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David Grimwade

Publications and source records attributed to David Grimwade.

17 recordsLinked to original sources
PubMed2006

Expression of T-lineage-affiliated transcripts and TCR rearrangements in acute promyelocytic leukemia: implications for the cellular target of t(15;17).

Acute promyelocytic leukemia (APL) is the most differentiated form of acute myeloid leukemia (AML) and has generally been considered to result from transformation of a committed myeloid progenitor. Paradoxically, APL has long been known to express the T-cell lymphoid marker, CD2. We searched for other parameters indicative of T-cell lymphoid specification in a cohort of 36 APL cases, revealing a frequent but asynchronous T-cell lymphoid program most marked in the hypogranular variant (M3v) subtype, with expression of PTCRA, sterile TCRA, and TCRG transcripts and TCRG rearrangement in association with sporadic cytoplasmic expression of CD3 or TdT proteins. Gene-expression profiling identified differentially expressed transcription factors that have been implicated in lymphopoiesis. These data carry implications for the hematopoietic progenitor targeted by the PML-RARA oncoprotein in APL and are suggestive of a different cellular origin for classic hypergranular (M3) and variant forms of the disease. They are also consistent with the existence and subsequent transformation of progenitor populations with lymphoid/myeloid potential.

Cell Lineage↗
PubMed2006

Monitoring CML patients responding to treatment with tyrosine kinase inhibitors: review and recommendations for harmonizing current methodology for detecting BCR-ABL transcripts and kinase domain mutations and for expressing results.

The introduction in 1998 of imatinib mesylate (IM) revolutionized management of patients with chronic myeloid leukemia (CML) and the second generation of tyrosine kinase inhibitors may prove superior to IM. Real-time quantitative polymerase chain reaction (RQ-PCR) provides an accurate measure of the total leukemiacell mass and the degree to which BCR-ABL transcripts are reduced by therapy correlates with progression-free survival. Because a rising level of BCR-ABL is an early indication of loss of response and thus the need to reassess therapeutic strategy, regular molecular monitoring of individual patients is clearly desirable. Here we summarize the results of a consensus meeting that took place at the National Institutes of Health (NIH) in Bethesda in October 2005. We make suggestions for (1) harmonizing the differing methodologies for measuring BCR-ABL transcripts in patients with CML undergoing treatment and using a conversion factor whereby individual laboratories can express BCR-ABL transcript levels on an internationally agreed scale; (2) using serial RQ-PCR results rather than bone marrow cytogenetics or fluorescence in situ hybridization (FISH) for the BCR-ABL gene to monitor individual patients responding to treatment; and (3) detecting and reporting Philadelphia (Ph) chromosome-positive subpopulations bearing BCR-ABL kinase domain mutations. We recognize that our recommendations are provisional and will require revision as new evidence emerges.

Drug Monitoring↗
PubMed2006

Minimal residual disease directed therapy for childhood acute myeloid leukaemia: the time is now.

The continued improvement in the prognosis of childhood acute myeloid leukaemia (AML) has been paralleled by the use of increasingly intensive therapy. This has led to attempts to develop risk-directed strategies in which the most intensive treatment is reserved for those at highest risk of relapse. Unfortunately, current approaches, which rely on cytogenetic sub-grouping and morphological assessment of response to therapy, are inaccurate. New prognostic factors are needed. This annotation proposes that the introduction of protocols based on the measurement of minimal residual disease (MRD) holds the key to progression from an era of 'cure at all costs' to a more individualised approach. However, the full potential of MRD technologies will only be realised through properly designed studies with scrupulous attention to logistics and quality assurance. The article illustrates which children may benefit most from MRD analysis in AML and explores practical issues that should be addressed in the design of clinical trials.

Child↗
PubMed2005

Rexinoid-triggered differentiation and tumor-selective apoptosis of acute myeloid leukemia by protein kinase A-mediated desubordination of retinoid X receptor.

