RESUMO
The vascular endothelial growth factor (VEGF)/VEGFR and hepatocyte growth factor (HGF)/c-MET signaling pathways act synergistically to promote angiogenesis. Studies indicate VEGF inhibition leads to increased levels of phosphorylated c-MET, bypassing VEGF-mediated angiogenesis and leading to chemoresistance. We conducted a phase 1 clinical trial with 32 patients with refractory solid tumors to evaluate the safety, pharmacokinetics, and pharmacodynamics of combinations of VEGF-targeting pazopanib and the putative c-MET inhibitor ARQ197 (tivantinib) at 5 dose levels (DLs). Patients either took pazopanib and tivantinib from treatment initiation (escalation phase) or pazopanib alone for 7 days, with paired tumor sampling, prior to starting combination treatment (expansion phase). Hypertension was the most common adverse event. No more than 1 dose limiting toxicity (DLT) occurred at any DL, so the maximum tolerated dose (MTD) was not determined; DL5 (800 mg pazopanib daily and 360 mg tivantinib BID) was used during the expansion phase. Twenty of 31 evaluable patients achieved stable disease lasting up to 22 cycles. Circulating VEGF, VEGFR2, HGF, and c-MET levels were assessed, and only VEGF levels increased. Tumor c-MET levels (total and phosphorylated) were determined in paired biopsies before and after 7 days of pazopanib treatment. Total intact c-MET decreased in 6 of 7 biopsy pairs, in contrast to previously reported c-MET elevation in response to VEGF inhibition. These results are discussed in the context of our previously reported analysis of epithelial-mesenchymal transition in these tumors.
Assuntos
Inibidores da Angiogênese/uso terapêutico , Indazóis/uso terapêutico , Neoplasias/tratamento farmacológico , Pirimidinas/uso terapêutico , Pirrolidinonas/uso terapêutico , Quinolinas/uso terapêutico , Sulfonamidas/uso terapêutico , Inibidores da Angiogênese/administração & dosagem , Inibidores da Angiogênese/efeitos adversos , Inibidores da Angiogênese/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica , Área Sob a Curva , Relação Dose-Resposta a Droga , Esquema de Medicação , Fator de Crescimento de Hepatócito/metabolismo , Humanos , Indazóis/administração & dosagem , Indazóis/efeitos adversos , Indazóis/farmacocinética , Dose Máxima Tolerável , Pessoa de Meia-Idade , Neoplasias/patologia , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Pirimidinas/administração & dosagem , Pirimidinas/efeitos adversos , Pirimidinas/farmacocinética , Pirrolidinonas/administração & dosagem , Pirrolidinonas/efeitos adversos , Pirrolidinonas/farmacocinética , Quinolinas/administração & dosagem , Quinolinas/efeitos adversos , Quinolinas/farmacocinética , Sulfonamidas/administração & dosagem , Sulfonamidas/efeitos adversos , Sulfonamidas/farmacocinética , Fator A de Crescimento do Endotélio Vascular/efeitos dos fármacosRESUMO
Integrase strand transfer inhibitors (INSTIs) are highly effective against HIV infections. Co-crystal structures of the prototype foamy virus intasome have shown that all three FDA-approved drugs, raltegravir (RAL), elvitegravir and dolutegravir (DTG), act as interfacial inhibitors during the strand transfer (ST) integration step. However, these structures give only a partial sense for the limited inhibition of the 3'-processing reaction by INSTIs and how INSTIs can be modified to overcome drug resistance, notably against the G140S-Q148H double mutation. Based on biochemical experiments with modified oligonucleotides, we demonstrate that both the viral DNA +1 and -1 bases, which flank the 3'-processing site, play a critical role for 3'-processing efficiency and inhibition by RAL and DTG. In addition, the G140S-Q148H (SH) mutant integrase, which has a reduced 3'-processing activity, becomes more active and more resistant to inhibition of 3'-processing by RAL and DTG in the absence of the -1 and +1 bases. Molecular modeling of HIV-1 integrase, together with biochemical data, indicate that the conserved residue Q146 in the flexible loop of HIV-1 integrase is critical for productive viral DNA binding through specific contacts with the virus DNA ends in the 3'-processing and ST reactions. The potency of integrase inhibitors against 3'-processing and their ability to overcome resistance is discussed.
