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1.
Cell ; 171(3): 696-709.e23, 2017 Oct 19.
Artigo em Inglês | MEDLINE | ID: mdl-28965760

RESUMO

The transcription factor NRF2 is a master regulator of the cellular antioxidant response, and it is often genetically activated in non-small-cell lung cancers (NSCLCs) by, for instance, mutations in the negative regulator KEAP1. While direct pharmacological inhibition of NRF2 has proven challenging, its aberrant activation rewires biochemical networks in cancer cells that may create special vulnerabilities. Here, we use chemical proteomics to map druggable proteins that are selectively expressed in KEAP1-mutant NSCLC cells. Principal among these is NR0B1, an atypical orphan nuclear receptor that we show engages in a multimeric protein complex to regulate the transcriptional output of KEAP1-mutant NSCLC cells. We further identify small molecules that covalently target a conserved cysteine within the NR0B1 protein interaction domain, and we demonstrate that these compounds disrupt NR0B1 complexes and impair the anchorage-independent growth of KEAP1-mutant cancer cells. Our findings designate NR0B1 as a druggable transcriptional regulator that supports NRF2-dependent lung cancers.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/química , Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/química , Neoplasias Pulmonares/genética , Proteoma/análise , Transcriptoma , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Linhagem Celular Tumoral , Cisteína/metabolismo , Receptor Nuclear Órfão DAX-1/metabolismo , Redes Reguladoras de Genes , Humanos , Proteína 1 Associada a ECH Semelhante a Kelch/genética , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Ligantes , Neoplasias Pulmonares/metabolismo
2.
Mol Cell ; 65(6): 959-960, 2017 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-28306509

RESUMO

The PARK2 gene encodes an ubiquitin E3 ligase that is involved in mitochondrial homeostasis and linked to Parkinson's disease. In this issue, Gupta et al. (2017) demonstrate that PARK2 expression is frequently reduced in human cancers and that this alteration leads to dysregulated PI3K signaling.


Assuntos
Fosfatidilinositol 3-Quinases , Ubiquitina-Proteína Ligases/genética , Humanos , Mitocôndrias , Neoplasias , Doença de Parkinson/genética , Ubiquitina
3.
AAPS PharmSciTech ; 19(3): 971-977, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29302872

RESUMO

Antibody-drug conjugates (ADCs) are an emerging class of biopharmaceuticals. As such, there are no specific guidelines addressing impurity limits and qualification requirements. The current ICH guidelines on impurities, Q3A (Impurities in New Drug Substances), Q3B (Impurities in New Drug Products), and Q6B (Specifications: Test Procedures and Acceptance Criteria for Biotechnological/Biological Products) do not adequately address how to assess small molecule impurities in ADCs. The International Consortium for Innovation and Quality in Pharmaceutical Development (IQ) formed an impurities working group (IWG) to discuss this issue. This white paper presents a strategy for evaluating the impact of small molecule impurities in ADCs. This strategy suggests a science-based approach that can be applied to the design of control systems for ADC therapeutics. The key principles that form the basis for this strategy include the significant difference in molecular weights between small molecule impurities and the ADC, the conjugation potential of the small molecule impurities, and the typical dosing concentrations and dosing schedule. The result is that exposure to small impurities in ADCs is so low as to often pose little or no significant safety risk.


Assuntos
Contaminação de Medicamentos , Imunoconjugados/química , Peso Molecular
4.
BMC Genomics ; 17: 455, 2016 06 13.
Artigo em Inglês | MEDLINE | ID: mdl-27296290

RESUMO

BACKGROUND: Genome-scale functional genomic screens across large cell line panels provide a rich resource for discovering tumor vulnerabilities that can lead to the next generation of targeted therapies. Their data analysis typically has focused on identifying genes whose knockdown enhances response in various pre-defined genetic contexts, which are limited by biological complexities as well as the incompleteness of our knowledge. We thus introduce a complementary data mining strategy to identify genes with exceptional sensitivity in subsets, or outlier groups, of cell lines, allowing an unbiased analysis without any a priori assumption about the underlying biology of dependency. RESULTS: Genes with outlier features are strongly and specifically enriched with those known to be associated with cancer and relevant biological processes, despite no a priori knowledge being used to drive the analysis. Identification of exceptional responders (outliers) may not lead only to new candidates for therapeutic intervention, but also tumor indications and response biomarkers for companion precision medicine strategies. Several tumor suppressors have an outlier sensitivity pattern, supporting and generalizing the notion that tumor suppressors can play context-dependent oncogenic roles. CONCLUSIONS: The novel application of outlier analysis described here demonstrates a systematic and data-driven analytical strategy to decipher large-scale functional genomic data for oncology target and precision medicine discoveries.


