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1.
Leukemia ; 23(9): 1658-66, 2009 Sep.
Article in English | MEDLINE | ID: mdl-19387464

ABSTRACT

Activation of the Wnt signaling pathway has been implicated recently in the pathogenesis of leukemia. We studied the function of epigenetic regulation of the Wnt pathway and its prognostic relevance in acute myelogenous leukemia (AML). We used a methylation-specific polymerase chain reaction approach to analyze the promoter methylation status of a panel of Wnt antagonists including sFRP1, sFRP2, sFRP4, sFRP5, DKK1 and DKK3. Aberrant methylation of Wnt antagonists was detected in four AML cell lines and in up to 64% of AML marrow samples. Treatment of the cell lines with 5-aza-2'-deoxycytidine induced reexpression of methylated Wnt antagonists and inactivation of the Wnt pathway by downregulating the Wnt pathway genes cyclin D1, TCF1 and LEF1 and reducing nuclear localization of beta-catenin. In a subgroup of patients 60 years and younger with newly diagnosed AML and intermediate-risk cytogenetics, abnormal methylation of Wnt antagonists was associated with decreased 4-year relapse-free survival (28 vs 61%, respectively, P=0.03). Our results indicate a function of the epigenetic regulation of the Wnt pathway in predicting relapse in a subgroup of AML patients.


Subject(s)
DNA Methylation , Epigenesis, Genetic , Leukemia, Myeloid, Acute/genetics , Signal Transduction , Wnt Proteins/antagonists & inhibitors , Adult , Aged , Ataxia Telangiectasia Mutated Proteins , Azacitidine/analogs & derivatives , Azacitidine/pharmacology , Cell Cycle/drug effects , Cell Cycle Proteins/genetics , Cell Line, Tumor , Decitabine , Female , Genes, bcl-1 , Humans , Intercellular Signaling Peptides and Proteins/genetics , Leukemia, Myeloid, Acute/mortality , Male , Membrane Proteins/genetics , Middle Aged , Prognosis , Promoter Regions, Genetic , Protein Serine-Threonine Kinases/genetics , Wnt Proteins/physiology
2.
Leukemia ; 21(7): 1413-22, 2007 Jul.
Article in English | MEDLINE | ID: mdl-17476281

ABSTRACT

Therapy-related myelodysplasia and acute myeloid leukemia (t-MDS/AML) is a malignancy occurring after exposure to chemotherapy and/or radiotherapy. Polymorphisms involved in chemotherapy/radiotherapy response genes could be related to an increased risk of developing this neoplasia. We have studied 11 polymorphisms in genes of drug detoxification pathways (NQO1, glutathione S-transferase pi) and DNA repair xeroderma pigmentosum, complementation group (3) (XPC(3), X-ray repair cross complementing protein (1)), Nijmegen breakage syndrome (1), excision repair cross-complementing rodent repair deficiency, complementation group (5) and X-ray repair cross complementing protein (3) and in the methylene tetrahydrofolate reductase gene (MTHFR(2), 677C>T, 1298A>C), involved in DNA synthesis. The analyzed groups were a t-MDS/AML patients group (n=81) and a matched control group (n=64) treated similarly, and they did not develop t-MDS/AML. We found no significant differences when the groups were compared globally. However, when analysis was carried out according to the primary neoplasia involved, a significant association was observed between the MTHFR haplotype (single nucleotide polymorphisms 677 and 1298) and the risk of developing t-MDS/AML in the breast cancer patients group (P=0.016) and cyclophosphamide-treated hematological disease group (P=0.005). Risk haplotype was different for each case, corresponding to the 677T1298A haplotype after breast cancer treatment and the 677C1298C haplotype after hematological malignancy treatment. We postulate that such differences are related to variations in chemotherapy schemes between hematological and breast cancers and their differential interaction with the MTHFR route.


Subject(s)
Haplotypes , Leukemia/genetics , Methylenetetrahydrofolate Reductase (NADPH2)/genetics , Antineoplastic Agents/adverse effects , Breast Neoplasms/drug therapy , Breast Neoplasms/pathology , Case-Control Studies , Hematologic Neoplasms/drug therapy , Hematologic Neoplasms/pathology , Humans , Leukemia/chemically induced , Leukemia/etiology , Middle Aged , Neoplasms, Second Primary/etiology , Neoplasms, Second Primary/genetics , Polymorphism, Single Nucleotide , Risk Factors
3.
Leukemia ; 21(3): 446-52, 2007 Mar.
Article in English | MEDLINE | ID: mdl-17205057

ABSTRACT

To determine prognosis of acute promyelocytic leukemia (APL) failing to front-line therapy with all-trans retinoic acid (ATRA) and anthracyclines, outcome of 52 patients (32 M/20 F; age: 37, 3-72) included in PETHEMA trials LPA96 and LPA99 who presented with either molecular failure (MOLrel, n=16) or hematological relapse (HEMrel, n=36) was analyzed. Salvage therapy consisted of ATRA and high-dose ara-C-based chemotherapy (HDAC) in most cases (83%), followed by stem-cell transplantation (autologous, 18; allogeneic, 10; syngeneic, 1). Fourteen patients with MOLrel (88%) achieved second molecular complete response (molCR), whereas 81% HEMrel patients responded to second-line treatment, with 58% molCR. After median follow-up of 45 months, four MOLrel and 18 HEMrel patients, respectively, experienced a second relapse. Outcome after MOLrel compared favorably to HEMrel, with longer survival (5-year survival: 64+/-14 vs 24+/-8%, P=0.01) and lower relapse risk (5-year relapse risk: 30+/-13 vs 64+/-9%; P=0.044). Additionally, age

Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Leukemia, Promyelocytic, Acute/drug therapy , Salvage Therapy , Adolescent , Adult , Aged , Antineoplastic Combined Chemotherapy Protocols/administration & dosage , Biomarkers, Tumor/blood , Child , Child, Preschool , Combined Modality Therapy , Cytarabine/administration & dosage , Etoposide/administration & dosage , Female , Hematopoietic Stem Cell Transplantation , Humans , Idarubicin/administration & dosage , Kaplan-Meier Estimate , Leukemia, Promyelocytic, Acute/blood , Leukemia, Promyelocytic, Acute/mortality , Leukemia, Promyelocytic, Acute/pathology , Leukemia, Promyelocytic, Acute/surgery , Liposomes/administration & dosage , Male , Middle Aged , Mitoxantrone/administration & dosage , Neoplasm, Residual , Oncogene Proteins, Fusion/blood , Prognosis , Recurrence , Remission Induction , Survival Analysis , Time Factors , Treatment Outcome , Tretinoin/administration & dosage
4.
Rev. diagn. biol ; 54(4): 334-336, oct.-dic. 2005. ilus, tab, graf
Article in Es | IBECS | ID: ibc-044012

ABSTRACT

Se estima que entre el 5-10% de todos los casos de cáncerde mama (CM) son causados por mutaciones en los genessupresores de tumores BRCA1y BRCA2. El análisis mutacionalcompleto de estos genes ha permitido la identificación demutaciones patogénicas así como variantes de efecto desconocido(VED). Las VED se caracterizan porque su implicaciónpatogénica es confusa, y por lo tanto su importancia clínica esincierta, disminuyendo el valor de los resultados genéticos. Elobjetivo de este estudio ha sido caracterizar la significación clínicade dos nuevas variantes detectadas en un estudio previode 48 familias de la Comunidad Valenciana analizadas en elHospital Universitario La Fe de Valencia (España).Se han realizado análisis genéticos y bioinformáticos y se hanutilizado criterios bioquímicos para establecer si las VED sonpatogénicas. Estos análisis han determinado que la variantec.5025delT es una mutación deletérea y la variante c.8038C>Tafecta a secuencias consenso, requiriéndose estudios complementariospara determinar con precisión su significado


It is estimated that 5% to 10% of all breast cancer (BC)cases are caused by inherited mutations in the tumour suppressorgenes, BRCA1 and BRCA2. Whole gene mutationanalyses of these genes have led to the identification of pathogenicmutations and others variants known as genetic variantsof uncertain significance (USV). The pathogenicity of thesegenetic variants is unclear, and therefore their clinical relevanceis uncertain, diminishing the value of genetic test results. Theaim of the study has been to characterize the clinical significanceof two novel variants detected in a previous study of 48families from the Valencian Community analyzed in the UniversityHospital La Fe of Valencia (Spain). A range of genetic andbioinformatics analyses were performed and several biochemicalcriteria were used to establish whether the genetics variantsof study were pathogenic. These assays showed that thec.5025delT variant was a deleterious mutation and thec.8038C>T variant required of complementary studies to characterizeits meaning. This work highlights the importance ofstudying the USV in order to clarify their pathogenic effect thenthis information is essential for providing efficient counsellingfor BC families


Subject(s)
Female , Middle Aged , Humans , Mutation/genetics , Genes, Tumor Suppressor , Breast Neoplasms/pathology , Genetic Predisposition to Disease/genetics , Genes, BRCA1 , Genes, BRCA2
5.
Leuk Lymphoma ; 42(4): 747-56, 2001 Aug.
Article in English | MEDLINE | ID: mdl-11697505

ABSTRACT

The persistence of the AML1-ETO rearrangement performed by reverse transcription polymerase chain reaction (RT-PCR) has been reported in acute myeloid leukemia (AML) patients in long-term complete remission (CR). This persistence, which is not associated with hematological relapse, limits the clinical use of qualitative RT-PCR. Here, we present a new quantitative real-time PCR method to detect AML1-ETO rearrangement using fluorescently labeled probes. Quantitative detection of AML1-ETO was performed in capillary tubes using two fluorescently labeled probes in the LightCycler equipment. The reliability of the method was checked in twenty-two bone marrow samples and one apheresis sample from eight patients with t(8;21) collected at diagnosis and during follow-up assessment. The regression coefficients obtained for standard curves of AML1-ETO and AML were all greater than 0.98. The sensitivity attained allowed the detection of rearrangements at a dilution of 10(-5) Kasumi-1 cDNA. The intra-assay coefficient of variation was 4% for AML1-ETO, and 7% for AML. The inter-assay coefficient of variation was 19% for AML1-ETO and 12% for AML. A log reduction from two to four in the AML1-ETO/AML ratio was evident after CR. The study of the method and first results obtained in patient samples support that quantitative real-time PCR with hybridization probes is a new reliable and sensitive method to monitor minimal residual disease in AML patients. Moreover, the fluorescent probes with the Light-Cycler technology offer the advantage of a rapid detection.


