BRPI1000099A2 - Potentially active mimetic peptide compounds against hepatitis c virus and pharmaceutical composition containing such compounds - Google Patents
Potentially active mimetic peptide compounds against hepatitis c virus and pharmaceutical composition containing such compounds Download PDFInfo
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- BRPI1000099A2 BRPI1000099A2 BRPI1000099A BRPI1000099A2 BR PI1000099 A2 BRPI1000099 A2 BR PI1000099A2 BR PI1000099 A BRPI1000099 A BR PI1000099A BR PI1000099 A2 BRPI1000099 A2 BR PI1000099A2
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Abstract
COMPOSTOS PEPTìDEOS MIMéTICOS POTENCIALMENTE ATIVOS CONTRA VìRUS DA HEPATITE C E COMPOSIçãO FARMACêUTICA CONTENDO TAIS COMPOSTOS. A presente invenção se refere à compostos peptídeo miméticos potencialmente ativos contra virus da hepatite C e composição farmacêutica contendo tais compostos, desenhados como inibidores de polimerases e serina protease do vírus da Hepatite C (HCV), sintéticos, caracterizados por possuir, na porção central, uma estrutura do tipo 1,4:3,6-dianidromanitol. As porções laterais são caracterizadas por possuir ligações peptideo miméticas provenientes do acopíamento com diversos aminoácidos N-protegidos.MIMETIC PEPTIDE COMPOUNDS POTENTIALLY ACTIVE AGAINST HEPATITIS C VIRUS AND PHARMACEUTICAL COMPOSITION CONTAINING SUCH COMPOUNDS. The present invention relates to peptide mimetic compounds potentially active against hepatitis C virus and pharmaceutical composition containing such compounds, designed as synthetic Hepatitis C virus (HCV) polymerase and serine protease inhibitors, characterized by having, in the central portion, a structure of type 1,4: 3,6-dianhydromanitol. The side portions are characterized by having mimetic peptide bonds from coupling with several N-protected amino acids.
Description
Relatório Descritivo de Patente de InvençãoPatent Invention Descriptive Report
Compostos Peptídeos Miméticos Potencialmente Ativos Contra Vírus da Hepatite C ε Composição Farmacêutica Contendo tais CompostosPotentially Active Mimetic Peptide Compounds Against Hepatitis C Virus Pharmaceutical Composition Containing Such Compounds
Campo da InvençãoField of the Invention
A presente invenção se refere à COMPOSTOS PEPTÍDEO MIMÉTICOS POTENCIALMENTE ATIVOS CONTRA VÍRUS DA HEPATITE C E COMPOSIÇÃO FARMACÊUTICA CONTENDO TAIS COMPOSTOS, desenhados como inibidores de polimerases e serina protease do vírus da Hepatite C (HCV), sintéticos, caracterizados por possuir, na porção central, uma estrutura do tipo 1,4:3,6- dianidromanitol. As porções laterais são caracterizadas por possuir ligações peptídeo miméticas provenientes do acoplamento com diversos aminoácidos N- protegidos. Esses novos compostos, se ativos contra o vírus da hepatite C, poderão ser a base para a preparação de formulações antivirais capazes de cessar a proliferação do HCV responsável pela flavivirose, a Hepatite C.The present invention relates to potentially active mimetic peptide compounds against heparitis c virus. 1,4: 3,6-dianhydromanitol type structure. Side portions are characterized by having mimetic peptide bonds from coupling with various N-protected amino acids. These new compounds, if active against the hepatitis C virus, could be the basis for the preparation of antiviral formulations capable of stopping HCV proliferation responsible for flavivirosis, hepatitis C.
Antecedentes da InvençãoBackground of the Invention
A família Flaviviridae compreende mais de 60 viroses, das quais muitas são patogênicas ao ser humano. Dentre estas viroses encontram-se aquelas relacionadas ao vírus da Hepatite C (HCV), da Febre do Oeste do Nilo (ou Febre do Nilo Ocidental), da febre amarela e da Dengue. No caso das hepatites virais, são pelo menos sete os tipos de vírus que já foram caracterizados: A, B, C, D, E, G e TT, que têm em comum o hepatotropismo e podem levar a inflamação e necrose hepática. Hepatite é o termo genérico, com origem grega hepato- (fígado) e -itis (inflamação), que designa a inflamação do fígado.The family Flaviviridae comprises more than 60 viruses, many of which are pathogenic to humans. These viruses include those related to Hepatitis C virus (HCV), West Nile Fever (or West Nile Fever), Yellow fever and Dengue fever. In the case of viral hepatitis, there are at least seven types of viruses that have already been characterized: A, B, C, D, E, G and TT, which have hepatotropism in common and may lead to liver inflammation and necrosis. Hepatitis is the generic term of Greek origin hepato- (liver) and -itis (inflammation), which means inflammation of the liver.
As hepatites virais, particularmente, são causadas por diferentes agentes etiológicos, de distribuição global dentre eles o HCV (Craxi, A.; Laffi, G.; Zignego, A.L. Mol. Aspects Med. 2008, 29, 85-95; Manns, M.P.; Foster, G.R.; Rockstroh, J.K.; Zeuzem, S.; Zoulim, F.; Houghton, M. Nat. Rev. Drug Discov. 2007, 6, 991-1000). Em termos estruturais o HCV é um vírus envelopado, possuindo um genoma RNA1 com aproximadamente 9.4Kb. O genoma possui uma única janela aberta de leitura que codifica três proteínas estruturais (cerne, E1, E2) e sete proteínas não estruturais (p7, NS2 a NS5B). Duas regiões não traduzidas (UTR) presentes em cada extremidade do genoma são importantes para as etapas de tradução e transcrição do genoma RNA. Foi descrita uma grande variedade na seqüência genômica do HCV. Os diferentes genótipos foram reunidos em seis grupos principais e vários subtipos. Há uma distribuição geográfica diferenciada em relação aos diferentes genótipos. No Brasil, os mais freqüentes são os genótipos 1, 2 e 3 com incidência de 70; 2,5 e 28 %, respectivamente.Viral hepatitis, in particular, are caused by different globally distributed etiological agents, including HCV (Craxi, A.; Laffi, G .; Zignego, AL. Mol. Aspects Med. 2008, 29, 85-95; Manns, MP ; Foster, GR; Rockstroh, JK; Zeuzem, S.; Zoulim, F.; Houghton, M. Nat. Rev. Drug Discov. 2007, 6, 991-1000). In structural terms HCV is an enveloped virus, having an RNA1 genome of approximately 9.4Kb. The genome has a single open reading window that encodes three structural proteins (heartwood, E1, E2) and seven nonstructural proteins (p7, NS2 to NS5B). Two untranslated regions (RTU) present at each end of the genome are important for the RNA genome translation and transcription steps. A wide variety has been described in the HCV genomic sequence. The different genotypes were grouped into six main groups and various subtypes. There is a different geographical distribution in relation to different genotypes. In Brazil, the most frequent are genotypes 1, 2 and 3 with an incidence of 70; 2.5 and 28%, respectively.
O genoma viral codifica uma única poliproteína longa de aproximadamente 3000 aminoácidos. Esta poliproteína é processada subseqüentemente por proteases codificadas pelo vírus e pelo hospedeiro, gerando 10 proteínas virais maduras. As proteínas de cápsula ("core") E1 e E2 são estruturais e estão envolvidas com a montagem da partícula viral, enquanto as proteínas NS2 a NS5B são chamadas não-estruturais e estão envolvidas com a propagação do vírus. Estas proteínas não- estruturais incluem a helicase vírus-específica NS3, a protease NS3-NS4A e a RNA polimerase RNA-dependente NS5B (RdRp, ou replicase).The viral genome encodes a single long polyprotein of approximately 3000 amino acids. This polyprotein is subsequently processed by proteases encoded by the virus and host, generating 10 mature viral proteins. The core proteins E1 and E2 are structural and are involved with viral particle assembly, while proteins NS2 through NS5B are called nonstructural and are involved with virus propagation. These nonstructural proteins include the NS3 virus-specific helicase, the NS3-NS4A protease and the NS5B RNA-dependent RNA polymerase (RdRp, or replicase).
A atual terapia basea-se no uso do interferon alfa combinado com ribavirina. O tratamento apenas com o IFN-alfa apresenta apenas 10-19% de resposta sustentada. A Ribavirina é um análogo sintético da guanosina que tem ação direta contra vírus de RNA e DNA, por provável mecanismo de inibição da DNA polimerase vírus-dependente. A Ribavirina sozinha, no entanto, não tem qualquer efeito sobre a hepatite C. A combinação do interferon-alfa com a Ribavirina melhora a resposta virológica sustentada para 38-43%, com correspondente melhora na análise histológica (biópsia) e, possivelmente, nas complicações em longo prazo da hepatite, contudo não há estudos prospectivos convincentes (Heathcote, E.J. J. Viral Hepat. 2007, 14, Suppl 1, 82-8; Koike, K.J. Infect. Chemother. 2006, 12, 227-32; Toniutto, P.; Fabris, C.; Pirisi, M. Expert. Opin. Pharmacother. 2006, 7, 2025-35).Current therapy is based on the use of interferon alfa combined with ribavirin. IFN-alpha treatment alone has only a 10-19% sustained response. Ribavirin is a synthetic guanosine analogue that has direct action against RNA and DNA viruses, by probable mechanism of inhibition of virus-dependent DNA polymerase. Ribavirin alone, however, has no effect on hepatitis C. Combination of interferon-alpha with Ribavirin improves sustained virological response to 38-43%, with corresponding improvement in histological analysis (biopsy) and possibly long-term complications of hepatitis, however there are no convincing prospective studies (Heathcote, EJJ Viral Hepat. 2007, 14, Suppl 1, 82-8; Koike, KJ Infect. Chemother. 2006, 12, 227-32; Toniutto, P. ; Fabris, C.; Pirisi, M. Expert. Opin. Pharmacother. 2006, 7, 2025-35).
Mesmo com este arsenal terapêutico, a infecção pelo HCV é responsável por mais de 80% dos transplantes de fígado no mundo, e no Brasil. (Consenso da Sociedade Paulista de Infectologia (SPI) para Manuseio e Terapia da Hepatite C, São Paulo, SP.) Diversos novos compostos têm sido avaliados para o tratamento das infecções pelo HCV. Esses compostos se encontram em fases iniciais de avaliação, ou seja, estudos in vitro ou pré-clínicos em animais. Porém, a maioria desses compostos ainda não foi analisada em fases de triagem envolvendo seres humanos (estudos clínicos). De forma geral, apenas 1 em cada 1.000 novos compostos chega a ser avaliado em etapas que envolvem seres humanos. Entre esses, somente 1 em 5 recebe a aprovação pelo Food and Drug Administration, FDA, escritório de alimentos e medicamentos do governo dos Estados Unidos da América.Even with this therapeutic arsenal, HCV infection is responsible for over 80% of liver transplants worldwide, and in Brazil. (Consensus of the Paulista Society of Infectology (SPI) for Hepatitis C Management and Therapy, São Paulo, SP.) Several new compounds have been evaluated for the treatment of HCV infections. These compounds are in early stages of evaluation, ie in vitro or preclinical animal studies. However, most of these compounds have not yet been analyzed in screening stages involving humans (clinical studies). In general, only 1 out of every 1,000 new compounds is ever evaluated in stages involving humans. Of these, only 1 in 5 is approved by the US Food and Drug Administration, FDA.
Vários esforços têm se concentrado na lista de tratamentos que tem se mostrado promissores em estudos clínicos envolvendo seres humanos. A Tabela 1 abaixo mostra alguns compostos com potencial de inibir a infecção pelo HCV e as fases clínicas que se encontram.Several efforts have focused on the list of treatments that have shown promise in clinical trials involving humans. Table 1 below shows some compounds with the potential to inhibit HCV infection and the clinical phases that are present.
Tabela 1 - Compostos com potencial de inibição da infecção pelo HCV e suas respectivas fases clínicas. (Soriano, V.; Madejon, A.; Vispo, E.; Labarga, P.; Garcia- Samaniego, J.; Martin-Carbonero, L.; Sheldon, J.; Bottecchia1 M.; Tuma, P.; Barreiro, P. Expert. Op. Emerg. Drugs 2008, 13, 1-19).Table 1 - Compounds with potential inhibition of HCV infection and their respective clinical phases. (Soriano, V .; Madejon, A.; Vispo, E.; Labarga, P.; Garcia-Samaniego, J.; Martin-Carbonero, L.; Sheldon, J.; Bottecchia1 M .; Tuma, P .; Barreiro , P. Expert Op. Emergency Drugs 2008, 13, 1-19).
<table>table see original document page 4</column></row><table> <table>table see original document page 5</column></row><table><table> table see original document page 4 </column> </row> <table> <table> table see original document page 5 </column> </row> <table>
* Inibidores da RNA polimerase* RNA polymerase inhibitors
** Inibidores da protease** Protease Inhibitors
Os alvos da terapia anti-HCV estão concentrados nas diferentes etapas de replicação do RNA viral, e além desses as novas terapias estão direcionadas nas etapas de tradução (ribozimas, oligos anti-senso, RNAi), processamento proteolítico (inibidores da protease viral) e de replicação do genoma RNA (inibidores da RNA polimerase viral).The targets of anti-HCV therapy are concentrated at the different stages of viral RNA replication, and in addition to these new therapies are targeted at the translation steps (ribozymes, antisense oligos, RNAi), proteolytic processing (viral protease inhibitors) and RNA genome replication (viral RNA polymerase inhibitors).
O Valopicitabine (Idenix Pharmaceuticals), já em fase avançada de testes clínicos, se caracteriza por ser um inibidor RNA-RNA polimerase viral, e a análise envolvendo 79 pacientes mostraram em duas semanas uma redução de 90% da carga viral, sendo que a maioria desses pacientes não respondia ao tratamento com interferon. O Valopicitabine será empregado como um terceiro medicamento no tratamento, junto com o interferon e a Ribavirina, para se obter uma maior resposta terapêutica. Duas outras drogas, das quais uma deverá ser utilizada como terceira droga no tratamento são os inibidores de protease: SCH-503034 (Schering-Plough) e o VX-950 (Vertex). Em estudo realizado pela Vertex em 34 voluntários com o VX- 950, mostrou que após duas semanas de tratamento a diminuição da carga viral foi 250 vezes maior do que o Interferon Peguilado combinado a Ribavirina na redução da carga viral.Valopicitabine (Idenix Pharmaceuticals), already at an advanced stage of clinical trials, is characterized as a viral RNA-RNA polymerase inhibitor, and analysis involving 79 patients showed a 90% reduction in viral load within two weeks, with the majority of these patients did not respond to interferon treatment. Valopicitabine will be used as a third treatment medicine, together with interferon and ribavirin, to achieve a greater therapeutic response. Two other drugs, one of which should be used as a third drug in treatment are protease inhibitors: SCH-503034 (Schering-Plow) and VX-950 (Vertex). In a study by Vertex in 34 volunteers with VX-950, it showed that after two weeks of treatment the decrease in viral load was 250 times greater than the pegylated interferon combined with ribavirin in reducing viral load.
O HCV tem dois alvos potenciais para desenvolvimento de novas drogas antivirais. Um é a molécula de (NS5) RNA Polimerase RNA dependente que consegue replicar uma molécula de RNA em outra. Esta é uma atividade única dos vírus de RNA, não se encontrando essa atividade nas células hospedeiras. Dessa forma a RNA Polimerase RNA dependente é um alvo muito específico para o HCV. Além da replicase viral ainda temos a NS3 (serina protease) que é responsável pelo processamento da poliproteína viral. Apesar de ser uma serina protease, a NS3 tem uma estrutura molecular muito diferente das serinas proteases das células humanas, o que possibilita o desenvolvimento de inibidores específicos para esta protease.HCV has two potential targets for the development of new antiviral drugs. One is the RNA-dependent (NS5) RNA Polymerase molecule that can replicate one RNA molecule into another. This is a unique activity of RNA viruses, and this activity is not found in host cells. Thus RNA dependent RNA polymerase is a very specific target for HCV. In addition to viral replicase we also have NS3 (serine protease) which is responsible for processing viral polyprotein. Despite being a serine protease, NS3 has a very different molecular structure than human cell serine proteases, which enables the development of inhibitors specific for this protease.
O peso molecular do monômero da protease NS3 do HCV é de 70 kDa. Sua estrutura ativa é um dímero e a estrutura terciária do monômero tem uma extensão na porção N-terminal, 2 domínios com 6 "beta-barrels" (conjunto de fitas beta- pregueadas) cada, assim como um domínio de troca e uma alfa hélice. O sítio ativo da protease está localizado na porção N-terminal. Comparando a seqüência da NS3 de outros flavivírus podemos identificar três resíduos conservados (His-57, Asp-81, and Ser-139) que compõem o centro catalítico desta protease. O centro ativo da protease NS3 contém um sítio estabilizador de um oxiânion tetraédrico, intermediário da reação, "oxyanion/stabilization loop", e uma fita E2 e precedida por uma fita E1. A NS3 cliva a poliproteína viral entre os resíduos de aminoácidos Cys/Ser (nos sítios 4B/5A e 5A/5B), Cys/Ala (no 4A/4B), ou Thr/Ser (no 3/4A).The molecular weight of HCV NS3 protease monomer is 70 kDa. Its active structure is a dimer and the tertiary structure of the monomer has an N-terminal portion extension, 2 domains with 6 beta-barrels each, as well as an exchange domain and an alpha helix. . The active protease site is located at the N-terminal portion. By comparing the NS3 sequence of other flaviviruses we can identify three conserved residues (His-57, Asp-81, and Ser-139) that make up the catalytic center of this protease. The active center of the NS3 protease contains a tetrahedral oxyon stabilizing site, reaction intermediate, oxyanion / stabilization loop, and an E2 tape and preceded by an E1 tape. NS3 cleaves viral polyprotein between amino acid residues Cys / Ser (at sites 4B / 5A and 5A / 5B), Cys / Ala (No 4A / 4B), or Thr / Ser (No 3 / 4A).
