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1.
J Org Chem ; 85(3): 1466-1475, 2020 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-31660743

RESUMO

The Myc transcription factor represents an "undruggable" target of high biological interest due to its central role in various cancers. An abbreviated form of the c-Myc protein, called Omomyc, consists of the Myc DNA-binding domain and a coiled-coil region to facilitate dimerization of the 90 amino acid polypeptide. Here we present our results to evaluate the synthesis of Omomyc using three complementary strategies: linear Fmoc solid-phase peptide synthesis (SPPS) using several advancements for difficult sequences, native chemical ligation from smaller peptide fragments, and a high-throughput bacterial expression and assay platform for rapid mutagenesis. This multifaceted approach allowed access to up to gram quantities of the mini-protein and permitted in vitro and in vivo SAR exploration of this modality. DNA-binding results and cellular activity confirm that Omomyc and analogues presented here, are potent binders of the E-box DNA engaged by Myc for transcriptional activation and that this 90-amino acid mini-protein is cell permeable and can inhibit proliferation of Myc-dependent cell lines. We also present additional results on covalent homodimerization through disulfide formation of the full-length mini-protein and show the coiled-coil region can be truncated while preserving both DNA binding and cellular activity. Altogether, our results highlight the ability of advanced peptide synthesis to achieve SAR tractability in a challenging synthetic modality.


Assuntos
DNA , Proteínas Proto-Oncogênicas c-myc , Linhagem Celular , DNA/metabolismo , Fragmentos de Peptídeos , Ligação Proteica , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo
2.
ACS Med Chem Lett ; 13(8): 1255-1261, 2022 Aug 11.
Artigo em Inglês | MEDLINE | ID: mdl-35978702

RESUMO

The combination of insulin and incretin-based therapies has emerged as a potential promising tactic for the treatment of diabetes. Here we report the first example of a unimolecular triagonist to simultaneously target insulin, GLP-1, and glucagon receptors, aiming for better glycemic control and superior weight loss. The strategy for constructing such a unimolecular triagonist is the conjugation of the insulin moiety and GLP-1R/GCGR coagonist peptide via alkyne-azide click chemistry. Two tractable series differentiated by insulin conjugation sites, B1F and B29K, were identified. Triagonist 13 prepared through the conjugation at insulin B1F and position 24 of GLP-1R/GCGR coagonist exhibited insulin activity comparable to that of insulin degludec and potent and balanced GLP-1R and GCGR activities. Pharmacokinetic profiles of 13 in both rat and minipig were also discussed.

3.
J Pharm Biomed Anal ; 210: 114566, 2022 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-35042144

RESUMO

Lipidation, a common strategy to improve half-life of therapeutic peptides, affects their tendency to oligomerize, their interaction with plasmatic proteins, and their catabolism. In this work, we have leveraged the use of NMR and SPR spectroscopy to elucidate oligomerization propensity and albumin interaction of different analogs of the two marketed lipidated GLP-1 agonists liraglutide and semaglutide. As most lipidated therapeutic peptides are administered by subcutaneous injection, we have also assessed in vitro their catabolism in the SC tissue using the LC-HRMS-based SCiMetPep assay. We observed that oligomerization had a shielding effect against catabolism. At the same time, binding to albumin may provide only limited protection from proteolysis due to the higher unbound peptide fraction present in the subcutaneous compartment with respect to the plasma. Finally, identification of catabolites in rat plasma after SC dosing of semaglutide showed a good correlation with the in vitro data, with Tyr19-Leu20 being the major cleavage site. Early characterization of the complex interplay between oligomerization, albumin binding, and catabolism at the injection site is essential for the synthesis of lipidated peptides with good pharmacokinetic profiles.


Assuntos
Diabetes Mellitus Tipo 2 , Peptídeo 1 Semelhante ao Glucagon , Albuminas , Animais , Meia-Vida , Hipoglicemiantes , Liraglutida , Peptídeos , Ratos
4.
ACS Med Chem Lett ; 13(8): 1248-1254, 2022 Aug 11.
Artigo em Inglês | MEDLINE | ID: mdl-35978682

RESUMO

Peptide-based analogues of the gut-derived incretin hormone, glucagon-like peptide 1 (GLP1), stimulate insulin secretion in a glucose-dependent manner. Currently marketed GLP1 receptor (GLP1R) agonists are safe and effective in the management of Type 2 diabetes but often offer only modest weight loss. This has prompted the search for safe and effective alternatives to enhance the weight loss component of these treatments. We have demonstrated that concomitant activation GLP1R and the glucagon receptor (GCGR) can improve glucose metabolism and provide superior weight loss when compared to selective GLP1R agonism in preclinical species. This paper will highlight chemistry structure-activity relationship optimization and summarize in vivo efficacy studies toward the discovery of a once daily balanced dual agonist 12 (MK-1462), which was advanced into clinical trials.