Assuntos
Biomarcadores Tumorais , Genoma Humano , Genômica , Neoplasias/genética , Medicina de Precisão , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Biologia Computacional/métodos , Descoberta de Drogas , Perfilação da Expressão Gênica , Genômica/métodos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Terapia de Alvo Molecular , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Medicina de Precisão/métodos , Transdução de Sinais/efeitos dos fármacos
5.
Mol Cancer Ther ; 22(7): 891-900, 2023 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-37186518

RESUMO

KRAS is one of the most commonly mutated oncogenes in lung, colorectal, and pancreatic cancers. Recent clinical trials directly targeting KRAS G12C presented encouraging results for a large population of non-small cell lung cancer (NSCLC), but resistance to treatment is a concern. Continued exploration of new inhibitors and preclinical models is needed to address resistance mechanisms and improve duration of patient responses. To further enable the development of KRAS G12C inhibitors, we present a preclinical framework involving translational, non-invasive imaging modalities (CT and PET) and histopathology in a conventional xenograft model and a novel KRAS G12C knock-in mouse model of NSCLC. We utilized an in-house developed KRAS G12C inhibitor (Compound A) as a tool to demonstrate the value of this framework in studying in vivo pharmacokinetic/pharmacodynamic (PK/PD) relationship and anti-tumor efficacy. We characterized the Kras G12C-driven genetically engineered mouse model (GEMM) and identify tumor growth and signaling differences compared to its Kras G12D-driven counterpart. We also find that Compound A has comparable efficacy to sotorasib in the Kras G12C-driven lung tumors arising in the GEMM, but like observations in the clinic, some tumors inevitably progress on treatment. These findings establish a foundation for evaluating future KRAS G12C inhibitors that is not limited to xenograft studies and can be applied in a translationally relevant mouse model that mirrors human disease progression and resistance.


Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Animais , Camundongos , Humanos , Xenoenxertos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Transplante Heterólogo , Modelos Animais de Doenças , Mutação
6.
J Pharm Sci ; 112(12): 2965-2980, 2023 12.
Artigo em Inglês | MEDLINE | ID: mdl-37741455

RESUMO

Antibody-drug conjugates unite the specificity and long circulation time of an antibody with the toxicity of a chemical cytostatic or otherwise active drug using appropriate chemical linkers to reduce systemic toxicity and increase therapeutic index. This combination of a large biological molecule and a small molecule creates an increase in complexity. Multiple production processes are required to produce the native antibody, the drug and the linker, followed by conjugation of afore mentioned entities to form the final antibody-drug conjugate. The connected processes further increase the number of points of control, resulting in necessity of additional specifications and intensified analytical characterization. By combining scientific understanding of the production processes with risk-based approaches, quality can be demonstrated at those points where control is required and redundant comparability studies, specifications or product characterization are avoided. Over the product development lifecycle, this will allow process qualification to focus on those areas critical to quality and prevent redundant studies. The structure of the module 3 common technical document for an ADC needs to reflect each of the production processes and the combined overall approach to quality. Historically, regulatory authorities have provided varied expectations on its structure. This paper provides an overview of essential information to be included and shows that multiple approaches work as long as adequate cross-referencing is included.


Assuntos
Imunoconjugados , Imunoconjugados/química , Anticorpos Monoclonais/química
7.
Mol Cancer Ther ; 8(1): 1-9, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19139107

RESUMO

The phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway is currently one of the most exciting drug targets in oncology. However, only a short time ago, the paradigm existed that drugs targeted to the four PI3K class I isoforms would be too toxic for use in cancer therapy due to effects on physiologic signaling. Since that time, studies have delineated the roles of these four isoforms in nonpathologic signaling as well as their roles in cancer. An extensive effort has gone into developing agents that inhibit one or more PI3K isoforms, as well as closely related proteins implicated in cancer. These agents have proved to be tolerable and therapeutically beneficial in animal studies, and a number are in clinical testing. The agents, their properties, and their molecular targets are discussed in this review.


