Prati Pal Singh

Synthesis, antiprotozoal, antimicrobial, β-hematin inhibition, cytotoxicity and methemoglobin (MetHb) formation activities of bis(8-aminoquinolines).

In continuing our search of potent antimalarials based on 8-aminoquinoline structural framework, three series of novel bis(8-aminoquinolines) using convenient one to four steps synthetic procedures were synthesized. The bisquinolines were evaluated for in vitro antimalarial (Plasmodiumfalciparum), antileishmanial (Leishmaniadonovani), antimicrobial (a panel of pathogenic bacteria and fungi), cytotoxicity, β-hematin inhibitory and methemoglobin (MetHb) formation activities. Several compounds exhibited superior antimalarial activities compared to parent drug primaquine. Selected compounds (44, 61 and 79) when tested for in vivo blood-schizontocidal antimalarial activity (Plasmodiumberghei) displayed potent blood-schizontocial activities. The bisquinolines showed negligible MetHb formation (0.2-1.2%) underlining their potential in the treatment of glucose-6-phosphate dehydrogenase deficient patients. The bisquinoline analogues (36, 73 and 79) also exhibited promising in vitro antileishmanial activity, and antimicrobial activities (43, 44 and 76) against a panel of pathogenic bacteria and fungi. The results of this study provide evidence that bis(8-aminoquinolines), like their bis(4-aminoquinolines) and artemisinin dimers counterparts, are a promising class of antimalarial agents.

Models of latent tuberculosis: their salient features, limitations, and development.

Latent tuberculosis is a subclinical condition caused by Mycobacterium tuberculosis, which affects about one-third of the population across the world. To abridge the chemotherapy of tuberculosis, it is necessary to have active drugs against latent form of M. tuberculosis. Therefore, it is imperative to devise in vitro and models of latent tuberculosis to explore potential drugs. In vitro models such as hypoxia, nutrient starvation, and multiple stresses are based on adverse conditions encountered by bacilli in granuloma. Bacilli experience oxygen depletion condition in hypoxia model, whereas the nutrient starvation model is based on deprivation of total nutrients from a culture medium. In the multiple stress model dormancy is induced by more than one type of stress. In silico mathematical models have also been developed to predict the interactions of bacilli with the host immune system and to propose structures for potential anti tuberculosis compounds. Besides these in vitro and in silico models, there are a number of in vivo animal models like mouse, guinea pig, rabbit, etc. Although they simulate human latent tuberculosis up to a certain extent but do not truly replicate human infection. All these models have their inherent merits and demerits. However, there is no perfect model for latent tuberculosis. Therefore, it is imperative to upgrade and refine existing models or develop a new model. However, battery of models will always be a better alternative to any single model as they will complement each other by overcoming their limitations.

Immunomodulation by morphine in Plasmodium berghei-infected mice.

The effect of morphine on immunomodulation and host defense have been investigated during Plasmodium berghei infection in BALB/c mice. A single low (5.0 mg/kg) subcutaneous dose of morphine strongly suppressed (sometimes completely eliminated) the parasitaemia, whereas a high dose (80.0 mg/kg) exerted mild potentiating effect. Mice treated with the low dose showed a significant (p < 0.05) increase in the total number of circulating leukocytes, the number (pool-size) of peritoneal macrophages, and the phagocytic activity of peritoneal macrophages, in vitro. Conversely, in mice treated with the high dose, all these parameters were diminished. Silica (3.0 mg/mouse), administered intravenously, abrogated the morphine-induced protective effects but did not affect its potentiation of the infection. Naloxone pretreatment (4.0 mg/kg) completely blocked the protective effects of morphine, suggesting the mediation via naloxone-sensitive opiate-receptors; paradoxically, it did not affect the potentiating effects. These observations indicate that morphine exerted a dose-dependent, biphasic effect on the course of P. berghei infection in mice, apparently by modulating the macrophage-mediated protective mechanisms.

