NK cells differentiated from bone marrow, cord blood and peripheral blood stem cells exhibit similar phenotype and functions

In the present study, we investigated the differentiation of human NK cells from bone marrow, cord blood and mobilized peripheral blood purified CD34+ stem cells using a potent culture system. Elutriated CD34+ stem cells were grown for several weeks in medium supplemented with stem cell factor (SCF) and IL-15 in the presence or absence of a murine stromal cell line (MS-5). Our data indicate that IL-15 induced the proliferation and maturation of highly positive CD56+ NK cells in both types of culture, although murine stromal cells slightly increased the proliferation of NK cells. NK cells differentiated in the presence of MS-5 were mostly CD56+ CD7 and a small subset expressed CD16. These in vitro differentiated CD56+ NK cells displayed cytolytic activity against the HLA class I- target K562. The CD56+ CD16+ subset also lysed NK-resistant Daudi cells. Neither of these NK subsets were shown to express Fas ligand. Total CD56+ cells expressed high amounts of transforming growth factor-beta and granulocyte-macrophage colony-stimulating factor, but no IFN-gamma. Investigation of NK receptor expression showed that most CD56+ cells expressed membrane CD94 and NKG2-A mRNA. PCR analysis revealed that p58 was also expressed in these cells. The role of CD94 in NK cell-mediated cytotoxicity was assessed on human HLA-B7-transfected murine L cells. While a low cytotoxic activity towards HLA-B7 cells was observed, the HLA-DR4 control cells were killed with high efficiency. These studies demonstrate that cytolytic and cytokine-producing NK cells may be derived from adult and fetal precursors by IL-15 and that these cells express a CD94 receptor which may influence their lytic potential.     

Effect of Oxime Ether Incorporation in Acyl Indole Derivatives on PPAR Subtype Selectivity

Compounds that simultaneously activate peroxisome proliferator‐activated receptor (PPAR) subtypes α and γ have the potential to effectively treat dyslipidemia and type 2 diabetes (T2D) in a single pharmaceutically active molecule. The frequently observed side effects of selective PPARγ agonists, such as edema and weight gain, were expected to be overcome by using additive PPARα activity, leading to dual PPARα/γ agonists with balanced activity for both subtypes. Herein we report the discovery, synthesis, and optimization of a new series of α‐ethoxyphenylpropionic acid bearing 5‐ or 6‐substituted indoles. The incorporation of oxime ethers on the carbonyl portion of the benzoyl group can bring the PPARα/γ potency ratio equal to or slightly greater than one, as is the case for compounds 20 c and 21 a . Compound 20 c shows high efficacy in an ob/ob mouse model of T2D and dyslipidemia, similar to that of rosiglitazone and tesaglitazar, but with a significant increase in body weight gain. In contrast, compound 21 a , less potent as a dual PPARα/γ activator than 20 c , showed an interesting pharmacological profile, as it elicits a decrease in body weight relative to reference compounds.     

Synthesis and Pharmacological Evaluation of a series of the Agomelatine Analogues as Melatonin MT1/MT2 Agonist and 5‐HT2C Antagonist

Agomelatine is a naphthalenic analogue of melatonin that is in clinical use for the treatment of major depressive disorders. Interestingly, while agomelatine exhibits potent affinity for melatonin receptors, it binds with only moderate affinity to the serotonin 5‐HT_2C receptor. Optimization of agomelatine toward this target could further potentiate its clinical efficacy. To explore this hypothesis and to access derivatives in which a key point of agomelatine metabolism is blocked, a series of naphthalenic derivatives was designed and synthesized as novel analogues of agomelatine. Most of the prepared compounds exhibited good binding affinity at the melatonin MT₁ and MT₂ receptor subtypes. Two compounds, an acetamide and an acrylamide derivative, exhibited good binding affinities at both the human melatonin (MT) receptors and the serotonin 5‐HT_2C receptor subtype, with pK_i values of 7.96 and 7.95 against MT₁, 7.86 and 8.68 against MT2, and 6.64 and 6.44 against 5‐HT_2C, respectively.     

Synthesis and biological evaluation of new 3-aralkylamino-2-aryl-2H-1, 2,4-pyridothiadiazine 1,1-dioxides as potential CCK-receptor ligands

A series of 2-aralkyl-4H-pyridothiadiazine 1,1-dioxides and 3-aralkylamino-2-aryl-2H-pyrido[4,3-e]-1,2,4-thiadiazine 1,1-dioxides structurally related to quinazolinone CCK receptor antagonists were synthesized and evaluated as CCK-A and CCK-B receptor ligands. The compounds were effective as cholecystokinin-ligands in the micromolar range of concentration, c.f. the cholecystokinin receptor antagonists asperlicin, lorglumide or benzotript, and were thus less potent than the best quinazolinones previously reported. Although the compounds were unsuitable for drug use, the work contributed to our understanding of the chemistry of unusual 2,3-disubstituted pyridothiadiazinedioxides.     