Assuntos
Neoplasias/tratamento farmacológico , Neoplasias/enzimologia , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase , Inibidores de Proteínas Quinases/uso terapêutico , Animais , Descoberta de Drogas , Humanos , Isoenzimas/antagonistas & inibidores , Isoenzimas/classificação , Isoenzimas/metabolismo , Fosfatidilinositol 3-Quinases/classificação , Inibidores de Proteínas Quinases/química , Transdução de Sinais
8.
Mol Cancer Ther ; 8(1): 94-100, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19139117

RESUMO

The phosphatidylinositol 3-kinase (PI3K)/Akt signaling cascade is an important component of the insulin signaling in normal tissues leading to glucose uptake and homeostasis and for cell survival signaling in cancer cells. Hyperglycemia is an on-target side effect of many inhibitors of PI3K/Akt signaling including the specific PI3K inhibitor PX-866. The peroxisome proliferator-activated receptor gamma agonist pioglitazone, used to treat type 2 diabetes, prevents a decrease in glucose tolerance caused by acute administration of PX-866. Our studies have shown that pioglitazone does not inhibit the antitumor activity of PX-866 in A-549 non-small cell lung cancer and HT-29 colon cancer xenografts. In vitro studies also showed that pioglitazone increases 2-[1-(14)C]deoxy-D-glucose uptake in L-6 muscle cells and prevents inhibition of 2-deoxyglucose uptake by PX-866. Neither pioglitazone nor PX-866 had an effect on 2-deoxyglucose uptake in A-549 lung cancer cells. In vivo imaging studies using [18F]2-deoxyglucose (FDG) positron emission tomography showed that pioglitazone increases FDG accumulation by normal tissue but does not significantly alter FDG uptake by A-549 xenografts. Thus, peroxisome proliferator-activated receptor gamma agonists may be useful in overcoming the increase in blood glucose caused by inhibitors of PI3K signaling by preventing the inhibition of normal tissue insulin-mediated glucose uptake without affecting antitumor activity.


Assuntos
Gonanos/farmacologia , Hiperglicemia/enzimologia , Hiperglicemia/prevenção & controle , PPAR gama/agonistas , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Tiazolidinedionas/farmacologia , Animais , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Progressão da Doença , Glucose/metabolismo , Proteínas Facilitadoras de Transporte de Glucose/metabolismo , Glucose-6-Fosfato/análogos & derivados , Glucose-6-Fosfato/metabolismo , Humanos , Camundongos , Neoplasias/tratamento farmacológico , Neoplasias/patologia , PPAR gama/metabolismo , Pioglitazona , Tiazolidinedionas/uso terapêutico , Ensaios Antitumorais Modelo de Xenoenxerto
9.
Mol Cancer Ther ; 7(9): 2621-32, 2008 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-18790745

RESUMO

AKT, a phospholipid-binding serine/threonine kinase, is a key component of the phosphoinositide 3-kinase cell survival signaling pathway that is aberrantly activated in many human cancers. Many attempts have been made to inhibit AKT; however, selectivity remains to be achieved. We have developed a novel strategy to inhibit AKT by targeting the pleckstrin homology (PH) domain. Using in silico library screening and interactive molecular docking, we have identified a novel class of non-lipid-based compounds that bind selectively to the PH domain of AKT, with "in silico" calculated K(D) values ranging from 0.8 to 3.0 micromol/L. In order to determine the selectivity of these compounds for AKT, we used surface plasmon resonance to measure the binding characteristics of the compounds to the PH domains of AKT1, insulin receptor substrate-1, and 3-phosphoinositide-dependent protein kinase 1. There was excellent correlation between predicted in silico and measured in vitro K(D)s for binding to the PH domain of AKT, which were in the range 0.4 to 3.6 micromol/L. Some of the compounds exhibited PH domain-binding selectivity for AKT compared with insulin receptor substrate-1 and 3-phosphoinositide-dependent protein kinase 1. The compounds also inhibited AKT in cells, induced apoptosis, and inhibited cancer cell proliferation. In vivo, the lead compound failed to achieve the blood concentrations required to inhibit AKT in cells, most likely due to rapid metabolism and elimination, and did not show antitumor activity. These results show that these compounds are the first small molecules selectively targeting the PH domain of AKT.