Subject(s)
Oncogene Proteins, Fusion/genetics , RNA, Neoplasm/analysis , Reverse Transcriptase Polymerase Chain Reaction/standards , Transcription Factors/genetics , Acute Disease , Adolescent , Adult , Child , Child, Preschool , Core Binding Factor Alpha 2 Subunit , DNA Probes , Female , Fluorescent Dyes , Humans , Leukemia, Myeloid/diagnosis , Leukemia, Myeloid/genetics , Male , Middle Aged , Neoplasm, Residual/diagnosis , Neoplasm, Residual/genetics , RUNX1 Translocation Partner 1 Protein , Reference Standards , Reproducibility of Results , Sensitivity and Specificity , Translocation, Genetic
6.
Blood ; 98(8): 2332-8, 2001 Oct 15.
Article in English | MEDLINE | ID: mdl-11588027

ABSTRACT

The potential role of unrelated donor cord blood transplantation (UD-CBT) in adults remains unclear. This study reports the results of UD-CBT in 22 adults with hematologic malignancies following conditioning with thiotepa, busulfan, cyclophosphamide, and antithymocyte globulin in 21, with thiotepa, fludarabine, and antithymocyte globulin in 1, and graft-versus-host disease (GVHD) prophylaxis with cyclosporine and prednisone. Median age was 29 years (range, 18-46 years), and median weight was 69.5 kg (range, 41-85 kg). HLA match was 6 of 6 in 1 case, 5 of 6 in 13 cases, and 4 of 6 in 8 cases. Median number of nucleated cells infused was 1.71 x 10(7)/kg (range, 1.01 x 10(7)/kg to 4.96 x 10(7)/kg). All 20 patients surviving more than 30 days had myeloid engraftment, and only 1, who received the lowest cell dose, developed secondary graft failure. Median time to reach an absolute neutrophil count of at least 0.5 x 10(9)/L was 22 days (range, 13-52 days). Median time to platelets numbered at least 20 x 10(9)/L was 69 days (range, 49-153 days). Seven patients (32%) developed acute GVHD above grade II, and 9 of 10 patients at risk developed chronic GVHD, which became extensive in 4 patients. Twelve patients remained alive and disease-free 3 to 45 months after transplantation. Disease-free survival (DFS) at 1 year was 53%. Age strongly influenced DFS (P =.01). For patients aged 30 years or younger, the DFS at 1 year was 73%. These preliminary results suggest that UD-CBT should be considered a reasonable alternative in young adults with hematologic malignancy and no appropriate bone marrow donor.


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Hematologic Neoplasms/therapy , Hematopoietic Stem Cell Transplantation , Adult , Antilymphocyte Serum/therapeutic use , Cyclophosphamide/therapeutic use , Female , Fetal Blood , Graft vs Host Disease/prevention & control , Hematologic Neoplasms/drug therapy , Hematologic Neoplasms/mortality , Histocompatibility Testing , Humans , Immunosuppressive Agents/therapeutic use , Infant, Newborn , Leukemia/drug therapy , Leukemia/mortality , Leukemia/therapy , Leukocyte Count , Male , Middle Aged , Myelodysplastic Syndromes/drug therapy , Myelodysplastic Syndromes/mortality , Myelodysplastic Syndromes/therapy , Platelet Count , Survival Rate , Thiotepa/therapeutic use , Treatment Failure
7.
Br J Haematol ; 114(1): 99-103, 2001 Jul.
Article in English | MEDLINE | ID: mdl-11472351

ABSTRACT

Of 167 newly diagnosed acute promyelocytic leukaemia patients, 83 patients were long (L)-form (50%), eight variable (V)-form (5%) and 76 short (S)-form (45%). The V-form and S-form groups presented a significantly higher percentage of patients with white blood cell counts > 10 x 10(9)/l (P < 0.05). The S-form cases displayed a significantly higher number of cases with M3v microgranular features (P = 0.005) and CD34 expression (P < 0.0001). There were no differences between the three isoforms in complete remission (CR) rate (overall CR 90%), but the 3-year disease-free survival was lower for V-form cases than it was for L- and S-form cases (62% vs. 94% and 89%, P = 0.056). We conclude that the V-form and S-form types are associated with some negative prognostic features at diagnosis. However, our data were only able to demonstrate an association with adverse prognosis in the V-form type and, moreover, as the number of cases was limited, needs to be confirmed in large, uniformly treated series.


Subject(s)
Leukemia, Promyelocytic, Acute/metabolism , Neoplasm Proteins/genetics , Oncogene Proteins, Fusion/genetics , Adolescent , Adult , Aged , Antigens, CD34/analysis , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Child , Child, Preschool , Disease-Free Survival , Female , Humans , Infant , Infant, Newborn , Leukemia, Promyelocytic, Acute/drug therapy , Leukemia, Promyelocytic, Acute/immunology , Leukocyte Count , Male , Middle Aged , Polymerase Chain Reaction/methods , Prognosis , Proportional Hazards Models , Protein Isoforms/genetics , Treatment Outcome , Tretinoin/therapeutic use
8.
Haematologica ; 86(6): 570-6, 2001 Jun.
Article in English | MEDLINE | ID: mdl-11418365