Neuroimmunomodulatory Effects of Morphine in Leishmania donovani-Infected Hamsters

Objective: The effect of morphine on host defense during Leishmania donovani infection in golden hamsters was studied. Methods: Hamsters were intracardially infected with L. donovani amastigotes and then monitored by spleen touch print microscopic examination. Morphine and naloxone were administered subcutaneously and intraperitoneally, respectively. Leukocytes were counted by a hemocytometer, and ex vivo phagocytosis was determined by the examination of stained adherent macrophages. Results: Low doses of morphine, 1.75 and 2.5 mg/kg × 2, administered subcutaneously on day 0 and day 15 significantly (p < 0.05) suppressed the infection, whereas high doses (20.0 and 50.0 mg/kg × 2) exacerbated the infection. On day 30, hamsters treated with low doses of morphine showed a significant (p < 0.05) increase in the number of circulating leukocytes and the pool size and phagocytic activity of peritoneal macrophages ex vivo; in hamsters treated with high doses, all these parameters appeared to be diminished. The bone marrow of morphine-treated hamsters showed a fall in total cellularity and no change in the number of monocytes; however, in those treated with low doses, the infection was completely eliminated by day 30, and paradoxically, a significant (p < 0.05) potentiation of infection was observed in hamsters treated with high doses. The spleens of hamsters treated with both low and high doses of morphine showed a significant (p < 0.05) decrease and increase in weight, respectively; treatment with low doses also caused an almost 2-fold increase in the percentage of monocytes. Morphine apparently exerted its protective effects via naloxone-sensitive opioid receptors; naloxone pretreatment did not affect the potentiation of infection. Conclusion: Conditional doses of morphine apparently biphasically modulated the course of L. donovani infection in hamsters, at least in part through macrophage-mediated mechanisms.

Macromolecular prodrugs. XII. Primaquine conjugates: synthesis and preliminary antimalarial evaluation.

Macromolecular prodrugs. XII. Primaquine conjugates: Synthesis and preliminary antimalarial evaluation New primaquine conjugates 5-7 with glucosamine and two polymers of polyaspartamide type, poly[α,β-(N-2-hydroxyethyl-DL-aspartamide)] (PHEA) and poly[α,β-(N-3-hydroxypropyl-DL-aspartamide)] (PHPA), were synthesized, characterized and screened for their antimalarial activity. The conjugates differed in the type of covalent bonding, length of the spacer between the polymeric carrier and drug, molecular mass and drug-loading. Blood-schizontocidal activity of the prepared conjugates was tested against Plasmodium berghei infection in Swiss mice. Polymeric conjugates showed better antimalarial activity than the glucosamine conjugate. Makromolekulski prolijekovi. XII. Konjugati primakina: Sinteza i preliminarno ispitivanje antimalarijskog djelovanja U radu je opisana sinteza, karakterizacija i ispitivanje antimalarijskog djelovanja novih konjugata primakina 5-7 s glukozaminom i dva polimera poliaspartamidnog tipa, poli[α,β-(N-2-hidroksietil-DL-aspartamidom)] (PHEA) i poli[α,β-(N-3-hidroksipropil-DL-aspartamidom)] (PHPA). Konjugati su se razlikovali po vrsti kovalentne veze, duljini razmaknice između polimernog nosača i ljekovite tvari, molekulskoj masi i količini vezanog lijeka. Šizontocidno djelovanje pripravljenih konjugata ispitano je na miševima inficiranima parazitom Plasmodium berghei. Polimerni konjugati pokazali su jače antimalarijsko djelovanje nego konjugat s glukozaminom.

Antimalarial activities of ring-substituted bioimidazoles

We report in vitro antimalarial activities against chloroquine sensitive and resistant Plasmodium falciparum strains, and in vivo activities against Plasmodium berghei in mice for four series of ring-substituted-L-histidines and histamines.

Lymphokines production by concanavalin A-stimulated mouse splenocytes: modulation by Met-enkephalin and a related peptide.

Methionine-enkephalin (ME) and its synthetic congener Tyr-D-Ala-Gly-Me-Phe-Gly-NH.C3H7-iso (82/205), in a concentration-dependent biphasic manner modulated the concanavalin A (Con A)-stimulated phagocytosis-promoting (PP)-activity elaboration in the culture supernatants of mouse splenocytes in vitro. Both these peptides at 1 x 10(-5) and 1 x 10(-6) M inhibited the production of PP activity; conversely, at 1 x 10(-7)-1 x 10(-9) M they augmented it. Peptide 82/205 was nearly 1.2-fold more inhibitory and approximately 1.8-fold more potent in augmenting the PP activity elaboration. The PP activity appeared to be due to lymphokines (LK) gamma interferon and interleukin-4 as the neutralizing concentrations of monoclonal antibodies against these LK significantly (p < 0.05) inhibited it. Cycloheximide (50.0 micrograms/ml) completely inhibited the production of LK indicating their de novo synthesis. The peptides appeared to exert their inhibitory and augmenting effects via delta- and mu-opioid receptors, respectively, as pretreatment of splenocytes with 100-fold higher (1 x 10(-3) M) concentration of naloxone was required to block their inhibitory effect; the augmenting effect was blocked by 1 x 10(-5) M only. None of the peptides or naloxone could directly stimulate the splenocytes for PP-LK elaboration.

