RESUMEN
PURPOSE: An understanding of how hematopoietic cells respond to therapy that causes myelosuppression will help develop approaches to prevent this potentially life-threatening toxicity. The goal of this study was to determine how human myeloid precursor cells respond to temozolomide (TMZ)-induced DNA damage. EXPERIMENTAL DESIGN: We developed an ex vivo primary human myeloid precursor cells model system to investigate the involvement of cell-death pathways using a known myelosuppressive regimen of O(6)-benzylguanine (6BG) and TMZ. RESULTS: Exposure to 6BG/TMZ led to increases in p53, p21, γ-H2AX, and mitochondrial DNA damage. Increases in mitochondrial membrane depolarization correlated with increased caspase-9 and -3 activities following 6BG/TMZ treatment. These events correlated with decreases in activated AKT, downregulation of the DNA repair protein O(6)-methylguanine-DNA methyltransferase (MGMT), and increased cell death. During myeloid precursor cell expansion, FAS/CD95/APO1(FAS) expression increased over time and was present on approximately 100% of the cells following exposure to 6BG/TMZ. Although c-flipshort, an endogenous inhibitor of FAS-mediated signaling, was decreased in 6BG/TMZ-treated versus control, 6BG-, or TMZ alone-treated cells, there were no changes in caspase-8 activity. In addition, there were no changes in the extent of cell death in myeloid precursor cells exposed to 6BG/TMZ in the presence of neutralizing or agonistic anti-FAS antibodies, indicating that FAS-mediated signaling was not operative. CONCLUSIONS: In human myeloid precursor cells, 6BG/TMZ-initiated apoptosis occurred by intrinsic, mitochondrial-mediated and not extrinsic, FAS-mediated apoptosis. Human myeloid precursor cells represent a clinically relevant model system for gaining insight into how hematopoietic cells respond to chemotherapeutics and offer an approach for selecting effective chemotherapeutic regimens with limited hematopoietic toxicity.
Asunto(s)
Metilación de ADN/efectos de los fármacos , Dacarbazina/análogos & derivados , Células Progenitoras Mieloides/efectos de los fármacos , Antineoplásicos Alquilantes/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/genética , Western Blotting , Proteína Reguladora de Apoptosis Similar a CASP8 y FADD/genética , Proteína Reguladora de Apoptosis Similar a CASP8 y FADD/metabolismo , Ciclo Celular/efectos de los fármacos , Células Cultivadas , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Daño del ADN , ADN Mitocondrial/genética , Dacarbazina/farmacología , Perfilación de la Expresión Génica , Guanina/análogos & derivados , Guanina/farmacología , Histonas/genética , Histonas/metabolismo , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Células Progenitoras Mieloides/metabolismo , O(6)-Metilguanina-ADN Metiltransferasa/genética , O(6)-Metilguanina-ADN Metiltransferasa/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Temozolomida , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Receptor fas/genética , Receptor fas/metabolismoRESUMEN
BACKGROUND: Preprocedural dietary restrictions have been identified as a common reason potential candidates for colorectal cancer screening do not undergo colonoscopy as recommended. OBJECTIVE: To study whether a low-residue diet impacts bowel preparation with oral sulfate solution. DESIGN: Endoscopist blinded, prospective, randomized controlled trial. SETTING: Community-based outpatient ambulatory surgical center. PATIENTS: Patients scheduled for outpatient colonoscopy. INTERVENTIONS: Subjects were randomized to ingest either a low-residue diet of specified foods for breakfast, lunch, and snack or a clear liquid diet the day before the colonoscopy. MAIN OUTCOME MEASUREMENTS: The quality of the bowel preparation was assessed using the Boston Bowel Preparation Scale. Subject satisfaction with bowel preparation, diet, and severity of side effects was measured by a visual analog scale. RESULTS: Two hundred thirty subjects were recruited (114 clear liquid and 116 low residue). Mean preparation scores were not statistically different in either their segmental scores or total score. Subjects in the low-residue arm reported significantly higher satisfaction with bowel preparation medication, diet, and entire preparation process. Observed rates of side effects were low, and there was no statistical difference between the two groups. The rate of procedural cancellation was significantly higher in the clear liquid group compared with the low-residue group (20% vs 9%, P = .03). LIMITATIONS: Single-center study. CONCLUSIONS: A low-residue diet did not impair the quality of bowel preparation achieved with split-dose oral sulfate solution but did improve patient satisfaction.
Asunto(s)
Catárticos/uso terapéutico , Colonoscopía/métodos , Dieta , Satisfacción del Paciente , Anciano , Femenino , Humanos , Masculino , Persona de Mediana EdadRESUMEN
Humanized bone-marrow xenograft models that can monitor the long-term impact of gene-therapy strategies will help facilitate evaluation of clinical utility. The ability of the murine bone-marrow microenvironment in NOD/SCID versus NOD/SCID/γ chain(null) mice to support long-term engraftment of MGMT(P140K)-transduced human-hematopoietic cells following alkylator-mediated in vivo selection was investigated. Mice were transplanted with MGMT(P140K)-transduced CD34(+) cells and transduced cells selected in vivo. At 4 months after transplantation, levels of human-cell engraftment, and MGMT(P140K)-transduced cells in the bone marrow of NOD/SCID versus NSG mice varied slightly in vehicle- and drug-treated mice. In secondary transplants, although equal numbers of MGMT(P140K)-transduced human cells were transplanted, engraftment was significantly higher in NOD/SCID/γ chain(null) mice compared to NOD/SCID mice at 2 months after transplantation. These data indicate that reconstitution of NOD/SCID/γ chain(null) mice with human-hematopoietic cells represents a more promising model in which to test for genotoxicity and efficacy of strategies that focus on manipulation of long-term repopulating cells of human origin.
RESUMEN
PURPOSE: Preclinical in vivo studies can help guide the selection of agents and regimens for clinical testing. However, one of the challenges in screening anticancer therapies is the assessment of off-target human toxicity. There is a need for in vivo models that can simulate efficacy and toxicities of promising therapeutic regimens. For example, hematopoietic cells of human origin are particularly sensitive to a variety of chemotherapeutic regimens, but in vivo models to assess potential toxicities have not been developed. In this study, a xenograft model containing humanized bone marrow is utilized as an in vivo assay to monitor hematotoxicity. EXPERIMENTAL DESIGN: A proof-of-concept, temozolomide-based regimen was developed that inhibits tumor xenograft growth. This regimen was selected for testing because it has been previously shown to cause myelosuppression in mice and humans. The dose-intensive regimen was administered to NOD.Cg-Prkdc(scid)IL2rg(tm1Wjl)/Sz (NOD/SCID/γchain(null)), reconstituted with human hematopoietic cells, and the impact of treatment on human hematopoiesis was evaluated. RESULTS: The dose-intensive regimen resulted in significant decreases in growth of human glioblastoma xenografts. When this regimen was administered to mice containing humanized bone marrow, flow cytometric analyses indicated that the human bone marrow cells were significantly more sensitive to treatment than the murine bone marrow cells and that the regimen was highly toxic to human-derived hematopoietic cells of all lineages (progenitor, lymphoid, and myeloid). CONCLUSIONS: The humanized bone marrow xenograft model described has the potential to be used as a platform for monitoring the impact of anticancer therapies on human hematopoiesis and could lead to subsequent refinement of therapies prior to clinical evaluation.