Adenosine (Rys. 1) is a purine nucleoside playing an important role in human body. It is involved in key pathways such as purinergic nucleic acid base synthesis, amino acid metabolism and modulation of cellular metabolic status [1,2]. Adenosine acts through the four types of adenosine receptors: A1, A2A, A2B and A3 belonging to the G protein-coupled receptor family [3]. In physiological conditions this nucleoside is present in a micromolar range [5]. However, when metabolic stress occurs extracellular level of adenosine raises revealing its protective properties. Depending on an activated receptor subtype, adenosine demonstrates cardioprotective and neuroprotective activity during hypoxia or ischemia, it stimulates the immunological system [6, 7]. Besides many potential applications, adenosine is used mainly for the treatment of paroxysmal supraventricular tachycardia. Limitations are linked to a very short blood half-time and no receptor specificity [8]. This review is focused on novel literature data about synthesis of adenosine analogues with interesting biological activities. In order to influence adenosine receptor selectivity and pharmacokinetic properties a nucleoside structure can be modified in purine [14, 15, 17, 22, 26, 27, 35] or sugar ring [29, 32]. New interesting compounds are also synthesized by cyclisation of adenosine [36]. Modification of adenosine structure allowed obtaining compounds with targeted action: antiarrhythmic [11, 12], antinociceptive [9], antilipolytic [13], antiviral [29] or anticancer [35].
Transplantology is getting more and more important in medicine. Development of surgical techniques and immunosuppressive treatment enabled to establish successful transplantations with various organs and tissues. However, allografts are recognized as foreign tissues and stimulate rejection, i.e. a strong immunological response which, if not stopped, results in complete destruction of the transplanted tissue. In order to prevent the rejection patients have to be treated with immunosuppressive drugs after transplantation. Unfortunately, such a damping of immune system poses a risk of cancer or severe infections. The treatment itself is also toxic, notably when applied in a long-term maintenance therapy. Currently, adverse effects of immunosuppressive drugs are recognized as the ones to be involved significantly in chronic rejection and limitation of long survival of grafted tissues. Whereas prevention of acute rejection is mostly successful, there is still no efficient treatment for chronic graft rejection. Reduction of a dose of immunosuppressive drugs or an invention of new active substances is considered the most promising solution. Nowadays, immunosuppressive drugs can be divided into the three main groups: agents which inhibit production of cytokines taking part in cells’ activation (glicocorticosteroids, calcineurine inhibitors, mTOR inhibitors), antiproliferative compounds (azathiopirin, mycophenolate mofetil, mycophenolic acid sodium salt), and antibodies. In this article we present new investigations towards immunosuppressive drugs, their structures and synthetic methods.
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Dryness of skin, lack of elasticity, laxity, pigmentation, fine lines and wrinkles are the effects of skin aging [1-11]. Researchers and technologists try to fight with these problems. Researchers are still looking for new biologically active substances, which would improve the appearance of skin and technologists are creating a special composition of skin care products. The following publication is a review of currently used biologically active peptides applied in cosmetic products for skin aging (Tab. 1). There are three main groups of peptides with different mechanism of action: neurotransmitter-affecting peptides, carrier peptides and signal peptides [16]. Neuropeptides are peptides, which mimic the action of botulinum neurotoxin (the structure of active toxin is presented in the Fig. 1). These peptides decrease muscle contraction through interactions at the neuromuscular junction [13, 16, 18]. Carrier peptides take part in delivering a cofactor required for wound healing and enzymatic processes into the dermis [16, 28, 29]. Instead, signal peptides are able to stimulate collagen synthesis and cause growth of human dermal skin fibroblasts [16, 18].
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Nowadays the battle with tumor diseases and bacterial infections is an important test for the scientists. Therefore a lot of attention is focused on obtaining novel, more effective, selectively acting and less toxic drugs. Because of that reason, compounds with wide range of action like acridines/acridones are of great interest for scientists. This study contains synthesis and biological activity of acridine/acridone derivatives which are used in medicine as an antitumor agents and those which are at clinical trials. Among the synthetic methods there are modifications of Ullmann synthesis [2, 3, 9]. Lately Belmont et al. [11, 12] described a new methodology for the synthesis of 1,3,7-trisubstituted acridines, 1-amino-acridine and tetrahydro-cyclopenta[c]acridine-2,5-diones (Scheme 5). The starting materials are commercially available quinolines. Among acridine/acridone group there are compounds with diffe-rent molecular targets, e.g. topoisomerase inhibitors [1, 19-33], telomerase inhibitors [1, 34], protein-kinases inhibitors [36-40], P-glycoproteine inhibitors [56, 57, 76], conjugates with metals [1] and hypoxia-selective derivatives [2, 41-55]. This paper also describes compounds useful in Alzheimer disease [5, 16], antibacterial [2, 5], antiparasites [4, 5] and antiviral infections [5, 13, 14].