ABSTRACT

BACKGROUND AND OBJECTIVES: The detection of PML-RAR by reverse transcription (RT) polymerase chain reaction (PCR) in acute promyelocytic leukemia (APL) patients who are in hematologic remission influences therapeutic decision making in several trials. In the light of this, the Spanish group has recently designed an external quality assessment program (EQAP) of RT-PCR detection of PML-RAR, which includes a study of sensitivity of the participating laboratories. DESIGN AND METHODS: Eighteen laboratories were involved in the program. Ten laboratories followed the method of Biondi et al., 5 employed that of Borrow et al. and the 3 remaining used other protocols. The sensitivity was studied in five rounds of quality control. The first two shipments consisted of dilutions of NB4 RNA into non-APL RNA. The third round consisted of serial dilutions of the NB4 cell line into HL60 cells. The fourth and five rounds consisted of plasmid dilutions containing the bcr1 and bcr3 PML-RAR isoforms. RESULTS: The results showed that the distinct methods allow detection of the PML-RAR hybrid up to a dilution of 10(-4), and exceptionally, up to 10(-5). The laboratories following the method of Biondi et al. usually detected the 10(-3) dilution and less frequently the 10(-4) one, whereas those using other methods usually detected PML-RAR transcript in the 10(-4) dilution, and less commonly in the 10(-5) dilution. However, each of the PCR methods used by EQAP participating laboratories successfully detected at least 50 copies of PML-RAR alpha fusion transcript in plasmid dilution controls. INTERPRETATION AND CONCLUSIONS: The results point to heterogeneous sensitivity amongst participating laboratories. This may reflect differences in methodology, although variations in sample quality may also account for discrepant findings.


Subject(s)
Laboratories/standards , Neoplasm Proteins/analysis , Oncogene Proteins, Fusion/analysis , Reverse Transcriptase Polymerase Chain Reaction/standards , Humans , Neoplasm Proteins/genetics , Observer Variation , Oncogene Proteins, Fusion/genetics , Quality Control , RNA, Messenger/metabolism , Reproducibility of Results , Tumor Cells, Cultured
9.
Bone Marrow Transplant ; 27(7): 693-701, 2001 Apr.
Article in English | MEDLINE | ID: mdl-11360108

ABSTRACT

The potential role of unrelated donor cord blood transplantation (UD-CBT) in adults is not well established. We report the results of UD-CBT in nine adult patients with chronic myeloid leukemia (CML). The median age was 27 years (range, 19-41 years), and the median weight was 62 kg (range, 45-78 kg). At transplant, six patients were in chronic phase (five in first, and one in second), two in blast crisis, and one in accelerated phase. Eight had received intensive chemotherapy, and three had undergone autologous peripheral blood hematopoietic stem cell transplantation. Four had received interferon with no cytogenetic response, and only three underwent UD-CBT within 1 year of diagnosis. After serological typing for class I antigens, and high-resolution DNA typing for DRB1, the degree of HLA match between patients and cord blood (CB) units was 4/6 in six cases and 5/6 in three cases. The median number of nucleated cells infused was 1.7 x 10(7)/kg (range, 1.2 to 4.9 x 10(7)/kg), and was above 2 x 10(7)/kg in only two cases. All patients received thiotepa, busulfan, cyclophosphamide and anti-thymocyte globulin as conditioning; cyclosporine and prednisone for graft-versus-host disease (GVHD) prophylaxis; and G-CSF from day +7 until engraftment. All seven evaluable cases engrafted. The median time to reach an absolute neutrophil count > or =0.5 x 10(9)/l and > or =1 x 10(9)/l was 22 days (range, 19-52 days) and 28 days (range, 23-64 days), respectively. In the four patients evaluable for platelet recovery time to levels of > or =20 x 10(9) platelets/l, > or =50 x 10(9) platelets/l, and > or =100 x 10(9) platelets/l, these ranged from 50 to 128 days, 60 to 139 days, and 105 to 167 days, respectively. Three patients developed acute GVHD above grade II, and three of the five patients at risk developed extensive chronic GVHD. Four patients, all transplanted in chronic phase, remain alive in molecular remission more than 18, 19, 24 and 42 months after transplantation. These preliminary results suggest that UD-CBT may be considered a reasonable alternative in adults with CML who lack an appropriate bone marrow donor.


Subject(s)
Blood Donors , Fetal Blood/cytology , Hematopoietic Stem Cell Transplantation/standards , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/therapy , Acute Disease , Adult , Chronic Disease , Disease-Free Survival , Graft vs Host Disease/etiology , Graft vs Host Disease/immunology , Hematopoietic Stem Cell Transplantation/adverse effects , Histocompatibility , Histocompatibility Testing , Humans , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/complications , Pilot Projects , Survival Rate , Treatment Outcome
10.
Haematologica ; 85(12): 1248-54, 2000 Dec.
Article in English | MEDLINE | ID: mdl-11114130

ABSTRACT

BACKGROUND AND OBJECTIVES: The limited value of qualitative reverse transcription polymerase chain-reaction (RT-PCR) for monitoring chronic myeloid leukemia (CML) patients has prompted the development of quantitative assays. We have developed a quantitative real-time PCR (QC-PCR) method in the LightCycler, based on the use of fluorescently labeled probes (HybProbes), to estimate BCR-ABL fusion gene transcripts in samples from CML patients. DESIGN AND METHODS: Fifty-two samples (45 peripheral blood, five bone marrow, and two apheresis product samples) from nine patients with CML were analyzed. Seven patients were studied at diagnosis and during follow-up after hematopoietic stem cell transplantation (HSCT), whereas two were evaluated only after HSCT. The PCR reaction was carried out in capillary tubes in a final volume of 10 microL, using 2 microL cDNA, the Mensik et al. primers, and two HybProbes. The results for BCR-ABL were normalized with reference to ABL. The PCR program is completed in only 45 min. RESULTS: The sensitivity attained allowed the detection of rearrangements at dilutions of between 5-10(-4) and 10(-5) K562 cDNA. The within-assay coefficient of variation was 11% for BCR-ABL, and 9% for ABL. A greater than 2 log reduction in the BCR-ABL/ABL ratio was evident shortly after transplantation in all allografted patients. INTERPRETATION AND CONCLUSIONS: We may conclude that the TaqMan probe technology can be easily adapted to HybProbes with equivalent results. Besides, the results of BCR-ABL quantification in the follow-up of patients clearly confirm that real-time PCR with HybProbes is a reliable and sensitive method for monitoring minimal residual leukemia after HSCT in CML patients.


