloroethyl) sulfide, SM), a chemical warfare agent, is classified as a class I human carcinogen by IARC. No effective antidote against this agent is available. The synthetic aminothiol, amifostine, earlier known as WR-2721, has been extensively used as a chemical radioprotector for normal tissues in cancer radiotherapy and chemotherapy. SM is a radiomimetic agent; this prompted us to evaluate the protective efficacy of amifostine and three of its analogs, DRDE-07 [S-2(2-aminoethylamino) ethyl phenyl sulphide], DRDE-30 [S-2(2-aminoethyl amino) ethyl propyl sulphide] and DRDE-35 [S-2(2-aminoethyl amino) ethyl butyl sulphide], against sulfur mustard-induced mutagenicity in the Ames Salmonella/microsome assay. The antidotes were also evaluated for possible mutagenic activity. DRDE-07 was mutagenic in strain TA104 in the absence of S9; DRDE-30 was mutagenic in strain TA100; amifostine and DRDE-35 did not show mutagenic activity in any of the five tester strains used. SM is mutagenic in strains TA97a and TA102, with or without S9 activation. In the antimutagenicity studies, DRDE-07 and DRDE-35 showed promising antimutagenic activity against SM in the absence of S9, in comparison to amifostine. DRDE-07 and DRDE-35 are promising protective agents against SM-induced mutagenicity.

Nanoencapsulation of DMSA monoester for better therapeutic efficacy of the chelating agent against arsenic toxicity.

AIMS: Exposure to toxic metals remains a widespread occupational and environmental problem in world. Chelation therapy is a mainstream treatment used to treat heavy metal poisoning. This paper describes the synthesis, characterization and therapeutic evaluation of monoisoamyl 2,3-dimercaptosuccinic acid (MiADMSA)-encapsulated polymeric nanoparticles as a detoxifying agent for arsenic poisoning. MATERIALS & METHODS: Polymeric nanoparticles entrapping the DMSA monoester, which can evade the reticulo-endothelial system and have a long circulation time in the blood, were prepared. Particle characterization was carried out by transmission electron microscopy and dynamic light scattering. An in vivo study was conducted to investigate the therapeutic efficacy of MiADMSA-encapsulated polymeric nanoparticles (nano- MiADMSA; 50 mg/kg orally for 5 days) and comparison drawn with bulk MiADMSA. Swiss albino mice exposed to sodium arsenite for 4 weeks were treated for 5 days to evaluate alterations in blood, brain, kidney and liver oxidative stress variables. The study also evaluated the histopathological changes in tissues and the chelating potential of the nanoformulation. RESULTS: Our results show that nano-MiADMSA have a narrow size distribution in the 50-nm range. We observed an enhanced chelating potential of nano-MiADMSA compared with bulk MiADMSA as evident in the reversal of biochemical changes indicative of oxidative stress and efficient removal of arsenic from the blood and tissues. Histopathological changes and urinary 8-OHdG levels also prove better therapeutic efficacy of the novel formulation for arsenic toxicity. CONCLUSION: The results from our study show better therapeutic efficacy of nano-MiADMSA in removing arsenic burden from the brain and liver.

Ameliorating effect of S-2(ω-aminoalkylamino) alkylaryl sulfide (DRDE-07) on sulfur mustard analogue, 2-chloroethyl ethyl sulfide-induced oxidative stress and inflammation.