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Mycophenolic acid (MPA) 1 is one of the most substituted phtalides. Its chemical structure incorporates a highly functionalized, hexasubstituted benzene ring [3, 4]. This compound is one of the oldest known antibiotics [1, 2]. MPA is the most potent uncompetitive inhibitor of inosine 5'-monophosphate dehydrogenase (IMPDH). This enzyme catalyzes a rate - limiting step in the de novo biosynthesis of purine nucleotides [13]. Mycophenolic acid as an IMPDH inhibitor functions as antifungal, antiviral, antibacterial and immunosupressive agent [5-11]. Its derivatives: mycophenolate mofetil (MMF; CellCept(r), Roche AG) and mycophenolate sodium (MPS; Myfortic(r), Novartis Pharma AG) are used in combination with corticosteroids and calcineurin inhibitors (cyclospo-rine A or tacrolimus) for the treatment and prophylaxis of organ rejection in solid organ transplants. The metabolic lability of mycophenolic acid and severe side effects in clinical treatment are the main reasons for the development of new synthetic pathways of its derivatives [14]. This paper reviews the most important approaches in mycophenolic acid synthesis and its derivatives and displays structure-reactivity relationships of these compounds. Synthesis of mycophenolic acid as one of the highest substituted phtalide is described [23-35]. The most common synthetic approach in preparation of highly substituted benzenes is by using benzene ring constructions with five or six required substituents [28-30]. First of these methods [25] is based on construction of the pentasubstituted resorcinol derivative via thermal addition of the alkynyl ether to the cyclobutenone. The synthetic strategy of the second method [28, 30] is depicted in Scheme 4 and the key step of this approach involves reaction between 16 and 17. Alternative approach to total mycophenolic acid synthesis is preparation of its intermediates [31]. Mycophenolic acid derivatives were divided into five groups, according to their chemical structure. For each of them synthetic pathway was shown and structure-biological activity relationships were described [40]. It has been found that replacement of the mycophenolic acid lactone ring with other cyclic groups resulted in loss of potency. A phenolic hydroxyl group and the aromatic methyl substituent were found to be essential for high activity. Replacement of the methoxy group with ethyl, vinyl or methyl resulted in compounds with higher activity than mycophenolic acid itself [41]. It has also been discovered that substitution with small alkyl groups in the ? position to the carboxylic group results in enhanced potency [46]. Furthermore monocyclic and indol derivatives were obtained and the carboxyamide derivative was selected for screening against prostate cancer [54]. Also new monocyclic analogues were obtained but they did not show any anticancer activity [55]. There have been synthesized several analogues of mycophenolic adenine dinucleotide [50-52] or mycophenolic adenine methylene-bis(sulfonamide)s [53] which showed inhibitory activity against IMPDH. Recently, a series of novel IMPDH inhibitors based on a methoxy-(5-oxazolyl)-phenyl (MOP) moiety have been designed [56].
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Bacterial cell wall peptidoglycan (PGN) is a potent immunostimulator and immune adjuvant. Numerous studies reported on immunoactivities of bacterial PGN, most of which have been reproduced by a chemically synthesized low-molecular PGN fragment, muramyldipeptide (MDP) 1 (Fig. 1) [2, 4, 5, 7, 8, 10, 13]. Another type of PGN fragment, desmuramylpeptides (DMPs), has also been chemically synthesized to mimic PGN containing meso-DAP, and the DMPs exerted similar bioactivities to MDP. In 1984 [18] reported that y-D-Glu-meso-DAP was the minimum structural unit capable of eliciting bioactivities induced by DMPs. Recently demonstrated that intracellular protein carrying a nucleotidebinding oligomerization domain (NOD), NOD2 an intracellular receptor for MDP and DMPs containing DAP was recognized by another NOD protein, NOD1 [8, 33]. Replacement of the N-acetylmuramyl moiety with various acyl groups thus represents an important approach to the design and synthesis of new immunologically active MDP analogues – desmuramylpeptides, e.g. FK-156 9, pimelautide 11 (Fig. 2), 7-(oxoacyl)-L-alanyl-D-isoglutamines, carbocyclic MDP analogues (Fig. 13) [3, 13] in which a more lipophilic cyclohexane ring is present instead of the polyhydroxy pyranose ring of D-glucosamine, and the adamantyl-substituted MDP analogue LK-415 (Fig. 8) [54]. The FK-156 isolated from Streptomyces olivaceogriseus [21, 22] and its synthetic analogue of FK-565 10 (Fig. 2) have been reported to be a potent stimulant of antibody production and free of pyrogenicity. These compounds with close structural resemblance to bacterial cell wall peptidoglycan peptides, exhibit very interesting biological activities. Both FK-156 10 and FK-565 11 (Fig. 3) enhance host defense ability against microbial infections, exhibit strong antiviral activity and remarkable antitumor potency [2, 13, 14, 18]. Also other acyl-DMPs were obtained and their activity described (Table 1). The most promising DMPs analogues were series of phthalimido-DMPs 46-53 (Fig. 7). In these compounds N-acetylmuramic acid residue was replaced by various N-phthaloylated amino acids [42–49] or phthalimido substituted aminoethoxyacetic acid to give immunologically active acyclic MDP analogues like LK-423 46 (LK-413 47, LK-511 48, LK-512 49, LK-508 50) (Fig. 7) [42, 47–49]. LK-423 has been selected for further studies to develop an anti-inflammatory pharmaceutical agent. In 2001 Gobec et al. [54] reported the synthesis of new adamantyl-DMPs LK-415 55 and LK-517 56 (Fig. 8) with 1-adamantyl-carboxamido moiety replacing N-acetylglucosamine fragment in MDP. Their efficiency in modulating the production of cytokines IL-12, TNFá, IFNă, IL-4, and IL-10 was measured in vitro in ionomycin and PMA activated cultures of PBMC, co-incubated with the analogues tested. The results were compared with the activity of MDP. All substances were strong regulators of IL-12 synthesis and IFNă synthesis as well. Introduction of diethyl phosphonate moiety into LK-517 was of great importance for augmented T-cell cytokine production. Dzierzbicka et al. [55] described synthesis of three analogues of DMPs 57a,b, 58 modified with an amino-acridine/acridone residue. The screening data indicate that the analogues 57a,b and 58 (Fig. 9) exhibit low cytotoxic activity. Uehara et al. [59] reported MDP and DAP-containing desmuramylpeptides in combination with chemically synthesized Toll-like receptor agonists (Fig. 10) synergistically induced production of IL-8 in a NOD2- and NOD1-dependent manner, respectively, in human monocytic cells in culture. In 2008 Kawasaki et al. [33] designed synthesis of DAP containing PGN fragments and tracheal cytotoxin (TCT) (Fig. 5) and investigated their biological activity. Recently, N-acetylglucosamine- 1,6-anhydro-N-acetylmuramylpentapeptide (Fig. 12) and evaluation of its turnover by AmpD from Escherichia coli has been reported [61]. The synergism of MDP and DMPs with other chemotherapeutics is also promising in the therapy of many infectious and anticancer diseases. This paper reviews the most important approaches to desmuramylpeptides (DMPs), their derivatives and displays structure-reactivity relationships of these compounds.
Novel 3'-N-(tuftsin or retro-tuftsin)-amino-combretastatin conjugates have been synthesized as potential anticancer compounds. We hope that the conjugation of immunomodulators like tuftsin derivatives with amino-combretastatin A-4 would improve the therapeutical properties of combretastatin A-4.
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This article described synthesis and biological activity of combretastatin A-4 (CA-4) and its analogues. Combretastatin A-4 (CA-4), a natural product isolated from the South African bush willow tree Combretum caffrum, binds to the colchicine binding site and inhibits the polymerization of microtubules. CA-4 exhibits potent cytotoxicity against a variety of human cancer cell lines including multidrug-resistant (MDR) cell lines [5-7]. The studies of structure-activity relationship (SAR) of CA-4 1 (Fig. 1) showed that 3,4,5-trimethoxy substitution on the A ring and the 4'-methoxy group on the B ring and the cis-olefin configuration are crucial for potent cytotoxicity, while the 3'-hydroxy group is optional [5-7]. A many of CA-4 analogues were synthesized where the double bond have been replaced by introduction of nonheterocyclic groups (e.g. ethers, olefins, ketones, sulfonates, sulfonamides, amide derivatives, amine, cyclopentanes) or heterocyclic groups containing five-membered rings (e.g. pyrazoles, thiazoles, triazoles, tetrazoles, oxazoles, furans, dioxolanes, thiophenes) and indoles [5, 7, 41, 56] (Fig. 9-12). Up to now, many CA-4 analogues and their biological activity have been extensively studied and three derivatives are currently in clinical trials: a water-soluble disodium phosphate derivative of CA-4 (CA-4P) 11c (Fig. 3); Oxi-4503, a water-soluble combretastatin A-1 (CA-1diP) 4a (Fig. 1); and AC7700 59e (Scheme 7) an aminocombretastatin prodrug developed in Japan in 1998 [5-10, 34].