Apart from PML-retinoic acid receptor-alpha (RARalpha) acute promyelocytic leukemia all other acute myeloid leukemias (AML) are unresponsive to retinoid differentiation therapy. However, elevating the levels of cyclic AMP (cAMP) confers onto retinoid X receptor (RXR)-selective agonists ("rexinoids") the ability to induce terminal granulocyte differentiation and apoptosis of all-trans retinoic acid-resistant and insensitive AML cells and patients' blasts. Protein kinase A activation leads to corepressor release from the RAR subunit of the RAR-RXR heterodimer, resulting in "desubordination" of otherwise silent RXR, which acquires transcriptional competence in response to cognate ligands. Rexinoid-cAMP induction of endogenous RARbeta is blunted in mouse embryo fibroblasts lacking RARs, but reintroduction of exogenous RARalpha reestablishes responsiveness, thus confirming that the RARalpha-RXR heterodimer is the rexinoid mediator. The apoptogenic effect of this treatment involves enhanced expression of the death receptor DR5 and its cognate ligand, tumor necrosis factor-related apoptosis inducing ligand, both of which are known to induce apoptosis in a tumor cell-selective manner and lead to the activation of initiator caspases. Immunohistochemistry confirmed induction of tumor necrosis factor-related apoptosis inducing ligand and DR5 in AML patient blasts cultured ex vivo. AML patients' blasts responded to rexinoid-cAMP combination treatment with induction of maturation and apoptosis, independent of karyotype, immunophenotype, and French-American-British classification status. Clonogenic assays revealed complete inhibition of blast clonogenicity in four out of five tested samples. Our results suggest that despite the genetic, morphologic, and clinical variability of this disease, the combination of rexinoids and cAMP-elevating drugs, such as phosphodiesterase inhibitors, might lead to a novel therapeutic option for AML patients by inducing a tumor-selective death pathway.

Acute Disease↗
PubMed2005

Relationship between FLT3 mutation status, biologic characteristics, and response to targeted therapy in acute promyelocytic leukemia.

The prognostic significance of FLT3 mutations in acute promyelocytic leukemia (APL) is not firmly established and is of particular interest given the opportunities for targeted therapies using FLT3 inhibitors. We studied 203 patients with PML-RARA-positive APL; 43% of the patients had an FLT3 mutation (65 internal tandem duplications [ITDs], 19 D835/I836, 4 ITD+D835/I836). Both mutations were associated with higher white blood cell (WBC) count at presentation; 75% of the patients with WBC counts of 10 x 10(9)/L or greater had mutant FLT3. FLT3/ITDs were correlated with M3v subtype (P < .001), bcr3 PML breakpoint (P < .001), and expression of reciprocal RARA-PML transcripts (P = .01). Microarray analysis revealed differences in expression profiles among patients with FLT3/ITD, D835/I836, and wild-type FLT3. Patients with mutant FLT3 had a higher rate of induction death (19% vs 9%; P = .04, but no significant difference in relapse risk (28% vs 23%; P = .5) or overall survival (59% vs 67%; P = .2) at 5 years. In in vitro differentiation assays using primary APL blasts (n = 6), the FLT3 inhibitor CEP-701 had a greater effect on cell survival/proliferation in FLT3/ITD+ cells, but this inhibition was reduced in the presence of ATRA. Furthermore, in the presence of CEP-701, ATRA-induced differentiation was reduced in FLT3/ITD+ cells. These data carry implications for the use of FLT3 inhibitors as frontline therapy for APL.

Adolescent↗
PubMed2005

DNA topoisomerase II in therapy-related acute promyelocytic leukemia.