Assuntos
Desenho de Fármacos , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/química , Tiepinas/farmacologia , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Sequência de Aminoácidos , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacocinética , Antineoplásicos/farmacologia , Ligação Competitiva/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Ensaio de Imunoadsorção Enzimática , Feminino , Células HT29 , Humanos , Proteínas Substratos do Receptor de Insulina , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos SCID , Dados de Sequência Molecular , Estrutura Terciária de Proteína , Alinhamento de Sequência , Tiepinas/síntese química , Tiepinas/química , Tiepinas/farmacocinética
10.
Cancer Res ; 79(12): 3100-3111, 2019 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-31040156

RESUMO

Cnk1 (connector enhancer of kinase suppressor of Ras 1) is a pleckstrin homology (PH) domain-containing scaffold protein that increases the efficiency of Ras signaling pathways, imparting efficiency and specificity to the response of cell proliferation, survival, and migration. Mutated KRAS (mut-KRAS) is the most common proto-oncogenic event, occurring in approximately 25% of human cancers and has no effective treatment. In this study, we show that selective inhibition of Cnk1 blocks growth and Raf/Mek/Erk, Rho and RalA/B signaling in mut-KRAS lung and colon cancer cells with little effect on wild-type (wt)-KRAS cells. Cnk1 inhibition decreased anchorage-independent mut-KRas cell growth more so than growth on plastic, without the partial "addiction" to mut-KRAS seen on plastic. The PH domain of Cnk1 bound with greater affinity to PtdIns(4,5)P2 than PtdIns(3,4,5)P3, and Cnk1 localized to areas of the plasma membranes rich in PtdIns, suggesting a role for the PH domain in the biological activity of Cnk1. Through molecular modeling and structural modification, we identified a compound PHT-7.3 that bound selectively to the PH domain of Cnk1, preventing plasma membrane colocalization with mut-KRas. PHT-7.3 inhibited mut-KRas, but not wild-type KRas cancer cell and tumor growth and signaling. Thus, the PH domain of Cnk1 is a druggable target whose inhibition selectively blocks mutant KRas activation, making Cnk1 an attractive therapeutic target in patients with mut-KRAS-driven cancer. SIGNIFICANCE: These findings identify a therapeutic strategy to selectively block oncogenic KRas activity through the PH domain of Cnk1, which reduces its cell membrane binding, decreasing the efficiency of Ras signaling and tumor growth.


Assuntos
Antineoplásicos/farmacologia , Neoplasias do Colo/tratamento farmacológico , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Mutação , Neoplasias Pancreáticas/tratamento farmacológico , Proteínas Proto-Oncogênicas p21(ras)/genética , Bibliotecas de Moléculas Pequenas/farmacologia , Proliferação de Células , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Humanos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Domínios de Homologia à Plecstrina , Células Tumorais Cultivadas
11.
Cancer Chemother Pharmacol ; 58(4): 444-50, 2006 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-16485116

RESUMO

BACKGROUND: The purpose of the study was to evaluate the use of phospho-Akt in mouse and human skin as a surrogate target for tumor phospho-Akt to measure the effect of antitumor inhibitors of phosphatidylinositol-3-kinase (PI-3-K)/Akt (protein kinase B) signaling. METHOD: The expression of phosphoSer473-Akt was quantitatively assessed by Western blotting in human HT-29 colon, MCF-7 breast, A-549 non small cell lung tumor xenografts in mice, and by immunohistochemistry in mouse skin and human hair. RESULTS: The pattern of PI-3-K isoforms in human hair keratinocytes was similar to that in tumor but mouse hair keratinocytes showed a different pattern. A high level of phospho-Akt staining was present in keratinocytes of the external root sheath of the hair and was inhibited by the PI-3-K inhibitor PX-866 administered to mice, and in human hair exposed to PX-866 in culture. The inhibition of phospho-Akt by PX-866 in mouse hair keratinocytes was greater than inhibition of phospho-Akt in HT-29 and A-549 xenografts in the same mice. Phospho-Akt in mouse hair keratinocytes was inhibited by the Akt inhibitor PX-316 to a lesser degree than in MCF-7 tumor xenografts. CONCLUSIONS: Hair offers a way of measuring the effects of PI-3-K signaling inhibitors and, in cancer patients, may provide a readily obtainable surrogate tissue for assessing PI-3-K and phospho-Akt inhibition in tumor.