Sulfur mustard (SM; 2,2'-dichloro diethyl sulfide), an alkylating chemical warfare agent, poses a major threat in both military conflict and chemical terrorism situations. 2-chloroethyl ethyl sulfide (CEES) is a monofunctional analogue of SM, frequently used in laboratory settings, therefore increasing chances of its exposure. S-2(ω-aminoalkylamino) alkylaryl sulfide (DRDE-07) is an analogue of amifostine reported to have protective effects against SM but its effect on CEES is largely unexplored. Therefore, this study was planned to explore the effects of DRDE-07 against CEES-induced toxicity. 0.75 LD50 (1068 mg/kg) of CEES was exposed percutaneously in the presence or absence of DRDE-07 (249 mg/kg p.o.) which is given prophylactically (before 30 minute) to male mice. Animals were sacrificed on 24 h, 7th day and 14th day of CEES exposure, and tissues were collected to study oxidative stress and inflammatory markers. CEES exposure depleted intracellular GSH level and activities of GSH-linked enzymes (GR, GPx and GST) which play a major role in GSH metabolism. CEES exposure augmented lipid peroxidation indicating severe oxidative stress. It also initiated inflammation causing an increase in proinflammatory (IL1-α, IL1-ß, IL-6, TNF-α and IFN-ϒ) and corresponding decrease in anti-inflammatory cytokines (IL-4 and IL-10). This was also accompanied by neutrophils infiltration indicated by higher than normal myeloperoxidase (MPO) levels. DRDE-07 efficiently reduced the oxidative stress and also facilitated to resolve inflammatory alterations. This study thus evaluated the beneficial role of DRDE-07 in ameliorating the deleterious effects of CEES and can be potentially used against SM/CEES poisoning.

The genus Plectranthus in India and its chemistry.

Phytochemical constituents isolated from Indian species of the genus Plectranthus reported up to 2009 are compiled. In India, the genus Plectranthus is found in all the habitats and altitudes, particularly in the Himalaya, the Southern Ghats, and the Nilgiri region. P. amboinicus, P. barbatus, P. caninus, P. mollis, P. coetsa, and P. incanus are the most common species found in India. Phytochemical studies of the genus revealed that Indian Plectranthus species are rich in essential oil, and that the most abundant secondary metabolites are diterpenoids, i.e., labdanes, abietanes, and ent-kauranes, as well as triterpenoids.

Analgesic and anti-inflammatory activity of amifostine, DRDE-07, and their analogs, in mice.

OBJECTIVES: To find out the analgesic and anti-inflammatory activity, if any, of Amifostine [S-2(3 amino propyl amino) ethyl phosphorothioate], DRDE-07 [S-2(3 amino ethyl amino) ethyl phenyl sulphide] and their analogs DRDE-30 and DRDE-35, the probable prophylactic agent for sulphur mustard (SM). MATERIALS AND METHODS: In order to find out the analgesic activities of the compounds two methods were employed, namely, acetic acid-induced writhing test and formalin-induced paw licking. The persistent pain model of formalin-induced hind paw licking was carried out to test the effect of the compounds on neurogenic pain or early phase (0 to 5 minutes) and on the peripheral pain or the late phase (15 to 30 minutes). To test the effect of the compound in acute inflammation, carrageenan-induced hind paw edema was carried out. This model of inflammation involves a variety of mediators of inflammation. RESULTS: DRDE-07 (81.7%) and DRDE-30 (79.4%) showed significant reduction in the acetic acid-induced writhing test. DRDE-07 (93.1%), DRDE-30 (82%), and DRDE-35 (61.3%) showed significant reduction in the second or late phase of formalin-induced paw licking. All the analogs (more than 60%) including amifostine (43.9%) showed significant reduction of paw edema in the carrageenan-induced paw edema in mice. CONCLUSION: The analgesic and anti-inflammatory activity of the antidotes were comparable with aspirin.

Ameliorative effect of DRDE 07 and its analogues on the systemic toxicity of sulphur mustard and nitrogen mustard in rabbit.

Despite extensive research efforts, there is no unanimous approval of any animal model to evaluate the toxicity of sulphur mustard [SM; bis (2-chloroethyl) sulphide] or nitrogen mustard [HN-3; tris-(2-chloroethyl) amine] and screening of various prophylactic and therapeutic agents against them. In this study, differential toxicity of mustard agents in higher animal model that is male rabbit was determined. Protective efficacy of DRDE 07 [S-2(2-aminoethylamino) ethyl phenyl sulphide] and its analogues were also evaluated against SM and HN-3 toxicity. Differential toxicity study of SM and HN-3 reveals that both the compounds were more toxic by percutaneous route as compared to subcutaneous route. Till date, there is no recommended drug to counteract SM induced toxicity or mortality in vivo. However, DRDE 07 (an amifostine analogue) and its analogues are found to be very effective protective agents against percutaneously exposed SM in rabbits. The present experiments also showed that SM does not cause skin injury alone but also can cause systemic toxicity as well. DRDE 07 and many of its analogues may prove as prototype compounds for the development of better prophylactic and therapeutic drugs to counter the toxicity of SM or HN-3. In conclusion, rodents and rabbits can be used for the screening of drugs against the blistering agents.

