Asunto(s)
Reacción de Fase Aguda/fisiopatología , Citocinas/fisiología , Sistema Hematopoyético/fisiología , Proteínas de Fase Aguda/biosíntesis , Proteínas de Fase Aguda/genética , Anemia/fisiopatología , Animales , Plaquetas/citología , Eritropoyesis/fisiología , Eritropoyetina/fisiología , Regulación de la Expresión Génica , Granulocitos/citología , Hematopoyesis/fisiología , Células Madre Hematopoyéticas/citología , Humanos , Inflamación/fisiopatología , Hierro/metabolismo , Hígado/metabolismo , Megacariocitos/citología , Modelos BiológicosRESUMEN
PURPOSE: Carmustine (BCNU) resistance has been correlated with tumor expression of the DNA repair enzyme O6-alkylguanine-DNA alkyltransferase (AT). It has been shown that streptozotocin will deplete AT activity of human colon cancer cells in vitro and potentiate BCNU cytotoxicity. This clinical trial was conducted to determine whether streptozotocin can be used as a modulator of AT in metastatic colorectal cancers and thereby overcome clinical resistance to BCNU. PATIENTS AND METHODS: Fifteen patients with fluorouracil-resistant metastatic colon or rectal cancers were treated sequentially with 2 g/m2 of streptozotocin followed 5 1/2 hours later by BCNU. Sequential biopsies of metastases before and after streptozotocin were conducted to determine whether streptozotocin depletes tumor AT. Peripheral-blood mononuclear cells (PBMCs) were evaluated as a surrogate tissue for prediction of baseline AT levels and streptozotocin posttreatment modulation of the AT in metastases. RESULTS: Streptozotocin treatment led to a 78% (range, 69% to 89%) decrease in the AT levels in colon cancer metastases; however, myelosuppression and hepatic toxicity limited the BCNU dose to 130 mg/m2. A similar decrease in AT levels of PBMCs was found; however, the absolute levels of AT in PBMCs at baseline and following streptozotocin were not predictive of the levels expressed in metastases from the same patient. Despite the decrease in tumor levels of AT, no clinical responses were observed. CONCLUSION: Streptozotocin decreases but does not fully deplete AT activity in metastatic colorectal cancers and the residual AT level in metastases is sufficient to maintain clinical resistance to BCNU. We have also demonstrated that sequential computed tomography (CT)-directed biopsies of colorectal cancer metastases can be used to evaluate strategies to investigate modulators of AT-directed repair. AT levels of PBMCs do not predict for the AT level or degree of modulation achieved in the metastatic tumor.
Asunto(s)
Carmustina/uso terapéutico , Neoplasias Colorrectales/enzimología , Reparación del ADN , Metiltransferasas/metabolismo , Estreptozocina/uso terapéutico , Adulto , Anciano , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/patología , Resistencia a Medicamentos , Quimioterapia Combinada , Femenino , Humanos , Leucocitos Mononucleares/enzimología , Masculino , Persona de Mediana Edad , Metástasis de la Neoplasia , O(6)-Metilguanina-ADN Metiltransferasa , Análisis de Regresión , Tomografía Computarizada por Rayos XRESUMEN
Synchronous cutaneous T-cell lymphoma and low-grade B-cell lymphoproliferative disorders have rarely been reported in the same patient. Coexpression of each phenotype in the same lymph node has not, to our knowledge, been previously documented. We describe an 86-year-old man with chronic pruritus and erythroderma and recent-onset peripheral lymphadenopathy and lymphocytosis. Lymph node biopsy provided morphological and immunohistochemical evidence of concurrent small B lymphocytic lymphoma and small pleomorphic T-cell lymphoma. Immunophenotyping of nodal lymphocytes demonstrated two distinct clones: IgM-kappa B-cells with CD5 positivity and CD7 negative T-helper cells. Both immunoglobulin (heavy and light chains) and T-cell receptor (beta I and beta II) gene rearrangements were detected by Southern blot analysis of the lymph node. In contrast, the immunophenotype of lymphocytes from peripheral blood and bone marrow was exclusively that of T-helper cells with atypical CD7 deletion. Electron microscopic examination of circulating lymphocytes revealed small cerebriform Sezary cells. This case demonstrates that small lymphocytic lymphoma may coexist intranodally with cutaneous T-cell lymphoma as a unique form of composite T- and B-cell lymphoma.