Assuntos
Inibidores Enzimáticos/farmacologia , Gonanos/farmacologia , Cabelo/enzimologia , Inibidores de Fosfoinositídeo-3 Quinase , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Pele/enzimologia , Animais , Linhagem Celular Tumoral , Humanos , Fosfatos de Inositol/farmacologia , Camundongos , Fosfatidilinositol 3-Quinases/metabolismo , Coelhos , Transdução de Sinais/efeitos dos fármacos
12.
Clin Cancer Res ; 11(12): 4338-40, 2005 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-15958615

RESUMO

PURPOSE: The purpose of the study was to evaluate the stability of phosphoprotein as a marker of signaling activity in human tumors using clinical samples and xenografts. EXPERIMENTAL DESIGN: The expression of phospho-Ser473-Akt (p-Akt) was assessed by immunohistochemistry in paraffin-embedded samples from patients enrolled in a Southwest Oncology Group clinical trial of gastroesophageal junction tumors and by immunohistochemistry and Western blotting in human colon tumor xenografts at various times after removal from the animal. RESULTS: Clinical samples had evaluable p-Akt staining only when obtained as biopsies (9 of 13) and no staining was observed in tumors obtained as surgically resected samples (0 of 15). In HT-29 colon cancer xenografts, p-Akt staining was present in fresh sample but not in tissue that had been allowed to stand for 30 minutes at room temperature. Western blotting of HT-29 tumor xenografts at room temperature showed a slow decrease in total Akt with a half-life of 180 minutes and a rapid decrease in p-Akt with a half-life of 20 minutes. CONCLUSIONS: Caution should be used when using phosphoprotein levels in human tumor specimens to measure intrinsic signaling activity or drug effects because of the potential for rapid dephosphorylation. Rapid processing of biopsies is essential and postoperative surgical samples may be of limited value because of the time to fixation.


Assuntos
Biomarcadores Tumorais/análise , Neoplasias do Colo/patologia , Fosfoproteínas/análise , Transdução de Sinais , Animais , Western Blotting , Neoplasias do Colo/metabolismo , Neoplasias do Colo/fisiopatologia , Células HT29 , Humanos , Imuno-Histoquímica , Camundongos , Camundongos SCID , Transplante de Neoplasias , Proteínas Serina-Treonina Quinases/análise , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/análise , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-akt , Serina/metabolismo , Transplante Heterólogo
13.
Mol Cancer Ther ; 4(9): 1349-57, 2005 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16170026

RESUMO

Epidermal growth factor receptor (EGFR) inhibitors such as gefitinib show antitumor activity in a subset of non-small cell lung cancer (NSCLC) patients having mutated EGFR. Recent work shows that phosphatidylinositol-3-kinase (PI3-K) is coupled to the EGFR only in NSCLC cell lines expressing ErbB-3 and that EGFR inhibitors do not inhibit PI3-K signaling in these cells. The central role PI3-K plays in cell survival suggests that a PI3-K inhibitor offers a strategy to increase the antitumor activity of EGFR inhibitors in resistant NSCL tumors that do not express ErbB-3. We show that PX-866, a PI3-K inhibitor with selectivity for p110alpha, potentiates the antitumor activity of gefitinib against even large A-549 NSCL xenografts giving complete tumor growth control in the early stages of treatment. A-549 xenograft phospho-Akt was inhibited by PX-866 but not by gefitinib. A major toxicity of PX-866 administration was hyperglycemia with decreased glucose tolerance, which was reversed upon cessation of treatment. The decreased glucose tolerance caused by PX-866 was insensitive to the AMP-activated protein kinase inhibitor metformin but reversed by insulin and by the peroxisome proliferator-activated receptor-gamma activator pioglitazone. Prolonged PX-866 administration also caused increased neutrophil counts. Thus, PX-866, by inhibiting PI3-K signaling, may have clinical use in increasing the response to EGFR inhibitors such as gefitinib in patients with NSCLC and possibly in other cancers who do not respond to EGFR inhibition.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Receptores ErbB/antagonistas & inibidores , Gonanos/farmacologia , Inibidores de Fosfoinositídeo-3 Quinase , Quinazolinas/farmacologia , Animais , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Inibidores Enzimáticos/farmacologia , Gefitinibe , Teste de Tolerância a Glucose , Humanos , Hiperglicemia/etiologia , Hipoglicemiantes/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Masculino , Metformina/farmacologia , Camundongos , Camundongos SCID , Neutrófilos/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Pioglitazona , Tiazolidinedionas/farmacologia , Transplante Heterólogo , Células Tumorais Cultivadas
14.
Mol Cancer Ther ; 3(7): 763-72, 2004 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15252137