BACKGROUND: Chromosomal translocations leading to chimeric oncoproteins are important in leukemogenesis, but how they form is unclear. We studied acute promyelocytic leukemia (APL) with the t(15;17) translocation that developed after treatment of breast or laryngeal cancer with chemotherapeutic agents that poison topoisomerase II. METHODS: We used long-range polymerase chain reaction and sequence analysis to characterize t(15;17) genomic breakpoints in therapy-related APL. To determine whether topoisomerase II was directly involved in mediating breaks of double-stranded DNA at the observed translocation breakpoints, we used a functional in vitro assay to examine topoisomerase II-mediated cleavage in the normal homologues of the PML and RARA breakpoints. RESULTS: Translocation breakpoints in APL that developed after exposure to mitoxantrone, a topoisomerase II poison, were tightly clustered in an 8-bp region within PML intron 6. In functional assays, this "hot spot" and the corresponding RARA breakpoints were common sites of mitoxantrone-induced cleavage by topoisomerase II. Etoposide and doxorubicin also induced cleavage by topoisomerase II at the translocation breakpoints in APL arising after exposure to these agents. Short, homologous sequences in PML and RARA suggested the occurrence of DNA repair by means of the nonhomologous end-joining pathway. CONCLUSIONS: Drug-induced cleavage of DNA by topoisomerase II mediates the formation of chromosomal translocation breakpoints in mitoxantrone-related APL and in APL that occurs after therapy with other topoisomerase II poisons.

Antineoplastic Agents↗
PubMed2005

Biology and management of relapsed acute myeloid leukaemia.

Disease relapse remains the major cause of treatment failure in adults with acute myeloid leukaemia (AML). This reflects both the failure of current salvage regimens and the absence of effective strategies to secure long-term disease-free survival in those patients who achieve a second remission. Recent progress in understanding the pathogenesis of relapsed disease has enabled the identification of a variety of dysregulated molecular pathways and these now provide a rational basis for the design of novel targeted therapies. At the same time, advances in allogeneic stem-cell transplantation have permitted the extension of the curative potential of allografting to patients in whom it was previously contraindicated. As a result, a range of novel drug and transplant therapies has become available in patients with relapsed AML, and it is realistic to anticipate that a co-ordinated assessment of their clinical and biological impact will provide the basis for the design of future, more effective treatments in relapsed disease.

Acute Disease↗
PubMed2004

Pathogenesis, diagnosis and monitoring of residual disease in acute promyelocytic leukaemia.

The clinical course of acute promyelocytic leukaemia (APL) has changed over the last 25 years from one that was fatal for the majority of patients to representing one of the most curable subtypes of acute myeloid leukaemia. Besides improved supportive care this has mainly been achieved through the introduction of novel targeted therapies in the form of all-trans retinoic acid (ATRA) and arsenic trioxide that specifically address the underlying molecular lesion. APL is characterized by chromosomal rearrangements of 17q21 leading to the formation of fusion proteins involving retinoic acid receptor alpha (RARA). To date five different fusion partners of RARA have been identified, but the vast majority of cases are characterized by the presence of the t(15;17)(q22;q12-21), which involves the promyelocytic leukaemia (PML) gene. The identification of different breakpoint microclusters within RARA intron 2 suggests that sequence-associated or structural factors play a role in the formation of the t(15;17). In addition, the comparison of forward and reverse genomic junctions has revealed microhomologies, deletions and/or duplications of either gene consistent with the hypothesis that the t(15;17) occurs by non-homologous recombination of DNA after processing of the double strand breaks by a dysfunctional DNA damage repair mechanism. The detection of the PML-RARA fusion gene by reverse-transcription polymerase chain reaction (RT-PCR) is routinely used for diagnosis and monitoring of minimal residual disease (MRD). In PML-RARA-positive APL about 70% of patients are expected to be cured with a combination of ATRA and anthracycline-based chemotherapy. However, relapse remains a major problem. The identification of patients at high risk of relapse and the development of risk-adapted treatment schedules are therefore clearly the most challenging tasks in the treatment of APL. Recent studies have shown that pre-emptive chemotherapy at the time of molecular relapse improves survival compared to treatment at the point of haematological relapse. Quantitative RT-PCR technology is expected to further improve the predictive value of MRD monitoring and therefore to guide therapy in order to reduce the rate of relapses and to increase rates of cure in high-risk patients.