Asunto(s)
Ganglios Linfáticos/ultraestructura , Linfocitos/ultraestructura , Linfoma de Células B/ultraestructura , Linfoma Cutáneo de Células T/ultraestructura , Neoplasias Primarias Múltiples/ultraestructura , Anciano , Anciano de 80 o más Años , Linfocitos B/ultraestructura , Citometría de Flujo , Reordenamiento Génico , Genes de Inmunoglobulinas , Humanos , Inmunohistoquímica , Inmunofenotipificación , Linfoma de Células B/genética , Linfoma de Células B/inmunología , Linfoma Cutáneo de Células T/genética , Linfoma Cutáneo de Células T/inmunología , Masculino , Microscopía Electrónica , Neoplasias Primarias Múltiples/genética , Neoplasias Primarias Múltiples/inmunología , Receptores de Antígenos de Linfocitos T/genética , Piel/ultraestructura , Linfocitos T Colaboradores-InductoresRESUMEN
METHODS: Between 1983 and 1989, 42 patients with Stage II, III, and IV, node-negative, squamous cell head and neck cancer were treated with concurrent 5-fluorouracil, cisplatin, and radiation therapy. Two courses of chemotherapy with 30 Gy of concurrent radiation therapy were to be followed in all patients by definitive surgery and then an additional 30 Gy of radiation therapy and one to two courses of chemotherapy. The patients who achieved a complete response to the initial induction treatment, however, did not undergo surgery. RESULTS: After the completion of all therapy, 41 of the 42 patients (98%) were considered disease-free. Only 4 of these 41 had relapses, for a projected Kaplan-Meier disease-free survival rate of 86%. Treatment failure occurred in no patients with Stage II, 1 of 17 patients with Stage III, and 4 of 14 patients with Stage IV disease. Of the 42 patients, 23 (55%) did not require surgery after achieving a complete response to induction therapy, and only 1 of these 23 patients subsequently had a relapse. CONCLUSIONS: Although the value of adding chemotherapy to conventional treatment remains unproven in squamous cell head and neck cancer, this treatment schedule appears promising in node-negative disease. Randomized trials will be necessary, however, to validate the efficacy of this approach and confirm the suggestion by the authors that surgery can be avoided in most patients with N0 disease.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Carcinoma de Células Escamosas/tratamiento farmacológico , Carcinoma de Células Escamosas/radioterapia , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Neoplasias de Cabeza y Cuello/radioterapia , Anciano , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Cisplatino/administración & dosificación , Cisplatino/efectos adversos , Terapia Combinada , Femenino , Fluorouracilo/administración & dosificación , Fluorouracilo/efectos adversos , Humanos , Masculino , Persona de Mediana Edad , Análisis de Supervivencia , Resultado del TratamientoRESUMEN
Although induction of sister chromatid exchanges (SCEs) following nitrosourea exposure may be greater during cell proliferation, the increase could be offset by the action of the DNA repair protein O6-alkylguanine DNA alkyltransferase (alkyltransferase). To evaluate these factors in resting and proliferating (phytohemagglutinin stimulated) human lymphocytes, we studied the effect of changes in alkyltransferase activity on 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU)-induced SCEs. Phytohemagglutinin stimulation resulted in induction of alkyltransferase activity (5.9 +/- 0.3 units, resting, versus 9.2 +/- 0.2 units, proliferating). In both resting and proliferating lymphocytes the alkyltransferase activity was inactivated by 85-88% after an 18-h exposure to 0.5 mM of the modified base O6methylguanine (O6mGua). However, 48 h after removal of O6mGua, proliferating lymphocytes recovered alkyltransferase activity while resting cells did not. In the absence of O6mGua, both resting and proliferating lymphocytes were equally sensitive to BCNU-induced SCEs. Following inactivation of the alkyltransferase by O6mGua, BCNU-induced SCEs were markedly increased, but the increase was much greater in resting than proliferating cells, 4-fold vs. 2.6-fold at each dose of BCNU (P less than 0.001). The factors providing partial protection against BCNU-induced SCEs in proliferating lymphocytes appear to include the proliferation-dependent increase in alkyltransferase activity and the ability of proliferating lymphocytes to rapidly recover alkyltransferase activity after its inactivation. Thus, the alkyltransferase appears to provide an important mechanism of resistance to SCE induction in human lymphocytes.