Assuntos
Domínio Catalítico , DNA Viral/metabolismo , Farmacorresistência Viral/efeitos dos fármacos , Inibidores de Integrase de HIV/farmacologia , Integrase de HIV/metabolismo , Biocatálise/efeitos dos fármacos , Guanina/metabolismo , Integrase de HIV/química , Inibidores de Integrase de HIV/química , Íons , Magnésio/farmacologia , Modelos Moleculares , Mutação/genética , Especificidade por Substrato/efeitos dos fármacosRESUMO
The vinylogous urea, NSC727447, was proposed to allosterically inhibit ribonuclease H (RNase H) activity of human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) by interacting with the thumb subdomain of its non-catalytic p51 subunit. Proximity of the p51 thumb to the p66 RNase H domain implied that inhibitor binding altered active site geometry, whereas protein footprinting suggested a contribution from α-helix I residues Cys-280 and Lys-281. To more thoroughly characterize the vinylogous urea binding site, horizontal alanine scanning mutagenesis between p51 residues Lys-275 and Thr-286 (comprising α-helix I and portions of the neighboring αH/αI and αI/αJ connecting loops) was combined with a limited vertical scan of Cys-280. A contribution from Cys-280 was strengthened by our observation that all substitutions at this position rendered selectively mutated, reconstituted p66/p51 heterodimers â¼45-fold less sensitive to inhibition. An â¼19-fold reduced IC(50) for p51 mutant T286A coupled with a 2-8-fold increased IC(50) when intervening residues were substituted supports our original proposal of p51 α-helix I as the vinylogous urea binding site. In contrast to these allosteric inhibitors, mutant enzymes retained equivalent sensitivity to the natural product α-hydroxytropolone inhibitor manicol, which x-ray crystallography has demonstrated functions by chelating divalent metal at the p66 RNase H active site. Finally, reduced DNA strand-transfer activity together with increased vinylogous urea sensitivity of p66/p51 heterodimers containing short p51 C-terminal deletions suggests an additional role for the p51 C terminus in nucleic acid binding that is compromised by inhibitor binding.
Assuntos
Inibidores Enzimáticos/química , HIV-1/enzimologia , Ribonuclease H do Vírus da Imunodeficiência Humana/antagonistas & inibidores , Ribonuclease H do Vírus da Imunodeficiência Humana/química , Tiofenos/química , Substituição de Aminoácidos , Linhagem Celular , Cristalografia por Raios X , HIV-1/genética , Humanos , Mutagênese , Mutação de Sentido Incorreto , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Ribonuclease H do Vírus da Imunodeficiência Humana/genéticaRESUMO
BACKGROUND: The recently approved anti-AIDS drug rilpivirine (TMC278, Edurant) is a nonnucleoside inhibitor (NNRTI) that binds to reverse transcriptase (RT) and allosterically blocks the chemical step of DNA synthesis. In contrast to earlier NNRTIs, rilpivirine retains potency against well-characterized, clinically relevant RT mutants. Many structural analogues of rilpivirine are described in the patent literature, but detailed analyses of their antiviral activities have not been published. This work addresses the ability of several of these analogues to inhibit the replication of wild-type (WT) and drug-resistant HIV-1. RESULTS: We used a combination of structure activity relationships and X-ray crystallography to examine NNRTIs that are structurally related to rilpivirine to determine their ability to inhibit WT RT and several clinically relevant RT mutants. Several analogues showed broad activity with only modest losses of potency when challenged with drug-resistant viruses. Structural analyses (crystallography or modeling) of several analogues whose potencies were reduced by RT mutations provide insight into why these compounds were less effective. CONCLUSIONS: Subtle variations between compounds can lead to profound differences in their activities and resistance profiles. Compounds with larger substitutions replacing the pyrimidine and benzonitrile groups of rilpivirine, which reorient pocket residues, tend to lose more activity against the mutants we tested. These results provide a deeper understanding of how rilpivirine and related compounds interact with the NNRTI binding pocket and should facilitate development of novel inhibitors.