RESUMO

We have developed biologically stable semisynthetic viridins as inhibitors of phosphoinositide (PtdIns)-3-kinases. The most active compound was PX-866 (acetic acid (1S,4E,10R,11R,13S,14R)-[4-diallylaminomethylene-6-hydroxy-1-methoxymethyl-10,13-dimethyl-3,7,17-trioxo-1,3,4,7,10,11,12,13,14,15,16,17-dodecahydro-2-oxa-cyclopenta[a]phenanthren-11-yl ester), which inhibited purified PtdIns-3-kinase with an IC50 of 0.1 nmol/L and PtdIns-3-kinase signaling measured by phospho-Ser473-Akt levels in HT-29 colon cancer cells with an IC50 of 20 nmol/L. PX-866 administered to mice at 10 mg/kg inhibited phospho-Ser473-Akt in HT-29 colon tumor xenografts up to 80% with recovery taking >48 hours after p.o. administration but more rapidly after i.v. or i.p. administration. PX-866 was eliminated from mouse plasma with a half-life of 18 minutes and a clearance of 360 mL/min/kg following i.v. administration and, when administered i.p. or p.o., showed first-pass metabolism with sequential N-deallylation. Synthetic standards of the N-deallylated metabolites of PX-866 inhibited PtdIns-3-kinase at low nanomolar per liter concentrations. PX-866 exhibited in vivo antitumor activity against s.c. OvCar-3 human ovarian cancer and A-549 human lung cancer xenografts in immunodeficient mice with log cell kills up to 1.2. PX-866 also increased the antitumor activity of cisplatin against A-549 xenografts and radiation treatment against OvCar-3 xenografts. The results show that PX-866 is a biologically stable broad-spectrum PtdIns-3-kinase inhibitor with good pharmacokinetics that causes prolonged inhibition of PtdIns-3-kinase signaling in human tumor xenografts. PX-866 exhibits single agent in vivo antitumor activity and increases the antitumor effects of cisplatin and radiation treatment.


Assuntos
Antineoplásicos/farmacologia , Inibidores Enzimáticos/farmacologia , Gonanos/farmacologia , Inibidores de Fosfoinositídeo-3 Quinase , Androstadienos/sangue , Androstadienos/farmacologia , Androstadienos/toxicidade , Androstenos/sangue , Androstenos/farmacologia , Androstenos/toxicidade , Animais , Anticorpos Fosfo-Específicos/imunologia , Antineoplásicos/química , Bacteriocinas/sangue , Bacteriocinas/farmacologia , Bacteriocinas/toxicidade , Linhagem Celular Tumoral , Cisplatino/farmacologia , Neoplasias do Colo/enzimologia , Inibidores Enzimáticos/química , Feminino , Gonanos/química , Humanos , Neoplasias Pulmonares/enzimologia , Camundongos , Camundongos SCID , Neoplasias Ovarianas/enzimologia , Neoplasias Ovarianas/radioterapia , Proteínas Serina-Treonina Quinases/análise , Proteínas Serina-Treonina Quinases/imunologia , Proteínas Proto-Oncogênicas/análise , Proteínas Proto-Oncogênicas/imunologia , Proteínas Proto-Oncogênicas c-akt , Wortmanina , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Oncol Res ; 14(10): 513-27, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15559765