A NS3 assume uma estrutura de dois conjuntos de fitas beta pregueadas. O primeiro localizado na porção N-terminal contém as fitas A1, B1, C1, D1, E1, e F1. O segundo barril beta é composto pelas fitas A2, B2, C2, D2, E2, e F2.The NS3 assumes a structure of two pleated beta tape sets. The first located in the N-terminal portion contains tapes A1, B1, C1, D1, E1, and F1. The second beta barrel consists of tapes A2, B2, C2, D2, E2, and F2.
Vários trabalhos têm sido relatados sobre inibidores de serina protease. Recentemente, derivados hexapeptídicos contendo uma função eletrofílica a-ceto- amida têm sido descritos como potentes inibidores da serina protease de HCV (Arasappan, A.; Njoroge, F.G.; Chan, T.-Y.; Bennett, F.; Bogen, S.L.; Chen, K.; Gu, H.; Hong1 L.; Jao, E.; Liu1 Y.-T.; Lovey, R.G.; Parekh1 T.; Pike1 R.E.; Pinto, P.; Santhanam, B.; Venkatraman, S.; Vaccaro, H.; Wang, H.; Yang, X.; Zhu, Z.; Mckittrick, B.; Saksena1 A.K.; Girijavallabhan, V.; Pichardo, J.; Butkiewicz, N.; Ingram, N.; Malcolm, B.; Prongay, A.; Yao, N.; Marten, B.; Madison, V.; Kemp, S.; Levy, O.; Lim-Wilby, M.; Tamura, S.; Ganguly, A.K. Bioorg. Med. Chem. Lett. 2005, 15, 4180-4184). Ainda dentro da classe das α-ceto-amidas, o VX-950 e o SCH503034 (Bocepravir), já mencionados anteriormente, encontram-se em desenvolvimento clínico. O VX-950 foi capaz de reduzir a carga viral plasmática de pacientes dosados com 750mg a cada 8h, e em alguns pacientes o vírus tornou-se indetectável após 14 dias de dosagem (Chen, K.X.; Vibulbhan, B.; Yang, W.; Cheng, K-C.; Liu1 R.; Pichardo, J.; Butkiewiez, N.; Njoroge, F.G. Bioorg. Med. Chem. 2008, doi:10.1016/j.bmc.2007.11.002. Perni, R.B.; Almquist, S.J.; Byrn, R.A.; Chandorkar, G.; Chaturvedi1 P.R.; Courtney1 L.F.; Decker1 C.J.; Dinehart1 K.; Gates1 C.A.; Harbeson1 S.L.; Heiser1 Α.; Kalkeri1 G.; Kolaczkowski, E.; Lin1 K.; Luong1 Y-P.; Rao1 Β.G.; Taylor, W.P.; Thomson, J.A.; Tung1 R.D.; Wei1 Y.; Kwong, A.D.; Lin1 C. Antimicrob. Agents Chemother. 2006, 50, 899-909).Several papers have been reported on serine protease inhibitors. Recently, hexapeptide derivatives containing an α-keto-amide electrophilic function have been described as potent inhibitors of HCV serine protease (Arasappan, A.; Njoroge, FG; Chan, T.-Y .; Bennett, F.; Bogen, SL Chen, K .; Gu, H.; Hong1 L.; Jao, E.; Liu1 Y.-T .; Lovey, RG; Parekh1 T.; Pike1 RE; Pinto, P .; Santhanam, B.; Venkatraman, Vaccaro H. Wang Yang H. Zhu Zhu Mckittrick B. Saksena AK Girijavallabhan V Pichardo J Butkiewicz N Ingram N ; Malcolm, B.; Prongay, A.; Yao, N.; Marten, B.; Madison, V.; Kemp, S .; Levy, O .; Lim-Wilby, M .; Tamura, S .; Ganguly, AK Bioorg, Med. Chem. Lett. 2005, 15, 4180-4184). Still within the α-keto amide class, VX-950 and SCH503034 (Bocepravir), already mentioned, are in clinical development. VX-950 was able to reduce the plasma viral load of 750mg dosed patients every 8h, and in some patients the virus became undetectable after 14 days of dosing (Chen, KX; Vibulbhan, B.; Yang, W. ; Cheng, KC .; Liu1 R .; Pichardo, J .; Butkiewiez, N.; Njoroge, FG Bioorg. Med. Chem. 2008, doi: 10.1016 / j.bmc.2007.11.002. Perni, RB; Almquist, SJ; ; Byrn, RA; Chandorkar, G .; Chaturvedi1 PR; Courtney1 LF; Decker1 CJ; Dinehart1 K .; Gates1 CA; Harbeson1 SL; Heiser1 Α .; Kolaczkowski, E .; Lin1 K .; Luong1 YP .; Taylor, WP; Thomson, JA; Tung1 RD; Wei Y; Kwong, AD; Lin1 C. Antimicrob. Agents Chemother. 2006, 50, 899-909).
Na literatura patentária foram encontrados documentos relacionados à presente invenção, porém sem infringir o escopo da mesma, sendo as mais relevantes descritas a seguir:In the patent literature, documents related to the present invention were found, but without infringing its scope, the most relevant being described below:
O documento W02008137126 descreve um método de tratamento de infecção por HCV utilizando uma combinação terapêutica compreendendo um inibidor de serina protease e um de polimerase. Não sendo um composto único como inibidor de ambas serina protease e polimerase do HCV1 e tais compostos descritos fazem parte do estado da técnica.WO2008137126 describes a method of treating HCV infection using a therapeutic combination comprising a serine protease inhibitor and a polymerase inhibitor. Not being a single compound as inhibitor of both HCV1 serine protease and polymerase and such disclosed compounds are part of the state of the art.
O documento W02007092616 descreve medicamentos, composições ou kits farmacêuticos e métodos, baseados em combinações de pelo menos um inibidor de protease de HCV, e um inibidor de polimerase de HCV, para serem administrados no tratamento de sintomas causados por HCV. Os compostos que atuam como inibidores de protease e polimerase também estão descritos.WO2007092616 describes pharmaceuticals, compositions or pharmaceutical kits and methods, based on combinations of at least one HCV protease inhibitor and one HCV polymerase inhibitor, to be administered in the treatment of symptoms caused by HCV. Compounds that act as protease and polymerase inhibitors are also described.
O documento US7105499 descreve compostos que atuam como inibidores de polimerase do HCV.US7105499 describes compounds that act as HCV polymerase inhibitors.
O documento US2007027071 descreve compostos macrocíclicos que atuam como inibidores de protease do HCV.US2007027071 describes macrocyclic compounds that act as HCV protease inhibitors.
O documento PI0401908-3, de titularidade da UFRJ1 descreve compostos inibidores de serina protease sintéticos, com um eixo de simetria C2, utilizados como base para a preparação de formulações antivirais capazes de cessar a proliferação dos vírus responsáveis pela maioria das flaviroses, como por exemplo, hepatite C1 Dengue e febre do oeste do Nilo.UFRJ1 owned document PI0401908-3 describes synthetic serine protease inhibitor compounds having a C2 axis of symmetry, used as a basis for the preparation of antiviral formulations capable of stopping the proliferation of viruses responsible for most flaviroses, such as , hepatitis C1 Dengue fever and West Nile fever.
Como pode ser observado existem diversos documentos que descrevem ou inibidores de protease ou inibidores de polimerase, ou ainda uma combinação de ambos, porém nenhum consegue descrever um composto capaz de atuar como inibidor de protease e polimerase, mais especificamente relacionado ao HCV. Sumário da InvençãoAs can be seen there are several documents describing either protease inhibitors or polymerase inhibitors, or a combination of both, but neither can describe a compound capable of acting as a protease and polymerase inhibitor, more specifically related to HCV. Summary of the Invention
O primeiro objeto da presente invenção refere-se a compostos inibidores da serina protease e potencialmente inibidores de RNA e DNA polimerases de fórmula geral (I) e (II):The first object of the present invention relates to serine protease inhibiting compounds and potentially RNA and DNA polymerase inhibitors of formula (I) and (II):
<formula>formula see original document page 8</formula><formula> formula see original document page 8 </formula>
O segundo objeto dessa invenção refere-se a uma composição farmacêutica contendo o dito composto de fórmula molecular (I),The second object of this invention relates to a pharmaceutical composition containing said compound of formula (I),
<formula>formula see original document page 8</formula><formula> formula see original document page 8 </formula>
assim como seus derivados e/ou seus sais farmaceuticamente aceitáveis, para a produção de um medicamento usados como inibidores de DNA e RNA polimerases e/ou serina proteases.as well as their derivatives and / or pharmaceutically acceptable salts thereof, for the manufacture of a medicament used as inhibitors of DNA and RNA polymerases and / or serine proteases.
O terceiro objeto dessa invenção trata-se de uma composição farmacêutica contendo o composto de fórmula molecular (I),The third object of this invention is a pharmaceutical composition containing the compound of formula (I),
<formula>formula see original document page 8</formula> assim como seus derivados e/ou seus sais farmaceuticamente aceitáveis, voltados para a produção de um medicamento usado no tratamento de doenças causadas pelos vírus da família Flaviviridae em mamíferos humanos e não humanos.<formula> formula see original document page 8 </formula> as well as derivatives and / or pharmaceutically acceptable salts thereof for the manufacture of a medicament for the treatment of diseases caused by the Flaviviridae family viruses in human and non-human mammals.
O quarto objeto dessa invenção trata-se de uma composição farmacêutica contendo o dito composto de fórmula molecular (II),The fourth object of this invention is a pharmaceutical composition containing said compound of formula (II),
<formula>formula see original document page 9</formula><formula> formula see original document page 9 </formula>
assim como seus derivados e/ou seus sais farmaceuticamente aceitáveis, usados para a produção de um medicamento usados como inibidores de DNA e RNA polimerases e/ou serina proteases.as well as their derivatives and / or pharmaceutically acceptable salts thereof used for the manufacture of a medicament used as inhibitors of DNA and RNA polymerases and / or serine proteases.
O último objeto dessa invenção trata-se de uma composição farmacêutica, contendo o dito composto de fórmula molecular (II),The last object of this invention is a pharmaceutical composition containing said compound of molecular formula (II).
<formula>formula see original document page 9</formula><formula> formula see original document page 9 </formula>
assim como seus derivados e/ou seus sais farmaceuticamente aceitáveis, voltados para a produção de um medicamento usado no tratamento de doenças causadas pelos vírus da família Flaviviridae em mamíferos humanos e não humanos.as well as derivatives and / or pharmaceutically acceptable salts thereof for the manufacture of a medicament used for the treatment of diseases caused by the Flaviviridae family viruses in human and non-human mammals.
Descrição das FigurasDescription of the Figures
A Figura 1 mostra através dos gráficos A e B os resultados da inibição da protease NS3/4A do HCV. Na avaliação do potencial inibitório do composto 22 na enzima NS3pro/4A, foram realizados ensaios de IC50% que é uma medida que indica a concentração necessária da droga para inibir 50% da atividade in vitro. Em A, está representada a curva de inibição da NS3pro/4A com o composto 22. Em B1 a curva de inibição apresenta diversos pontos que representam diversas concentrações do composto (0; 7,8; 15,6; 31,2; 62,5; 125,0; 250,0; 500,0; e 1000,ΟμΜ).Figure 1 shows through graphs A and B the results of HCV NS3 / 4A protease inhibition. In the evaluation of the inhibitory potential of compound 22 in the enzyme NS3pro / 4A, IC50% assays were performed which is a measure that indicates the concentration of the drug required to inhibit 50% of in vitro activity. In A, the inhibition curve of NS3pro / 4A with compound 22 is shown. In B1 the inhibition curve has several points representing different concentrations of the compound (0.7.8; 15.6; 31.2; 62.). 5, 125.0, 250.0, 500.0, and 1000, ΟμΜ).
A Figura 2 avalia se o composto 22 promove aumento ou redução da atividade proteolítica, realizado a partir de uma nova síntese química do composto. Um novo lote do composto 22 foi resuspendido em DMSO a 100% e os ensaios cinéticos realizados no mesmo dia. No gráfico A, cada ponto representa diversas concentrações do composto (0; 7,8; 15,6; 31,2; 62,5; 125,0; 250,0; 500,0; e 1000,0μΜ). No gráfico B, curva de inibição da NS3pro/4A com o composto 22 recém sintetizado mostra em Cada ponto a média de 4 medidas independentes de 4 replicatas em diversas concentrações (0; 0,08; 0,16; 0,63; 1,25; 2,5; 5,0 e 10,ΟμΜ) do composto 22.Figure 2 assesses whether compound 22 promotes increased or decreased proteolytic activity, performed from a new chemical synthesis of the compound. A new batch of compound 22 was resuspended in 100% DMSO and the same day kinetic assays. In graph A, each point represents various concentrations of the compound (0.7.8, 15.6, 31.2, 62.5, 125.0, 250.0, 500.0, and 1000.0μΜ). In graph B, NS3pro / 4A inhibition curve with newly synthesized compound 22 shows at each point the average of 4 independent measurements of 4 replicates at various concentrations (0; 0.08; 0.16; 0.63; 1, 25, 2.5, 5.0 and 10, ΟμΜ) of compound 22.
Descrição Detalhada da InvençãoDetailed Description of the Invention
A presente invenção refere-se aos compostos de fórmula geral (I) e (II), desenhados como inibidores de polimerases e serina proteases.The present invention relates to compounds of formula (I) and (II) designed as polymerase and serine protease inhibitors.
Estes compostos são potencialmente ativos em vírus da família flaviviridae, preferencialmente; tais compostos são ativos na inibição da serina protease do HCV e vírus da dengue.These compounds are potentially active in flaviviridae viruses, preferably; Such compounds are active in inhibiting HCV serine protease and dengue virus.
Para os compostos inibidores de polimerases e serina da presente invenção de fórmula geral (I) e (II),For the polymerase and serine inhibiting compounds of the present invention of formula (I) and (II),
<formula>formula see original document page 10</formula><formula> formula see original document page 10 </formula>
X é definido como aminoácidos protegidos com grupamentos carboxi-f-butila (-Boc) e/ou carboxibenzila (-Cbz) que serão utilizados na síntese dos pseudopeptídeos. Sendo Z igual a benzila (-Bn), carboxibenzila (-Cbz) ou carboxi-t- butila (-Boc); capazes de conferir rigidez ao inibidor, e que por sua natureza também podem atuar em polimerases, como DNA ou RNA polimerases, inibindo-as.X is defined as amino acids protected with carboxy-f-butyl (-Boc) and / or carboxybenzyl (-Cbz) groups that will be used in the synthesis of pseudopeptides. Z being benzyl (-Bn), carboxybenzyl (-Cbz) or carboxy-t-butyl (-Boc); capable of imparting rigidity to the inhibitor, and which by their nature may also act on polymerases, such as DNA or RNA polymerases, by inhibiting them.
Os ditos compostos de fórmula molecular (I) e (II), assim como seus derivados e/ou sais farmaceuticamente aceitáveis são usados em uma composição farmacêutica como inibidores de DNA e RNA polimerases e serina proteases.Said compounds of formula (I) and (II), as well as their pharmaceutically acceptable salts and / or derivatives thereof are used in a pharmaceutical composition as inhibitors of DNA and RNA polymerases and serine proteases.
Tal composição farmacêutica apresenta uma concentração que varia na faixa de 10μΜ a 150μΜ do dito composto de fórmula molecular I e/ou II, assim como seus derivados e/ou sais farmaceuticamente aceitáveis.Such pharmaceutical composition has a concentration ranging from 10μΜ to 150μΜ of said compound of molecular formula I and / or II, as well as their pharmaceutically acceptable salts and / or derivatives.
Para fins dessa invenção os excipientes são todos aqueles compostos químicos que tem como objetivo dar forma, volume, estabilidade a composição farmacêutica. Todos os excipientes que aqui serão usados são aqueles das indústrias farmacêuticas tais como: adjuvantes, edulcorantes, solventes, estabilizantes, antioxidantes, conservantes, tampões, flavorizantes, tensoativos, umectantes, lubrificantes, dispersantes e todos aqueles conhecidos por um técnico na área.For purposes of this invention the excipients are all those chemical compounds which are intended to give the pharmaceutical composition form, volume, stability. All excipients to be used herein are those from the pharmaceutical industries such as adjuvants, sweeteners, solvents, stabilizers, antioxidants, preservatives, buffers, flavorants, surfactants, humectants, lubricants, dispersants and all those known to one skilled in the art.