Assuntos
Infecções por HIV/virologia , Transcriptase Reversa do HIV/genética , HIV-1/efeitos dos fármacos , HIV-1/enzimologia , Nitrilas/farmacologia , Pirimidinas/farmacologia , Inibidores da Transcriptase Reversa/farmacologia , Linhagem Celular , Cristalografia , Infecções por HIV/tratamento farmacológico , Transcriptase Reversa do HIV/antagonistas & inibidores , Transcriptase Reversa do HIV/metabolismo , HIV-1/genética , HIV-1/fisiologia , Humanos , Modelos Moleculares , Estrutura Molecular , Mutação , Nitrilas/síntese química , Nitrilas/química , Pirimidinas/síntese química , Pirimidinas/química , Inibidores da Transcriptase Reversa/síntese química , Inibidores da Transcriptase Reversa/química , RilpivirinaRESUMO
The design of novel integrase (IN) inhibitors has been aided by recent crystal structures revealing the binding mode of these compounds with a full-length prototype foamy virus (PFV) IN and synthetic viral DNA ends. Earlier docking studies relied on incomplete structures and did not include the contribution of the viral DNA to inhibitor binding. Using the structure of PFV IN as the starting point, we generated a model of the corresponding HIV-1 complex and developed a molecular dynamics (MD)-based approach that correlates with the in vitro activities of novel compounds. Four well-characterized compounds (raltegravir, elvitegravir, MK-0536, and dolutegravir) were used as a training set, and the data for their in vitro activity against the Y143R, N155H, and G140S/Q148H mutants were used in addition to the wild-type (WT) IN data. Three additional compounds were docked into the IN-DNA complex model and subjected to MD simulations. All three gave interaction potentials within 1 standard deviation of values estimated from the training set, and the most active compound was identified. Additional MD analysis of the raltegravir- and dolutegravir-bound complexes gave internal and interaction energy values that closely match the experimental binding energy of a compound related to raltegravir that has similar activity. These approaches can be used to gain a deeper understanding of the interactions of the inhibitors with the HIV-1 intasome and to identify promising scaffolds for novel integrase inhibitors, in particular, compounds that retain activity against a range of drug-resistant mutants, making it possible to streamline synthesis and testing.
Assuntos
DNA Viral/metabolismo , Infecções por HIV/tratamento farmacológico , Inibidores de Integrase de HIV/farmacologia , Integrase de HIV/metabolismo , HIV-1/efeitos dos fármacos , Simulação de Dinâmica Molecular , Spumavirus/química , Substituição de Aminoácidos , Sítios de Ligação , DNA Viral/química , Desenho de Fármacos , Farmacorresistência Viral , Infecções por HIV/virologia , Integrase de HIV/química , Inibidores de Integrase de HIV/química , Inibidores de Integrase de HIV/metabolismo , HIV-1/química , HIV-1/genética , Compostos Heterocíclicos com 3 Anéis/química , Compostos Heterocíclicos com 3 Anéis/metabolismo , Compostos Heterocíclicos com 3 Anéis/farmacologia , Humanos , Mutação , Oxazinas , Piperazinas , Ligação Proteica/efeitos dos fármacos , Piridonas , Pirrolidinonas/química , Pirrolidinonas/metabolismo , Pirrolidinonas/farmacologia , Relação Quantitativa Estrutura-Atividade , Quinolonas/química , Quinolonas/metabolismo , Quinolonas/farmacologia , Raltegravir Potássico , Spumavirus/genética , TermodinâmicaRESUMO
Although an extensive body of scientific and patent literature exists describing the development of HIV-1 integrase (IN) inhibitors, Merck's raltegravir and Gilead's elvitegravir remain the only IN inhibitors FDA-approved for the treatment of AIDS. The emergence of raltegravir-resistant strains of HIV-1 containing mutated forms of IN underlies the need for continued efforts to enhance the efficacy of IN inhibitors against resistant mutants. We have previously described bicyclic 6,7-dihydroxyoxoisoindolin-1-ones that show good IN inhibitory potency. This report describes the effects of introducing substituents into the 4- and 5-positions of the parent 6,7-dihydroxyoxoisoindolin-1-one platform. We have developed several sulfonamide-containing analogs that enhance potency in cell-based HIV assays by more than two orders-of-magnitude and we describe several compounds that are more potent than raltegravir against the clinically relevant Y143R IN mutant.