Ketoximes to N-substituted thioamides via PSCl3 mediated Beckmann rearrangement.

N-Substituted thioamides were accessed from ketoximes by utilising PSCl3 as a uniquely capable reagent to induce Beckmann rearrangement as well as to capture the intermediate nitrilium ion.

Expeditious microwave-assisted thionation with the system PSCl3/H2O/Et3N under solvent-free condition.

A novel thionation protocol for carbonyl compounds, with the system PSCl3/H2O/Et3N has been discovered. Clean, rapid, and efficient synthesis of a variety of thiocarbonyl compounds such as thioamides, thiolactams, thioketones, thioxanthones, and thioacridone can be achieved through this simple and convenient method under solventless condition with microwave irradiation.

Essential metal status, prooxidant/antioxidant effects of MiADMSA in male rats: age-related effects.

Thiols are known to act as protectants in the biological system for their involvement in a number of metabolic regulations. In this study, we investigated the effect of a new and potent thiol-chelating agent, monoisoamyl 2,3-dimercaptosuccinic acid (MiADMSA), an analog of meso 2,3-dimercaptosuccinic acid, to find out if it could act as a prooxidant (because of its lipophilic character) or antioxidant (because of thiol moiety) that could supplement its chelating properties in different age groups of male rats (young, adult, and old rats) and produce effective clinical recoveries in the treatment of metal intoxication. Animals were treated with 25, 50, and 100 mg/kg of MiADMSA, i.p, once daily for 1 week to assess the effect on the antioxidant system in major organs based on sensitive biochemical variables indicative of oxidative stress. Results suggested that MiADMSA administration increased the activity of d-aminolevulinic acid dehydratase in all the age groups and increased blood glutathione (GSH) levels in young rats. MiADMSA also potentiated the synthesis of metallothioneine in liver and kidneys and GSH levels in liver and brain. Apart from this it also significantly reduced the glutathione disulfide levels in tissues. However, administration of MiADMSA caused some concern over the copper loss. MiADMSA was found to be safe in rats of all ages.

Beneficial role of monoesters of meso-2,3-dimercaptosuccinic acid in the mobilization of lead and recovery of tissue oxidative injury in rats.

We investigated the therapeutic efficacy of meso-2,3-dimercaptosuccinic acid (DMSA) and two of its analogues, monomethyl dimercaptosuccinic acid (MmDMSA) and mono-cyclohexyl dimercaptosuccinic acid (MchDMSA) in reducing lead concentration in blood and soft tissues, and in recovering lead induced oxidative stress in rats. Male wistar rats were exposed to lead acetate in drinking water for 20 weeks, followed by 5 days of oral treatment with DMSA (100mg/kg, oral, once daily), MmDMSA or MchDMSA (50 and 100mg/kg). Biochemical variables indicative of oxidative stress along with lead, zinc and copper concentration were evaluated in blood and other soft tissues. Exposure to lead caused a significant decrease in blood delta-aminolevulinic acid dehydratase (ALAD) activity and glutathione (GSH) level. These changes were accompanied by inhibition of kidney ALAD and an increase in delta-aminolevulinic acid synthatase (ALAS) activity in liver and kidneys. Also seen were a pronounced depletion of brain GSH, glutathione peroxidase (GPx), glutathione-S-transferase (GST) and decreased superoxide dismutase (SOD) activity and an increase in thiobarbituric acid reactive substances (TBARS) and reactive oxygen species (ROS) levels. These biochemical changes were correlated with an increased uptake of lead in blood and soft tissues. Blood and kidneys zinc concentration decreased significantly following lead exposure while, copper concentration remained unchanged. No effect of chelation on hepatic zinc concentration was noted, only liver copper concentration showed significant depletion on treatment with DMSA and MmDMSA (100mg/kg). Treatment with DMSA, MmDMSA and MchDMSA provided significant recovery in altered biochemical variables and brain DNA damage besides significant depletion of tissue lead burden. Among the chelating agents used, MchDMSA and MmDMSA provided better recovery in altered biochemical variables and depletion of lead concentration in tissues compared to DMSA. The above results suggest DMSA monoesters to be a better treatment option than DMSA in eliciting recovery to the altered biochemical variables and in the depletion of body lead burden.