Asunto(s)
Carmustina/farmacología , Metiltransferasas/fisiología , Intercambio de Cromátides Hermanas/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Daño del ADN , Reparación del ADN , Guanina/análogos & derivados , Guanina/farmacología , Humanos , Activación de Linfocitos , Linfocitos/efectos de los fármacos , Linfocitos/ultraestructura , O(6)-Metilguanina-ADN Metiltransferasa , Fitohemaglutininas/farmacologíaRESUMEN
Drug resistance in myeloid leukemias may be mediated by an increased capacity to repair chemotherapy-induced DNA damage. Some tumor cell lines that are resistant to nitrosoureas contain the DNA repair protein O6-alkylguanine-DNA alkyltransferase (alkyltransferase). This protects cells by removing cytotoxic, nitrosourea-induced O6-alkylguanine adducts. We measured the level of alkyltransferase activity in myeloid leukemic cells freshly obtained from patients to determine whether the alkyltransferase was an important factor in nitrosourea resistance in these cells and whether inactivation of this protein could sensitize leukemic cells to nitrosoureas. Myeloid leukemic cells from patients with acute nonlymphocytic leukemia and chronic myelogenous leukemia had higher levels of alkyltransferase than did myeloid precursors from normal donors (P less than .01). This difference did not appear to be due to the state of differentiation of the leukemic or normal cells. To show that this repair protein mediated nitrosourea resistance in leukemic cells, cells were treated with the modified base O6-methylguanine to selectively and irreversibly inactivate the alkyltransferase and then exposed to 1,3-bis (2-chloroethyl)-1-nitrosourea (BCNU). An 18-hour incubation in 0.5 mmol/L O6-methylguanine caused an 87% +/- 3.6% decrease in alkyltransferase activity in leukemic cells and a 73% +/- 8.6% decrease in normal myeloid precursors. After treatment with O6-methylguanine, clonogenic leukemic cells from ten different donors became much more sensitive to BCNU, with a decrease in the dose needed to reduce colony survival by 50% (LD50) of 6.3 +/- 1.4-fold. A lesser effect was seen on CFU-GM, BFU-E, and CFU-GEM where the LD50 decreased two- to threefold. These studies show that nitrosourea resistance in myeloid leukemic cells can be abrogated by inactivation of the DNA repair protein O6-alkylguanine-DNA alkyltransferase. This method of biochemical modulation of DNA repair will sensitize leukemic cells to nitrosoureas in vitro and has the potential of increasing the therapeutic index of nitrosoureas in this disease.
Asunto(s)
Carmustina/farmacología , Células Madre Hematopoyéticas/efectos de los fármacos , Leucemia Mieloide Aguda/enzimología , Leucemia Mieloide/enzimología , Metiltransferasas/metabolismo , Proteínas de Neoplasias/metabolismo , Células Madre Neoplásicas/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Factores Estimulantes de Colonias/farmacología , Reparación del ADN , Resistencia a Medicamentos , Guanina/análogos & derivados , Guanina/farmacología , Células Madre Hematopoyéticas/enzimología , Humanos , Metiltransferasas/antagonistas & inhibidores , Células Madre Neoplásicas/enzimología , O(6)-Metilguanina-ADN Metiltransferasa , Células Tumorales Cultivadas/efectos de los fármacos , Células Tumorales Cultivadas/enzimologíaRESUMEN
The HL-60 promyelocytic leukemia cell line is resistant to nitrosoureas and contains high levels of the DNA repair protein O6-alkylguanine-DNA alkyltransferase (alkyltransferase). We examined the protective role of the alkyltransferase in the nitrosourea resistance observed in this myeloid leukemia cell line to determine whether inactivation of the alkyltransferase with the modified base, O6-methylguanine (O6mGua), could sensitize these cells to nitrosoureas. The HL-60 cells were sensitized approximately 3.0-fold to five different nitrosoureas when the alkyltransferase was inactivated by 88% following a 24-h preincubation in 0.5 mM O6mGua. No effect of O6mGua preincubation was observed in the K562 chronic myelogenous leukemia cell line which is sensitive to nitrosoureas and has low levels of alkyltransferase activity. When regeneration of HL-60 alkyltransferase activity after exposure to nitrosoureas was prevented by maintaining cells in O6mGua, HL-60 became even more sensitive (3.7- to 8.5-fold) to nitrosoureas but remained slightly more resistant than K562. Next, we compared the dose of methyl- and chloroethylnitrosoureas which were cytotoxic in HL-60 with the dose which caused repair-induced inactivation of the alkyltransferase. Both methyl- and chloroethyl-nitrosoureas caused the dose-dependent inactivation of the alkyltransferase and with both, cytotoxicity was increased with O6mGua exposure. However, chloroethylnitrosoureas, which form a variety of O6 alkylation adducts, some of which are poorly repaired, exhibited 7-12 times more cytotoxicity relative to repair-induced inactivation of the alkyltransferase whereas methylnitrosoureas became cytotoxic only when the alkyltransferase had been inactivated. These data suggest that leukemic cells are sensitized to both methyl- and chloroethylnitrosoureas when O6mGua is used to persistently inactivate the alkyltransferase. However, the alkyltransferase provides more efficient protection from methylnitrosoureas than chloroethylnitrosoureas most likely because the latter form adducts which are poorly repaired by the protein and which if unrepaired may become cytotoxic cross-links.