Assuntos
Inibidores de Integrase de HIV/farmacologia , Integrase de HIV/metabolismo , Indóis/farmacologia , Sulfonamidas/farmacologia , Síndrome da Imunodeficiência Adquirida/tratamento farmacológico , Síndrome da Imunodeficiência Adquirida/virologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Integrase de HIV/genética , Inibidores de Integrase de HIV/síntese química , Inibidores de Integrase de HIV/química , HIV-1/efeitos dos fármacos , HIV-1/enzimologia , Humanos , Indóis/síntese química , Indóis/química , Testes de Sensibilidade Microbiana , Estrutura Molecular , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/químicaRESUMO
Preclinical studies provide valuable data in the early development of novel drugs for patients with cancer. Many cancer treatment regimens now utilize multiple agents with different targets to delay the emergence of drug-resistant tumor cells, and experimental agents are often evaluated in combination with FDA-approved drugs. The Biological Testing Branch (BTB) of the U.S. NCI has evaluated more than 70 FDA-approved oncology drugs to date in human xenograft models. Here, we report the first release of a publicly available, downloadable spreadsheet, ROADMAPS (Responses to Oncology Agents and Dosing in Models to Aid Preclinical Studies, dtp.cancer.gov/databases_tools/roadmaps.htm), that provides data filterable by agent, dose, dosing schedule, route of administration, tumor models tested, responses, host mouse strain, maximum weight loss, drug-related deaths, and vehicle formulation for preclinical experiments conducted by the BTB. Data from 70 different single targeted and cytotoxic agents and 140 different xenograft models were included. Multiple xenograft models were tested in immunocompromised mice for many cancer histologies, with lung cancer as the most broadly tested (24 models). Many of the dose levels and schedules used in these experiments were comparable with those tolerated in humans. Targeted and cytotoxic single agents were included. The online spreadsheet will be updated periodically as additional agent/dose/model combinations are evaluated. ROADMAPS is intended to serve as a publicly available resource for the research community to inform the design of clinically relevant, tolerable single and combinatorial regimens in preclinical mouse models. SIGNIFICANCE: ROADMAPS includes data that can be used to identify tolerable dosing regimens with activity against a variety of human tumors in different mouse strains, providing a resource for planning preclinical studies.