Subject(s)
Fusion Proteins, bcr-abl/genetics , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/genetics , Reverse Transcriptase Polymerase Chain Reaction/standards , Adolescent , Adult , Bone Marrow/metabolism , Female , Fluorescent Dyes , Follow-Up Studies , Humans , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/diagnosis , Male , Middle Aged , RNA, Messenger/blood , RNA, Messenger/metabolism
11.
Blood ; 96(4): 1247-53, 2000 Aug 15.
Article in English | MEDLINE | ID: mdl-10942364

ABSTRACT

Preliminary independent reports of the Italian GIMEMA and the Spanish PETHEMA trials for newly diagnosed acute promyelocytic leukemia (APL) indicated a similarly high antileukemic efficacy in terms of complete remission and disease-free survival rates. To better investigate these studies and the prognostic factors influencing relapse risk, this study analyzed the updated results of 217 patients with PML/RAR alpha-positive APL enrolled in GIMEMA (n = 108) and PETHEMA (n = 109). All patients received identical induction (AIDA schedule) and maintenance. For consolidation, GIMEMA patients received 3 courses including idarubicin/cytarabine, mitoxantrone/etoposide, and idarubicin/cytarabine/thioguanine, whereas PETHEMA patients received the same drugs and dose schedule of idarubicin and mitoxantrone with the omission of nonintercalating agents. Depending on whether molecular relapses were classified as censored or uncensored events, the 3-year Kaplan-Meier estimates of relapse-free survival (RFS) for the combined series were 90 +/- 2% and 86 +/- 2%, respectively. Minor differences observed between the 2 patient cohorts were negligible. Multivariate regression analysis of RFS showed that initial leukocyte (WBC) and platelet counts were the only variables with independent prognostic value. The resulting predictive model for RFS demonstrated its capability of segregating patients into low-risk (WBC count 40 x 10(9)/L), intermediate-risk (WBC count 10 x 10(9)/L) groups, with distinctive RFS curves (P <.0001). The conclusions are that omission of nonanthracycline drugs from the AIDA regimen is not associated with reduced antileukemic efficacy and a simple predictive model may be used for risk-adapted therapy in this disease. (Blood. 2000;96:1247-1253)


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/administration & dosage , Leukemia, Promyelocytic, Acute/drug therapy , Leukemia, Promyelocytic, Acute/pathology , Adolescent , Adult , Aged , Cohort Studies , Cytarabine/administration & dosage , Etoposide/administration & dosage , Female , Humans , Idarubicin/administration & dosage , Male , Middle Aged , Mitoxantrone/administration & dosage , Multivariate Analysis , Predictive Value of Tests , Recurrence , Risk , Thioguanine/administration & dosage , Tretinoin/administration & dosage
12.
Haematologica ; 85(7): 699-703, 2000 Jul.
Article in English | MEDLINE | ID: mdl-10897121

ABSTRACT

BACKGROUND AND OBJECTIVES: The detection of CBFbeta/MYH11 transcripts by RT-PCR has became a valuable and widely used technique in the accurate cytogenetic and molecular classification of acute myeloid leukemia (AML), but the clinical value of RT-PCR for monitoring minimal residual disease (MRD) during follow-up remains unclear. DESIGN AND METHODS: We analyzed the factors predicting relapse and the value of MRD monitoring by RT-PCR in a series of 16 patients with CBFb/MYH11-positive AML (15 M4Eo; 1 M4). Fifteen were newly diagnosed cases (CR1) and one was studied after first relapse (CR2). Eight patients had clinical relapse at 6 to 19 months from the achievement of CR. RESULTS: Presenting WBC count had a significant prognostic influence on disease-free survival (p=0.001). All four patients with a WBC count >100x10(9)/L relapsed, while only four additional relapses occurred among the eleven patients who had an initial WBC count below 100x10(9)/L. With regards to molecular monitoring, all relapses but one occurred in patients who showed persistent RT-PCR positivity during hematologic remission. By contrast, conversion to a repeatedly PCR-negative status was observed in the seven patients who remained in CR1 after a median follow-up of 48 months (range 31-79 months), as well as in the transplanted patient who was monitored in CR2. In these patients a PCR-positivity could be detected up to 24 months after diagnosis (median time to conversion to PCR-negative: 8 months). INTERPRETATION AND CONCLUSIONS: In conclusion, marked hyperleukocytosis (>100x10(9)/L) confers poor prognosis to the patient with CBFbeta/MYH11-positive AML. In addition, slow kinetics of molecular remission was observed in this subset of AML, but the CBFb/MYH11 fusion transcript is no longer detectable in long-term survivors, indicating that molecular remission is an important therapeutic goal.