RESUMO

Akt, a serine/threonine kinase that promotes cell survival, is activated by binding of its pleckstrin homology (PH) domain to membrane phosphatidylinositol (PtdIns)-3-phosphates formed by PtdIns-3-kinase. D-3-Deoxy-phosphatidyl-myo-inositols that cannot be phosphorylated on the 3-position of the myo-inositol group are inhibitors of the Akt PH domain. The most active compound is D-3-deoxy-phosphatidyl-myo-inositol 1-[(R)-2-methoxy-3-octadecyloxypropyl hydrogen phosphate] (PX-316). PX-316 administered intraperitoneally to mice at 150 mg/kg inhibits Akt activation in HT-29 human tumor xenografts up to 78% at 10 h with recovery to 34% at 48 h. Phosphorylation of GSK-3beta, a downstream target of Akt, is also inhibited. There is no decrease in PtdIns(3,4,5)-trisphosphate levels by PX-316, showing it is not an inhibitor of PtdIns-3-K in vivo. Gene expression profiling of HT-29 tumor xenografts shows many similarities between the effects of PX-316 and the PtdIns-3-K inhibitor wortmannin, with downregulation of several ribosomal-related genes, while PX-316 uniquely increases the expression of a group of mitochondrial-related genes. PX-316 has antitumor activity against early human MCF-7 breast cancer and HT-29 colon cancer xenografts in mice. PX-316 formulated in 20% hydroxypropyl-beta-cyclodextrin for intravenous administration is well tolerated in mice and rats with no hemolysis and no hematological toxicity. Thus, PX-316 is the lead compound of a new class of potential agents that inhibit Akt survival signaling.


Assuntos
Inibidores Enzimáticos/farmacologia , Regulação Neoplásica da Expressão Gênica , Fosfatos de Inositol/farmacologia , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , 2-Hidroxipropil-beta-Ciclodextrina , Androstadienos/farmacologia , Animais , Antineoplásicos/farmacologia , Sítios de Ligação , Linhagem Celular Tumoral , Sobrevivência Celular , Relação Dose-Resposta a Droga , Regulação para Baixo , Ativação Enzimática , Humanos , Concentração Inibidora 50 , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos SCID , Mitocôndrias/metabolismo , Modelos Químicos , Modelos Moleculares , Transplante de Neoplasias , Análise de Sequência com Séries de Oligonucleotídeos , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Ligação Proteica , Conformação Proteica , Estrutura Terciária de Proteína , Proteínas Proto-Oncogênicas c-akt , Serina/química , Transdução de Sinais , Fatores de Tempo , Wortmanina , beta-Ciclodextrinas/química
18.
J Natl Cancer Inst ; 104(3): 228-39, 2012 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-22247021

RESUMO

BACKGROUND: Mutations in the v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) play a critical role in cancer cell growth and resistance to therapy. Most mutations occur at codons 12 and 13. In colorectal cancer, the presence of any mutant KRas amino acid substitution is a negative predictor of patient response to targeted therapy. However, in non-small cell lung cancer (NSCLC), the evidence that KRAS mutation is a predictive factor is conflicting. METHODS: We used data from a molecularly targeted clinical trial for 215 patients with tissues available out of 268 evaluable patients with refractory NSCLC to examine associations between specific mutant KRas proteins and progression-free survival and tumor gene expression. Transcriptome microarray studies of patient tumor samples and reverse-phase protein array studies of a panel of 67 NSCLC cell lines with known substitutions in KRas and in immortalized human bronchial epithelial cells stably expressing different mutant KRas proteins were used to investigate signaling pathway activation. Molecular modeling was used to study the conformations of wild-type and mutant KRas proteins. Kaplan-Meier curves and Cox regression were used to analyze survival data. All statistical tests were two-sided. RESULTS: Patients whose tumors had either mutant KRas-Gly12Cys or mutant KRas-Gly12Val had worse progression-free survival compared with patients whose tumors had other mutant KRas proteins or wild-type KRas (P = .046, median survival = 1.84 months) compared with all other mutant KRas (median survival = 3.35 months) or wild-type KRas (median survival = 1.95 months). NSCLC cell lines with mutant KRas-Gly12Asp had activated phosphatidylinositol 3-kinase (PI-3-K) and mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK) signaling, whereas those with mutant KRas-Gly12Cys or mutant KRas-Gly12Val had activated Ral signaling and decreased growth factor-dependent Akt activation. Molecular modeling studies showed that different conformations imposed by mutant KRas may lead to altered association with downstream signaling transducers. CONCLUSIONS: Not all mutant KRas proteins affect patient survival or downstream signaling in a similar way. The heterogeneous behavior of mutant KRas proteins implies that therapeutic interventions may need to take into account the specific mutant KRas expressed by the tumor.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Genes ras , Neoplasias Pulmonares/metabolismo , Terapia de Alvo Molecular , Mutação , Transdução de Sinais , Ácido Aspártico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Linhagem Celular Tumoral , Ensaios Clínicos Fase II como Assunto , Cisteína , Intervalo Livre de Doença , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Genes ras/genética , Vetores Genéticos , Glicina , Humanos , Immunoblotting , Imunoprecipitação , Estimativa de Kaplan-Meier , Lentivirus , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/mortalidade , Análise em Microsséries , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ensaios Clínicos Controlados Aleatórios como Assunto , Serina-Treonina Quinases TOR/metabolismo , Resultado do Tratamento , Valina
19.
Curr Opin Drug Discov Devel ; 13(1): 41-9, 2010 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-20047145