Humans↗
PubMed2003

Global down-regulation of HOX gene expression in PML-RARalpha + acute promyelocytic leukemia identified by small-array real-time PCR.

Acute promyelocytic leukemia (APL) is associated with a reciprocal and balanced translocation involving the retinoic acid receptor-alpha (RARalpha). All-trans retinoic acid (ATRA) is used to treat APL and is a potent morphogen that regulates HOX gene expression in embryogenesis and organogenesis. HOX genes are also involved in hematopoiesis and leukemogenesis. Thirty-nine mammalian HOX genes have been identified and classified into 13 paralogous groups clustered on 4 chromosomes. They encode a complex network of transcription regulatory proteins whose precise targets remain poorly understood. The overall function of the network appears to be dictated by gene dosage. To investigate the mechanisms involved in HOX gene regulation in hematopoiesis and leukemogenesis by precise measurement of individual HOX genes, a small-array real-time HOX (SMART-HOX) quantitative polymerase chain reaction (PCR) platform was designed and validated. Application of SMART-HOX to 16 APL bone marrow samples revealed a global down-regulation of 26 HOX genes compared with normal controls. HOX gene expression was also altered during differentiation induced by ATRA in the PML-RARalpha(+) NB4 cell line. PML-RARalpha fusion proteins have been reported to act as part of a repressor complex during myeloid cell differentiation, and a model linking HOX gene expression to this PML-RARalpha repressor complex is now proposed.

Bone Marrow↗
PubMed2003

Genomic anatomy of the specific reciprocal translocation t(15;17) in acute promyelocytic leukemia.

The genomic breakpoints in the t(15;17)(q22;q21), associated with acute promyelocytic leukemia (APL), are known to occur within three different PML breakpoint cluster regions (bcr) on chromosome 15 and within RARA intron 2 on chromosome 17; however, the precise mechanism by which this translocation arises is unclear. To clarify this mechanism, we (i). assembled the sequence of RARA intron 2, (ii). amplified and sequenced the genomic PML-RARA junction sequences from 37 APL patients, and (iii). amplified and sequenced the reverse RARA-PML genomic fusion in 29 of these cases. Three significant breakpoint microclusters within RARA intron 2 were identified, suggesting that sequence-associated or structural factors play a role in the formation of the t(15;17). There was no evidence that the location of a breakpoint in PML had any relationship to the location of the corresponding breakpoint in RARA. Although some sequence motifs previously implicated in illegitimate recombinations were found in the microcluster regions, these associations were not significant. Comparison of forward and reverse genomic junctions revealed microhomologies, deletions, and/or duplications of either gene in all but one case, in which a complex rearrangement with inversion of the PML-derived sequence was found. These findings are consistent with the hypothesis that the t(15;17) occurs by nonhomologous recombination of DNA after processing of the double-strand breaks by a dysfunctional DNA damage-repair mechanism.

Chromosome Breakage↗
PubMed2003

The molecular pathogenesis of acute promyelocytic leukaemia: implications for the clinical management of the disease.