Synthesis of new conjugates of muramyl dipeptide (MDP) and nor-muramyl dipeptide (nor-MDP) with retro-tuftsin derivatives is presented. The corresponding protected retro-tuftsin derivatives were also synthesized. The synthesis of a greater number of conjugates will enable structure-activity relation ship studies.
This article reviews synthesis of 2,6-diaminopimelic acid, an important amino acid biosynthesized by bacteria and higher plants. Anumber of peptidoglycan fragments containing the diaminopimelic acid (DAP) residue exhibit antitumor and/or immunostimulant activity. DAP is a versatile building block of many natural and synthetic compounds with a number of potentialmedicinal applications. The synthesis of immunostimulants FK-156 and FK-565 is also presented.
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This article concerns methods of synthesis and biological activity of peptidyl anthraquinones. The group of these components have a big influence on a present medicine. Among components, that were presented in this paper, many number of them show antine-oplastic activity. These peptidyl anthraquinones demonstrate special mechanism of action, stabilizing cleavable complex consisting of topoisomerase, convalently attached to DNA and stop correct replication and transcription. Using of topoisomerases inhibitors in therapy of cancer have promising results. Anthraquinones analogues with peptidyl chain including KCR motif very important in AP-1 protein binding to DNA. This group of compounds can be used as potential transcription factor inhibitors, because they show DNA intercalative binding and recognize specific sequences of DNA binding domains. Synthetic GnRH analogues including rings of anthraquinone present interesting drugs using in reproductive diseases, for instance prostate, ovarian and breast cancer. The mechanism of action of GnRH analogues count on binding to GnRH receptors, which we can find in cells of reproductive organs. MMP-9 activity can be exploited to activate prodrugs and be a target the tumour cells in BM. Cleavage of the prodrugs with MMP-9 resulsts in the liberation of the active form of drug which can be an inhibitor of topoisomerase. Tripeptide derivatives of anthraquinone show the activity as histochemical reagents for detection of TTP I activity. Peptidyl anthraquinones can be also used for protein purification and labeled peptides as biochemical probes for DNA detection.
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Microtubule targeting drugs being in the late preclinical or early clinical development are described in this article. New semisynthetic paclitaxel analogues, natural compounds of diverse structure such as epothilones, combretastatins, colchino-ids or dolastatins and synthetic compounds of low molecular weight such as heterocombretastatins, sulfonamides, phenstatins, indoles and quinolones belong to this category of anticancer medicines. Microtubules are hollow tubes consisting of L- and B-tubulin heterodimer proteins that polymerize parallelly to a cylindrical axis. The targeting of microtubules is an important mechanism in cancer chemotherapy for such drugs as the vinca alkaloids (vincristine (1), vin-blastine (la)), podophilotoxin, their semisynthetic analogues and taxanes (paclitaxel (2), docetaxel (3)) known of their great usefulness in the anticancer therapy. These agents may stabilize microtubules, as the taxanes do, or destabilize them, as it is in the case of the vinca alkaloids. Today, more than 30 compounds targeting tubulin, either stabilizing or destabilizing microtubule dynamics, are in late preclinical or early clinical development. Despite of more than 30 years after the discovery of paclitaxel microtubule inhibitors they are still of the topic of interest of all over the world. In the end of 1990 and up to 2005 year survey articles on the microtubule inhibitors were published .We expect now that the paper which presents last results study may be useful.