Os tais excipientes podem ser empregados em diferentes formas farmacêuticas, tais como pós, comprimidos, cápsulas, nanocápsulas, nanoesferas, processos de hot-melt extrusion, co-cristais, ampolas, soluções orais, adesivos transdérmicos, soluções injetáveis musculares ou parenterais, pomadas, cremes, géis, suspensões, emulsões, aerosóis, tinturas, elexires, pastas, pastilhas, supositórios, ugüentos, óvulos, e todas aquelas conhecidas por um técnico da arte.Such excipients may be employed in different pharmaceutical forms such as powders, tablets, capsules, nanocapsules, nanospheres, hot melt extrusion processes, co-crystals, ampoules, oral solutions, transdermal patches, muscle or parenteral injectable solutions, ointments, creams, gels, suspensions, emulsions, aerosols, dyes, elexirs, pastes, lozenges, suppositories, urges, eggs, and all those known to a person skilled in the art.
Tal composição farmacêutica que contém os compostos (I) e/ou (II), assim como seus derivados e sais farmaceuticamente aceitáveis, é empregada para a produção de um medicamento no tratamento de doenças causadas pelos vírus da família Flaviviridae, compreendendo os vírus dessa família os vírus da hepatite C, da dengue, da febre amarela, da febre do oeste ou febre do Nilo, sendo preferencialmente empregado no tratamento de doenças provocadas pelo vírus da dengue e da hepatite C. No caso da hepatite C estão compreendidos os seguintes vírus A, B, C, D, E, G e TT.Such a pharmaceutical composition containing the compounds (I) and / or (II), as well as their pharmaceutically acceptable salts and derivatives, is employed for the manufacture of a medicament in the treatment of diseases caused by Flaviviridae viruses, comprising viruses of this family. Hepatitis C, Dengue, Yellow Fever, West Fever or Nile Fever viruses, and is preferably used for the treatment of diseases caused by Dengue and Hepatitis C virus. , B, C, D, E, G and TT.
Compostos de fórmula geral (I) e (II) foram produzidos a partir do isomanídeo como análogos dos previamente publicados pelo grupo, com a analogia adicional a pró-nucleosídios, isto é, compostos capazes de inibir polimerases. Sua analogia estrutural com dipeptídeos rígidos cíclicos o faz um excelente chassi, cerne rígido, "scaffold". Além disto, pelo fato de apresentar um eixo C2 de simetria, pode ser homologado, em duas ou em uma posição, mantendo ou não o eixo de simetria, via reação com diferentes aminoácidos e diidroamino ácidos, por exemplo, via acoplamento direto catalisado por EDC, HOBt, e aminoácidos.Compounds of formula (I) and (II) were produced from the isomanide as analogues to those previously published by the group, with the additional analogy to pro-nucleosides, that is, compounds capable of inhibiting polymerases. Its structural analogy with cyclic rigid dipeptides makes it an excellent chassis, rigid core, "scaffold". Moreover, because it has a C2 axis of symmetry, it can be homologated, in two or in one position, maintaining or not the symmetry axis, via reaction with different amino acids and dihydroamino acids, for example, via EDC-catalyzed direct coupling. , HOBt, and amino acids.
A partir dos dados obtidos anteriormente passou-se, então, a etapa de síntese dos inibidores da serina protease previstos.From the data obtained above, the synthesis step of the predicted serine protease inhibitors was then passed.
A primeira etapa na obtenção dos derivados de fórmula geral I consistiu na conversão do isomanídeo (1) no derivado mono-tosilado (2) por reação com cloreto de p-toluenosulfonila em piridina por 12h a temperatura ambiente. O tratamento do mono-tosilado (2) com cloreto de benzila em meio básico usando TBAB como catalisador de transferência de fases forneceu o derivado benzilado (3). Este reagiu com azida de sódio em [bmim]BF4 a 120°C por 12h forneceu a azida (4), com inversão de configuração. Redução da azida (4) com paládio sobre carbono em etanol por 12h utilizando uma pressão de 40psi resultou na amina (5), que foi o substrato para a formação de novas amidas derivadas do isomanideo (10-22) (Esquema 1).The first step in obtaining the derivatives of formula I was the conversion of isomanide (1) to the mono-tosylated derivative (2) by reaction with p-toluenesulfonyl chloride in pyridine for 12h at room temperature. Treatment of mono-tosylate (2) with benzyl chloride in basic medium using TBAB as phase transfer catalyst provided the benzylated derivative (3). This reacted with sodium azide in [bmim] BF4 at 120 ° C for 12h provided azide (4) with reversal of configuration. Reduction of palladium azide (4) on carbon in ethanol for 12h using a pressure of 40psi resulted in amine (5), which was the substrate for the formation of new isomanide-derived amides (10-22) (Scheme 1).
Esquema 1. Síntese dos compostos intermediários para a obtenção da amina (5).Scheme 1. Synthesis of intermediate compounds to obtain the amine (5).
<formula>formula see original document page 12</formula> Diferentes solventes podem ser usados na etapa de substituição nucleofílica, tais como, DMF e DMSO. Porém, o [bmim]BF4, líquido iônico, apresenta muitas vantagens em relação aos solventes orgânicos convencionais, além de ter apresentado um melhor rendimento reacional. Os líquidos iônicos não possuem pressão de vapor mensurável à temperatura ambiente, e mesmo a temperaturas bastante elevadas degradam apenas acima de 400°C, podendo ser usados em vácuo sem que haja perda de solvente.<formula> formula see original document page 12 </formula> Different solvents may be used in the nucleophilic substitution step such as DMF and DMSO. However, [bmim] BF4, ionic liquid, has many advantages over conventional organic solvents, besides having a better reaction yield. Ionic liquids have no measurable vapor pressure at room temperature, and even at very high temperatures they only degrade above 400 ° C and can be used in a vacuum without solvent loss.
O líquido iônico, [bmim]BF4 (9) foi sintetizado a partir do N-metil imidazol (7). Este reagiu com o 1-bromobutano formando o derivado (8) que, por troca iônica com NaBF4, originou (7). O líquido iônico foi purificado por coluna cromatográfica em sílica gel tendo como eluente o diclorometano, sendo obtido em 83% de rendimento como mostrado no esquema 2.The [bmim] BF4 (9) ionic liquid was synthesized from N-methyl imidazole (7). This reacted with 1-bromobutane forming derivative (8) which upon ion exchange with NaBF4 gave rise to (7). The ionic liquid was purified by silica gel column chromatography eluting with dichloromethane and was obtained in 83% yield as shown in scheme 2.
Esquema 2. Síntese do liquido iônico [Bmim]+[BF4]"Scheme 2. Ionic Liquid Synthesis [Bmim] + [BF4] "
<formula>formula see original document page 13</formula><formula> formula see original document page 13 </formula>
A síntese dos inibidores da serina protease previstos pela fórmula geral II iniciou-se pela mono benzilação do isomanídeo usando cloreto de benzila em meio básico formando o composto monobenzilado (6).The synthesis of the serine protease inhibitors provided for by formula II began by mono benzylation of the isomanide using benzyl chloride in basic medium forming the monobenzylated compound (6).
Esquema 3. Síntese do derivado mono benzilado (6).Scheme 3. Synthesis of the mono benzylated derivative (6).
<formula>formula see original document page 13</formula> A última etapa para obtenção dos compostos de fórmula geral I e Il consistiu na reação de acoplamento da amina O-benzilada (5) e do mono benzil isomanídeo (6) com diversos amino ácidos (D- ou L-) contendo grupos de proteção N-BOC e/ou N-Cbz usando DCC como reagente de acoplamento e DMAP como catalisador em diclorometano (Esquema 4). A estrutura dos amino ácidos N-protegidos bem como os rendimentos das reações é mostrada na tabela 1.<formula> formula see original document page 13 </formula> The last step to obtain the compounds of formula I and II was the coupling reaction of O-benzyl amine (5) and mono benzyl isomanide (6) with various amino acids. (D- or L-) acids containing N-BOC and / or N-Cbz protecting groups using DCC as coupling reagent and DMAP as catalyst in dichloromethane (Scheme 4). The structure of the N-protected amino acids as well as the reaction yields are shown in table 1.
Esquema 4. Síntese dos compostos (10-22) e (23-31)Scheme 4. Synthesis of compounds (10-22) and (23-31)
<formula>formula see original document page 14</formula><formula> formula see original document page 14 </formula>
Tabela 1- Estruturas e rendimentos dos produtos (10-22) e (23-31)Table 1- Structures and yields of products (10-22) and (23-31)
<table>table see original document page 14</column></row><table> <table>table see original document page 15</column></row><table> A presente invenção será descrita detalhadamente através dos exemplos apresentados abaixo. É necessário frisar que a invenção não está limitada a esses exemplos e inclui variações e modificações dentro dos limites nos quais ela se aplica.<table> table see original document page 14 </column> </row> <table> <table> table see original document page 15 </column> </row> <table> The present invention will be described in detail through the examples presented. below, down, beneath, underneath, downwards, downhill. It should be noted that the invention is not limited to these examples and includes variations and modifications within the limits to which it applies.
Exemplo 1: Ensaio de atividade e inibição da protease recombinante NS3pro/4A do HCV.Example 1: Test of activity and inhibition of HCV NS3pro / 4A recombinant protease.
A avaliação da atividade proteolítica do HCV foi realizada com o kit SensoLyte®520 HCV Protease Assay Kit *Fluorimetric* (AnaSpec1CA1USA). Este ensaio possibilita realizar a triagem de inibidores anti-HCV-NS3pro/4A com maior eficiência e sensibilidade, baseado no fenômeno de FRET (fluorecence ressonance energy transfer), no qual o peptídeo derivado do sítio de clivagem NS4A/NS4B encontra-se marcado com fluoróforo 5-FAM/QXL™520, sendo a fluorescência do 5- FAM suprimida pela QXL™520. A quantificação da atividade proteolítica ocorre quando o peptídeo FRET sofre a clivagem em dois fragmentos pela NS3pro/4A, com isso, a fluorescência do 5-FAM pode ser monitorada em excitação/emissão igual a 490nm/520nm. A grande vantagem na utilização do peptídeo marcado com 5-FAM, é que a emissão de fluorescência ocorre em longo comprimento de onda, sofrendo menos interferência da autofluorescência dos compostos testados.HCV proteolytic activity was evaluated with the SensoLyte®520 HCV Protease Assay Kit * Fluorimetric * (AnaSpec1CA1USA). This assay makes it possible to screen anti-HCV-NS3pro / 4A inhibitors with greater efficiency and sensitivity, based on the FRET (fluorecence resonance energy transfer) phenomenon, in which the peptide derived from the NS4A / NS4B cleavage site is labeled with 5-FAM / QXL ™ 520 fluorophore, with 5 FAM fluorescence suppressed by QXL ™ 520. Quantification of proteolytic activity occurs when the FRET peptide is cleaved into two fragments by NS3pro / 4A, so 5-FAM fluorescence can be monitored at 490nm / 520nm excitation / emission. The great advantage of using the 5-FAM-labeled peptide is that the fluorescence emission occurs at long wavelength, suffering less interference from the autofluorescence of the tested compounds.
Exemplo 2: Metodologia utilizada na dosagem da capacidade inibitória dos compostos inibidores de polimerases e serina proteasesExample 2: Methodology used for dosing inhibitory capacity of polymerase and serine protease inhibitor compounds
Os compostos Iiofilizados foram resuspendidos em DMSO (100%) a uma concentração de 20mM . Posteriormente, 100μΜ dos compostos foram pré- incubados a 25°C por 15 min com 10ng da enzima HCV-NS3pro/4A em volume final de 50μΙ_ do tampão de reação do kit SensoLyte®520 HCV Protease Assay Kit *Fluorimetric* (AnaSpec1CA1USA), seguindo o protocolo do fabricante. A reação foi iniciada com a adição do substrato Peptídeo 5-FAM/QXL™520 FRET, cuja clivagem do peptídeo foi monitorada no leitor de placas SpectraMax M2e (Molecular Devices) em excitação/emissão igual a 490nm/520nm medindo a fluorescência gerada a cada 30 segundos por 60 min.The lyophilized compounds were resuspended in DMSO (100%) at a concentration of 20mM. Subsequently, 100μΜ of the compounds were preincubated at 25 ° C for 15 min with 10ng of the enzyme HCV-NS3pro / 4A in a final volume of 50μΙ_ of the SensoLyte®520 HCV Protease Assay Kit * Fluorimetric * (AnaSpec1CA1USA) reaction buffer, following the manufacturer's protocol. The reaction was initiated by the addition of 5-FAM / QXL ™ 520 FRET Peptide substrate, whose peptide cleavage was monitored on the SpectraMax M2e plate reader (Molecular Devices) at excitation / emission equal to 490nm / 520nm by measuring the fluorescence generated at each 30 seconds for 60 min.
Os parâmetros cinéticos de velocidade inicial (Vi) das reações foram calculados por regressões linear e/ou polinomial de 3a ordem utilizando-se o programa SigmaPlot versão 10.0 e Excel 2007, respectivamente, sendo os valores de Vi convertidos em atividade residual (%).The initial velocity (Vi) reaction kinetic parameters were calculated by 3rd order linear and / or polynomial regressions using the SigmaPlot version 10.0 and Excel 2007 software, respectively, and the Vi values converted to residual activity (%).
O teste cinético de 22 compostos (10 a 31) está mostrado na Tabela 3. Podemos verificar que os compostos 19, 20, 21, 22, e 23 tiveram atividade inibitória para a protease NS3/4A do HCV, sendo que o composto 22 foi o mais ativo, com uma capacidade inibitória de 64%.The kinetic test of 22 compounds (10 to 31) is shown in Table 3. We can verify that compounds 19, 20, 21, 22, and 23 had inhibitory activity for HCV NS3 / 4A protease, and compound 22 was the most active, with an inhibitory capacity of 64%.
A inibição parcial da NS3pro/4A em altas concentrações do composto 22, pode ser explicada devido a ausência do domínio helicase da NS3 e/ou do complexo replicativo (ensaio in vitro). Eles podem estar contribuindo para a correta orientação da molécula e/ou acentuando a interação do composto 22 com a protease NS3/4A (ensaio in vivo). Na Figura 1A ocorreu uma completa inibição da NS3pro/4A com o inibidor padrão (Ι05ο%=0,80±0,37μΜ). Entretanto, o composto 22 em altas concentrações inibiu parcialmente (-45%) a atividade proteolítica da NS3/4A.Partial inhibition of NS3pro / 4A at high concentrations of compound 22 can be explained due to the absence of the NS3 helicase domain and / or replicative complex (in vitro assay). They may be contributing to the correct orientation of the molecule and / or enhancing the interaction of compound 22 with the NS3 / 4A protease (in vivo assay). In Figure 1A a complete inhibition of NS3pro / 4A occurred with the standard inhibitor (Ι05ο% = 0.80 ± 0.37μΜ). However, compound 22 at high concentrations partially inhibited (-45%) the proteolytic activity of NS3 / 4A.
A inibição parcial da NS3pro/4A em altas concentrações do composto 22, pode ser explicada devido a ausência do domínio helicase da NS3 e/ou do complexo replicativo (ensaio in vitro). Eles podem estar contribuindo para a correta orientação da molécula e/ou acentuando a interação do composto 22 com a protease NS3/4A (ensaio in vivo). Contudo, quando realizamos os ensaios in vitro com o composto 22 armazenado a -20°C não observamos essa ativação (Figura 1A). Esses dados sugerem que o composto 22 não seja estável quando armazenado a 4°C, e que seu produto aumente a atividade proteolítica da NS3/4A.Partial inhibition of NS3pro / 4A at high concentrations of compound 22 can be explained due to the absence of the NS3 helicase domain and / or replicative complex (in vitro assay). They may be contributing to the correct orientation of the molecule and / or enhancing the interaction of compound 22 with the NS3 / 4A protease (in vivo assay). However, when we performed in vitro assays with compound 22 stored at -20 ° C we did not observe this activation (Figure 1A). These data suggest that compound 22 is not stable when stored at 4 ° C, and that its product increases the proteolytic activity of NS3 / 4A.
É importante ressaltar que quando iniciamos a avaliar, in vitro, a atividade inibitória do composto 22 armazenada a 4°C, observamos que o mesmo em baixas concentrações, aumentava a atividade proteolítica da NS3/4A em 2,5X(Figura 2A). Contudo, quando realizamos os ensaios in vitro com o composto 22 armazenado a - 20°C não observamos essa ativação (Figura 1A). Esses dados sugerem que o composto 22 não seja estável quando armazenado a 4°C, e que seu produto aumente a atividade proteolítica da NS3/4A.Importantly, when we began to evaluate, in vitro, the inhibitory activity of compound 22 stored at 4 ° C, we observed that it at low concentrations increased the proteolytic activity of NS3 / 4A by 2.5X (Figure 2A). However, when we performed in vitro assays with compound 22 stored at -20 ° C we did not observe this activation (Figure 1A). These data suggest that compound 22 is not stable when stored at 4 ° C, and that its product increases the proteolytic activity of NS3 / 4A.