6.
Bioorg Med Chem Lett ; 19(15): 4042-5, 2009 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-19553107

RESUMO

PARP inhibitors have been demonstrated to retard intracellular DNA repair and therefore sensitize tumor cells to cytotoxic agents or ionizing radiation. We report the identification of a novel class of PARP1 inhibitors, containing a pyrrolo moiety fused to a dihydroisoquinolinone, derived from virtual screening of the proprietary collection. SAR exploration around the nitrogen of the aminoethyl appendage chain of 1 led to compounds that displayed low nanomolar activity in a PARP1 enzymatic assay.


Assuntos
Inibidores de Poli(ADP-Ribose) Polimerases , Quinolonas/química , Antineoplásicos/farmacologia , Sítios de Ligação , Química Farmacêutica/métodos , Cristalografia por Raios X/métodos , Reparo do DNA , Desenho de Fármacos , Humanos , Ligação de Hidrogênio , Concentração Inibidora 50 , Ligantes , Modelos Químicos , Polímeros/química , Relação Estrutura-Atividade
8.
Mol Cell Biol ; 39(22)2019 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-31501275

RESUMO

The MYC oncogene is upregulated in human cancers by translocation, amplification, and mutation of cellular pathways that regulate Myc. Myc/Max heterodimers bind to E box sequences in the promoter regions of genes and activate transcription. The MYC inhibitor Omomyc can reduce the ability of MYC to bind specific box sequences in promoters of MYC target genes by binding directly to E box sequences as demonstrated by chromatin immunoprecipitation (CHIP). Here, we demonstrate by both a proximity ligation assay (PLA) and double chromatin immunoprecipitation (ReCHIP) that Omomyc preferentially binds to Max, not Myc, to mediate inhibition of MYC-mediated transcription by replacing MYC/MAX heterodimers with Omomyc/MAX heterodimers. The formation of Myc/Max and Omomyc/Max heterodimers occurs cotranslationally; Myc, Max, and Omomyc can interact with ribosomes and Max RNA under conditions in which ribosomes are intact. Taken together, our data suggest that the mechanism of action of Omomyc is to bind DNA as either a homodimer or a heterodimer with Max that is formed cotranslationally, revealing a novel mechanism to inhibit the MYC oncogene. We find that in vivo, Omomyc distributes quickly to kidneys and liver and has a short effective half-life in plasma, which could limit its use in vivo.


Assuntos
Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Genes myc , Fragmentos de Peptídeos/genética , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Sequência de Aminoácidos , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Linhagem Celular , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina/métodos , DNA/metabolismo , Proteínas de Ligação a DNA/metabolismo , Feminino , Células HCT116 , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Fragmentos de Peptídeos/metabolismo , Fragmentos de Peptídeos/farmacologia , Regiões Promotoras Genéticas , Ligação Proteica , Proteínas Proto-Oncogênicas c-myc/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-myc/farmacologia , Proteínas Recombinantes/farmacologia , Transcrição Gênica , Ativação Transcricional
9.
J Med Chem ; 51(4): 861-74, 2008 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-18217703

RESUMO

HIV integrase is one of the three enzymes encoded by HIV genome and is essential for viral replication, but integrase inhibitors as marketed drugs have just very recently started to emerge. In this study, we show the evolution from the N-methylpyrimidinone structure to bicyclic pyrimidinones. Introduction of a suitably substituted amino moiety modulated the physical-chemical properties of the molecules and conferred nanomolar activity in the inhibition of spread of HIV-1 infection in cell culture. An extensive SAR study led to sulfamide (R)- 22b, which inhibited the strand transfer with an IC50 of 7 nM and HIV infection in MT4 cells with a CIC95 of 44 nM, and ketoamide (S)- 28c that inhibited strand transfer with an IC50 of 12 nM and the HIV infection in MT4 cells with a CIC95 of 13 nM and exhibited a good pharmacokinetic profile when dosed orally to preclinical species.