Novel and selective partial agonists of 5-HT3 receptors. 2. Synthesis and biological evaluation of piperazinopyridopyrrolopyrazines, piperazinopyrroloquinoxalines, and piperazinopyridopyrroloquinoxalines

In continuation of our previous work on piperazinopyrrolothienopyrazine derivatives, three series of piperazinopyridopyrrolopyrazines, piperazinopyrroloquinoxalines, and piperazinopyridopyrroloquinoxalines were prepared and evaluated as 5-HT3 receptor ligands. The chemical modifications performed within these new series led to structure-activity relationships regarding both high affinity and selectivity for the 5-HT3 receptors that are in agreement with those established previously for the pyrrolothienopyrazine series. The best compound (8a) obtained in these new series is in the picomolar range of affinity for 5-HT3 receptors with a selectivity higher than 10(6). Four of the high-affinity 5-HT3 ligands (8a, 15a,b, and 16d) were selected in both the pyridopyrrolopyrazine and the pyrroloquinoxaline series and were characterized in vitro and in vivo as agonists or partial agonists. Compound 8a was also evaluated in the light/dark test where it showed potential anxiolytic-like activity at very low doses per os.     

6-aminoalkyloxazolo[4,5-b]pyridin-2(3H)-ones: synthesis and evaluation of antinoceptive activity

Series of 6-aminoalkyloxazolo[4,5-b]pyridin-2(3H)-ones incorporating structural modifications both in the alkyl chain and basic amino moiety were tested for their analgesic efficacy and safety in mice and rats. Two of the synthesised compounds, 4a (3-methyl-6-[(4-phenyl-1-piperazinyl)methyl]oxazolo[4,5-b]pyridin-2(3H)-one) and 12a (3-methyl-6?1-[2-(4-phenyl-1-piperazinyl)ethan-1-ol]?oxazolo[4,5-b]pyridin- 2(3H)-one) were found to be more potent than aspirin with ED50 values of 26 (16.1-42.4) and 15.5 (11.4-21.2) mg/kg po (mouse, phenylquinone writhing test) respectively and 6 (3.1-9.8) and 5.5 (3.5-8.8) mg/kg po (rat, acetic acid writhing test). Compounds 4a and 12a proved to be potent nonopioid nonantiinflammatory analgesics but unfortunately have sedative properties at relatively low doses (respectively 64 and 16 mg/kg po, mice).     

Synthesis and structure-activity relationships of novel naphthalenic and bioisosteric related amidic derivatives as melatonin receptor ligands

A previous paper reported the synthesis of melatonin receptor ligands. In order to complete the structure-activity relationships and to obtain antagonists to the melatonin receptor, a new series of naphthalenic analogues of melatonin have been synthesized. Modifications include deletion of the 7-methoxy group, replacement of the ethylene moiety, replacement of the amidic function by bioisosteres, and replacement of the naphthalenic nucleus by other bicyclic rings. Almost all the structural modifications lead to decreased affinity for the melatonin receptor. However, the N-n propyl urea derivative (27) is a very potent ligand at this receptor (pKi = 14.3). Most interestingly deletion of the methoxy group resulted in the first antagonist in this series. This molecule, compound 12, or N-[2-(1-naphthyl)-ethyl]cyclobutyl carboxamide has been selected for preclinical development.     

New Radioligands for Describing the Molecular Pharmacology of MT1 and MT2 Melatonin Receptors

Melatonin receptors have been studied for several decades. The low expression of the receptors in tissues led the scientific community to find a substitute for the natural hormone melatonin, the agonist 2-[¹²⁵I]-iodomelatonin. Using the agonist, several hundreds of studies were conducted, including the discovery of agonists and antagonists for the receptors and minute details about their molecular behavior. Recently, we attempted to expand the panel of radioligands available for studying the melatonin receptors by using the newly discovered compounds SD6, DIV880, and {"type":"entrez-nucleotide","attrs":{"text":"S70254","term_id":"546525","term_text":"S70254"}}S70254. These compounds were characterized for their affinities to the hMT₁ and hMT₂ recombinant receptors and their functionality in the classical GTPγS system. SD6 is a full agonist, equilibrated between the receptor isoforms, whereas {"type":"entrez-nucleotide","attrs":{"text":"S70254","term_id":"546525","term_text":"S70254"}}S70254 and DIV880 are only partial MT₂ agonists, with K_i in the low nanomolar range while they have no affinity to MT₁ receptors. These new tools will hopefully allow for additions to the current body of information on the native localization of the receptor isoforms in tissues.     

Synthesis and SAR Studies of Isoquinoline and Tetrahydroisoquinoline Derivatives as Melatonin Receptor Ligands

In our constant search for new successors of agomelatine, we report herein a new series of compounds resulting from bioisosteric modulation of the naphthalene ring. The isoquinoline and tetrahydroisoquinoline derivatives were synthesized and pharmacologically evaluated. This isosteric replacement of the naphthalene group of agomelatine has led to potent agonist and partial agonist compounds with nanomolar melatonergic binding affinities. Overall, the presence of a nitrogen atom was accompanied with a decrease in the binding affinity toward both MT₁ and MT₂ and the loss of 5HT_2C response, especially for tetrahydroisoquinoline in comparison with the parent compound. Interestingly, due to the presence of this nitrogen atom, a notable improvement in the pharmacokinetic properties was observed for all compounds.