Com o objetivo de avaliar se o composto 22 promove aumento ou redução da 30 atividade proteolítica, foi realizada uma nova síntese química do composto. Esse novo lote do composto 22 foi resuspendido em DMSO a 100% e os ensaios cinéticos realizados no mesmo dia. Nossos dados revelam que em baixas concentrações do composto 22 (recém sintetizado) a atividade proteolítica da NS3/4A aumentou aproximadamente 15% (Figura 2B).In order to evaluate whether compound 22 promotes increase or decrease in proteolytic activity, a new chemical synthesis of the compound was performed. This new batch of compound 22 was resuspended in 100% DMSO and the same day kinetic assays. Our data reveal that at low concentrations of newly synthesized compound 22, the proteolytic activity of NS3 / 4A increased approximately 15% (Figure 2B).
Tabela 3 - Atividade inibitória dos 22 compostos da patente frente à protease NS3/4A do HCV numa reação in vitro com o kit SensoLyte® 520 HCV ProteaseTable 3 - Inhibitory activity of 22 compounds of the patent against HCV NS3 / 4A protease in an in vitro reaction with SensoLyte® 520 HCV Protease Kit
Assay Kit (Anaspec, CA, USA).Assay Kit (Anaspec, CA, USA).
<table>table see original document page 18</column></row><table><table> table see original document page 18 </column> </row> <table>
Exemplo 3: Obtenção do 1,4:3,6-dianidro-mono(4-metillbenzenosulfonato)-D- manitol (2)Example 3: Obtaining 1,4: 3,6-dianhydro-mono (4-methylbenzenesulfonate) -D-mannitol (2)
A uma solução do isomanideo (1g, 6mmol) e piridina (1,1 OmL) em CH2CI2 (10mL) foi adicionado cloreto de p-toluenosulfonila (1,49g, 7mmol) a 0°C. A reação foi agitada a temperatura ambiente por 12h e após foi diluída com CH2CI2 (20mL). A solução foi lavada com água, HCI 1N e solução saturada de cloreto de sódio. Após evaporação do solvente, o sólido foi recristalizado com uma mistura de metanol/isopropanol fornecendo o produto como um sólido branco em 40% de rendimento. PF: 103-104°C.To a solution of isomanide (1g, 6mmol) and pyridine (1.1 OmL) in CH 2 Cl 2 (10mL) was added p-toluenesulfonyl chloride (1.49g, 7mmol) at 0 ° C. The reaction was stirred at room temperature for 12h and then diluted with CH 2 Cl 2 (20mL). The solution was washed with water, 1N HCl and saturated sodium chloride solution. After evaporation of the solvent, the solid was recrystallized with a methanol / isopropanol mixture providing the product as a white solid in 40% yield. MP: 103-104 ° C.
RMN1H δ ppm (CDCI3, 300MHz): 7,84 (d, 2H, J = 8,1 Hz); 7,34 (d, 2H, J = 8,1 Hz); 4,91 (dd, 1H, J = 6,6/ 5,5 Hz); 4,42 (t, 1H, J = 5,1 Hz); 4,49 (t, 1H, J = 4,8 Hz); 4,29- 4,25 (m, 1H); 4,04-3,95 (m, 2H); 3,79 (t, 1H, J = 7,8 Hz); 3,55 (dd, 1H, J = 7,2/1,8 Hz); 2,46 (s, 3H).1 H NMR δ ppm (CDCl 3, 300MHz): 7.84 (d, 2H, J = 8.1 Hz); 7.34 (d, 2H, J = 8.1 Hz); 4.91 (dd, 1H, J = 6.6 / 5.5 Hz); 4.42 (t, 1H, J = 5.1 Hz); 4.49 (t, 1H, J = 4.8 Hz); 4.29-4.25 (m, 1H); 4.04-3.95 (m, 2H); 3.79 (t, 1H, J = 7.8 Hz); 3.55 (dd, 1H, J = 7.2 / 1.8 Hz); 2.46 (s, 3H).
IV ν cm"1(KBr): 3526, 2933, 2866, 1596, 1359, 1189, 1173, 1050, 1019, 818, 663. Exemplo 4: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-(4- metilbenzenosulfonato)-D-manitol (3)IR ν cm -1 (KBr): 3526, 2933, 2866, 1596, 1359, 1189, 1173, 1050, 1019, 818, 663. Example 4: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2- (4-methylbenzenesulfonate) -D-mannitol (3)
A uma solução de (2) (1 Og1 33mmol), solução aquosa de KOH 50% e TBAB (0,322g, 1mmol) em CH2CI2 (80mL), foi adicionado cloreto de benzila (6,54g, 38mmol). Após agitação por 12h, a mistura foi diluída com água (10mL) e a fase orgânica foi separada. A fase aquosa foi extraída com CH2CI2 e seca com Na2SO4. O produto foi purificado por coluna cromatográfica em sílica gel tendo como eluente uma mistura de hexano e acetato de etila, fornecendo o produto como um óleo incolor em 75% de rendimento.To a solution of (2) (1 Og1 33mmol), 50% aqueous KOH and TBAB (0.322g, 1mmol) in CH2 Cl2 (80mL) was added benzyl chloride (6.54g, 38mmol). After stirring for 12h, the mixture was diluted with water (10mL) and the organic phase was separated. The aqueous phase was extracted with CH 2 Cl 2 and dried with Na 2 SO 4. The product was purified by silica gel column chromatography eluting with a mixture of hexane and ethyl acetate, providing the product as a colorless oil in 75% yield.
RMN1H δ ppm (CDCI3, 300MHz): 7,81 (d, 2H, J = 8,4 Hz); 7,35-7,32 (m, 7H); 4,91- 4,89 (m, 1H); 4,69 (d, 1H, J= 12,0 Hz); 4,50 (d, 1H, J = 12,0 Hz); 4,47 (t, 2H, J = 2,1 Hz); 4,04-3,79 (m, 4H); 3,62 (t, 1H, J= 8,4 Hz); 2,44 (s, 3H).1 H NMR δ ppm (CDCl 3, 300MHz): 7.81 (d, 2H, J = 8.4 Hz); 7.35-7.32 (m, 7H); 4.91-4.89 (m, 1H); 4.69 (d, 1H, J = 12.0 Hz); 4.50 (d, 1H, J = 12.0 Hz); 4.47 (t, 2H, J = 2.1 Hz); 4.04-3.79 (m, 4H); 3.62 (t, 1H, J = 8.4 Hz); 2.44 (s, 3H).
IVv cm"1 (filme): 3063, 2977, 2950, 2879, 1598, 1454, 1366, 1190, 1178, 1141, 1027, 853.IRv cm -1 (film): 3063, 2977, 2950, 2879, 1598, 1454, 1366, 1190, 1178, 1141, 1027, 853.
ao20 = +98 (c, 0.1) DMSO.α 20 = + 98 (c, 0.1) DMSO.
Exemplo 5: Obtenção do 1,4:3,6-dianidro-2-azido-2-deoxi-5-0-(benzil)-D- Glucitol (4)Example 5: Obtaining 1,4: 3,6-Dianhydro-2-azido-2-deoxy-5-0- (benzyl) -D-Glucitol (4)
Uma mistura de (3) (1,86g, 4mmol), [bmim]+[BF4]" previamente seco sob vácuo a 90°C (4,8mL, 23mmol) e NaN3 (0,93g, 14mmol) foi aquecida a 120°C por 12h. Adicionou-se água e a fase aquosa foi extraída com éter etílico. O produto foi purificado por coluna cromatográfica flash (hexano/acetato de etila), obtendo um óleo amarelo claro em 73% de rendimento.A mixture of (3) (1.86g, 4mmol), [bmin] + [BF4] "previously dried under vacuum at 90 ° C (4.8mL, 23mmol) and NaN3 (0.93g, 14mmol) was heated to 120 ° C. Water was added and the aqueous phase was extracted with ethyl ether The product was purified by flash column chromatography (hexane / ethyl acetate) to give a pale yellow oil in 73% yield.
RMN1H δ ppm (CDCI3, 300MHz): 7,36-7,26 (m, 5H); 4,75 (d, 1H, J = 11,7 Hz); 4,66 (t, 1H, J = 4,5 Hz); 4,54 (d, 1H, J = 11,7 Hz); 4,48 (d, 1H, J = 4,2 Hz); 4,15-4,00 (m, 4H); 3,86 (dd, 1H,J = 6,3/2,7 Hz); 3,66 (t, 1H, J = 7,8 Hz).1 H NMR δ ppm (CDCl 3, 300MHz): 7.36-7.26 (m, 5H); 4.75 (d, 1H, J = 11.7 Hz); 4.66 (t, 1H, J = 4.5 Hz); 4.54 (d, 1H, J = 11.7 Hz); 4.48 (d, 1H, J = 4.2 Hz); 4.15-4.00 (m, 4H); 3.86 (dd, 1H, J = 6.3 / 2.7 Hz); 3.66 (t, 1H, J = 7.8 Hz).
RMN 13C δ ppm (CDCI3, 75,4 MHz): 137,4 (-C), 128,3 (-CH), 127,8 (-CH), 127,7 (- CH), 86,4 (-CH), 80,5 (-CH), 78,8 (-CH), 72,6 (-CH2), 72,4 (-CH2), 70,7 (-CH2), 66,2 (- CH).13 C-NMR δ ppm (CDCl 3, 75.4 MHz): 137.4 (-C), 128.3 (-CH), 127.8 (-CH), 127.7 (-CH), 86.4 (- CH), 80.5 (-CH), 78.8 (-CH), 72.6 (-CH2), 72.4 (-CH2), 70.7 (-CH2), 66.2 (-CH) .
IV ν cm-1 (filme): 3063, 3031, 2946, 2878, 2102, 1455, 1320, 1256, 1135, 1100, 1083,1021,739.IR ν cm -1 (film): 3063, 3031, 2946, 2878, 2102, 1455, 1320, 1256, 1135, 1100, 1083,1021,739.
HRMS calculada para C13H15N3O3Na, 284,1011; encontrada, 284,1025. oco20 = +92 (c, 0.1) DMSO.HRMS calcd for C 13 H 15 N 3 O 3 Na, 284.1011; found 284.1025. hollow20 = + 92 ° (c, 0.1) DMSO.
Exemplo 6: Obtenção do 1,4:3,6-dianidro-2-amino-2-deoxi-5-0-(benzil)-D- Glucitol (5)Example 6: Obtaining 1,4: 3,6-Dianhydro-2-amino-2-deoxy-5-0- (benzyl) -D-Glucitol (5)
A uma solução de (4) (0,870g, 3mmol) em etanol (25ml_) foi adicionado Pd/C 5% (0,3mmol) e a mistura foi hidrogenada sob 40psi for 5h. Após esse tempo a suspensão foi filtrada em XAD-4 e lavada com etanol. Evaporado o solvente obteve- se um óleo incolor em rendimento quantitativo.To a solution of (4) (0.870g, 3mmol) in ethanol (25ml) was added 5% Pd / C (0.3mmol) and the mixture was hydrogenated under 40psi for 5h. After this time the suspension was filtered on XAD-4 and washed with ethanol. Evaporating the solvent gave a colorless oil in quantitative yield.
RMN1H δ ppm (CDCI3, 300MHz): 7,37-7,35 (m, 5H); 4,76 (d, 1H, J= 11,4 Hz); 4,68 (t, 1H, J = 4,2 Hz); 4,54 (d, 1H, J = 12,0 Hz); 4,27 (d, 1H, J = 4,2 Hz); 4,06-4,02 (m, 1H); 3,85 (dd, 1H, J = 6,3/2,4 Hz); 3,76 (d, 1H, J = 9,3 Hz); 3,64 (t, 1H, J = 7,8 Hz); 3,53 (d, IH1J = 4,2 Hz); 3,45 (s, 1H).1 H NMR δ ppm (CDCl 3, 300MHz): 7.37-7.35 (m, 5H); 4.76 (d, 1H, J = 11.4 Hz); 4.68 (t, 1H, J = 4.2 Hz); 4.54 (d, 1H, J = 12.0 Hz); 4.27 (d, 1H, J = 4.2 Hz); 4.06-4.02 (m, 1H); 3.85 (dd, 1H, J = 6.3 / 2.4 Hz); 3.76 (d, 1H, J = 9.3 Hz); 3.64 (t, 1H, J = 7.8 Hz); 3.53 (d, 1H H = 4.2 Hz); 3.45 (s, 1H).
RMN 13C δ ppm (CDCI3l 75,4 MHz): 137,9 (-C), 128,6 (-CH), 128,0 (-CH), 127,8 (- CH), 90,0 (-CH), 80,1 (-CH), 79,4 (-CH), 76,3 (-CH2), 72,6 (-CH2), 70,5 (-CH2), 58,8 (- CH).13 C-NMR δ ppm (CDCl3 75.4 MHz): 137.9 (-C), 128.6 (-CH), 128.0 (-CH), 127.8 (-CH), 90.0 (-CH ), 80.1 (-CH), 79.4 (-CH), 76.3 (-CH2), 72.6 (-CH2), 70.5 (-CH2), 58.8 (-CH).
IV ν cm-1 (filme): 3360, 2876, 1605, 1455, 1369, 1209, 1065, 1017, 751, 700. HRMS calculada para C13H17NO3Na, 236,1287; encontrada, 236,1284.IR ν cm -1 (film): 3360, 2876, 1605, 1455, 1369, 1209, 1065, 1017, 751, 700. HRMS calculated for C 13 H 17 NO 3 Na, 236.1287; found, 236.1284.
Exemplo 7: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-(hidroxi)-D-manitol (6) Uma mistura de isomanideo (1g, 6,85 mmol), KOH (0,44g, 6,85 mmol) e água (3,5 mL) foi refluxada por 20min. Após esse tempo a reação foi resfriada a temperatura ambiente e foi adicionado cloreto de benzila (0,78 mL, 6,85 mmol). A mistura foi refluxada por 3h. Ao final desse tempo, foi adicionado HCI 2N (4 mL) e o produto foi extraído com acetato de etila. O produto foi obtido em 45% de rendimento após purificação por coluna cromatográfica. Pf = 93°C. RMN1H (300MHz, CDCI3): 7,36-7,27 (m, 5H); 4,79 (d, 1H, J = 12,0 Hz); 4,58 (d, 1H, J = 12,0 Hz); 4,57-4,53 (m, 1H); 4,48 (t, 1H, J = 5,1 Hz); 4,27 (br s, 1H); 4,11-3,95 (m, 3H); 3,76-3,68 (m, 2H); 2,82 (br s, 1H). RMN13C (50 MHz, CDCI3): 137,5; 128,4; 127,9; 127,8; 81,7; 80,6; 78,9; 74,7; 72,5; 72,2; 71,3.Example 7: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2- (hydroxy) -D-mannitol (6) A mixture of isomanoid (1g, 6.85 mmol), KOH ( 0.44g, 6.85 mmol) and water (3.5 mL) was refluxed for 20min. After this time the reaction was cooled to room temperature and benzyl chloride (0.78 mL, 6.85 mmol) was added. The mixture was refluxed for 3h. At the end of this time, 2N HCl (4 mL) was added and the product was extracted with ethyl acetate. The product was obtained in 45% yield after purification by column chromatography. Mp = 93 ° C. 1 H NMR (300MHz, CDCl 3): 7.36-7.27 (m, 5H); 4.79 (d, 1H, J = 12.0 Hz); 4.58 (d, 1H, J = 12.0 Hz); 4.57-4.53 (m, 1H); 4.48 (t, 1H, J = 5.1 Hz); 4.27 (br s, 1H); 4.11-3.95 (m, 3H); 3.76-3.68 (m, 2H); 2.82 (br s, 1H). 13 C-NMR (50 MHz, CDCl 3): 137.5; 128.4; 127.9; 127.8; 81.7; 80.6; 78.9; 74.7; 72.5; 72.2; 71.3.
IV ν (Cm"1) (KBr): 3459, 3404, 2920, 2865, 1459, 1419, 1329, 1266, 1200, 1118, 1068, 1021, 820.IR ν (Cm -1) (KBr): 3459, 3404, 2920, 2865, 1459, 1419, 1329, 1266, 1200, 1118, 1068, 1021, 820.
Exemplo 8: Obtenção do Líquido lônico [Bmim]+BF4' (composto 9).Example 8: Obtaining Lonic Liquid [Bmim] + BF4 '(compound 9).
A uma solução de N-metil imidazol (200mmol, 16mL) em tolueno (25mL), a 0°C, foi adicionado 1-Bromobutano (230mmol, 25mL) lentamente e com agitação. A mistura foi deixada em refluxo (110oC) por 36 h. Foram formadas duas fases: a mais densa (viscosa) e a mais leve. Elas foram separadas. A fase viscosa foi lavada com éter etílico (3 x 50mL) e acetato de etila (3x50mL). Logo após, foi adicionado CH2Cb (150mL) e NaBF4 (164mmol, 18g). A mistura ficou agitando por 24h à temperatura ambiente. Após este tempo, a mistura foi filtrada 2 vezes, adicionado o carvão ativo e deixada por mais 24h sob agitação. Ao final, a mistura foi filtrada, evaporada e por 25 fim filtrada em alumina neutra lavando bem com CH2Cl2. O produto foi obtido como um óleo amarelo claro em 83% de rendimento.To a solution of N-methyl imidazole (200mmol, 16mL) in toluene (25mL) at 0 ° C was added 1-Bromobutane (230mmol, 25mL) slowly and with stirring. The mixture was allowed to reflux (110 ° C) for 36 h. Two phases were formed: the densest (viscous) and the lightest. They were separated. The viscous phase was washed with ethyl ether (3 x 50mL) and ethyl acetate (3x50mL). Soon after, CH2Cb (150mL) and NaBF4 (164mmol, 18g) were added. The mixture was stirred for 24h at room temperature. After this time, the mixture was filtered 2 times, the activated charcoal added and left for a further 24h under stirring. At the end, the mixture was filtered, evaporated and finally filtered on neutral alumina by washing well with CH 2 Cl 2. The product was obtained as a light yellow oil in 83% yield.