Arsenic antagonism studies with monoisoamyl DMSA and zinc in male mice.

Administration of zinc either alone or in combination with monoisoamyl dimercaptosuccinic acid (DMSA) during and post-arsenic exposure was investigated in male mice. The animals were administered 2mgkg(-1) arsenic as sodium arsenite, intraperitoneally, once daily for 5 days either alone or in combination with 10mgkg(-1), zinc (as zinc acetate, orally), 50mgkg(-1) monoisoamyl dimercaptosuccinic acid (MiADMSA) given orally (p.o.), 2h after arsenic administration. Another group of arsenic treated animals was given both zinc (10mgkg(-1)) and MiADMSA (50mgkg(-1), p.o.). Animals were sacrificed 24h after the last dose. In another set of experimentation, arsenic pre-exposed mice (2mgkg(-1), i.p. for 5 days) were treated with saline, zinc, MiADMSA or zinc plus MiADMSA for next 3 days and sacrificed thereafter. Exposure to arsenic led to a significant inhibition of blood δ-aminolevulinic acid dehydratase (ALAD), depletion of glutathione (GSH) level and marginal elevations of zinc protoporphyrin (ZPP). Arsenic exposure caused a significant decrease in hepatic and renal GSH level and an increase in liver oxidized glutathione (GSSG) and liver and kidney thiobarbituric acid reactive substance (TBARS) levels. Concomitant administration of zinc with arsenic provided significant protection to blood ALAD activity while, GSH and ZPP levels remained unaltered. Co-administration of MiADMSA with arsenic significantly prevented accumulation of arsenic in blood, liver and kidney while, zinc had no effect on tissue arsenic concentration. Combined administration of zinc and MiADMSA had no major additional beneficial effects over their individual effects. Interestingly, post-arsenic exposure treatment with MiADMSA provided significant recovery in blood ALAD activity while, zinc supplementation alone had no effect. The best results however, were obtained when MiADMSA was administered along-with zinc. Most of the biochemical variables indicative of hepatic oxidative stress responded favorably to MiADMSA treatment while, zinc administration had no effect. Administration of MiADMSA significantly depleted arsenic concentration from the soft tissues while, combined zinc and MiADMSA had no additional beneficial effect over the individual effect of MiADMSA. The results thus lead us to conclude that in order to achieve best effects of chelation therapy, co-administration of zinc with chelator might be preferred. However, detailed experimental studies with variable doses and after chronic arsenic exposure are required.

Novel S-substituted aminoalkylamino ethanethiols as potential antidotes against sulfur mustard toxicity.

Sulfur mustard (SM) is a highly toxic chemical warfare agent. A satisfactory treatment regimen is not yet available for this toxicant. In a search for an effective antidote against SM, a series of novel S-2(omega-aminoalkylamino)ethyl alkyl/aryl thioethers [H(2)N(CH(2))(n)()NHCH(2)CH(2)SR], where R = alky, alicyclic, aryl, and heterocyclic substituents, have been designed and synthesized as candidate antidotes against SM toxicity. These compounds were screened for their protective efficacy through the oral route against dermally applied sulfur mustard in female mice measured on the basis of percent survival following percutaneous administration of SM. A number of compounds demonstrated significant protection.