Effects of morphine during Mycobacterium tuberculosis H37Rv infection in mice

The effects of opiates in various infections are well known; however, very little is known about tuberculosis infection. Therefore, in the present study, we report for the first time, the effects of morphine during murine tuberculosis. Mice were infected intravenously with Mycobacterium tuberculosis H37Rv, administered morphine (0.1-100 mg/kg subcutaneously on day 0 and day +15) and sacrificed on day +30 for CFU enumeration in lungs and spleen. Morphine exerted maximum suppression of infection at 5 mg/kg, and sometimes completes elimination of infection; naloxone, silica and aminoguanidine blocked the protective effect of morphine. In vitro, morphine lacked direct antimycobacterial activity up to 1x10(-4) M concentration, as assessed by radiometric BACTEC method. In macrophage model of infection, morphine showed maximal killing at 1x10(-7) M concentration, the activity was blocked by naloxone and aminoguanidine. These observations suggest that morphine exerts a dose-dependent effect in murine tuberculosis, the protective effect being naloxone-reversible and may involve macrophage-mediated protective mechanisms. These results may be helpful in developing new opioid-like chemical entities against tuberculosis infection.

Extended side chain analogues of 8-aminoquinolines: Synthesis and evaluation of antiprotozoal, antimicrobial, β-hematin inhibition, and cytotoxic activities

We report the synthesis of double, triple and quadruple extended side chain analogues of the antimalarial drug primaquine and some other 8-aminoquinolines. The synthesized analogues have exhibited potent antimalarial activities in vitro against both the drug-sensitive D6 strain (IC50 = 0.19–0.92 μg mL−1) and the drug-resistant W2 strain (IC50 = 0.12–0.82 μg mL−1) of P. falciparum and in vivo against drug-sensitive P. berghei infected mice (100% curative at 25 mg kg−1 day−1, and resulted in either 4/6 or 5/6 cures at 10 mg kg−1 day−1) for the most promising structures. These analogues were also found to be free of cytotoxic effects at the highest test concentration of 23.8 μg mL−1 in a panel consisting of six cell lines. The promising 8-aminoquinolines inhibited β-hematin (IC50 = 9.6–20.8 μM) in vitro underlining the disruption of the heme catabolism pathway in the malaria parasite as their potential biochemical pathway for antimalarial action. The analogues also displayed potent antileishmanial activities in vitro against L. donovani promastigotes (IC50 = 1.6–32 μg mL−1; IC90 = 4–40 μg mL−1) and moderate in vitro antimicrobial activities against a panel of bacteria and fungi.

Evaluation of BACTEC 460 TB system for rapid in vitro screening of drugs against latent state Mycobacterium tuberculosis H37Rv under hypoxia conditions.

Mycobacterium tuberculosis exhibits latent state due to its ability to survive for extended periods under oxygen depletion conditions. The conventional plating method employed for in vitro screening of antitubercular agents against latent state M. tuberculosis is laborious and time consuming exercise. Towards this end, BACTEC 460 TB system of antitubercular screening was evaluated for testing efficacy of antimicrobial agents in hypoxia induced model of latent tuberculosis, in vitro. In this study, drugs like isoniazid, metronidazole and rifampin were tested at concentrations- 2, 10 and 50 microg/ml for their antilatency activity by using BACTEC and conventional methods. Results obtained from both the methods were comparable (P>0.05) and a good correlation was observed between colony forming units and growth index values. Further, time to determine mycobacterial growth was significantly reduced (P<or=0.001) in BACTEC method (4-7 days) as compared to plating method (26-30 days). BACTEC method was found to be faster, cost effective and more sensitive as compared to plating method. Being rapid, reliable and reproducible, this method can be a promising alternative to conventional plating method for screening of potential antitubercular agents in in vitro models of latent tuberculosis.