Subject(s)
Leukemia, Myeloid/diagnosis , Leukemia, Myeloid/genetics , Oncogene Proteins, Fusion/genetics , Acute Disease , Adolescent , Adult , Aged , Child , Chromosome Aberrations/diagnosis , Chromosome Aberrations/genetics , Chromosome Aberrations/pathology , Chromosome Disorders , Chromosomes, Human, Pair 16 , Disease-Free Survival , Female , Gene Rearrangement , Humans , Kinetics , Leukemia, Myeloid/blood , Leukocyte Count , Male , Middle Aged , Neoplasm, Residual/diagnosis , Oncogene Proteins, Fusion/blood , Prognosis , RNA, Messenger/blood , Recurrence , Reverse Transcriptase Polymerase Chain Reaction
13.
Med Clin (Barc) ; 114(8): 281-5, 2000 Mar 04.
Article in Spanish | MEDLINE | ID: mdl-10774514

ABSTRACT

BACKGROUND: Molecular assay commonly used to detect the PML/RAR alpha rearrangement in acute promyelocytic leukemia (APL) has the limited sensitivity in comparison with the higher sensitivity of RAR alpha/PML detection. This prompted us to perform both assays in parallel to monitor a group of APL. PATIENTS AND METHODS: The study included 56 APL patients mainly treated according with the PETHEMA LPA-96 protocol. The PML/RAR alpha was detected according with Biondi's et al method and the RAR alpha/PML following the Grimwade's et al RT-PCR method (Human Press Inc.). RESULTS: RAR alpha/PML rearrangement was detected in 90% (20/22) of the patients at diagnosis positives for PML/RAR alpha. RAR alpha/PML was detected in 74% (14/19) of post-induction samples versus 37% (7/19) of positives for PML/RAR alpha. Likewise RAR alpha/PML rearrangement was detected in some post-consolidation samples (2/11) that all were PMI/RAR alpha negatives. In patients in maintenance regimen a greater proportion of RAR alpha/PML positives (6/28) versus PML/RAR alpha (2/28) were observed. In a patient in complete remission RAR alpha/PML preceded the positivity of PML/RAR alpha and persisted after PMI/RAR alpha negativization. The results of the patients monitored since the diagnosis showed that RAR alpha/PML revert to negative one month after PML/RAR alpha negativization. CONCLUSIONS: RAR alpha/PML rearrangement is not expressed in the totality of the APL patients, but in only a 90% of them. RAR alpha/PML rearrangement was detected in a greater proportion of samples than PML/RAR alpha. RAR alpha/PML rearrangement lasted longer than PML/RAR alpha after treatment.


Subject(s)
Gene Rearrangement/genetics , Leukemia, Promyelocytic, Acute/genetics , Neoplasm Proteins/genetics , Oncogene Proteins, Fusion/genetics , Humans , Neoplasm, Residual , Sensitivity and Specificity
14.
Blood ; 94(9): 3015-21, 1999 Nov 01.
Article in English | MEDLINE | ID: mdl-10556184

ABSTRACT

The Spanish PETHEMA group designed a protocol for newly diagnosed PML/RARalpha-positive acute promyelocytic leukemia (APL) in which induction and consolidation followed the original AIDA regimen, except for the omission of cytarabine and etoposide from consolidation. Induction consisted of 45 mg/m(2) all-trans retinoic acid (ATRA) daily until complete remission (CR) and 12 mg/m(2) idarubicin on days 2, 4, 6, and 8. Patients in CR received 3 monthly chemotherapy courses: idarubicin 5 mg/m(2)/d x 4 (course no. 1), mitoxantrone 10 mg/m(2)/d x 5 (course no. 2), and idarubicin 12 mg/m(2)/d x 1 (course no. 3). Maintenance therapy consisted of 90 mg/m(2)/d mercaptopurine orally, 15 mg/m(2)/wk methotrexate intramuscularly, and, intermittently, 45 mg/m(2)/d ATRA for 15 days every 3 months. Between November 1996 and December 1998, 123 patients with newly diagnosed PML/RARalpha-positive APL from 39 centers were enrolled. A total of 109 patients achieved CR (89%; 95% confidence interval [CI], 83 to 95), 12 died of early complications, and the remaining 2 were resistant. Consolidation treatment was associated with very low toxicity and no deaths in remission were recorded. Molecular assessment of response by reverse transcriptase-polymerase chain reaction (RT-PCR) showed conversion to PCR-negative in 48 of 99 (51%) and 82 of 88 patients (93%) after induction and consolidation, respectively. The 2-year Kaplan-Meier estimates of overall survival and event-free survival were 82% +/- 4% and 79% +/- 4%, respectively. For patients who achieved CR, the 2-year disease-free survival (DFS) was 92% +/- 3%. These data indicate that a significant reduction in toxicity might be obtained in APL using a less intensive consolidation without apparently compromising the antileukemic effect. These results also suggest a minor role for cytarabine and etoposide in the treatment of newly diagnosed PML/RARalpha-positive APL patients.


