RESUMO
BACKGROUND: The prognosis of patients with different gastrointestinal cancers varies widely. Despite advances in treatment strategies, such as extensive resections and the addition of new drugs to chemotherapy regimens, conventional treatment strategies have failed to improve survival for many tumours. Although promising, the clinical application of molecularly guided personalized treatment has proven to be challenging. This narrative review focuses on the personalization of cancer therapy using patient-derived three-dimensional 'organoid' models. METHODS: A PubMed search was conducted to identify relevant articles. An overview of the literature and published protocols is presented, and the implications of these models for patients with cancer, surgeons and oncologists are explained. RESULTS: Organoid culture methods have been established for healthy and diseased tissues from oesophagus, stomach, intestine, pancreas, bile duct and liver. Because organoids can be generated with high efficiency and speed from fine-needle aspirations, biopsies or resection specimens, they can serve as a personal cancer model. Personalized treatment could become a more standard practice by using these cell cultures for extensive molecular diagnosis and drug screening. Drug sensitivity assays can give a clinically actionable sensitivity profile of a patient's tumour. However, the predictive capability of organoid drug screening has not been evaluated in prospective clinical trials. CONCLUSION: High-throughput drug screening on organoids, combined with next-generation sequencing, proteomic analysis and other state-of-the-art molecular diagnostic methods, can shape cancer treatment to become more effective with fewer side-effects.
Assuntos
Neoplasias Gastrointestinais/terapia , Modelos Anatômicos , Organoides , Medicina de Precisão/métodos , Técnicas de Cultura de Células/métodos , HumanosRESUMO
BACKGROUND: The modest benefits of gemcitabine (dFdC) therapy in patients with pancreatic ductal adenocarcinoma (PDAC) are well documented, with drug delivery and metabolic lability cited as important contributing factors. We have used a mouse model of PDAC: KRAS(G12D); p53(R172H); pdx-Cre (KPC) that recapitulates the human disease to study dFdC intra-tumoural metabolism. METHODS: LC-MS/MS and NMR were used to measure drug and physiological analytes. Cytotoxicity was assessed by the Sulphorhodamine B assay. RESULTS: In KPC tumour tissue, we identified a new, Kennedy pathway-linked dFdC metabolite (gemcitabine diphosphate choline (GdPC)) present at equimolar amounts to its precursor, the accepted active metabolite gemcitabine triphosphate (dFdCTP). Utilising additional subcutaneous PDAC tumour models, we demonstrated an inverse correlation between GdPC/dFdCTP ratios and cytidine triphosphate (CTP). In tumour homogenates in vitro, CTP inhibited GdPC formation from dFdCTP, indicating competition between CTP and dFdCTP for CTP:phosphocholine cytidylyltransferase (CCT). As the structure of GdPC precludes entry into cells, potential cytotoxicity was assessed by stimulating CCT activity using linoleate in KPC cells in vitro, leading to increased GdPC concentration and synergistic growth inhibition after dFdC addition. CONCLUSIONS: GdPC is an important element of the intra-tumoural dFdC metabolic pathway in vivo.