Assuntos
Aminopiridinas/síntese química , Azepinas/síntese química , Compostos Bicíclicos Heterocíclicos com Pontes/síntese química , Inibidores de Integrase de HIV/síntese química , Integrase de HIV/metabolismo , Pirimidinonas/síntese química , Administração Oral , Aminopiridinas/farmacocinética , Aminopiridinas/farmacologia , Animais , Azepinas/farmacocinética , Azepinas/farmacologia , Disponibilidade Biológica , Compostos Bicíclicos Heterocíclicos com Pontes/farmacocinética , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Linhagem Celular , Cães , Integrase de HIV/genética , Inibidores de Integrase de HIV/farmacocinética , Inibidores de Integrase de HIV/farmacologia , HIV-1/efeitos dos fármacos , Humanos , Macaca mulatta , Microssomos Hepáticos/metabolismo , Pirimidinonas/farmacocinética , Pirimidinonas/farmacologia , Ratos , Estereoisomerismo , Relação Estrutura-Atividade
10.
Sci Rep ; 8(1): 585, 2018 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-29330364

RESUMO

A promising emerging area for the treatment of obesity and diabetes is combinatorial hormone therapy, where single-molecule peptides are rationally designed to integrate the complementary actions of multiple endogenous metabolically-related hormones. We describe here a proof-of-concept study on developing unimolecular polypharmacy agents through the use of selection methods based on phage-displayed peptide libraries (PDL). Co-agonists of the glucagon (GCG) and GLP-1 receptors were identified from a PDL sequentially selected on GCGR- and GLP1R-overexpressing cells. After two or three rounds of selection, 7.5% of randomly picked clones were GLP1R/GCGR co-agonists, and a further 1.53% were agonists of a single receptor. The phages were sequenced and 35 corresponding peptides were synthesized. 18 peptides were potent co-agonists, 8 of whom showed EC50 ≤ 30 pM on each receptor, comparable to the best rationally designed co-agonists reported in the literature. Based on literature examples, two sequences were engineered to stabilize against dipeptidyl peptidase IV cleavage and prolong the in vivo half-life: the engineered peptides were comparably potent to the parent peptides on both receptors, highlighting the potential use of phage-derived peptides as therapeutic agents. The strategy described here appears of general value for the discovery of optimized polypharmacology paradigms across several metabolically-related hormones.


Assuntos
Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Peptídeos/síntese química , Peptídeos/farmacologia , Receptores de Glucagon/agonistas , Diabetes Mellitus/tratamento farmacológico , Dipeptidil Peptidase 4/metabolismo , Humanos , Obesidade/tratamento farmacológico , Biblioteca de Peptídeos , Peptídeos/genética , Polimedicação , Análise de Sequência de DNA
11.
J Med Chem ; 50(9): 2225-39, 2007 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-17428043

RESUMO

Human immunodeficiency virus type-1 (HIV-1) integrase, one of the three constitutive viral enzymes required for replication, is a rational target for chemotherapeutic intervention in the treatment of AIDS that has also recently been confirmed in the clinical setting. We report here on the design and synthesis of N-benzyl-5,6-dihydroxypyrimidine-4-carboxamides as a class of agents which exhibits potent inhibition of the HIV-integrase-catalyzed strand transfer process. In the current study, structural modifications on these molecules were made in order to examine effects on HIV-integrase inhibitory potencies. One of the most interesting compounds for this series is 2-[1-(dimethylamino)-1-methylethyl]-N-(4-fluorobenzyl)-5,6-dihydroxypyrimidine-4-carboxamide 38, with a CIC95 of 78 nM in the cell-based assay in the presence of serum proteins. The compound has favorable pharmacokinetic properties in preclinical species (rats, dogs, and monkeys) and shows no liabilities in several counterscreening assays, highlighting its potential as a clinically useful antiviral agent.