RMN 1H δ ppm (CDCl3, 300 MHz): 9,27 (s, 1H); 7,38 (s, 1H); 7,32 (s, 1H); 4,22 (t, 2H, J = 7,2 Hz); 3,99 (s, 3H); 1,91-1,80 (m, 2H); 1,43-1,30 (m, 2H); 0,94 (t, 3H, J = 7,2 Hz).1 H NMR δ ppm (CDCl 3, 300 MHz): 9.27 (s, 1H); 7.38 (s, 1H); 7.32 (s, 1H); 4.22 (t, 2H, J = 7.2 Hz); 3.99 (s, 3H); 1.91-1.80 (m, 2H); 1.43-1.30 (m, 2H); 0.94 (t, 3H, J = 7.2 Hz).
IV ν cm-1(KBr): 3606, 3418, 3161, 3119, 2964, 2939, 2877, 1634, 1574, 1467, 1385, 1338, 1286, 1170, 1062, 848, 754, 623. Exemplo 9: Processo geral para obtenção das amidas (10-22) e ésteres (23-31)IR ν cm -1 (KBr): 3606, 3418, 3161, 3119, 2964, 2939, 2877, 1634, 1574, 1467, 1385, 1338, 1286, 1170, 1062, 848, 754, 623. Example 9: General Procedure for obtaining amides (10-22) and esters (23-31)
A uma solução da amina benzilada (5) ou isomanideo mono benzilado (6) (1,27 mmol) em CH2CI2 (10 mL) foram adicionados o respectivo aminoácido N- protegido (1,27 mmol), e a O0C1 DCC (1,27 mmol) e DMAP (0,12 mmol). Após agitação por 12h a temperatura ambiente, a mistura foi filtrada e o filtrado foi seco em vácuo. O produto foi purificado por coluna cromatográfica usando hexano e acetato de etila como solvente.To a solution of the benzylated amine (5) or mono benzylated isomanoid (6) (1.27 mmol) in CH 2 Cl 2 (10 mL) was added the respective N-protected amino acid (1.27 mmol), and to OCOCl DCC (1, 27 mmol) and DMAP (0.12 mmol). After stirring for 12h at room temperature, the mixture was filtered and the filtrate was dried in vacuo. The product was purified by column chromatography using hexane and ethyl acetate as solvent.
Exemplo 10: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(N-í-butoxicarbonil- L-prolina)-D-glucitol (10)Example 10: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-N- (N-t-butoxycarbonyl-L-proline) -D-glucitol (10)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,40-7,25 (m, 5H); 4,77-4,74 (m, 1H); 4,66 (t, 1H, j = 4,8 Hz); 4,57 (d, 1H, J = 11,4 Hz); 4,41-4,40 (m, 1H); 4,25-4,24 (m, 1H); 4,16- 4,12 (m, 2H); 4,10-4,01 (m, 1H); 3,87-3,82 (m, 1H); 2,21-1,92 (m, 2H); 1,92-1,86 (m, 2H); 1,41 (s, 9H).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.40-7.25 (m, 5H); 4.77-4.74 (m, 1H); 4.66 (t, 1H, j = 4.8 Hz); 4.57 (d, 1H, J = 11.4 Hz); 4.41-4.40 (m, 1H); 4.25-4.24 (m, 1H); 4.16 - 4.12 (m, 2H); 4.10-4.01 (m, 1H); 3.87-3.82 (m, 1H); 2.21-1.92 (m, 2H); 1.92-1.86 (m, 2H); 1.41 (s, 9H).
RMN13C (50 MHz, CDCI3): 175,7; 155,9; 139,4; 129,4; 129,0; 128,8; 88,4; 81,9; 81,4; 80,7; 74,0; 73,5; 71,7; 61,4; 58,5; 32,6; 28,7; 25,4; 24.7. IV ν (cm"1) filme (CH2CI2): 3055, 2986, 1684, 1604, 1423, 1159, 987, 896, 741. Ot020= +91 (c 0,1 DMSO). Aspecto: óleo incolor. HRMS calculada para C23H32N2O6Na1 455,2158; Encontrada, 455,2164. Exemplo 11: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(N-f-butoxicarbonil- L-valina)-D-glucitol (11)13 C-NMR (50 MHz, CDCl 3): 175.7; 155.9; 139.4; 129.4; 129.0; 128.8; 88.4; 81.9; 81.4; 80.7; 74.0; 73.5; 71.7; 61.4; 58.5; 32.6; 28.7; 25.4; 24.7. IR ν (cm -1) film (CH 2 Cl 2): 3055, 2986, 1684, 1604, 1423, 1159, 987, 896, 741. Ot020 = +91 (c 0.1 DMSO) Appearance: colorless oil HRMS calculated for C 23 H 32 N 2 O 6 Na 1 455.2158 Found 455.2164 Example 11 Obtaining 1,4: 3,6-Dianhydro-5-0- (benzyl) -2-N- (Nf-butoxycarbonyl-L-valine) -D- glucitol (11)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,38-7,24 (m, 5H); 6,23 (br s, 1H); 5,01 (br s, 1H); 4,76 (d, 1H, J = 11,7 Hz); 4,60-4,53 (m, 2H); 4,42 (q, 2H, J = 3,9 Hz); 4,08-4,02 (m, 2H); 3,88-3,81 (m 3H); 3,69-3,63 (m, 1H); 2,15-2,09 (m, 1H); 1,44 (s, 9H); 0,95 (d, 3H, J = 6,9 Hz); 0,91 (d, 3H, J = 6,9 Hz).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.38-7.24 (m, 5H); 6.23 (br s, 1H); 5.01 (br s, 1H); 4.76 (d, 1H, J = 11.7 Hz); 4.60-4.53 (m, 2H); 4.42 (q, 2H, J = 3.9 Hz); 4.08-4.02 (m, 2H); 3.88-3.81 (m 3H); 3.69-3.63 (m, 1H); 2.15-2.09 (m, 1H); 1.44 (s, 9H); 0.95 (d, 3H, J = 6.9 Hz); 0.91 (d, 3H, J = 6.9 Hz).
RMN13C (50 MHz, CDCI3): 171,4; 155,9; 137,6; 128,4; 127,9; 127,8; 86,9; 80,3; 80,2; 80,0; 79,0; 72,5; 70,8; 59,9; 56,8; 30,6; 28,2; 19,3; 17.8.13 C-NMR (50 MHz, CDCl 3): 171.4; 155.9; 137.6; 128.4; 127.9; 127.8; 86.9; 80.3; 80.2; 80.0; 79.0; 72.5; 70.8; 59.9; 56.8; 30.6; 28.2; 19.3; 17.8.
IV ν (cm"1) KBr: 3335, 2968, 2934, 2879, 1687, 1654, 1529, 1461, 1368, 1299, 1250, 1169, 1092, 1052, 1018, 738. PF = 102-103°CIR ν (cm -1) KBr: 3335, 2968, 2934, 2879, 1687, 1654, 1529, 1461, 1368, 1299, 1250, 1169, 1092, 1052, 1018, 738. MP = 102-103 ° C
ao20 = +67 (c 0,1 DMSO). Aspecto: sólido branco. HRMS calculada para C23H34N2O6Na1 457,2315; Encontrada, 457,2324. Exemplo 12: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(N-f-butoxicarbonil- 0-benzil-L-serina)-D-glucitol (12)α 20 = +67 (c 0.1 DMSO). Appearance: white solid. HRMS calcd for C23H34N2O6Na1 457.2315; Found 457.2324. Example 12: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-N- (N-t-butoxycarbonyl-0-benzyl-L-serine) -D-glucitol (12)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,35-7,27 (m, 10H); 4,73 (d, 1H, J = 12 Hz); 4,54- 4,45 (m, 3H); 4,40-4,38 (m, 2H); 4,32-4,31 (m, 1H); 4,23-4,21 (m, 1H); 4,05-3,95 (m, 2H); 3,91-3,81 (m, 3H); 3,67-3,57 (m, 1H); 3,64 (dd, 1H, J = 7,5; 8,7 Hz); 3,61-3,54 (m, 1H); 1,44 (s, 9H).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.35-7.27 (m, 10H); 4.73 (d, 1H, J = 12 Hz); 4.54-4.45 (m, 3H); 4.40-4.38 (m, 2H); 4.32-4.31 (m, 1H); 4.23-4.21 (m, 1H); 4.05-3.95 (m, 2H); 3.91-3.81 (m, 3H); 3.67-3.57 (m, 1H); 3.64 (dd, 1H, J = 7.5; 8.7 Hz); 3.61-3.54 (m, 1H); 1.44 (s, 9H).
RMN13C (50 MHz, CDCI3): RMN13C: 169,9; 155,7; 137,6; 137,3; 128,5; 128,4; 128,0; 127,9; 127,8; 86,8; 80,4; 80,1; 79,1; 73,5; 73,2; 72,4; 70,7; 69,8; 56,8; 51,9; 28,2. IV ν (cm"1) KBr: 3289, 2968, 2974, 2873, 1690, 1650, 1553, 1511, 1299, 1310, 1243, 1168, 1133, 1049, 864, 742.13 C-NMR (50 MHz, CDCl 3): 13 C-NMR: 169.9; 155.7; 137.6; 137.3; 128.5; 128.4; 128.0; 127.9; 127.8; 86.8; 80.4; 80.1; 79.1; 73.5; 73.2; 72.4; 70.7; 69.8; 56.8; 51.9; 28.2. IR ν (cm -1) KBr: 3289, 2968, 2974, 2873, 1690, 1650, 1553, 1511, 1299, 1310, 1243, 1168, 1133, 1049, 864, 742.
PF = 116-117°CMP = 116-117 ° C
aD20 = +57 (c 0,1 DMSO). Aspecto: sólido branco.α D 20 = +57 (c 0.1 DMSO). Appearance: white solid.
HRMS calculada para C28H36N2O7Na, 535,2420; Encontrada, 535,2400.HRMS calcd for C 28 H 36 N 2 O 7 Na, 535.2420; Found 535.2400.
Exemplo 13: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(N-f-butoxicarbonil- L-metionina)-D-glucitol (13)Example 13: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-N- (N-t-butoxycarbonyl-L-methionine) -D-glucitol (13)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,35-7,30 (m, 5H); 6,55 (br s, 1H); 5,16 (brs, 1H); 4,77 (d, 1H, J = 12 Hz); 4,59-4,52 (m, 2H); 4,42-4,38 (m, 2H); 4,24-4,17 (m, 1H); 20 4,08-4,02 (m 2H); 3,87-3,82 (m, 2H); 3,67 (dd, 1H, J = 7,5; 9,0 Hz); 2,55-2,52 (m, 2H); 2,10 (s, 3H); 2,06-2,03 (m, 2H); 1,44 (s, 9H).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.35-7.30 (m, 5H); 6.55 (br s, 1H); 5.16 (brs, 1H); 4.77 (d, 1H, J = 12 Hz); 4.59-4.52 (m, 2H); 4.42-4.38 (m, 2H); 4.24-4.17 (m, 1H); 4.08-4.02 (m 2H); 3.87-3.82 (m, 2H); 3.67 (dd, 1H, J = 7.5; 9.0 Hz); 2.55-2.52 (m, 2H); 2.10 (s, 3H); 2.06-2.03 (m, 2H); 1.44 (s, 9H).
RMN13C (50 MHz, CDCI3): 171,3; 155,6; 137,6; 128,4; 127,8; 127,8; 86,9; 80,4; 80,2; 73,2; 72,4; 70,8; 56,8; 53,4; 31,2; 30,2; 28,2; 15.2.13 C-NMR (50 MHz, CDCl 3): 171.3; 155.6; 137.6; 128.4; 127.8; 127.8; 86.9; 80.4; 80.2; 73.2; 72.4; 70.8; 56.8; 53.4; 31.2; 30.2; 28.2; 15.2.
IVv (cm'1) KBr: 3297, 2974, 2931, 2873, 1660, 1650, 1530, 1452, 1368, 1249, 1168, 1092,860,742. PF = 91-93°CIRv (cm -1) KBr: 3297, 2974, 2931, 2873, 1660, 1650, 1530, 1452, 1368, 1249, 1168, 1092,860,742. MP = 91-93 ° C
aD20 = +34 (c 0,1 CH2CI2). Aspecto: sólido branco.α D 20 = + 34 ° (c 0.1 CH 2 Cl 2). Appearance: white solid.
HRMS calculada para C23H32N2O6NaS, 489,2035; Encontrada, 489,2038.HRMS calcd for C23H32N2O6NaS, 489.2035; Found 489.2038.
Exemplo 14: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(N-í-butoxicarbonil- L-triptofano)-D-glucitol (14) RMN 1H δ ppm (CDCI3, 300 MHz): 8,24 (br s, 1H); 7,64 (d, 1 Η, J = 7,8 Hz); 7,36-7,29 (m, 6H); 7,19-7,07 (m, 3H); 5,70 (br s, 1H); 5,29 (br s, 1H); 4,48 (q, 1H, J = 12 Hz); 4,39-4,36 (m, 1H); 4,23-4,21 (m, 1H); 3,84-3,65 (m, 5H); 3,55-3,49 (m 3H); 3,29 (dd, 1H, J= 5,7; 14,4 Hz); 3,10 (dd, IH1J= 8,4; 14,4 Hz); 1,44 (s, 9H).Example 14: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-N- (N-t-butoxycarbonyl-L-tryptophan) -D-glucitol (14) 1 H NMR δ ppm ( CDCl3, 300 MHz): 8.24 (br s, 1H); 7.64 (d, 1 Η, J = 7.8 Hz); 7.36-7.29 (m, 6H); 7.19-7.07 (m, 3H); 5.70 (br s, 1H); 5.29 (br s, 1H); 4.48 (q, 1H, J = 12 Hz); 4.39-4.36 (m, 1H); 4.23-4.21 (m, 1H); 3.84-3.65 (m, 5H); 3.55-3.49 (m 3H); 3.29 (dd, 1H, J = 5.7; 14.4 Hz); 3.10 (dd, 1H H = 8.4; 14.4 Hz); 1.44 (s, 9H).
RMN13C (50 MHz, CDCI3): 171,4; 155,4; 137,6; 136,1; 128,4; 127,9; 127,7; 127,3; 123,2; 122,2; 119,7; 118,9; 111,2; 110,6; 86,3; 80,1; 79,6; 79,0; 72,6; 72,3; 70,4; 56,4; 55,6; 28,6; 28,3.13 C-NMR (50 MHz, CDCl 3): 171.4; 155.4; 137.6; 136.1; 128.4; 127.9; 127.7; 127.3; 123.2; 122.2; 119.7; 118.9; 111.2; 110.6; 86.3; 80.1; 79.6; 79.0; 72.6; 72.3; 70.4; 56.4; 55.6; 28.6; 28.3.
IV ν (cnT1) KBr: 3321, 2974, 2933, 2881, 1685, 1646, 1523, 1455, 1366, 1250, 1169, 1095, 1018, 863, 743. PF = 161-162°CIR ν (cnT1) KBr: 3321, 2974, 2933, 2881, 1685, 1646, 1523, 1455, 1366, 1250, 1169, 1095, 1018, 863, 743. MP = 161-162 ° C
aD20 = +44 (c 0,1 CH2CI2). Aspecto: sólido branco.α D 20 = + 44 ° (c 0.1 CH 2 Cl 2). Appearance: white solid.
HRMS calculada para C20H35N3O6Na1 544,2424; Encontrada, 544,2408.HRMS calcd for C20H35N3O6Na1 544.2424; Found 544.2408.