W pracy opisano syntezę nowych połączeń muramylodipeptydu (MDP), nor-muramylodipeptydu (nor-MDP) oraz desmuramylopeptydu z aminowymi pochodnymi 1-nitroakrydyny/4-nitroakrydonu, N-(hydroksyalkilo)-4-karboksyamido-akrydyny/9-akrydonu, batracyliną i jej pochodnymi oraz tuftsyną i tuftsyną o odwróconej sekwencji. Pochodne MDP i nor-MDP (estry ten-butylowe 4,6-0-benzylideno-1-O-benzylo-N-acetylo-muramylo-(lub nor-muramylopeptydów) - substraty do syntezy koniugatów - otrzymywano metodami literaturowymi z pewnymi modyfikacjami, np. usuwanie z funkcji karboksylowej osłony ten-butylowej za pomocą 90% TFA umożliwiło w temperaturze pokojowej równocześnie odszczepienie grupy benzylidenowej z C6 i C4. Tak otrzymanymi pochodnymi 1-O-benzylo-MDP i 1-O-benzylo-nor-MDP, bez oczyszczania, acylowano aminowe czy hydroksylowe pochodne akrydyny, akrydonu, batracyliny oraz tuftsyny. Pośród prezentowanych 56 nowych koniugatów znajdują się 24 połączenia otrzymane przez utworzenie wiązania amidowego pomiędzy grupą karboksylową izoglutaminy a aminowymi pochodnymi 1-nitroakrydyny/4-nitroakrydonu. Reakcje prowadzono w bezwodnym DMF wg trzech procedur: DPP Ą mieszanych bezwodników i EEDQ. Kolejne 3 koniugaty to połączenia desmuramylopeptydów z pochodnymi amino-1-nitroakrydyny lub 9-akrydonu. Dwa z nich zawierały kwas mezodiaminopimelinowy, a w trzecim L-alaninę zastąpiono L-waliną zaś kwas mezo-diaminopimelinowy L-lizyną. Kwas mezo-diaminopimelinowy potrzebny do syntezy desmuramylopeptydów otrzymano z całkowicie chronionego kwasu pimelinowego, wykorzystując a-chymotrypsynę do selektywnej hydrolizy grupy estrowej przy centrum S. Pochodne amino-1-nitroakrydyny/4-nitroakrydonu otrzymano znanymi sposobami w wyniku kondensacji UlImana kwasu o-chlorobenzoesowego z m-nitroaniliną lub 3-amino-4-nitrochlorobenzenem w obecności pyłu miedzi jako katalizatora, cyklizacji i łączeniu z odpowiednimi alkiloaminami. Dziewięć nowych koniugatów MDP lub nor-MDP zsyntetyzowano, modyfikując ich C-koniec przez utworzenie wiązania estrowego z pochodnymi N-(hydroksyalkilo)-4-karboksyamido-akrydyna/9akrydonu. Do syntezy wykorzystano HBTU lub EDCI jako odczynniki kondensujące. Nadmiar jednego z substratów ułatwił otrzymywanie produktu z umiarkowaną wydajnością. Kwas 9-akrydono-4karboksylowy i kwas akrydyno-4-karboksylowy, substraty do otrzymania tego typu koniugatów zsyntetyzowano wg znanych procedur i łączono je z odpowiednimi aminoalkoholami, stosując DPP A lub CDI, otrzymując pochodne N-(hydroksyalkilo)-4-karboksyamido-akrydyny/9-akrydonu. Podjęto również próby zastosowania reakcji Mitsunobu do tworzenia wiązania estrowego pomiędzy MDP lub nor-MDP a pochodnymi N-(hydroksyalkilo)-4-karboksyamido-9-akrydonu. Próby zakończyły się niepowodzeniem pomimo wielu doświadczeń w zmienianych warunkach reakcji, stosowania różnych nadmiarów molowych substratów i zmiennej kolejności dozowania reagentów. Koniugaty zawierające pochodne akrydyny/akrydonu przekazano do badań aktywności cytotoksycznej w Narodowym Instytucie Rakowym (NCI, Bethesda, USA). Siedem połączeń MDP lub norMDP z pochodnymi amino-I-nitroakrydyny wykazało wysoką aktywność cytotoksyczną i zostało zakwalifikowane do badań in vivo w teście Hollow Fiber. Najbardziej aktywne koniugaty działają cytotoksycznie na komórki nowotworu piersi, nerki, jajnika, czerniaka, okrężnicy, płuc i prostaty. Na podstawie testów in vivo Hollow Fiber trzy