Assuntos
Carcinoma Ductal Pancreático/tratamento farmacológico , Desoxicitidina/análogos & derivados , Neoplasias Pancreáticas/tratamento farmacológico , Animais , Antimetabólitos Antineoplásicos/farmacocinética , Antimetabólitos Antineoplásicos/farmacologia , Carcinoma Ductal Pancreático/metabolismo , Colina/metabolismo , Desoxicitidina/metabolismo , Desoxicitidina/farmacocinética , Desoxicitidina/farmacologia , Difosfatos/metabolismo , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Pancreáticas/metabolismo , Células Tumorais Cultivadas , GencitabinaRESUMO
The past few decades have seen virtually no treatment advances for patients with metastatic pancreatic cancer. Clinical hallmark features of pancreatic ductal adenocarcinoma (PDA) include late symptom onset, invasive growth, early liver and lymph node metastasis, and resistance to available chemotherapies. nab-Paclitaxel (Abraxane®) is generated through high-pressure homogenization of human albumin and conventional paclitaxel resulting in non-covalently bound, water-soluble albumin-paclitaxel particles with an approximate diameter of 130ânm. Results from the recently completed Metastatic Pancreatic Adenocarcinoma Trial (MPACT) (phase III trial) showed a significant survival benefit for patients treated with nab-paclitaxel in combination with gemcitabine, and this treatment regimen is currently being implemented in national and international guidelines for PDA patients. Therefore, this regimen provides a much needed vantage point of attack for this recalcitrant tumor offering potential new hope for our patients. Mechanisms such as stromal depletion, selective intratumoral accumulation, synergism with gemcitabine metabolism and secreted protein acidic and rich in cysteine (SPARC) mediated anti-tumor activity have been suggested for nab-paclitaxel. This review discusses the clinical and experimental advances of nab-paclitaxel in pancreatic cancer.
Assuntos
Albuminas/uso terapêutico , Recidiva Local de Neoplasia/mortalidade , Recidiva Local de Neoplasia/prevenção & controle , Paclitaxel/uso terapêutico , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/mortalidade , Antineoplásicos Fitogênicos/uso terapêutico , Intervalo Livre de Doença , Medicina Baseada em Evidências , Humanos , Neoplasias Pancreáticas/diagnóstico , Prevalência , Fatores de Risco , Taxa de Sobrevida , Resultado do TratamentoRESUMO
During tumor initiation, oncogene-induced senescence (OIS) is proposed to limit the progression of preneoplasms to invasive carcinoma unless circumvented by the acquisition of certain tumor suppressor mutations. Using a variety of biomarkers, OIS has been previously reported in a wide range of human and murine precursor lesions, including the pancreas, lung, colon and skin. Here, we have characterized a panel of potential OIS biomarkers in human and murine pancreatic intraepithelial neoplasia (PanIN), and found that only senescence-associated ß-galactosidase (SAßgal) activity is specifically enriched in these precursors, compared with pancreatic ductal adenocarcinoma (PDA). Indeed, many of the other proposed OIS biomarkers are detected in actively proliferating PanIN epithelium and in cells within the microenvironment. Surprisingly, acinar to ductal metaplasia (ADM), a distinct preneoplasm that is potentially a precursor for PanIN, also exhibits SAßgal activity and contains a higher content of p21 and p53 than PanIN. Therefore, SAßgal activity is the only biomarker that accurately identifies a small and heterogeneous population of non-proliferating premalignant cells in the pancreas, and the concomitant expression of p53 and p21 in ADM supports the possibility that PanIN and ADM each exhibit discrete senescence blocks.
Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Progressão da Doença , Neoplasias Pancreáticas/metabolismo , beta-Galactosidase/metabolismo , Animais , Carcinoma Ductal Pancreático/patologia , Senescência Celular , Humanos , Metaplasia/patologia , Camundongos , Mutação , Neoplasias Pancreáticas/patologia , Lesões Pré-Cancerosas/metabolismoRESUMO
Pancreatic ductal adenocarcinoma (PDA) eludes early detection and resists current therapies, earning its distinction as the most lethal malignancy by organ site in the western world. This dire reality prompted extensive yet generally disappointing efforts to generate transgenic mouse models of this malignancy. Recently, mutant mice that develop pancreatic intraepithelial neoplasms (PanIN), the presumed preinvasive stage of PDA, were produced by conditionally expressing an endogenous oncogenic Kras allele in the developing murine pancreas. Mice with PanIN demonstrated promise in the pursuit of biomarkers of early pancreatic cancer, and, importantly, such mice eventually developed and succumbed to PDA after a long latency, establishing PanINs as true precursors to the invasive disease. Furthermore, the incorporation of conditional mutations in tumor suppressor alleles known to be altered in human PDA synergized with oncogenic Kras to produce advanced PDA with a short latency, recapitulating central pathophysiological events in human PDA. These models facilitate a variety of biological and clinical investigations such as explorations of the cellular origins of PDA and the development of treatment strategies for advanced PanIN and PDA. In addition, lessons from modeling PDA may be applicable to other tumor types and illuminate general principles of carcinogenesis.