RESUMO

The four isoforms of class I phosphatidylinositol-3-kinase (PI3K) were originally thought to be redundant in function; however, further research and new technologies have revealed that each subunit has distinct characteristics. In the past decade the number of PI3K inhibitors has increased from a few agents with unacceptable promiscuity and pharmacological properties, to a family of selective agents that are either progressing through experimental development or are in clinical trials. These agents, with two notable exceptions, target multiple members of the PI3K class I isoforms. As data become increasingly available, the concept that inhibiting a single PI3K isoform may offer improved therapeutic benefit, while eliminating the potentially negative effects of pan-isoform inhibition, is driving efforts to develop more specific inhibitors. However, questions remain regarding the best isoform to inhibit for maximum benefit in different pathological settings, and whether increased specificity may lead to a loss in efficacy as a result of isoform redundancy in some settings. This review discusses the current understanding of individual PI3K isoforms in physiology and pathological states, as well as the status of PI3K inhibitors in preclinical and clinical development.


Assuntos
Inibidores Enzimáticos/farmacologia , Fosfatidilinositol 3-Quinases/fisiologia , Inibidores de Fosfoinositídeo-3 Quinase , Animais , Inibidores Enzimáticos/química , Inibidores Enzimáticos/uso terapêutico , Oncologia , Modelos Biológicos , Fosfatidilinositol 3-Quinases/classificação , Isoformas de Proteínas/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia
20.
Mol Aspects Med ; 31(2): 135-44, 2010 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-20176047

RESUMO

The phosphatidylinositol-3-kinase (PI3K) signaling pathway is implicated in multiple aspects of tumorigenesis and tumor maintenance, and recent years have seen significant efforts towards developing agents to inhibit the pathway. However, the development of such agents raises issues such as what specific member or members in the PI3K family should be inhibited to achieve maximal therapeutic benefit, and can specific inhibitors be developed with the necessary pharmacologic properties to allow them to proceed to clinical trials? The number of PI3K inhibitors has gone from a handful of archetypal inhibitors which largely determined how the pathway was initially defined through their inhibition of PI3K, but also due to their off target properties, to a much larger number of inhibitors of not only PI3K but also other members of the PI3K family. The question remains to be answered whether greater therapeutic efficacy will be obtained through the use of inhibitors with increased specificity, or through inhibitors that target a spectrum of targets within the pathway. This review will cover the development of agents targeting the pathway, and will discuss current issues surrounding the development of such agents.


Assuntos
Antineoplásicos/farmacologia , Inibidores Enzimáticos/farmacologia , Neoplasias/tratamento farmacológico , Inibidores de Fosfoinositídeo-3 Quinase , Animais , Antineoplásicos/uso terapêutico , Inibidores Enzimáticos/uso terapêutico , Humanos , Neoplasias/enzimologia , Transdução de Sinais
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