Subject(s)
Anthracyclines/administration & dosage , Antineoplastic Combined Chemotherapy Protocols/administration & dosage , Leukemia, Promyelocytic, Acute/drug therapy , Neoplasm Proteins , Oncogene Proteins, Fusion , Adolescent , Adult , Aged , Anthracyclines/adverse effects , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Female , Humans , Idarubicin/administration & dosage , Idarubicin/adverse effects , Male , Middle Aged , Treatment Outcome , Tretinoin/administration & dosage , Tretinoin/adverse effects
15.
Leuk Res ; 23(9): 851-4, 1999 Sep.
Article in English | MEDLINE | ID: mdl-10475625

ABSTRACT

CML with exclusive expression of ALL-type bcr/abl has only been rarely described. In some cases, the presence of this fusion gene has been associated to a differentiated subtype of CML that share some features with CMML, while in another case this molecular hallmark has been associated to a bad prognosis of the disease with a blast phase as clinical presentation or an early transformation to blast phase. We report a case of a 30-year-old woman who was diagnosed of CML in chronic phase in May 1989. She received treatment first with busulfan, achieving hematological remission and afterwards with interferon and Hydroxiurea. In February 1998, she was admitted at our hospital for an ABSCT. Then, molecular studies were performed. Multiplex PCR revealed the presence of a 481 bp product identified as the ela2 bcr/abl transcript and confirmed by sequencing. After 9 years from diagnosis, the patient remains in hematological remission and in good clinical condition.


Subject(s)
Fusion Proteins, bcr-abl/genetics , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/genetics , Precursor Cell Lymphoblastic Leukemia-Lymphoma/genetics , RNA, Messenger/biosynthesis , Adult , Female , Humans
16.
Cancer Genet Cytogenet ; 110(2): 111-4, 1999 Apr 15.
Article in English | MEDLINE | ID: mdl-10214358

ABSTRACT

The inv(16) and t(16;16) characterize a subgroup of acute myelomonocytic leukemia (AML) with distinct morphological features and a favorable prognosis. Both cytogenetic abnormalities result in a fusion of CBF beta at 16q22 and MYH11 gene at 16p13, whose detection by PCR and fluorescence in situ hybridization (FISH) is useful for diagnosis and monitoring of the disease. Variant translocations of inv(16)/t(16;16) are very rare and whether they are also associated with a favorable prognosis is unknown. We report a patient presenting with typical AML-M4Eo and a three-way translocation of inv(16) involving 16p13, 16q22, and 3q22. FISH studies on bone marrow (BM) chromosomes using CBFB and MYH11 DNA probes revealed a fusion of CBFB and MYH11 on 16q of the der(16), as well as a signal from MYH11 on 16p but not from CBFB; normal signals for both probes were present on the normal 16. Neither of these labeled probes was on the der(3), but the translocation between the der(3) and der(16) was confirmed by using a chromosome 16 painting probe. Molecular analysis of BM cells using RT-PCR identified a CBFB-MYH11 fusion transcript type D. After achieving complete remission, the patient relapsed. We conclude that FISH and PCR are feasible tools to distinguish cases with variant abnormalities of inv(16) from cases with other chromosome 16 abnormalities. Variant abnormalities of inv(16) may be not associated with favorable prognosis.


Subject(s)
Chromosome Inversion , Chromosomes, Human, Pair 16 , Leukemia, Myelomonocytic, Acute/genetics , Translocation, Genetic , Adult , Eosinophils/pathology , Female , Humans , In Situ Hybridization, Fluorescence , Male , Reverse Transcriptase Polymerase Chain Reaction
17.
Leukemia ; 12(12): 2024-8, 1998 Dec.
Article in English | MEDLINE | ID: mdl-9844933

ABSTRACT

To standardize the results obtained in PML/RAR alpha RT-PCR detection by laboratories of hospitals involved in the Spanish Program for Treatment of Hematological Malignancies (PETHEMA) LPA-96, designed for the treatment of acute promyelocytic leukemia (APL), cDNA samples obtained by reverse transcription of RNA from bone marrow samples of patients with APL were sent to participating laboratories. During the first year of this external quality assessment trial nine samples were tested by a maximum of 12 laboratories. The control gene was satisfactorily amplified in 90% of the samples (62 of 69 samples), supporting the adequacy of the cDNA to be used as control sample. There was an 83% concordance between laboratories for PML/RAR alpha detection with similar results for the type of PML/RR alpha rearrangements. However, 17% disagreement still remained, attributable to low sensitivity or inadequacy of methods followed. The results stressed the need for implementation of an external quality assessment scheme to ensure the standardization of the results.


Subject(s)
Leukemia, Promyelocytic, Acute/genetics , Neoplasm Proteins/analysis , Oncogene Proteins, Fusion/analysis , Reverse Transcriptase Polymerase Chain Reaction/standards , Gene Amplification , Humans , Quality Control , Reproducibility of Results , Spain
18.
Leukemia ; 12(6): 992-5, 1998 Jun.
Article in English | MEDLINE | ID: mdl-9639431

ABSTRACT

The contribution of residual malignant cells contaminating the autologous graft with the occurrence of post-transplant relapse in acute myeloid leukemia (AML) is still unclear. The presence of a specific molecular marker (the PML/RAR alpha rearrangement) in acute promyelocytic leukemia (APL) offers the opportunity to investigate better the pathogenesis of disease recurrence after transplant. We report an APL patient who received high-dose chemotherapy and peripheral blood stem cell (PBSC) autograft in second hematologic remission. Two leukaphereses that tested PML/RAR alpha positive by RT-PCR were obtained during the post-reinduction hematopoietic recovery, while the patient also tested PCR positive in the BM, and was reinfused after myeloablative chemotherapy (BUCY4), when the patient had spontaneously converted to PCR negative in the marrow. At present, he remains in continuous molecular and hematologic remission 22 months after PBSC transplantation. This is the second report of an APL patient who was transplanted in molecular remission with a PML/RAR alpha-positive PBSC autograft. As in the previous report, the prolonged clinical and molecular remission experienced post-transplant suggests that autologous PBSC infusion is still worthy of consideration for patients with APL in spite of the detection of PML/RAR alpha-positive cells in the PBSC collections. Possible underlying mechanisms and the potential role of molecular monitoring of the graft, as well as the host, before and after transplant, in patients with APL undergoing autologous HSCT are also discussed.