Assuntos
Carcinoma Ductal Pancreático/genética , Neoplasias Pancreáticas/genética , Animais , Carcinoma Ductal Pancreático/diagnóstico , Carcinoma Ductal Pancreático/terapia , Modelos Animais de Doenças , Humanos , Camundongos , Camundongos Mutantes , Neoplasias Pancreáticas/diagnóstico , Neoplasias Pancreáticas/terapia , Lesões Pré-Cancerosas/genéticaRESUMO
Adenocarcinoma of the lung is the most common form of lung cancer, but the cell of origin and the stages of progression of this tumor type are not well understood. We have developed a new model of lung adenocarcinoma in mice harboring a conditionally activatable allele of oncogenic K-ras. Here we show that the use of a recombinant adenovirus expressing Cre recombinase (AdenoCre) to induce K-ras G12D expression in the lungs of mice allows control of the timing and multiplicity of tumor initiation. Through the ability to synchronize tumor initiation in these mice, we have been able to characterize the stages of tumor progression. Of particular significance, this system has led to the identification of a new cell type contributing to the development of pulmonary adenocarcinoma.
Assuntos
Adenocarcinoma/genética , Genes ras/genética , Neoplasias Pulmonares/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Animais , Códon , Progressão da Doença , Expressão Gênica , Técnicas Imunoenzimáticas , Integrases/metabolismo , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Knockout , Modelos Animais , Mutação , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Fatores de Tempo , Proteínas Virais/metabolismoRESUMO
Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract, and they are generally resistant to chemotherapy and radiation therapy. Most GISTs express the KIT receptor tyrosine kinase protein, and a subset of GISTs contain activating mutations within the KIT juxtamembrane region. We evaluated 48 GISTs, including 10 benign, 10 borderline, and 28 malignant cases, to determine whether KIT expression and activation are general properties of these tumors. Immunohistochemical KIT expression was demonstrated in each case. Somatic KIT mutations were found in 44 tumors (92%), of which 34 (71%) had juxtamembrane region mutations. Other GISTs had KIT mutations in the extracellular region (n = 6) and in two different regions in the tyrosine kinase domain (n = 4). Contrary to previous reports, KIT mutations were not identified preferentially in higher-grade tumors: indeed, they were found in each of 10 histologically benign GISTs. Notably, mutations in all KIT domains were associated with high-level KIT activation/phosphorylation, and KIT activation was also demonstrated in the four GISTs that lacked detectable KIT genomic and cDNA mutations. These studies underscore the role of KIT activation in GIST pathogenesis, and they suggest that activated KIT might represent a universal therapeutic target in GISTs.