z nich zostały wyselekcjonowane, przez Komitet Oceny Biologicznej NCI, do bardziej zaawansowanych testów in vivo na ksenoprzeszczepach (xenograft assays). Analogi desmuramylopeptydów z pochodnymi amino-I-nitroakrydyn oraz analogi MDP lub nor-MDP zawierające pochodne N-(hydroksyalkilo)-4-karboksyamido-akrydyny lub akrydonu okazały się nieaktywne. Zsyntetyzowano 8 koniugatów MDP lub nor-MOP z batracyliną lub jej pochodnymi. Reakcję acylowania prowadzono w bezwodnym DMF według dwóch procedur: DPPA i EEDQ. Opracowano udoskonaloną metodę syntezy batracyliny. Jako substrat wykorzystano komercyjną p-fenylenodiaminę, którą przeprowadzono w pochodną symetrycznie chronioną grupami acetylowymi lub uretanowymi, po czym poddano reakcji Czerniaka-Einhorna i w wyniku późniejszego usunięcia grup ochronnych uzyskano batracylinę (BAT) z dobrymi wydajnościami. Ta modyfikacja zmniejszyła liczbę etapów i ułatwiła skomplikowaną dotychczas syntezę batracyliny. Pochodne batracyliny [N-(N-Boc-aminokwas)]-BAT otrzymano, stosując DCC lub EEDQ w bezwodnym chlorku metylenu. Koniugaty MDP zawierające batracylinę zostały skierowane do badań aktywności cytotoksycznej w Narodowym Instytucie Rakowym (NCI, Bethesda, USA) oraz w Katedrze Histologii i Immunologii Akademii Medycznej w Gdańsku. Z oznaczeń przeprowadzonych w NCI wynika, że związki te w badanym zakresie stężeń 10^-4 - 10^-8 M są nieaktywne w rutynowych testach tego ośrodka. Celem badań w Katedrze Histologii i Immunologii Akademii Medycznej w Gdańsku było oznaczenie immunomodulacyjnych właściwości tych koniugatów wobec subpopulacji leukocytów krwi obwodowej hodowanych w obecności komórek linii nowotworowych (K562, WEHI 164, Ab). Stwierdzono, że 3 koniugaty właściwościami hamowania rozwoju komórek wybranych linii nowotworowych przewyższają batracylinę, wydają się więc obiecującymi związkami, które w przyszłości mogą znaleźć zastosowanie jako leki. Ich skuteczność musi być jednak zweryfikowana na modelu zwierzęcym. Zsyntetyzowano 6 koniugatów MDP lub nor-MDP z tuftsyną (H- Thr-Lys-Pro-Arg-OH) i 6 koniugatów MDP lub nor-MDP z pochodnątuftsyny o odwróconej sekwencji (H-Arg-Pro-Lys-Thr-OMe). Do syntezy zarówno pochodnych tuftsyny, jak i ich koniugatów z MDP lub nor-MDP wybrano azydofosforan difenylu (DPP A) jako odczynnik kondensujący oraz metodę mieszanych bezwodników. W Katedrze Histologii i Immunologii Akademii Medycznej w Gdańsku przeprowadzono badania koniugatów MDP z tuftsyną, tuftsyny handlowej (firmy Bachem), pochodnej nitrotuftsyny (H-Thr-Lys-Pro-Arg(NOz)-OH) i nor-MDP, których celem było: ocena wpływu koniugatów na żywotność mononuklearnych komórek krwi obwodowej (PBMC), limfocytów krwi obwodowej (PBL) i monocytów, ocena sekrecji czynnika martwicy nowotworu (TNF.J i interleukiny 6 (IL6) w hodowlach komórek stymulowanych badanymi związkami, zbadanie wpływu na cytotoksyczność naturalnych komórek zabójczych (NK) oraz ocena wybuchu tlenowego w subpopulacjach krwi obwodowej pod wpływem badanych związków. Badania wykazały, że zaletą testowanych związków (w szczególności nitrotuftsyny) było ich szybsze działanie i większa skuteczność w porównaniu do tuftsyny. Planowane testy in vivo na modelu zwierzęcym mogą potwierdzić potencjalną użyteczność tych połączeń. Struktura otrzymanych koniugatów została potwierdzona na podstawie widm NMR (500 MHz), analizy elementarnej i jakościowej analizy na płytkach TLC.