Subject(s)
Hematopoietic Stem Cell Transplantation , Leukemia, Promyelocytic, Acute/therapy , Neoplasm Proteins/genetics , Nuclear Proteins , Receptors, Retinoic Acid/genetics , Transcription Factors/genetics , Adult , Gene Rearrangement , Humans , Leukemia, Promyelocytic, Acute/genetics , Male , Neoplasm, Residual , Polymerase Chain Reaction , Promyelocytic Leukemia Protein , Recurrence , Retinoic Acid Receptor alpha , Transplantation, Autologous , Tumor Suppressor Proteins
19.
Clin Chem Lab Med ; 36(3): 137-42, 1998 Mar.
Article in English | MEDLINE | ID: mdl-9589800

ABSTRACT

Two reverse transcription-polymerase chain reaction methods to detect the AML1/ETO rearrangement in the M2 subtype of acute myeloid leukaemia those of Downing et al. (Blood 1993; 81: 2860-5) and Satake et al. (Br J Haematol 1995; 91: 892-8) were evaluated. Bone marrow samples, one at diagnosis and two in complete remission from a patient with M2 subtype of acute myeloid leukaemia, with t(8;21), were analysed using both methods. The Kasumi-1 cell line was used as a positive control and a patient with M3 subtype of acute myeloid leukaemia as a negative control. To confirm the feasibility of Satake's method a group of 35 patients with subtypes of acute myeloid leukaemia at diagnosis were studied. The method of Downing requires Southern blotting and hybridization with a specific probe because it often generates non-specific amplification products. By contrast, the method of Satake yields only a single amplification product, using one single round of PCR in samples at diagnosis, or two rounds in complete remission samples. The sensitivity of this method allows the detection of a single Kasumi-1 cell in 10(6) normal cells. The AML1/ETO rearrangement was observed in 5 of the 35 cases of acute myeloid leukaemia at diagnosis (14.3%) and in 3 of the 14 cases of M2 subtype of acute myeloid leukaemia (21.4%). The two remaining positive cases corresponded to the acute myeloid leukaemia subtypes M4 and M6. The results indicate that the method of Satake better meets the requirements of the clinical laboratory due to its greater simplicity, specificity, sensitivity and feasibility, thus making it more appropriate for use in diagnosing and monitoring minimal residual disease.


Subject(s)
DNA-Binding Proteins/genetics , Gene Rearrangement/genetics , Leukemia, Myeloid, Acute/genetics , Polymerase Chain Reaction/methods , Proto-Oncogene Proteins , Transcription Factors/genetics , Base Sequence , Blotting, Southern , DNA-Binding Proteins/chemistry , Female , Humans , Leukemia, Erythroblastic, Acute/genetics , Leukemia, Myelomonocytic, Acute/genetics , Leukemia, Promyelocytic, Acute/genetics , Molecular Sequence Data , RUNX1 Translocation Partner 1 Protein , Transcription Factors/chemistry
20.
Clin Chim Acta ; 262(1-2): 99-119, 1997 Jun 27.
Article in English | MEDLINE | ID: mdl-9204213

ABSTRACT

The aim of this study is to evaluate the prognostic significance of c-erbB-2/neu amplification and epidermal growth factor receptor (EGFR) expression in primary breast cancer (BC) and their prognostic implications when combined with estradiol receptor (ER) status. In this work, 825 BCs were studied. Neu amplification was evaluated by dot-blot and EGFR expression was evaluated by ligand binding assay using I125-EGF. Neu, EGFR, estradiol and progesterone receptors (ER and PR) had a marked influence on disease free survival (DFS) in univariate analysis. In node-negative (NO) cases only neu was associated with short DFS (p = 0.005). However, in node-positive (N+) cases both EGFR (p = 0.005) and neu (p = 0.002) influenced DFS. None of the biological markers were significant predictors for overall survival (OS) in NO/BC. On the contrary, in N+/BC, EGFR + (p = 0.003) was associated with short OS. The EGFR + /neu/phenotype represented a sub-group with an even worse prognosis with respect to DFS (p = 0.0034) as well as EGFR + /ER-tumors (p = 0.005). Moreover, neu + /ER-patients also had a high probability of relapse (p = 0.0000) and death (p = 0.006). C-erbB-2/neu, EGFR, histological grade, pN, pT and ER were subjected to a Cox multivariate regression analysis: neu was the most important parameter in predicting recurrence, and EGFR was a significant predictor for OS.


Subject(s)
Breast Neoplasms/genetics , Breast Neoplasms/metabolism , ErbB Receptors/metabolism , Gene Amplification , Genes, erbB-2 , Receptors, Estradiol/metabolism , Adult , Aged , Aged, 80 and over , Breast Neoplasms/pathology , Disease-Free Survival , ErbB Receptors/biosynthesis , ErbB Receptors/genetics , Female , Humans , Lymph Nodes/pathology , Middle Aged , Multivariate Analysis , Prognosis , Survival Analysis
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