Assuntos
Inibidores de Integrase de HIV/síntese química , HIV-1/efeitos dos fármacos , Piridinas/síntese química , Pirimidinas/síntese química , Animais , Disponibilidade Biológica , Proteínas Sanguíneas/metabolismo , Linhagem Celular Tumoral , Cães , Inibidores de Integrase de HIV/farmacocinética , Inibidores de Integrase de HIV/farmacologia , Meia-Vida , Humanos , Macaca mulatta , Ligação Proteica , Piridinas/química , Piridinas/farmacologia , Pirimidinas/farmacocinética , Pirimidinas/farmacologia , Ratos , Relação Estrutura-Atividade , Replicação Viral
12.
J Med Chem ; 50(20): 4953-75, 2007 Oct 04.
Artigo em Inglês | MEDLINE | ID: mdl-17824681

RESUMO

The human immunodeficiency virus type-1 (HIV-1) encodes three enzymes essential for viral replication: a reverse transcriptase, a protease, and an integrase. The latter is responsible for the integration of the viral genome into the human genome and, therefore, represents an attractive target for chemotherapeutic intervention against AIDS. A drug based on this mechanism has not yet been approved. Benzyl-dihydroxypyrimidine-carboxamides were discovered in our laboratories as a novel and metabolically stable class of agents that exhibits potent inhibition of the HIV integrase strand transfer step. Further efforts led to very potent compounds based on the structurally related N-Me pyrimidone scaffold. One of the more interesting compounds in this series is the 2-N-Me-morpholino derivative 27a, which shows a CIC95 of 65 nM in the cell in the presence of serum. The compound has favorable pharmacokinetic properties in three preclinical species and shows no liabilities in several counterscreening assays.


Assuntos
Inibidores de Integrase de HIV/síntese química , Integrase de HIV/química , HIV-1/efeitos dos fármacos , Morfolinas/síntese química , Pirimidinonas/síntese química , Administração Oral , Animais , Disponibilidade Biológica , Proteínas Sanguíneas/metabolismo , Linhagem Celular Tumoral , Cães , Inibidores de Integrase de HIV/farmacocinética , Inibidores de Integrase de HIV/farmacologia , HIV-1/enzimologia , HIV-1/fisiologia , Humanos , Macaca mulatta , Morfolinas/farmacocinética , Morfolinas/farmacologia , Ligação Proteica , Pirimidinonas/farmacocinética , Pirimidinonas/farmacologia , Ratos , Estereoisomerismo , Relação Estrutura-Atividade , Replicação Viral/efeitos dos fármacos
13.
J Med Chem ; 51(18): 5843-55, 2008 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-18763751

RESUMO

Human immunodeficiency virus type-1 (HIV-1) integrase is one of the three virally encoded enzymes required for replication and therefore a rational target for chemotherapeutic intervention in the treatment of HIV-1 infection. We report here the discovery of Raltegravir, the first HIV-integrase inhibitor approved by FDA for the treatment of HIV infection. It derives from the evolution of 5,6-dihydroxypyrimidine-4-carboxamides and N-methyl-4-hydroxypyrimidinone-carboxamides, which exhibited potent inhibition of the HIV-integrase catalyzed strand transfer process. Structural modifications on these molecules were made in order to maximize potency as HIV-integrase inhibitors against the wild type virus, a selection of mutants, and optimize the selectivity, pharmacokinetic, and metabolic profiles in preclinical species. The good profile of Raltegravir has enabled its progression toward the end of phase III clinical trials for the treatment of HIV-1 infection and culminated with the FDA approval as the first HIV-integrase inhibitor for the treatment of HIV-1 infection.


Assuntos
Infecções por HIV/tratamento farmacológico , Inibidores de Integrase de HIV/farmacologia , Pirrolidinonas/farmacologia , Administração Oral , Área Sob a Curva , Disponibilidade Biológica , Inibidores de Integrase de HIV/administração & dosagem , Inibidores de Integrase de HIV/farmacocinética , Inibidores de Integrase de HIV/uso terapêutico , Meia-Vida , Humanos , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Pirrolidinonas/administração & dosagem , Pirrolidinonas/farmacocinética , Pirrolidinonas/uso terapêutico , Raltegravir Potássico
14.
Bioorg Med Chem Lett ; 16(10): 2748-52, 2006 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-16503141

RESUMO

A series of aryltetrazolylacetanilides was synthesized and evaluated as HIV-1 non-nucleoside reverse transcriptase inhibitors on wild-type virus and on the clinically relevant K103N mutant strain. Extensive SAR investigation led to potent compounds, with nanomolar activity on K103N, and orally bioavailable in rats.


Assuntos
Acetanilidas/farmacologia , Transcriptase Reversa do HIV/antagonistas & inibidores , HIV-1/efeitos dos fármacos , Mutação , Inibidores da Transcriptase Reversa/farmacologia , Acetanilidas/química , Animais , HIV-1/enzimologia , HIV-1/genética , Ratos , Inibidores da Transcriptase Reversa/química , Relação Estrutura-Atividade
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