Assuntos
Neoplasias Gastrointestinais/enzimologia , Neoplasias Gastrointestinais/genética , Proteínas Proto-Oncogênicas c-kit/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Sequência de Aminoácidos , DNA Complementar/genética , DNA de Neoplasias/genética , Ativação Enzimática , Feminino , Neoplasias Gastrointestinais/patologia , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Mutação , Fosforilação , Proteínas Proto-Oncogênicas c-kit/biossíntese , Proteínas Proto-Oncogênicas c-kit/metabolismo , Homologia de Sequência de Aminoácidos , Células Estromais/enzimologia , Células Estromais/patologiaRESUMO
Mutations in the c-KIT receptor occur somatically in many sporadic Gastrointestinal Stromal Tumors (GIST), and similar mutations have been identified at the germline level in kindreds with multiple GISTs. These mutations activate the tyrosine kinase activity of c-KIT and induce constitutive signaling. To investigate the function of activated c-KIT in GIST, we established a human GIST cell line, GIST882, which expresses an activating KIT mutation (K642E) in the first part of the cytoplasmic split tyrosine kinase domain. Notably, the K642E substitution is encoded by a homozygous exon 13 missense mutation, and, therefore, GIST882 cells do not express native KIT. GIST882 c-KIT protein is constitutively tyrosine phosphorylated, but tyrosine phosphorylation was rapidly and completely abolished after incubating the cells with the selective tyrosine kinase inhibitor STI571. Furthermore, GIST882 cells evidenced decreased proliferation and the onset of apoptotic cell death after prolonged incubation with STI571. Similar results were obtained after administering STI571 to a primary GIST cell culture that expressed a c-KIT exon 11 juxtamembrane mutation (K558NP). These cell-culture-based studies support an important role for c-KIT signaling in GIST and suggest therapeutic potential for STI571 in patients afflicted by this chemoresistant tumor.
Assuntos
Inibidores Enzimáticos/farmacologia , Neoplasias Gastrointestinais/etiologia , Proteínas Oncogênicas/fisiologia , Piperazinas/farmacologia , Pirimidinas/farmacologia , Células Estromais , Apoptose , Benzamidas , Divisão Celular/efeitos dos fármacos , Neoplasias Gastrointestinais/metabolismo , Neoplasias Gastrointestinais/patologia , Humanos , Mesilato de Imatinib , Mutação , Proteínas Oncogênicas/genética , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-kit , Células Tumorais CultivadasRESUMO
Investigations into the molecular alterations in sarcomas have made substantial progress during the past decade. Classical linkage analysis and the direct sequencing of chromosomal translocation fusions have identified candidate genes in many different sarcomas. A large group of these genes participate in signal transduction pathways and represent potential sites of disease intervention with targeted therapies. This review will discuss five types of sarcoma that display aberrant tyrosine kinase pathway signaling: gastrointestinal stromal tumor, inflammatory myofibroblastic tumor, congenital fibrosarcoma and mesoblastic nephroma, dermatofibrosarcoma protuberans, and desmoplastic small round cell tumor; one sarcoma predisposition syndrome with specific dysregulation of the ras pathway--neurofibromatosis--will also be discussed.
Assuntos
Transdução de Sinais , Neoplasias de Tecidos Moles/tratamento farmacológico , Antineoplásicos/uso terapêutico , Humanos , Proteínas Tirosina Quinases/metabolismo , Neoplasias de Tecidos Moles/enzimologiaRESUMO
About 30% of human tumours carry ras gene mutations. Of the three genes in this family (composed of K-ras, N-ras and H-ras), K-ras is the most frequently mutated member in human tumours, including adenocarcinomas of the pancreas ( approximately 70-90% incidence), colon ( approximately 50%) and lung ( approximately 25-50%). To construct mouse tumour models involving K-ras, we used a new gene targeting procedure to create mouse strains carrying oncogenic alleles of K-ras that can be activated only on a spontaneous recombination event in the whole animal. Here we show that mice carrying these mutations were highly predisposed to a range of tumour types, predominantly early onset lung cancer. This model was further characterized by examining the effects of germline mutations in the tumour suppressor gene p53, which is known to be mutated along with K-ras in human tumours. This approach has several advantages over traditional transgenic strategies, including that it more closely recapitulates spontaneous oncogene activation as seen in human cancers.