Acute promyelocytic leukaemia (APL) is characterised by chromosomal rearrangements of 17q21, leading to fusion of the gene encoding retinoic acid receptor alpha (RARalpha) to a number of alternative partner genes (X), the most frequent of which are PML (>95%), PLZF (0.8%) and NPM (0.5%). Over the last few years, it has been established that the X-RARalpha fusion proteins play a key role in the pathogenesis of APL through recruitment of co-repressors and the histone deacetylase (HDAC)-complex to repress genes implicated in myeloid differentiation. Paradoxically, the X-RARalpha fusion protein has the potential to mediate myeloid differentiation at pharmacological doses of its ligand (all trans-retinoic acid (ATRA)), which is dependent on the dissociation of the HDAC/co-repressor complex. Arsenic compounds have also been shown to be promising therapeutic agents, leading to differentiation and apoptosis of APL blasts. It is now apparent that the nature of the RARalpha-fusion partner is a critical determinant of response to ATRA and arsenic, underlining the importance of cytogenetic and molecular characterisation of patients with suspected APL to determine the most appropriate treatment approach. Standard protocols involving ATRA combined with anthracycline-based chemotherapy, lead to cure of approximately 70% patients with PML-RARalpha-associated APL. Patients at high risk of relapse can be identified by minimal residual disease monitoring. The challenge for future studies is to improve complete remission rates through reduction of induction deaths, particularly due to haemorrhage, identification of patients at high risk of relapse who would benefit from additional therapy, and identification of a favourable-risk group, for which treatment intensity could be reduced, thereby reducing risks of treatment toxicity and development of secondary leukaemia/myelodysplasia. With the advent of ATRA and arsenic, APL has already provided the first example of successful molecularly targeted therapy; it is hoped that with further understanding of the pathogenesis of the disease, the next decade will yield further improvements in the outlook for these patients.

Animals↗
PubMed2002

The T-lineage-affiliated CD2 gene lies within an open chromatin environment in acute promyelocytic leukemia cells.

The nature of hemopoietic progenitors subject to leukemic transformation in acute myeloid leukemia (AML) has not been clearly defined. To address this issue, we have used DNase I hypersensitivity assays to study the chromatin structure surrounding the T-lineage-affiliated CD2 gene in the acute promyelocytic subtype of AML (APL). Upstream and downstream flanking regions of CD2 were found to be hypersensitive to DNase I in primary APL blasts, with an identical pattern of hypersensitive sites to those detected in cells of T-lineage. All of the sites were confirmed to be inaccessible to DNase I in B-lineage leukemia cells. The demonstration of T-cell-associated chromatin features in primary APL blasts has implications for the origin of APL that may arise in more primitive progenitors than previously considered to be the case.

CD2 Antigens↗
PubMed2002

The significance of minimal residual disease in patients with t(15;17).

Acute promyelocytic leukaemia (APL) is characterized by the t(15;17)(q22;q21) leading to the formation of PML-RARalpha and RARalpha-PML fusion genes which provide suitable targets for the assessment of minimal residual disease (MRD). Studies have focused upon detection of PML-RARalpha because, although assays for RARalpha-PML transcripts are more sensitive, they are not applicable to 25% of cases. Among patients receiving standard therapy (ATRA and anthracycline-based chemotherapy), qualitative assays using a nested reverse transcriptase-polymerase chain reaction (RT-PCR), which typically achieve sensitivities of 1 in 10(4), have been found to provide independent prognostic information suitable for directing an approach to treatment. Detection of PML-RARalpha at the end of consolidation, or subsequent recurrence of PCR positivity, heralds relapse, which may, however, be averted by additional therapy leading to improvements in survival for this "high-risk" subgroup of patients. MRD analysis has also proved of value in predicting response to autologous transplant procedures undertaken in second complete remission and in directing the need for additional therapy in the post-transplantation setting. Overall, these studies undertaken within the context of a relatively homogeneous disease entity confirm that MRD monitoring provides independent prognostic information, serving as a valuable model for improving treatment strategy in other molecularly defined subsets of acute myeloid leukaemia (AML). Nevertheless, conventional nested RT-PCR assays fail to detect residual disease in a significant proportion of patients who ultimately relapse, which may be a reflection of RNA quality and/or assay sensitivity. Therefore, it is hoped that "real-time" quantitative RT-PCR technology (RQ-PCR) which permits quantification of fusion gene transcripts in relation to endogenous control genes will be even more predictive of outcome and achieve greater standardization of MRD detection in the context of large-scale clinical trials.

Chromosomes, Human, Pair 15↗