Assuntos
Antineoplásicos , Neoplasias , Animais , Antineoplásicos/efeitos adversos , Humanos , Camundongos , Neoplasias/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Ras/Raf/MEK/ERK (MAPK) and PI3K/AKT signaling pathways influence several cell functions involved in oncogenesis, making them attractive drug targets. We describe a novel multiplex immunoassay to quantitate isoform-specific phosphorylation of proteins in the PI3K/AKT and MAPK pathways as a tool to assess pharmacodynamic changes. Isoform-specific assays measuring total protein and site-specific phosphorylation levels of ERK1/2, MEK1/2, AKT1/2/3, and rpS6 were developed on the Luminex platform with validated antibody reagents. The multiplex assay demonstrated satisfactory analytic performance. Fit-for-purpose validation was performed with xenograft models treated with selected agents. In PC3 and HCC70 xenograft tumors, the PI3Kß inhibitor AZD8186 suppressed phosphorylation of AKT1, AKT2, and rpS6 for 4 to 7 hours post single dose, but levels returned to baseline by 24 hours. AKT3 phosphorylation was suppressed in PC3 xenografts at all doses tested, but only at the highest dose in HCC70. The AKT inhibitor MK-2206 reduced AKT1/2/3 phosphorylation in SW620 xenograft tumors 2 to 4 hours postdose, and the MEK inhibitor selumetinib reduced MEK1/2 and ERK1/2 phosphorylation by up to 50% and >90%, respectively. Clinical utility was demonstrated by analyzing biopsies from untreated patients with plexiform neurofibromas enrolled in a clinical trial of selumetinib (NCT02407405). These biopsies showed MEK and ERK phosphorylation levels sufficient for measuring up to 90% inhibition, and low AKT and rpS6 phosphorylation. This validated multiplex immunoassay demonstrates the degree and duration of phosphorylation modulation for three distinct classes of drugs targeting the PI3K/AKT and MAPK pathways.
Assuntos
Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Animais , Feminino , Humanos , Camundongos , Camundongos Nus , Fosforilação , Isoformas de Proteínas , Transdução de Sinais , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Differential responses to tamoxifen may be due to inter-patient variability in tamoxifen metabolism into pharmacologically active Z-endoxifen. Z-endoxifen administration was anticipated to bypass these variations, increasing active drug levels, and potentially benefitting patients responding sub-optimally to tamoxifen. MATERIALS AND METHODS: Patients with treatment-refractory gynecologic malignancies, desmoid tumors, or hormone receptor-positive solid tumors took oral Z-endoxifen daily with a 3+3 phase 1 dose escalation format over 8 dose levels (DLs). Safety, pharmacokinetics/pharmacodynamics, and clinical outcomes were evaluated. RESULTS: Thirty-four of 40 patients were evaluable. No maximum tolerated dose was established. DL8, 360 mg/day, was used for the expansion phase and is higher than doses administered in any previous study; it also yielded higher plasma Z-endoxifen concentrations. Three patients had partial responses and 8 had prolonged stable disease (≥ 6 cycles); 44.4% (8/18) of patients at dose levels 6-8 achieved one of these outcomes. Six patients who progressed after tamoxifen therapy experienced partial response or stable disease for ≥ 6 cycles with Z-endoxifen; one with desmoid tumor remains on study after 62 cycles (nearly 5 years). CONCLUSIONS: Evidence of antitumor activity and prolonged stable disease are achieved with Z-endoxifen despite prior tamoxifen therapy, supporting further study of Z-endoxifen, particularly in patients with desmoid tumors.
RESUMO
Hourly rounding by nursing staff helps to proactively manage patient needs and minimize the number of unscheduled calls from patients. The focus of this study was to determine if an increased emphasis on hourly rounding had an effect on call bell usage on an oncology unit. Patient call bell usage requests, such as asking for water or repositioning, and the total number of all alarms, such as bed exit alarms and lavatory assist alarms, decreased. Subsequent patient satisfaction surveys showed an increase in patient perception of how quickly help was received.