Exemplo 15: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(N-carbobenziloxi- prolina-fenilalanina)-D-glucitol (15)Example 15: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-N- (N-carbobenzyloxyproline-phenylalanine) -D-glucitol (15)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,27-7,02 (m, 15H); 6,61 (br s, 1H); 6,29 (br s, 1H); 5,06-4,90 (m, 2H); 4,66 (d, 2H, J= 12 Hz); 4,45 (d, 2H, J= 12 Hz); 4,31-4,27 (m, 2H); 4,18-4,15 (m, 1H); 3,99-3,91 (m, 2H); 3,78-3,72 (m, 2H); 3,56 (t, 1H, J= 8,1 Hz); 3,44-3,30 (m, 3H); 3,10-3,03 (m 1H); 1,93-1,60 (m, 4H). RMN13C (50 MHz, CDCI3): 171,2; 170,2; 155,9; 137,6; 136,4; 135,9; 129,0; 128,6; 128,5; 128,3; 128,2; 127,7; 127,0; 86,8; 80,1; 79,0; 73,4; 72,3; 70,4; 67,5; 61,2; 57,0; 53,4; 47,0; 37,3; 29,3; 24,3.1 H NMR δ ppm (CDCl 3, 300 MHz): 7.27-7.02 (m, 15H); 6.61 (br s, 1H); 6.29 (br s, 1H); 5.06-4.90 (m, 2H); 4.66 (d, 2H, J = 12 Hz); 4.45 (d, 2H, J = 12 Hz); 4.31-4.27 (m, 2H); 4.18-4.15 (m, 1H); 3.99-3.91 (m, 2H); 3.78-3.72 (m, 2H); 3.56 (t, 1H, J = 8.1 Hz); 3.44-3.30 (m, 3H); 3.10-3.03 (m 1H); 1.93-1.60 (m, 4H). 13 C-NMR (50 MHz, CDCl 3): 171.2; 170.2; 155.9; 137.6; 136.4; 135.9; 129.0; 128.6; 128.5; 128.3; 128.2; 127.7; 127.0; 86.8; 80.1; 79.0; 73.4; 72.3; 70.4; 67.5; 61.2; 57.0; 53.4; 47.0; 37.3; 29.3; 24.3.
IV ν (cm'1) KBr: 3279, 3069, 2949, 2878, 1704, 1649, 1554, 1448, 1415, 1355, 1247, 1093, 1023, 914, 743. PF = 141-143°CIR ν (cm -1) KBr: 3279, 3069, 2949, 2878, 1704, 1649, 1554, 1448, 1415, 1355, 1247, 1093, 1023, 914, 743. MP = 141-143 ° C
ao20 = -15 (c 0,1 CH2CI2). Aspecto: sólido branco.α 20 = -15 (c 0.1 CH 2 Cl 2). Appearance: white solid.
HRMS calculada para C35H39N3O7Na, 636,2686; Encontrada, 636,2697.HRMS calcd for C35 H39 N3 O7 Na, 636.2686; Found 636.2697.
Exemplo 16: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(N-carbobenziloxi- D-prolina)-D-glucitol (16) RMN 1H δ ppm (CDCI3l 300 MHz): 7,36-7,26 (m, 6H); 5,15-5,13 (m, 2H); 4,75 (d, 1Η, J= 11,7 Hz); 4,56 (d, 1 Η, J= 12 Hz); 4,30-4,26 (m, 2H); 4,00-3,91 (m, 2H); 3,85-3,80 (m, 2H); 3,66-3,62 (m, 1H); 3,61-3,44 (m, 4H); 2,34-1,85 (m, 4H). RMN13C (50 MHz, CDCI3): 171,1; 156,5; 137,6; 136,4; 128,5; 128,4; 128,1; 127,8; 86,9; 80,1; 78,9; 73,4; 72,4; 70,6; 67,3; 60,2; 56,8; 47,0; 27,9; 24,6.Example 16: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-N- (N-carbobenzyloxy-D-proline) -D-glucitol (16) 1 H NMR δ ppm (CDCl3 300 MHz): 7.36-7.26 (m, 6H); 5.15-5.13 (m, 2H); 4.75 (d, 1Η, J = 11.7 Hz); 4.56 (d, 1 Η, J = 12 Hz); 4.30-4.26 (m, 2H); 4.00-3.91 (m, 2H); 3.85-3.80 (m, 2H); 3.66-3.62 (m, 1H); 3.61-3.44 (m, 4H); 2.34-1.85 (m, 4H). 13 C-NMR (50 MHz, CDCl 3): 171.1; 156.5; 137.6; 136.4; 128.5; 128.4; 128.1; 127.8; 86.9; 80.1; 78.9; 73.4; 72.4; 70.6; 67.3; 60.2; 56.8; 47.0; 27.9; 24.6.
IVv (cm1) KBr 3345, 3034, 2974, 2926, 2873, 1679, 1531, 1451, 1421, 1356, 1253, 1208, 1122, 1091, 952, 866, 758.IRv (cm -1) KBr 3345, 3034, 2974, 2926, 2873, 1679, 1531, 1451, 1421, 1356, 1253, 1208, 1122, 1091, 952, 866, 758.
PF = 89-90°CMP = 89-90 ° C
αD20 = +94 (c 0,1 CH2Cl2). Aspecto: sólido branco, HRMS calculada para CaeH30N2O6Na1 489,2002; Encontrada, 489,2011.α D 20 = +94 (c 0.1 CH 2 Cl 2). Appearance: white solid, HRMS calcd for CaeH30N2O6Na1 489.2002; Found 489.2011.
Exemplo 17: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(Na-f-butoxicarbonil- Ne-carbobenziloxi-L-lisina)-D-glucitol(17)Example 17: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-N- (Na-t-butoxycarbonyl-N-carbobenzyloxy-L-lysine) -D-glucitol (17)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,34-7,30 (m, 10H); 6,55 (br s, 1H); 5,29 (s, 2H); 5,09-5,02 (m, 1H); 4,75 (d, 1H, J= 11,7 Hz); 4,58 (t, 1H, J = 4,5 Hz); 4,51 (d, 1H, J = 12 Hz); 4,41-4,36 (m, 2H); 4,07-4,00 (m, 2H); 3,86-3,81 (m, 2H); 3,65 (dd, 1H, J = 7,5; 8,7 Hz); 3,17 (q, 2H, J = 6,0 Hz); 1,84-1,82 (m, 2H); 1,64-1,50 (tn, 2H); 1,43 (s, 9H); 1,39-1,34 (m, 2H).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.34-7.30 (m, 10H); 6.55 (br s, 1H); 5.29 (s, 2H); 5.09-5.02 (m, 1H); 4.75 (d, 1H, J = 11.7 Hz); 4.58 (t, 1H, J = 4.5 Hz); 4.51 (d, 1H, J = 12 Hz); 4.41-4.36 (m, 2H); 4.07-4.00 (m, 2H); 3.86-3.81 (m, 2H); 3.65 (dd, 1H, J = 7.5; 8.7 Hz); 3.17 (q, 2H, J = 6.0 Hz); 1.84-1.82 (m, 2H); 1.64-1.50 (tn, 2H); 1.43 (s, 9H); 1.39-1.34 (m, 2H).
RMN13C (50 MHz1 CDCI3): 172,0; 156,6; 155,8; 137,6; 136,5; 128,4; 128,3; 128,0; 127,9; 127,8; 86,9; 80,2; 80,1; 79,0; 73,2; 72,4; 70,7; 66,6; 56,8; 54,3; 40,1; 31,5; 29,4; 28,2; 22,4.13 C-NMR (50 MHz 1 CDCl 3): 172.0; 156.6; 155.8; 137.6; 136.5; 128.4; 128.3; 128.0; 127.9; 127.8; 86.9; 80.2; 80.1; 79.0; 73.2; 72.4; 70.7; 66.6; 56.8; 54.3; 40.1; 31.5; 29.4; 28.2; 22.4.
IV ν (cm'1) KBr: 3360, 2972, 2937, 2879, 1694, 1659, 1459, 1366, 1254, 1168, 1087, 1015, 908, 749.IR ν (cm -1) KBr: 3360, 2972, 2937, 2879, 1694, 1659, 1459, 1366, 1254, 1168, 1087, 1015, 908, 749.
PF = 62-63°CMP = 62-63 ° C
αD20 = +23 (c 0,1 CH2CI2). Aspecto: sólido branco.α D 20 = + 23 (c 0.1 CH 2 Cl 2). Appearance: white solid.
HRMS calculada para C32H43N3O8Na, 620,2948; Encontrada, 620,2958.HRMS calcd for C 32 H 43 N 3 O 8 Na, 620.2948; Found 620.2958.
Exemplo 18: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(Na-f-butoxicarbonil- N -tosil-L-histidina)-D-glucitol (18)Example 18: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-N- (Na-t-butoxycarbonyl-N-tosyl-L-histidine) -D-glucitol (18)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,93 (s, 1H); 7,81 (d, 2H, J = 8,4 Hz); 7,35-7,29 (m, 7H); 7,08 (s, 1H); 6,77 (br s, 1H); 5,87 (br s, 1H); 4,71 (d, 1H1 J = 11,7 Hz); 4,55 (d, 1H1 J = 11,7 Hz); 4,46 (t, 1H, J = 4,5 Hz); 4,34-4,23 (m, 3H), 4,04 (q, IH1J = 6,9 Hz); 3,92-3,80 (m, 2H); 3,65-3,59 (m, 2H); 3,04 (dd, 1H, J = 5,1; 14,7 Hz); 2,85 (dd, IH1J = 6,3; 14,7 Hz); 2,42 (s, 3H); 1,42 (s, 9H).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.93 (s, 1H); 7.81 (d, 2H, J = 8.4 Hz); 7.35-7.29 (m, 7H); 7.08 (s, 1H); 6.77 (br s, 1H); 5.87 (br s, 1H); 4.71 (d, 1H1 J = 11.7 Hz); 4.55 (d, 1H1 J = 11.7 Hz); 4.46 (t, 1H, J = 4.5 Hz); 4.34-4.23 (m, 3H); 4.04 (q, 1HH = 6.9 Hz); 3.92-3.80 (m, 2H); 3.65-3.59 (m, 2H); 3.04 (dd, 1H, J = 5.1; 14.7 Hz); 2.85 (dd, 1H H = 6.3; 14.7 Hz); 2.42 (s, 3H); 1.42 (s, 9H).
RMN13C (50 MHz1 CDCI3): 170,8; 155,4; 146,4; 140,4; 137,6; 134,5; 130,4; 128,3; 127,8; 127,3; 114,9; 86,7; 81,7; 80,1; 79,1; 72,9; 72,4; 70,7; 56,6; 53,9; 30,4; 28,2; 21,6.13 C-NMR (50 MHz 1 CDCl 3): 170.8; 155.4; 146.4; 140.4; 137.6; 134.5; 130.4; 128.3; 127.8; 127.3; 114.9; 86.7; 81.7; 80.1; 79.1; 72.9; 72.4; 70.7; 56.6; 53.9; 30.4; 28.2; 21.6.
IVv (cm-1) KBr 3290, 3073, 2974, 2934, 2879, 1661, 1527, 1376, 1300, 1248, 1174, 1088, 814, 744. PF = 91-93°CIRv (cm -1) KBr 3290, 3073, 2974, 2934, 2879, 1661, 1527, 1376, 1300, 1248, 1174, 1088, 814, 744. MP = 91-93 ° C
α20 = +55 (c 0,1 CH2CI2). Aspecto: sólido branco, HRMS calculada para C3IH38N4O8NaS, 649,2308; Encontrada, 649,2321.α 20 = + 55 ° (c 0.1 CH 2 Cl 2). Appearance: white solid, HRMS calcd for C 31 H 38 N 4 O 8 Na 2 S, 649.2308; Found, 649.2321.
Exemplo 19: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(N-carbobenziloxi- valina-fenilalanina)-D-glucitol (19)Example 19: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-N- (N-carbobenzyloxyvaline phenylalanine) -D-glucitol (19)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,33-7,12 (m, 15H); 6,77 (br s, 1H); 5,59-5,57 (m, 1H); 5.12-5,01 (m, 2H); 4,80 (q, 1H, J = 7,8 Hz); 4,65 (dd, 1H, J = 4,5; 12 Hz); 4,45- 4,38 (m, 1H); 4,32-4,27 (m, 2H); 4,01-3,80 (m, 3H); 3,77-3,70 (m, 2H); 3,66-3,57 (m, 1H); 3,10-2,98 (m, 2H); 2,11-1,95 (m, 1H); 0,91 (d, 3H, J = 6,6 Hz); 0,84 (d, 3H, J = 6,6 Hz).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.33-7.12 (m, 15H); 6.77 (br s, 1H); 5.59-5.57 (m, 1H); 5.12-5.01 (m, 2H); 4.80 (q, 1H, J = 7.8 Hz); 4.65 (dd, 1H, J = 4.5; 12 Hz); 4.45 - 4.38 (m, 1H); 4.32-4.27 (m, 2H); 4.01-3.80 (m, 3H); 3.77-3.70 (m, 2H); 3.66-3.57 (m, 1H); 3.10-2.98 (m, 2H); 2.11-1.95 (m, 1H); 0.91 (d, 3H, J = 6.6 Hz); 0.84 (d, 3H, J = 6.6 Hz).
RMN13C (50 MHz, CDCI3): 171,6; 170,5; 156,6; 137,8; 136,3; 136,2; 129,4; 129,3; 128,5; 128,4; 128,3; 128,1; 127,7; 126,9; 126,7; 87,0; 80,3; 78,9; 72,9; 72,2; 70,8; 67,1; 60,7; 56,6; 53,7; 39,2; 31,2; 19,3; 18,1.13 C-NMR (50 MHz, CDCl 3): 171.6; 170.5; 156.6; 137.8; 136.3; 136.2; 129.4; 129.3; 128.5; 128.4; 128.3; 128.1; 127.7; 126.9; 126.7; 87.0; 80.3; 78.9; 72.9; 72.2; 70.8; 67.1; 60.7; 56.6; 53.7; 39.2; 31.2; 19.3; 18.1.
IV ν (cm-1) KBr: 3290, 3066, 2962, 1708, 1646, 1544, 1393, 1241, 1132, 1091, 1023, 914, 741.IR ν (cm -1) KBr: 3290, 3066, 2962, 1708, 1646, 1544, 1393, 1241, 1132, 1091, 1023, 914, 741.
PF = 81-83°CMP = 81-83 ° C
ao20 = +26 (c 0,1 CH2CI2). Aspecto: sólido branco.α 20 = + 26 ° (c 0.1 CH 2 Cl 2). Appearance: white solid.
HRMS calculada para C35H4IN7O7Na, 638,2842; Encontrada, 638,2862.HRMS calcd for C35H4IN7O7Na, 638.2842; Found 638.2862.
Exemplo 20: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(N-f-butoxicarbonil- L-isoserina)-D-glucitol (20)Example 20: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-N- (N-t-butoxycarbonyl-L-isoserine) -D-glucitol (20)
30 RMN 1H δ ppm (CDCI3, 300 MHz): 7,36-7,18 (m, 5H); 5,37 (br s, 1H); 5,20 (brs, 1H); 4,77 (d, 1H, J = 12 Hz); 4,64 (t, 1H, J = 4,5 Hz); 4,53 (d, 1H, J = 12 Hz); 4,43-4,41 (m, 2H); 4,17 (br s, 1H); 4,11-4,02 (m, 2H); 3,88-3,81 (m, 2H); 3,70-3,64 (m, 1H); 3,61-3,44 (m, 2H); 1,74 (br s, 1H); 1,43 (s, 9H).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.36-7.18 (m, 5H); 5.37 (br s, 1H); 5.20 (brs, 1H); 4.77 (d, 1H, J = 12 Hz); 4.64 (t, 1H, J = 4.5 Hz); 4.53 (d, 1H, J = 12 Hz); 4.43-4.41 (m, 2H); 4.17 (br s, 1H); 4.11-4.02 (m, 2H); 3.88-3.81 (m, 2H); 3.70-3.64 (m, 1H); 3.61-3.44 (m, 2H); 1.74 (br s, 1H); 1.43 (s, 9H).
RMN13C (50 MHz, CDCI3): 171,8; 158,9; 137,6; 128,4; 127,8; 87,0; 80,7; 80,3; 79,0; 73,6; 73,4; 72,4; 70,7; 56,5; 44,7; 28,2.13 C-NMR (50 MHz, CDCl 3): 171.8; 158.9; 137.6; 128.4; 127.8; 87.0; 80.7; 80.3; 79.0; 73.6; 73.4; 72.4; 70.7; 56.5; 44.7; 28.2.
IV ν (cm1) KBr: 3398, 3315, 2975, 2936, 2889, 1708, 1640, 1547, 1515, 1399, 1325, 1256, 1171, 1110, 1043, 882, 731.IR ν (cm -1) KBr: 3398, 3315, 2975, 2936, 2889, 1708, 1640, 1547, 1515, 1399, 1325, 1256, 1171, 1110, 1043, 882, 731.
PF = 133-134°CMP = 133-134 ° C
αD20 = +4 (c 0,1 CH2CI2). Aspecto: sólido branco.α D 20 = + 4 (c 0.1 CH 2 Cl 2). Appearance: white solid.
HRMS calculada para C21H30N2O7Na, 445,1951; Encontrada, 445,1947.HRMS calcd for C21H30N2O7Na, 445.1951; Found 445.1947.
Exemplo 21: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(N-f-butoxicarbonil- L-treonina)-D-glucitol (21)Example 21: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-N- (N-t-butoxycarbonyl-L-threonine) -D-glucitol (21)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,35-7,29 (m, 5H); 6,92 (br s, 1H); 5,51 (br s, 1H); 4,73 (d, 1H, J= 12 Hz); 4,63-4,61 (m, 1H); 4,56 (d, 1H, J = 12 Hz); 4,43-4,35 (m, 3H); 4,08-4,04 (m, 2H); 4,00 (dd, 1H, J = 2,1; 8,1 Hz); 3,88-3,81 (m, 2H); 3,67 (dd, 1H, J = 7,5; 9,0 Hz); 1,46 (s, 9H); 1,17 (d, 3H, J = 6,3 Hz).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.35-7.29 (m, 5H); 6.92 (br s, 1H); 5.51 (br s, 1H); 4.73 (d, 1H, J = 12 Hz); 4.63-4.61 (m, 1H); 4.56 (d, 1H, J = 12 Hz); 4.43-4.35 (m, 3H); 4.08-4.04 (m, 2H); 4.00 (dd, 1H, J = 2.1; 8.1 Hz); 3.88-3.81 (m, 2H); 3.67 (dd, 1H, J = 7.5; 9.0 Hz); 1.46 (s, 9H); 1.17 (d, 3H, J = 6.3 Hz).