EN
56 new conjugates of muramyl dipeptide (MDP), nor-muramyl dipeptide (nor-MDP) or desmuramylpeptides with compounds of determine or potential biological activity (anticancer or immunostimulant), e.g. amino-l-nitroacridinel4-nitro-9-acridone derivatives 56a-z, 72a,b, 77, N-(hydroxyalkil)-4-carboxyamide-acridine/9-acridone 7Sa-i, batracylin and its derivatives 122a-h or tuftsin and its derivative 126a-f, 129a-f have been synthesized. Practically for every group of the conjugates it was necessar to develop specific conditions for synthesis and purification of the reaction products. MDP or nor-MDP derivatives, needed for the synthesis of the conjugates, were prepared according to the method proposed by Flowers and Jeanloz [25] with some modifications described in chapter 5.1. (Scheme 7). It was an essential fragment of my work to prove that MDP derivatives presented in the dissertation contain isoglutamine residue. The literature data shows that removing the ester group type OBn and OMe from the [gama]- or [alfa]-carboxylic group of the isoglutamine or glutamine residues under alkaline hydrolysis. conditions leads to cyclic glutarimide. In fact, the hydrolysis of isoglutamine ester under anhydrous conditions (IN KOH/MeOH in dry 1,4-dioxane) gives the glutarimide derivative 50 (Scheme 7). However, it turned out that the use of such derivatives for acylation of amines or alcohols under the conditions described in this work gave a structure containing the isoglutamine residue. This was proven by structure analysis of the products employing high resolution NMR spectroscopy, 2D-COSY, heterocorrelated HMBC and HSQC spectra. I also obtained the same products using MDP containing free carboxyl group - a product of hydrolysis of tert-butyl MDP esters. Treating the protected MDP or nor-MDP tert-butyl esters with 90% TFA for 20 min caused the removal of both tert-butyl and 4,6-benzylidene groups (Scheme 7). 1-Benzyl-MDP or 1-benzyl-nor-MDP without purification were used for synthesis of conjugates with amine or hydroxy-acridine/acridone derivatives, batracylin or tuftsin. In my dissertation I present the synthesis of 24 conjugates of MDP or nor-MDP modified at the C-terminal part by formation of an amide bond between the isoglutamine carboxylic group and the amine function of acridinelacridone derivatives 56a-z (Table 2). Synthesis of three analogues of desmuramyl-peptides, 72a,b, 77 (Table 4), modified at C-end with an amino-acridinelacridone residue was also reported. Moreover, two other analogues, 72a,b, containing meso-diaminopimelic acid; and one more analogue with L-alanyl instead of L-valine and meso-diaminopimelic acid instead of L-lysine were described 77 (Table 4). The screening data indicate that the analogues 72a,b, 77, and 78a-i exhibit low cytotoxic activity, whereas a large number of others (56a-z) are potent in vitro cytotoxic agents against a panel of human cell lines. Seven compounds of this group, 56b,c,e,g,h,i,I, were selected by the NCI Biological Evaluation Committee for evaluations by mean of in vivo Hollow Fiber assay and three of them, 56b,e,h, passed the test for further evaluation in subcutaneous human tumor xenograft assays. I developed a convenient procedure for preparation of 7-methyl or p-nitrobenzyl ester of (6R)Z-(2S)-Boc-meso-A2pm, a key substrate for synthesis of several biological active peptides. Dibenzy-loxycarbonyl-meso-diaminopimelic acid 15 [126], after convertion into dimethyl or di-p-nitrobenzyl ester 52 (Scheme 8), was selectively hydrolyzed at the S-center using a-chymotrypsin, followed by transformation into N-carboxyanhydride 54. Then the acidic hydrolysis with 10% AcOH in THF and Boc-protection of the amino group at the S-center allowed me to obtain the selectively protected meso-A2pm 55 with free carboxyl group, ready for coupling at the S-center [53]. Compound 55 was used for the synthesis of the conjugates of desmuramylpeptides with amino-1-nitro-acridine/acridone derivatives 72a,b (Scheme 12, Reaction A). Furthermore, 1 synthesized 9 new analogues of MDP or nor-MDP modified at the C-end with the 4-carboxamide-hydroxyalkyl-acridine/9-acridone derivatives 78a-i. Unexpectedly, the formation of the ester bond between the carboxyl group of isoglutamine belonging to MDP molecule and the hydroxyl group of 4-carboxamide-hydroxyalkyl-acridine/9-acridone turned out to be very difficult. When popular coupling reagents such us DCC, EEDQ, CCBT, CCMT were used, the reaction yields hardly exceeded 5%. After many attempts I have found reaction conditions enabling the preparation of these products with a yield of about 35-40%. The results have been achieved thanks to the utilization of HBTU or EDCI as coupling reagents and the application of 2-fold surplus of the hydroxy component (Scheme 13). The 4-carboxamide-acridine/9-acridone derivatives were obtained according to Scheme 14. Acridine-4-carboxylic acid 82 was prepared by reduction of the corresponding 9-acridone-4-carboxylic acid 81 with aluminumlmercury amalgam, followed by FeCh reoxidation of the resulting acridans. Both acids (81 and 82) were condensed with amino alcohols in DMF by means of either DPP A or 1,1' -carbonyldiimidazole. Structures of all products were established on the basis of NMR spectra and microanalyses (Table 5). Conjugates 78a-i containing 4-carboxamide acridine or 9-acridone display