Assuntos
Alarmes Clínicos/normas , Recursos Humanos de Enfermagem Hospitalar/normas , Enfermagem Oncológica/normas , Segurança do Paciente/normas , Satisfação do Paciente/estatística & dados numéricos , Visitas de Preceptoria/normas , Adulto , Idoso , Idoso de 80 Anos ou mais , Alarmes Clínicos/estatística & dados numéricos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Recursos Humanos de Enfermagem Hospitalar/estatística & dados numéricos , Enfermagem Oncológica/estatística & dados numéricos , Segurança do Paciente/estatística & dados numéricos , Visitas de Preceptoria/estatística & dados numéricos , Fatores de Tempo , Estados UnidosRESUMO
Integrase (IN) inhibitors are the newest class of antiretroviral agents developed for the treatment of HIV-1 infections. Merck's Raltegravir (RAL) (October 2007) and Gilead's Elvitegravir (EVG) (August 2012), which act as IN strand transfer inhibitors (INSTIs), were the first anti-IN drugs to be approved by the FDA. However, the virus develops resistance to both RAL and EVG, and there is extensive cross-resistance to these two drugs. New "2nd-generation" INSTIs are needed that will have greater efficacy against RAL- and EVG-resistant strains of IN. The FDA has recently approved the first second generation INSTI, GSK's Dolutegravir (DTG) (August 2013). Our current article describes the design, synthesis, and evaluation of a series of 1,8-dihydroxy-2-oxo-1,2-dihydroquinoline-3-carboxamides, 1,4-dihydroxy-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamides, and 1-hydroxy-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamides. This resulted in the identification of noncytotoxic inhibitors that exhibited single digit nanomolar EC50 values against HIV-1 vectors harboring wild-type IN in cell-based assays. Importantly, some of these new inhibitors retain greater antiviral efficacy compared to that of RAL when tested against a panel of IN mutants that included Y143R, N155H, G140S/Q148H, G118R, and E138K/Q148K.
Assuntos
Fármacos Anti-HIV/química , Fármacos Anti-HIV/farmacologia , Integrase de HIV/genética , HIV-1/efeitos dos fármacos , Piridinas/química , Piridinas/farmacologia , Pirrolidinonas/farmacologia , Linhagem Celular , Farmacorresistência Viral , Humanos , Espectroscopia de Ressonância Magnética , Mutação , Raltegravir Potássico , Espectrometria de Massas por Ionização por ElectrosprayRESUMO
Although there are structures of the different domains of human immunodeficiency virus type 1 (HIV-1) integrase (IN), there is no structure of the entire protein. The recently determined crystal structures of the prototype foamy virus (PFV) IN tetramer, in complexes with viral DNA, led to the generation of models of full-length HIV-1 IN. These models were generated, in part, by superimposing the structures of the domains of HIV-1 IN onto the structure of full-length PFV IN. We developed a model for HIV-1 IN-based solely on its sequence alignment with PFV IN-that differs in several ways from the previous models. Specifically, in our model, the junction between the catalytic core domain and C-terminal domain adopts a helix-loop-helix motif that is similar to the corresponding segment of PFV IN and differs from the crystal structures of these two HIV-1 IN domains. The alignment of residues in the C-terminal domain also differs from the previous models. Our model can be used to explain the phenotype of previously published HIV-1 IN mutants. We made additional mutants, and the behavior of these new mutants provides additional support for the model.
Assuntos
Integrase de HIV/química , Integrase de HIV/metabolismo , Mutagênese Sítio-Dirigida , Mutação de Sentido Incorreto , Sequência de Aminoácidos , Integrase de HIV/genética , HIV-1/química , HIV-1/genética , Modelos Moleculares , Dados de Sequência Molecular , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Conformação Proteica , Alinhamento de Sequência , Spumavirus/química , Spumavirus/genéticaRESUMO
On the basis of a series of lactam and phthalimide derivatives that inhibit HIV-1 integrase, we developed a new molecule, XZ-259, with biochemical and antiviral activities comparable to raltegravir. We determined the crystal structures of XZ-259 and four other derivatives in complex with the prototype foamy virus intasome. The compounds bind at the integrase-Mg(2+)-DNA interface of the integrase active site. In biochemical and antiviral assays, XZ-259 inhibits raltegravir-resistant HIV-1 integrases harboring the Y143R mutation. Molecular modeling is also presented suggesting that XZ-259 can bind in the HIV-1 intasome with its dimethyl sulfonamide group adopting two opposite orientations. Molecular dynamics analyses of the HIV-1 intasome highlight the importance of the viral DNA in drug potency.