Assuntos
Genes ras , Neoplasias Pulmonares/genética , Alelos , Animais , Linhagem Celular , Modelos Animais de Doenças , Regulação da Expressão Gênica , Marcação de Genes , Genes p53 , Humanos , Neoplasias Pulmonares/patologia , Linfoma/genética , Camundongos , Camundongos Endogâmicos C57BL , Mutação , Papiloma/genética , Recombinação Genética , Neoplasias Cutâneas/genética , Neoplasias do Timo/genéticaRESUMO
Lung cancer kills more Americans yearly than any other neoplastic process. Mortality rates have changed little over the past several decades, despite improvements in surgical techniques, radiation therapy and chemotherapy. The identification of mutations in oncogenes and tumor suppressor genes in human lung tumor specimens, including K-ras, p53, p16INK4a and Rb, offers molecular explanations for tumor development and resistance to therapy. Mouse models of human lung cancer may advance our understanding of this disease. The examination of mice which develop lung cancer either spontaneously or due to carcinogen exposure, and the creation of mouse strains harboring the specific genetic mutations found in human lung cancer are among strategies being pursued.
Assuntos
Modelos Animais de Doenças , Neoplasias Pulmonares/genética , Animais , Humanos , Neoplasias Pulmonares/patologia , CamundongosRESUMO
Antigen receptors on B and T lymphocytes transduce signals by activating nonreceptor protein tyrosine kinases (PTKs). A family of receptor PTKs contains kinase insert regions with the sequence tyrosine-X-X-methionine (where X is any amino acid) that when phosphorylated mediate the binding and activation of phosphatidylinositol 3-kinase (PI 3-kinase). The CD19 membrane protein of B cells enhances activation through membrane immunoglobulin M (mIgM) and was found to contain a functional analog of the kinase insert region. Ligation of mIgM induced phosphorylation of CD19 and association with PI 3-kinase. Thus, CD19 serves as a surrogate kinase insert region for mIgM by providing the means for PI 3-kinase activation by nonreceptor PTKs.
Assuntos
Antígenos CD/metabolismo , Antígenos de Diferenciação de Linfócitos B/metabolismo , Linfócitos B/imunologia , Imunoglobulina M/metabolismo , Fosfotransferases/metabolismo , Sequência de Aminoácidos , Antígenos CD19 , Linfócitos B/metabolismo , Sequência de Bases , Humanos , Dados de Sequência Molecular , Fosfatidilinositol 3-Quinases , Fosfatidilinositóis/metabolismo , Fosforilação , Células Tumorais Cultivadas , Tirosina/metabolismoRESUMO
We have investigated the mechanism by which the membrane protein complex of the B lymphocyte that contains CD19 and CR2 activates phospholipase C (PLC) to induce a rise in [CA2+]i. The CD19 complex resembled the membrane IgM complex in that three protein tyrosine kinase inhibitors suppressed increases in [Ca2+]i and inositol bisphosphate and inositol triphosphate generation. However, the activation of PLC by the CD19 complex could be distinguished from that by the membrane IgM complex by slower kinetics of generation of inositol phosphates, resistance to inhibition by activators of protein kinase C, and different pattern of tyrosine-phosphorylated cellular substrates. Western blot analysis of lysates from cells stimulated by the CD19 complex demonstrated a single new phosphotyrosine-containing protein of 85 kDa, whereas multiple other phosphotyrosine-containing proteins were present in cells activated by the mIgM complex. In particular, PLC-gamma 1, which is a substrate for the protein tyrosine kinase activated by the mIgM complex, was not tyrosine-phosphorylated in cells stimulated by the CD19 complex. Cross-linking the two complexes together caused a synergistic increase in [CA2+]i which was neither suppressed by activation of protein kinase C nor associated with increased tyrosine-phosphorylation of PLC, characteristic of the CD19 pathway. Therefore, the B cell has two signal transduction complexes, associated with membrane IgM and CD19, that activate PLC by different mechanisms and that can synergistically interact to enhance this function by the CD19 pathway.