no anticancer activity. In an attempt to use the Mitsunobu reaction for the acylation of N-(hydroxyalkyl)-9-acridone-4-carboxamide with muramyl dipeptide (MDP) or nor-muramyl dipeptide (nor-MDP), heterocyclic derivatives, oxazoles and oxazines were isolated instead of the expected esters. Although various molar proportions and orders of adding of the reagents were applied, ester formation was not observed. Oxazoles and oxazines were, in my opinion, formed as a result of intramolecular cyclization independently of the presence of an acidic component, Le. MDP or nor-MDP, in the reaction mixture. The mechanism of the reaction has been proposed. I have synthesized MDP and nor-MDP conjugates modified at the peptide residue with batracylin or its derivatives [60]. These conjugates, 122, were synthesized from partially protected MDP or nor-MDP 51 by means of DPP A as a coupling agent in the presence of TEA (Scheme 25, Table 11). I present a considerable modification of the batracylin synthesis (Scheme 24). In the improved method symmetrically protected l,4-phenylenediamine was used as a starting reagent. This made it possible to eliminate the most difficult steps of the hitherto existing synthesis and increase the product yield to 70 - 80% (after crystallization) [176]. The activity of these conjugates was evaluated at the Medical University of Gdansk on subpopulations of leukocytes isolated from venous blood cultures in the presence of tumour cells: K562 (human leukaemia), WEHl (mouse fibrosarcoma), Ab/Ma (hamster melanoma). The intensity of tumour cell death was measured by means of flow cytometry using subG 1 peak. Differentiation between apoptosis and necrosis was assessed using gel electrophoresis of DNA eluted from dead cells. Three conjugates, 122c, 122e, 122h, seem to be promising immunomodulators that could be used as medicines in the future. However, their effectiveness has to be verified on the animal model. I have also synthesized six conjugates of muramyl dipeptide (MOP) or nor-muramyl dipeptide (nor-MDP) with tuftsin (H-Thr-Lys-Pro-Arg-OH) (Scheme 27, Table 13) [194,195], and six conjugates of MOP or nor-MOP with retro-tuftsin (H-Arg-Pro-Lys-Thr-OMe) [196] (Scheme 28, Table 15). Also a tuftsin analog (H-Thr-Lys-Pro-Arg(NO2)-OH), 124, was obtained. The idea of the preparation of these conjugates was based on the assumption that the activity of the product would combine the biological activities of both tuftsin and MOP, or even enhance them, thanks to synergistic properties of MOP. The activity of tuftsin is generally directed toward the activation of non-specific elements of the immune system. It mainly stimulates the synthesis of free radicals by granulocytes and, to a lesser extent, by monocytes. Another action of tuftsin is connected with the secretion of cytokines by monocytes. On the other hand, MOP is much more deeply engaged in the stimulation of monocytes. It is believed that MOP, a fragment of bacterial cell wall peptidoglycan, acts via lipopolisaccharide receptor CO14 and via Toll-like receptors to stimulate a wide range of immune system elements. So, it was my intention to combine the effects of both these immunostimulants together. All the synthesized conjugates 1263-f, as well as nor-MOP and 124, were tested at the Department of Histology and Immunology, Medical University of Gdansk, Poland. The biological activity of the examined compounds was estimated using in vitro cultures of human monocytes and lymphocytes. The subtances displayed cytotoxic effects, as was revealed in the performed viability tests. The effects were most probably mediated by induction of the oxidative burst in monocytes and stimulation of redox enzymes in lymphocytes. In addition, the analogues turned out to be efficient stimulators of TNFa and IL6 secretion by monocytes and Iymphocytes. Nevertheless, the secretion of cytokines did not affect the viability of the leukocyte population used in the experiments. The beneficial properties of the examined compounds (mainly 124, 1263 and 126c), which imply their usefulness as potential therapeutic agents, are connected with their quick start of action and greater effectiveness as compared with tuftsin alone. An assay in vivo on animal models will be performed. The final products were isolated and purified by means of column chromatography on silica gel or radial chromatography. In some cases, when further purification was needed a preparative TLC was employed. The composition of the conjugates was confirmed by the analysis of NMR spectra, elemental analysis, and by TLC quantitative amino acid analysis.
The synthesis of retro-tuftsin analogue of sequence Arg-Pro-Lys-ThrOMe (Scheme 1) and it's conjugates containing MDP (muramyl dipeptide) or nor-MDP (nor-muramyl dipeptide) (Scheme 2) are described.
Combretastatin A-4 is a potent antineoplastic and antiangiogenesis natural substance isolated from the South African treeCombretum caffrum. This article concerns synthesis and biological activity of combretastatin A-4 and its analogues which are considered as prodrugs.
Muramyl peptides are fragments of peptidoglycan from the cell walls of bacteria. This article concerns muramyl peptides, and new analogues which are considered as prodrugs. Their synthesis and biological activity are also presented.
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