RMN13C (50 MHz, CDCI3): 171,2; 156,6; 137,6; 128,4; 127,9; 127,8; 86,9; 80,5; 80,3; 78,9; 73,2; 72,4; 70,5; 66,6; 58,2; 56,9; 28,2; 18,3.13 C-NMR (50 MHz, CDCl 3): 171.2; 156.6; 137.6; 128.4; 127.9; 127.8; 86.9; 80.5; 80.3; 78.9; 73.2; 72.4; 70.5; 66.6; 58.2; 56.9; 28.2; 18.3.
IV ν (cm-1) film (CH2CI2): 3944, 3755, 3691, 3056, 2987, 1682, 1604, 1424, 1265, 1159,986,896,742.IR ν (cm -1) film (CH 2 Cl 2): 3944, 3755, 3691, 3056, 2987, 1682, 1604, 1424, 1265, 1159,986,896,742.
αD20 = +6 (c 0,1 CH2CI2). Aspecto: óleo incolor.α D 20 = + 6 (c 0.1 CH 2 Cl 2). Appearance: colorless oil.
HRMS calculada para C22H32N2O7Na, 459,2107; Encontrada, 459,2085.HRMS calcd for C 22 H 32 N 2 O 7 Na, 459.2107; Found 459.2085.
Exemplo 22: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-N-(Na-f-butoxicarbonil- Nδ,Nω-di-carbobenziloxi-L-arginina)-D-glucitol (22)Example 22: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-N- (Na-t-butoxycarbonyl-Nδ, Nω -dicarbobenzyloxy-L-arginine) -D-glucitol (22)
RMN 1H δ ppm (CDCI3, 300 MHz): 9,42 (br s, 1H); 7,43-7,29 (m, 15H); 6,68 (br s, 1H); 5,64 (br s, 1H); 5,29 (s, 2H); 5,24 (s, 2H); 5,19-5,10 (m, 2H); 4,73 (d, 1H, J= 12 Hz); 4,52 (d, 1H, J = 12 Hz); 4,33-4,20 (m, 3H); 3,96-3,90 (m, 2H); 3,78 (dd, 1H, J = 7,5; 9,0 Hz); 3,63-3,57 (m, 1H); 1,94-1,70 (m, 2H); 1,68-1,58 (m, 4H); 1,43 (s, 9H). RMN13C (50 MHz, CDCI3): 171,8; 163,4; 160,7; 155,8; 155,6; 137,7; 136,6; 134,5; 128,8; 128,7; 128,5; 128,4; 128,3; 127,9; 127,8; 127,7; 86,9; 80,0; 79,9; 78,9; 72,9; 72,3; 70,4; 68,9; 67,0; 56,9; 49,0; 43,9; 28,4; 25,5; 24,7. IVv (cm"1) KBr: 3387, 3328, 2929, 2857, 1717, 1656, 1615, 1510, 1451, 1376, 1252, 1093, 744. PF = 81-83°C1 H NMR δ ppm (CDCl 3, 300 MHz): 9.42 (br s, 1H); 7.43-7.29 (m, 15H); 6.68 (br s, 1H); 5.64 (br s, 1H); 5.29 (s, 2H); 5.24 (s, 2H); 5.19-5.10 (m, 2H); 4.73 (d, 1H, J = 12 Hz); 4.52 (d, 1H, J = 12 Hz); 4.33-4.20 (m, 3H); 3.96-3.90 (m, 2H); 3.78 (dd, 1H, J = 7.5; 9.0 Hz); 3.63-3.57 (m, 1H); 1.94-1.70 (m, 2H); 1.68-1.58 (m, 4H); 1.43 (s, 9H). 13 C-NMR (50 MHz, CDCl 3): 171.8; 163.4; 160.7; 155.8; 155.6; 137.7; 136.6; 134.5; 128.8; 128.7; 128.5; 128.4; 128.3; 127.9; 127.8; 127.7; 86.9; 80.0; 79.9; 78.9; 72.9; 72.3; 70.4; 68.9; 67.0; 56.9; 49.0; 43.9; 28.4; 25.5; 24.7. IRv (cm -1) KBr: 3387, 3328, 2929, 2857, 1717, 1656, 1615, 1510, 1451, 1376, 1252, 1093, 744. MP = 81-83 ° C
«D20 = +34 (c 0,1 CH2CI2). Aspecto: sólido branco. HRMS calculada para C4OH4SN5Oi0Na, 782,3377; Encontrada, 782,3405.D 20 = + 34 ° (c 0.1 CH 2 Cl 2). Appearance: white solid. HRMS calcd for C 4 OH 4 SN 5 O 10 Na, 782.3377; Found, 782.3405.
Exemplo 23: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-0-(N-f-butoxicarbonil- L-prolina)-D-manitol (23)Example 23: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-0- (N-t-butoxycarbonyl-L-proline) -D-mannitol (23)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,30-7,21 (m, 5H); 5,06-4,95 (m, 1H); 4,76-4,68 (m, 2H); 4,57 (s, 1H); 4,53-4,46 (m, 1H); 4,39-4,30 (m, 1H); 4,07-3,99 (m, 1H); 3,95- 3,84 (m, 2H); 3,51-3,35 (m, 2H); 2,23-2,19 (m, 2H); 2,04-1,86 (m, 2H); 1,40 (s, 9H). RMN13C (50 MHz, CDCI3): 172,5; 153,7; 137,5; 128,4; 127,9; 80,6; 80,4; 79,9; 78,8; 74,3; 72,5; 70,8; 70,5; 59,0; 46,5; 30,9; 28,3; 24,2.1 H NMR δ ppm (CDCl 3, 300 MHz): 7.30-7.21 (m, 5H); 5.06-4.95 (m, 1H); 4.76-4.68 (m, 2H); 4.57 (s, 1H); 4.53-4.46 (m, 1H); 4.39-4.30 (m, 1H); 4.07-3.99 (m, 1H); 3.95-3.84 (m, 2H); 3.51-3.35 (m, 2H); 2.23-2.19 (m, 2H); 2.04-1.86 (m, 2H); 1.40 (s, 9H). 13 C-NMR (50 MHz, CDCl 3): 172.5; 153.7; 137.5; 128.4; 127.9; 80.6; 80.4; 79.9; 78.8; 74.3; 72.5; 70.8; 70.5; 59.0; 46.5; 30.9; 28.3; 24.2.
IV ν (cm-1) filme (CH2CI2): 2974, 2880, 1746, 1697, 1454, 1398, 1261, 1128, 1090, 1028,885,819.IR ν (cm -1) film (CH 2 Cl 2): 2974, 2880, 1746, 1697, 1454, 1398, 1261, 1128, 1090, 1028,885,819.
ao20 = +91 (c, 0.1 DMSO). Aspecto: óleo incolor.α 20 = + 91 ° (c, 0.1 DMSO). Appearance: colorless oil.
HRMS calculada para C23H3iN07Na, 456,1998; Encontrada, 456,2009.HRMS calcd for C23 H31 N07 Na, 456.1998; Found 456.2009.
Exemplo 24: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-0-(N-f-butoxicarbonil- L-valina)-D-manitol (24)Example 24: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-0- (N-t-butoxycarbonyl-L-valine) -D-mannitol (24)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,40-7,25 (m, 5H); 5,14 (q, 1H, J = 6,0 Hz); 5,01- 4,98 (m, 1H); 4,76-4,68 (m, 2H); 4,58 (s, 1H); 4,49 (t, 1H, J = 4,8 Hz); 4,31-4,26 (m, 1H); 4,13-4,00 (m, 2H); 3,95-3,90 (m, 2H); 3,62 (t, 1H, J = 8,5 Hz); 1,44 (s, 9H); 0,98 (d, 3H, J = 6,9 Hz); 0,90 (d, 3H, J = 6,9 Hz). RMN13C (50 MHz, CDCI3): 171,7; 155,5; 137,5; 128,4; 127,9; 80,5; 80,3; 79,7; 78,7; 74,8; 72,5; 71,0; 70,5; 58,5; 31,2; 28,3; 18,9; 17,4.1 H NMR δ ppm (CDCl 3, 300 MHz): 7.40-7.25 (m, 5H); 5.14 (q, 1H, J = 6.0 Hz); 5.01-4.98 (m, 1H); 4.76-4.68 (m, 2H); 4.58 (s, 1H); 4.49 (t, 1H, J = 4.8 Hz); 4.31-4.26 (m, 1H); 4.13-4.00 (m, 2H); 3.95-3.90 (m, 2H); 3.62 (t, 1H, J = 8.5 Hz); 1.44 (s, 9H); 0.98 (d, 3H, J = 6.9 Hz); 0.90 (d, 3H, J = 6.9 Hz). 13 C-NMR (50 MHz, CDCl 3): 171.7; 155.5; 137.5; 128.4; 127.9; 80.5; 80.3; 79.7; 78.7; 74.8; 72.5; 71.0; 70.5; 58.5; 31.2; 28.3; 18.9; 17.4.
IVv (cm1) filme (CH2CI2): 3350, 2970, 2880, 1713, 1502, 1461, 1367, 1251, 1161, 1090, 1023, 866, 740.IR (cm -1) film (CH 2 Cl 2): 3350, 2970, 2880, 1713, 1502, 1461, 1367, 1251, 1161, 1090, 1023, 866, 740.
CtD20 = +107 (c, 0.1 DMSO). Aspecto: óleo incolor. HRMS calculada para C23H33NO7Na, 458,2155; Encontrada, 458,2163. Exemplo 25: Obtenção do 1,4:3,6-dianidro-5-O-(benzil)-2-O-(N-f-butoxicarbonil- O-benzil-L-serina)-D-manitol (25)C t D 20 = +107 (c, 0.1 DMSO). Appearance: colorless oil. HRMS calcd for C23H33NO7Na, 458.2155; Found 458.2163. Example 25: Obtaining 1,4: 3,6-dianhydro-5-O- (benzyl) -2-O- (N-t-butoxycarbonyl-O-benzyl-L-serine) -D-mannitol (25)
RMN 1H δ ppm (CDCl3, 300 MHz): 7,36-7,27 (m, 10H); 5,43-5,40 (m, 1H); 5,13 (q, 1H, J = 5,7 Hz); 4,74-4,70 (m, 2H); 4,57-4,43 (m, 4H); 4,04-3,88 (m, 5H); 3,86-3,60 (m, 1H); 3,51-3,46 (m, 1H), 1,44 (s, 9H).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.36-7.27 (m, 10H); 5.43-5.40 (m, 1H); 5.13 (q, 1H, J = 5.7 Hz); 4.74-4.70 (m, 2H); 4.57-4.43 (m, 4H); 4.04-3.88 (m, 5H); 3.86-3.60 (m, 1H); 3.51-3.46 (m, 1H), 1.44 (s, 9H).
RMN13C (50 MHz, CDCl3): 170,1; 155,3; 137,5; 137,4; 128,4; 128,3; 127,9; 127,8; 127,7; 127,6; 80,5; 80,3; 79,9; 78,7; 75,1; 73,2; 72,5; 71,0; 70,4; 69,8; 53,9; 28,3. IV ν (cm-1) filme (CH2Cl2): 3328, 2974, 2928, 2876, 1749, 1713, 1499, 1363, 1245, 1166, 1098, 1028, 859, 742. αD20 = +92 (c, 0.1 DMSO). Aspecto: óleo incolor.13 C-NMR (50 MHz, CDCl 3): 170.1; 155.3; 137.5; 137.4; 128.4; 128.3; 127.9; 127.8; 127.7; 127.6; 80.5; 80.3; 79.9; 78.7; 75.1; 73.2; 72.5; 71.0; 70.4; 69.8; 53.9; 28.3. IR ν (cm -1) film (CH 2 Cl 2): 3328, 2974, 2928, 2876, 1749, 1713, 1499, 1363, 1245, 1166, 1098, 1028, 859, 742. α D 20 = +92 (c, 0.1 DMSO) . Appearance: colorless oil.
HRMS calculada para C28H35NO8Na, 536,2260; Encontrada, 536,2278.HRMS calcd for C28H35NO8Na, 536.2260; Found 536.2278.
Exemplo 26: Obtenção do 1,4:3,6-dianidro-5-O-(benzil)-2-O-(N-f-butoxicarbonil- L-metionina)-D-manitol (26)Example 26: Obtaining 1,4: 3,6-dianhydro-5-O- (benzyl) -2-O- (N-t-butoxycarbonyl-L-methionine) -D-mannitol (26)
RMN 1H δ ppm (CDCl3, 300 MHz): 7,36-7,32 (m, 5H); 5,13 (q, 2H, J = 5,7 Hz); 4,76- 4,69 (m, 2H); 4,59 (s, 1H); 4,49-4,46 (m, 1H); 4,07-4,04 (m, 3H); 3,95-3,90 (m, 1H); 3,60 (t, 1H, J = 8,5 Hz); 2,57-2,51 (m, 2H); 2,10-2,05 (m, 2H); 2,08 (s, 3H); 1,44 (s, 9H).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.36-7.32 (m, 5H); 5.13 (q, 2H, J = 5.7 Hz); 4.76-4.69 (m, 2H); 4.59 (s, 1H); 4.49-4.46 (m, 1H); 4.07-4.04 (m, 3H); 3.95-3.90 (m, 1H); 3.60 (t, 1H, J = 8.5 Hz); 2.57-2.51 (m, 2H); 2.10-2.05 (m, 2H); 2.08 (s, 3H); 1.44 (s, 9H).
RMN13C (50 MHz, CDCl3): 171,5; 155,1; 137,4; 128,4; 127,9; 127,8; 80,5; 80,3; 79,9; 78,6; 75,0; 72,5; 71,1; 70,4; 52,8; 32,0; 29,7; 28,2; 15,4. IV ν (cm-1) filme (CH2Cl2): 3339, 2974, 2923, 2876, 1710, 1509, 1450, 1366, 1253, 1166, 1069, 1028, 864, 743. αD20 = +116 (c 0,1 CH2Cl2). Aspecto: óleo incolor.13 C-NMR (50 MHz, CDCl 3): 171.5; 155.1; 137.4; 128.4; 127.9; 127.8; 80.5; 80.3; 79.9; 78.6; 75.0; 72.5; 71.1; 70.4; 52.8; 32.0; 29.7; 28.2; 15.4. IR ν (cm -1) film (CH 2 Cl 2): 3339, 2974, 2923, 2876, 1710, 1509, 1450, 1366, 1253, 1166, 1069, 1028, 864, 743. α D 20 = +116 (c 0.1 CH 2 Cl 2 ). Appearance: colorless oil.
HRMS calculada para C22H33NO7NaS, 490,1875; Encontrada, 490,1895.HRMS calcd for C22H33NO7NaS, 490.1875; Found, 490.1895.
Exemplo 27: Obtenção do 1,4:3,6-dianidro-5-O-(benzil)-2-O-(N-f-butoxicarbonil- L-triptofano)-D-manitol (27)Example 27: Obtaining 1,4: 3,6-dianhydro-5-O- (benzyl) -2-O- (N-t-butoxycarbonyl-L-tryptophan) -D-mannitol (27)
RMN 1H δ ppm (CDCl3, 300 MHz): 7,56 (d, 1H); 7,35-7,25 (m, 7H); 7,17-7,02 (m, 2H); 4,99 (q, 1H, J = 5,7 Hz); 4,75-4,46 (m, 5H); 4,13-4,01 (m, 1H); 3,90-3,85 (m, 2H); 3,80-3,67 (m, 1H); 3,58 (t, 1H, J = 8,4 Hz); 3,31-3,90 (m, 2H); 1,42 (s, 9H). RMN13C (50 MHz1 CDCI3): 171,7; 155,0; 137,5; 135,9; 128,4; 127,9; 127,6; 127,5; 122,9; 121,9; 119,4; 118,6; 111,1; 110,0; 81,7; 80,6; 80,5; 80,2; 79,7; 78,6; 74,9; 72,5; 60,3; 54,3; 28,2.1 H NMR δ ppm (CDCl 3, 300 MHz): 7.56 (d, 1H); 7.35-7.25 (m, 7H); 7.17-7.02 (m, 2H); 4.99 (q, 1H, J = 5.7 Hz); 4.75-4.46 (m, 5H); 4.13-4.01 (m, 1H); 3.90-3.85 (m, 2H); 3.80-3.67 (m, 1H); 3.58 (t, 1H, J = 8.4 Hz); 3.31-3.90 (m, 2H); 1.42 (s, 9H). 13 C-NMR (50 MHz 1 CDCl 3): 171.7; 155.0; 137.5; 135.9; 128.4; 127.9; 127.6; 127.5; 122.9; 121.9; 119.4; 118.6; 111.1; 110.0; 81.7; 80.6; 80.5; 80.2; 79.7; 78.6; 74.9; 72.5; 60.3; 54.3; 28.2.