Assuntos
Antígenos CD/fisiologia , Antígenos de Diferenciação de Linfócitos B/fisiologia , Linfócitos B/fisiologia , Cálcio/fisiologia , Imunoglobulina M/fisiologia , Ativação Linfocitária , Proteínas Tirosina Quinases/fisiologia , Receptores de Antígenos de Linfócitos B/fisiologia , Receptores de Complemento/fisiologia , Fosfolipases Tipo C/fisiologia , Antígenos CD19 , Ativação Enzimática , Humanos , Técnicas In Vitro , Fosfatos de Inositol/metabolismo , Substâncias Macromoleculares , Peso Molecular , Ésteres de Forbol/farmacologia , Fosfoproteínas/química , Fosfoproteínas/metabolismo , Fosfotirosina , Agregação de Receptores , Receptores de Complemento 3d , Transdução de Sinais , Tirosina/análogos & derivados , Tirosina/metabolismoRESUMO
The complement system augments the humoral immune response to low concentrations of antigen. This effect may be partly mediated by complement receptors on the surface of B lymphocytes that bind immunogenic complexes bearing fragments of C3 and C4. We have shown by immunoprecipitation analysis that the two complement receptors expressed by B lymphocytes, complement receptor 1 (CR1) and CR2, form a detergent-sensitive complex on the surface of tonsillar B lymphocytes and on K562 erythroleukemia cells that were co-transfected with cDNAs encoding CR1 and CR2. The CR1/CR2 complex is distinct from the CR2/CD19 complex and may assist B cell activation by efficiently capturing C3b-containing immunogens and maintaining such immunogens on the B cell after CR1 and factor I-mediated cleavage to iC3b and C3dg. The complement activating immunogen may then trigger signal transduction by the CR1/CR2 complex, the CR2/CD19 complex, or membrane immunoglobulin.
Assuntos
Antígenos de Diferenciação de Linfócitos B/metabolismo , Linfócitos B/metabolismo , Receptores de Complemento/metabolismo , Antígenos CD19 , Antígenos de Diferenciação de Linfócitos B/genética , Antígenos de Diferenciação de Linfócitos B/fisiologia , Linfócitos B/fisiologia , Linfócitos B/ultraestrutura , DNA/genética , Humanos , Leucemia Eritroblástica Aguda/metabolismo , Leucemia Eritroblástica Aguda/patologia , Leucemia Eritroblástica Aguda/fisiopatologia , Tonsila Palatina/citologia , Testes de Precipitina , Receptores de Complemento/genética , Receptores de Complemento/fisiologia , Receptores de Complemento 3b , Receptores de Complemento 3d , Transdução de Sinais/fisiologia , Transfecção/genéticaRESUMO
The complement system augments the humoral immune response, possibly by a mechanism that involves the B lymphocyte membrane receptor, CR2, which binds the C3dg fragment of C3 and triggers several B cell responses in vitro. The present study demonstrates that CR2 associates with a complex of membrane proteins that may mediate signal transduction by ligated CR2. Monoclonal antibodies to CR2 immunoprecipitated from digitonin lysates of Raji B lymphoblastoid cells a membrane complex containing CR2, approximately equimolar amounts of CD19, which is a member of the immunoglobulin superfamily, and three unidentified components: p130, p50, and p20. The complex, which was immunoprecipitated also with anti-CD19, could be dissociated by Nonidet P-40, accounting for its absence in previous studies of CR2. Expression of recombinant CR2 and CD19 in K562 erythroleukemia cells led to formation of a complex that contained not only these two proteins but also p130, p50, and p20, and another component, p14. These unidentified components of the CR2/CD19 complex coimmunoprecipitated with CD19 and not with CR2 from singly transfected cells, indicating primary association with the former. CD19 replicated the capacity of CR2 to interact synergistically with mIgM for increasing free intracellular Ca2+, suggesting that the complex mediates this function of CR2. Therefore, CR2 associates directly with CD19 to become a ligand-binding subunit of a pre-existing signal transduction complex of the B cell that may be representative of a family of membrane protein complexes. This interaction between the complement and immune systems differs from that between immunoglobulin and Clq by involving membrane rather than plasma proteins, and by having complement involved in the afferent phase of the immune response.