Asunto(s)
Leucemia Mieloide Aguda/enzimología , Metiltransferasas/fisiología , Compuestos de Nitrosourea/farmacología , Supervivencia Celular/efectos de los fármacos , Reparación del ADN , Relación Dosis-Respuesta a Droga , Resistencia a Medicamentos , Sinergismo Farmacológico , Guanina/análogos & derivados , Guanina/farmacología , Humanos , Leucemia Mieloide Aguda/patología , Metiltransferasas/análisis , Metiltransferasas/antagonistas & inhibidores , O(6)-Metilguanina-ADN Metiltransferasa , Células Tumorales CultivadasAsunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/toxicidad , Neoplasias Colorrectales/tratamiento farmacológico , Adulto , Anciano , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Neoplasias Colorrectales/patología , Evaluación de Medicamentos , Femenino , Fluorouracilo/administración & dosificación , Humanos , Leucovorina/administración & dosificación , Masculino , Persona de Mediana Edad , Metástasis de la NeoplasiaRESUMEN
O6-Alkylguanine, a DNA adduct formed by nitrosoureas, becomes the site of a point mutation during DNA synthesis by preferentially base mispairing with thymine rather than correctly base pairing with cytosine. To repair this adduct, cells contain a limited amount of O6-alkylguanine-DNA alkyltransferase (alkyltransferase), a protein which removes the alkyl group in a stoichiometric reaction. To prevent mutations, repair must occur before DNA replication takes place. Consequently, formation of point mutations is related inversely to the number of alkyltransferase molecules and directly to the rate of DNA synthesis. Bone marrow hematopoietic precursors, the target for nitrosourea-induced leukemia, are deficient in alkyltransferase activity. We questioned whether regenerating bone marrow is more susceptible to nitrosoureas than other organs due to persistently low levels of alkyltransferase activity during periods of increased cell proliferation and DNA synthesis. Following syngeneic bone marrow transplantation, murine hematopoietic cells underwent rapid cell proliferation but alkyltransferase activity remained well below the activity in liver. After N-nitrosomethylurea exposure, [3H]thymidine incorporation in rat bone marrow increased 3-fold and stem cell proliferation over 10-fold within 2 days of exposure, but alkyltransferase activity remained low. The relative susceptibility of bone marrow to mutagenic damage from O6-alkylguanine adducts was determined by comparing the ratio of alkyltransferase activity to [3H]thymidine incorporation in marrow, kidney, and liver. In untreated animals, the ratio was lowest in bone marrow and decreased further 48 h after N-nitrosomethylurea exposure to only 21% that of kidney and 1% that of liver. Thus, proliferating hematopoietic precursors appear more likely to form point mutations following nitrosourea exposure than other rodent tissues because they undergo rapid proliferation soon after DNA damage and before O6-alkylguanine adducts can be repaired. The combination of rapid cell proliferation and low DNA repair capacity may be the mechanism of nitrosourea induced leukemic transformation of the bone marrow.
Asunto(s)
Reparación del ADN , ADN/biosíntesis , Guanina/análogos & derivados , Células Madre Hematopoyéticas/metabolismo , Animales , Médula Ósea/efectos de los fármacos , Trasplante de Médula Ósea , Guanina/metabolismo , Masculino , Metilnitrosourea/toxicidad , Metiltransferasas/análisis , Ratones , Ratones Endogámicos , Mutación , O(6)-Metilguanina-ADN Metiltransferasa , Ratas , Ratas EndogámicasRESUMEN
O6-Alkylguanine-DNA alkyltransferase (alkyltransferase) is the repair protein for O6-alkylguanine, a pre-mutagenic adduct formed by a variety of alkylating agents. Previous comparisons of the repair capacity of O6-alkylguanine in different tissues have expressed the alkyltransferase activity relative to total protein, and have asserted that tissues with low levels of activity were at greater risk for mutagenic damage than tissues with higher levels of activity. Because the alkyltransferase uses DNA as substrate, and because tissues vary greatly in protein content, comparisons of tissue alkyltransferase activity may be more appropriately based on cellular DNA content. We compared alkyltransferase activity relative to tissue DNA content with the activity related to protein content in human, rat and mouse tissues. In each species, liver containing the highest level of activity using either method. In agreement with the findings of others, low levels of alkyltransferase activity relative to protein were seen in human brain, rat brain and small intestine, and mouse kidney. However, based on alkyltransferase activity relative to DNA content, low levels of activity were seen in human bone marrow myeloid precursors, rat bone marrow, brain and intestine, and mouse spleen and bone marrow. The range of activity between tissues was 18-fold in human, 15-fold in rat and 8-fold in mouse. In general, the rank of alkyltransferase activity relative to DNA for each tissue was human greater than rat greater than mouse. These results suggest that the mouse is more susceptible to nitrosoureas than rat or human. In each species, the organs with low levels of alkyltransferase activity relative to tissue DNA content would appear to be targets for mutagenic damage following nitrosourea exposure.