IV ν (cm-1) filme (CH2CI2): 3344, 2974, 2933, 2876, 1740, 1706, 1500, 1455, 1366, 1247, 1166, 1068, 1023, 858, 743.IR ν (cm -1) film (CH 2 Cl 2): 3344, 2974, 2933, 2876, 1740, 1706, 1500, 1455, 1366, 1247, 1166, 1068, 1023, 858, 743.
αD20 = +114 (c 0,1 CH2CI2). Aspecto: óleo incolor.α D 20 = +114 (c 0.1 CH 2 Cl 2). Appearance: colorless oil.
HRMS calculada para C29H34N2O7Na, 545,2264; Encontrada, 545,2283.HRMS calcd for C 29 H 34 N 2 O 7 Na, 545.2264; Found 545.2283.
Exemplo 28: Obtenção do 1,4:3,6-dianidro-5-O-(benzil)-2-O-(N-carbobenziloxi- prolina-fenila!anina)-D-manitol (28)Example 28: Obtaining 1,4: 3,6-dianhydro-5-O- (benzyl) -2-O- (N-carbobenzyloxyproline-phenylamine) -D-mannitol (28)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,40-7,00 (m, 15H); 5,12-5,09 (m, 2H); 4,90 (q, 1H, J = 7,2 Hz); 4,75-4,72 (m, 2H); 4,54-4,46 (m, 2H); 4,35-4,29 (m, 1H); 4,07-3,90 (m, 4H); 3,70-3,60 (m, 1H); 3,49-3,38 (m, 3H); 3,20-3,02 (m, 2H); 1,90-1,65 (m, 4H). RMN13C (50 MHz, CDCI3): 171,0; 170,7; 155,9; 137,4; 136,3; 135,9; 129,2; 128,4; 15 127,8; 126,9; 126,8; 80,3; 80,1; 78,6; 74,9; 72,4; 70,9; 70,7; 70,4; 67,2; 60,1; 53,0; 46,7; 37,7; 27,7; 23,3.1 H NMR δ ppm (CDCl 3, 300 MHz): 7.40-7.00 (m, 15H); 5.12-5.09 (m, 2H); 4.90 (q, 1H, J = 7.2 Hz); 4.75-4.72 (m, 2H); 4.54-4.46 (m, 2H); 4.35-4.29 (m, 1H); 4.07-3.90 (m, 4H); 3.70-3.60 (m, 1H); 3.49-3.38 (m, 3H); 3.20-3.02 (m, 2H); 1.90-1.65 (m, 4H). 13 C-NMR (50 MHz, CDCl 3): 171.0; 170.7; 155.9; 137.4; 136.3; 135.9; 129.2; 128.4; 15 127.8; 126.9; 126.8; 80.3; 80.1; 78.6; 74.9; 72.4; 70.9; 70.7; 70.4; 67.2; 60.1; 53.0; 46.7; 37.7; 27.7; 23.3.
IV ν (cm"1) filme (CH2CI2): 3413, 3316, 3060, 2952, 2881, 1744, 1694, 1524, 1448, 1416, 1355, 1200, 1122, 1029, 918, 738. αD20 = +53 (c 0,1 CH2CI2). Aspecto: óleo incolor. HRMS calculada para C35H38N2O8Na, 637,2526; Encontrada, 637,2542.IR ν (cm -1) film (CH 2 Cl 2): 3413, 3316, 3060, 2952, 2881, 1744, 1694, 1524, 1448, 1416, 1355, 1200, 1122, 1029, 918, 738. ΑD20 = +53 (c 0.1 CH 2 Cl 2) Appearance: colorless oil HRMS calcd for C 35 H 38 N 2 O 8 Na 637.2526 Found 637.2542.
Exemplo 29: Obtenção do 1,4:3,6-dianidro-5-O-(benzil)-2-O-(N-carbobenziloxi- D-prolina)-D-manitol (29)Example 29: Obtaining 1,4: 3,6-dianhydro-5-O- (benzyl) -2-O- (N-carbobenzyloxy-D-proline) -D-mannitol (29)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,36-7,29 (m, 10H); 5,18-5,07 (m, 1H); 4,98 (q, 1H, J = 5,7 Hz); 4,79-4,64 (m, 2H); 4,54 (dd, 1H,J = 2,4; 11,7 Hz); 4,49-4,42 (m, 1H); 4,38 (dd, 1H, J = 3,9; 8,4 Hz); 4,06-3,89 (m, 2H); 3,88-3,80 (m, 2H); 3,70-3,45 (m, 4H); 2,19-1,96 (m, 2H); 1,94-1,68 (m, 2H).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.36-7.29 (m, 10H); 5.18-5.07 (m, 1H); 4.98 (q, 1H, J = 5.7 Hz); 4.79-4.64 (m, 2H); 4.54 (dd, 1H, J = 2.4; 11.7 Hz); 4.49-4.42 (m, 1H); 4.38 (dd, 1H, J = 3.9; 8.4 Hz); 4.06-3.89 (m, 2H); 3.88-3.80 (m, 2H); 3.70-3.45 (m, 4H); 2.19-1.96 (m, 2H); 1.94-1.68 (m, 2H).
RMN13C (50 MHz, CDCI3): 172,3; 160,1; 137,5; 136,6 128,5; 138,4; 128,3; 127,9; 127,8; 127,7; 80,6; 80,2; 78,6; 74,5; 72,5; 71,4; 70,1; 66,9; 58,8; 46,4; 30,8; 23,4. IV ν (cm1) (KBr): 3405, 2964, 2869, 1739, 1708, 1465, 1411, 1351, 1197, 1124, 1071, 880, 746.13 C-NMR (50 MHz, CDCl 3): 172.3; 160.1; 137.5; 136.6 128.5; 138.4; 128.3; 127.9; 127.8; 127.7; 80.6; 80.2; 78.6; 74.5; 72.5; 71.4; 70.1; 66.9; 58.8; 46.4; 30.8; 23.4. IR ν (cm -1) (KBr): 3405, 2964, 2869, 1739, 1708, 1465, 1411, 1351, 1197, 1124, 1071, 880, 746.
Pf = 71-72°C.Mp = 71-72 ° C.
αD20= + 158 (c 0,1 CH2CI2). Aspecto: sólido branco. HRMS calculada para C26H2QNO7Na, 490,1842; Encontrada, 490,1824.α D 20 = + 158 (c 0.1 CH 2 Cl 2). Appearance: white solid. HRMS calcd for C 26 H 20 NO 7 Na, 490.1842; Found, 490.1824.
Exemplo 30: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-0-(Na-f- butoxicarbonil-NE-carbobenziloxi-L-lisina)-D-manitol (30)Example 30: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-0- (Na-t-butoxycarbonyl-NE-carbobenzyloxy-L-lysine) -D-mannitol (30)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,35-7,27 (m, 10H); 5,15-5,08 (m, 2H); 4,89 (br s, 1H); 4,74-4,68 (m, 2H); 4,57 (d, 1H1 J= 11,7 Hz); 4,47-4,44 (m, 1H); 4,32 (br s, 1H); 4,03-3,89 (m, 4H); 3,60 (t, 1H, J = 8,4 Hz); 3,18 (q, 2H, J = 6,3 Hz); 1,90-1,181 (m, 2H); 1,66-1,52 (m, 2H); 1,50 (s, 9H); 1,48-1,42 (m, 2H).1 H NMR δ ppm (CDCl 3, 300 MHz): 7.35-7.27 (m, 10H); 5.15-5.08 (m, 2H); 4.89 (br s, 1H); 4.74-4.68 (m, 2H); 4.57 (d, 1H1 J = 11.7 Hz); 4.47-4.44 (m, 1H); 4.32 (br s, 1H); 4.03-3.89 (m, 4H); 3.60 (t, 1H, J = 8.4 Hz); 3.18 (q, 2H, J = 6.3 Hz); 1.90-1.181 (m, 2H); 1.66-1.52 (m, 2H); 1.50 (s, 9H); 1.48-1.42 (m, 2H).
RMN13C (50 MHz, CDCI3): 171,9; 156,4; 155,3; 137,5; 136,5; 128,5; 128,4; 128,0; 127,9; 127,8; 127,7; 80,5; 80,2; 79,8; 78,6; 74,8; 72,5; 71,0; 70,4; 66,5; 53,3; 40,5; 32,2; 29,9; 28,2; 22,2.13 C-NMR (50 MHz, CDCl 3): 171.9; 156.4; 155.3; 137.5; 136.5; 128.5; 128.4; 128.0; 127.9; 127.8; 127.7; 80.5; 80.2; 79.8; 78.6; 74.8; 72.5; 71.0; 70.4; 66.5; 53.3; 40.5; 32.2; 29.9; 28.2; 22.2.
IV ν (cm"1) filme (CH2CI2): 3344, 3033, 2938, 2874, 1710, 1522, 1455, 1366, 1255, 1165, 1091, 1024, 859, 737. ao20 = +79 (c 0,1 CH2CI2). Aspecto: óleo incolor. HRMS calculada para C32H42N2O9Na, 621,2788; Encontrada, 621.2784.IR ν (cm -1) film (CH 2 Cl 2): 3344, 3033, 2938, 2874, 1710, 1522, 1455, 1366, 1255, 1165, 1091, 1024, 859, 737. Ao 20 = +79 (c 0.1 CH 2 Cl 2 Appearance: colorless oil HRMS calcd for C 32 H 42 N 2 O 9 Na 621.2788 Found 621.2784.
Exemplo 31: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-0-(N-f-butoxicarbonil- L-ácido aspártico éster benzílico)-D-manitol (31)Example 31: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-0- (N-t-butoxycarbonyl-L-aspartic acid benzyl ester) -D-mannitol (31)
RMN 1H δ ppm (CDCI3, 300 MHz): 7,36-7,31 (m, 10H); 5,51 (d, 1H, J= 8,1 Hz); 5,11- 5,06 (m, 3H); 4,75 (d, 1H, J = 12 Hz); 4,70-4,61 (m, 2H); 4,57 (d, 1H, J = 11,7 Hz); 4,53 (t, 1H, J = 4,8 Hz); 4,05-3,99 (m, 1H); 3,93-3,81 (m, 3H); 3,56 (t, 1H, J= 8,4 Hz); 3,02 (dd, IH1J = 4,5; 17 Hz); 2,93 (dd, IH1J= 4,5; 17 Hz); 1,43 (s, 9H). RMN13C (50 MHz, CDCI3): 170,0; 170,4; 155,2; 137,5; 135,4; 128,5; 128,4; 128,3; 128,2; 127,9; 127,8; 80,5; 80,2; 80,0; 78,6; 75,2; 72,5; 70,9; 70,3; 66,7; 49,8; 36,7; 28,2.1 H NMR δ ppm (CDCl 3, 300 MHz): 7.36-7.31 (m, 10H); 5.51 (d, 1H, J = 8.1 Hz); 5.11 - 5.06 (m, 3H); 4.75 (d, 1H, J = 12 Hz); 4.70-4.61 (m, 2H); 4.57 (d, 1H, J = 11.7 Hz); 4.53 (t, 1H, J = 4.8 Hz); 4.05-3.99 (m, 1H); 3.93-3.81 (m, 3H); 3.56 (t, 1H, J = 8.4 Hz); 3.02 (dd, 1H H = 4.5; 17 Hz); 2.93 (dd, 1H H = 4.5; 17 Hz); 1.43 (s, 9H). 13 C-NMR (50 MHz, CDCl 3): 170.0; 170.4; 155.2; 137.5; 135.4; 128.5; 128.4; 128.3; 128.2; 127.9; 127.8; 80.5; 80.2; 80.0; 78.6; 75.2; 72.5; 70.9; 70.3; 66.7; 49.8; 36.7; 28.2.
IVv (cm"1) filme (CH2CI2): 3361, 2975, 2881, 1732, 1501, 1457, 1362, 1212, 1158, 1089, 1065, 1028, 859, 744. Ot020= +92 (c 0,1 CH2CI2). Aspecto: óleo incolor. HRMS calculada para C29H35NO7Na, 564,2210; Encontrada, 564,2190.IR (cm -1) film (CH 2 Cl 2): 3361, 2975, 2881, 1732, 1501, 1457, 1362, 1212, 1158, 1089, 1065, 1028, 859, 744. Ot020 = +92 (c 0.1 CH 2 Cl 2) Appearance: colorless oil HRMS calculated for C 29 H 35 NO 7 Na 566.2210 Found 564.2190.
Exemplo 32: Obtenção do 1,4:3,6-dianidro-5-0-(benzil)-2-0-(N-f-butoxicarbonil- L-ácido aspártico éster benzílico)-D-manitol (31) RMN 1H δ ppm (CDCI3, 300 MHz): 7,36-7,31 (m, 10H); 5,51 (d, 1 Η, J = 8,1 Hz); 5,11- 5,06 (m, 3H); 4,75 (d, 1H, J = 12 Hz); 4,70-4,61 (m, 2H); 4,57 (d, 1H, J = 11,7 Hz); 4,53 (t, 1H, J = 4,8 Hz); 4,05-3,99 (m, 1H); 3,93-3,81 (m, 3H); 3,56 (t, 1H,J = 8,4 Hz); 3,02 (dd, 1H,J = 4,5; 17 Hz); 2,93 (dd, 1H, J = 4,5; 17 Hz); 1,43 (s, 9H). RMN13C (50 MHz1 CDCI3): 170,0; 170,4; 155,2; 137,5; 135,4; 128,5; 128,4; 128,3; 128,2; 127,9; 127,8; 80,5; 80,2; 80,0; 78,6; 75,2; 72,5; 70,9; 70,3; 66,7; 49,8; 36,7; 28,2.Example 32: Obtaining 1,4: 3,6-dianhydro-5-0- (benzyl) -2-0- (Nf-butoxycarbonyl-L-aspartic acid benzyl ester) -D-mannitol (31) 1 H NMR δ ppm (CDCl3, 300 MHz): 7.36-7.31 (m, 10H); 5.51 (d, 1 Η, J = 8.1 Hz); 5.11 - 5.06 (m, 3H); 4.75 (d, 1H, J = 12 Hz); 4.70-4.61 (m, 2H); 4.57 (d, 1H, J = 11.7 Hz); 4.53 (t, 1H, J = 4.8 Hz); 4.05-3.99 (m, 1H); 3.93-3.81 (m, 3H); 3.56 (t, 1H, J = 8.4 Hz); 3.02 (dd, 1H, J = 4.5; 17 Hz); 2.93 (dd, 1H, J = 4.5; 17 Hz); 1.43 (s, 9H). 13 C-NMR (50 MHz 1 CDCl 3): 170.0; 170.4; 155.2; 137.5; 135.4; 128.5; 128.4; 128.3; 128.2; 127.9; 127.8; 80.5; 80.2; 80.0; 78.6; 75.2; 72.5; 70.9; 70.3; 66.7; 49.8; 36.7; 28.2.
IV ν (cm"1) filme (CH2Cl2): 3361, 2975, 2881, 1732, 1501, 1457, 1362, 1212, 1158, 1089, 1065, 1028, 859, 744. αD20 = (c,). Aspecto: óleo incolor.IR ν (cm -1) film (CH 2 Cl 2): 3361, 2975, 2881, 1732, 1501, 1457, 1362, 1212, 1158, 1089, 1065, 1028, 859, 744. αD20 = (c,). colorless.
HRMS calculada para C29H35NO7Na, 564,2210; Encontrada, 564,2190.HRMS calcd for C 29 H 35 NO 7 Na, 564.2210; Found 564.2190.
A descrição acima da presente invenção bem como os exemplos que foram apresentados atua com o propósito de ilustração e não limitam a invenção à forma aqui revelada e exemplificada. Em conseqüência; variações e modificações compatíveis com os ensinamentos acima; e a habilidade ou conhecimento da técnica relevante; estão dentro do escopo da presente invenção. As modalidades acima descritas e exemplificadas têm a intenção de melhor explicar os modos conhecidos para a prática da invenção e para permitir que os técnicos na área utilizem a invenção em tais; ou outras; modalidades e com várias modificações necessárias pelas aplicações específicas ou usos da presente invenção. É a intenção que a presente invenção inclua todas as modificações e variações da mesma; dentro do escopo descrito no relatório e nas reivindicações anexas.The above description of the present invention as well as the examples which have been presented are for illustration purposes and do not limit the invention to the form disclosed and exemplified herein. In consequence; variations and modifications consistent with the above teachings; and the skill or knowledge of the relevant technique; are within the scope of the present invention. The above described and exemplified embodiments are intended to further explain known methods for practicing the invention and to enable those skilled in the art to use the invention in such; or others; embodiments and with various modifications required by the specific applications or uses of the present invention. It is intended that the present invention include all modifications and variations thereof; within the scope described in the report and the appended claims.
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| US9540391B2 (en) | 2013-01-17 | 2017-01-10 | Sanofi | Isomannide derivatives as inhibitors of soluble epoxide hydrolase |
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| US9540391B2 (en) | 2013-01-17 | 2017-01-10 | Sanofi | Isomannide derivatives as inhibitors of soluble epoxide hydrolase |
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