Investigation on the mechanisms underlying the chromosomal translocations in therapy-related acute myeloid leukemias

La leucemia acuta promielocitica (APL) è caratterizzata dalla traslocazione t(15;17) con la formazione del gene chimerico PML-RARa. La APL secondaria a trattamento con inibitori della topoisomerasi II (t-APL) rappresenta una nota complicanza del trattamento chemioterapico in pazienti affetti da cancro. Tuttavia negli ultimi anni sono stati descritti parecchi casi di t-APL in pazienti affetti da sclerosi multipla (SM) e trattati con mitoxantrone. In 12 pazienti affetti da MS con t-APL secondaria all’uso di mitoxantrone, l’analisi genomica ha mostrato un’alterata distribuzione all’interno dell’introne 6 del gene PML dei punti di rottura sul cromosoma 15 rispetto alle APL de novo (11/12, 92% vs 622/1022, 61%: p=0.035). Infatti, nonostante l’introne 6 abbia una dimenzione di circa 1kb, in 5 pazienti con t-apl secondaria a MS il punto di rottura nel cromosoma 15 cade in una regione di 8 paia di basi (hotspot) precedentemente desritta in pazienti affetti da t-APL secondaria a carcinoma della mammella (KM). Inoltre, all’interno del gene RARa , che si estende per circa 17 kb abbiamo identificato un altro punto di rottura comune a due pazienti con t-APL secondaria a trattamento con mitoxantrone per MS e KM. In 4 casi, l’utilizzo di saggi funzionali ci ha permesso di confermare sia in PML che in RARa la presenza di loci che sono risultati essere siti preferenziali di taglio da parte della topoisomerasi IIa in presenza di mitoxantrone. Questo studio conferma ulteriormente la presenza nei geni PML e RARa di porzioni di DNA particolarmente sensibili al danno indotto da mitoxantrone che possono spiegare la propensione a sviluppare questo la t-APL in pazienti trattati con questo chemioterapico. Abbiamo esteso l’analisi genomica anche in un caso con leucemia acuta mieloide con traslocazione t(16;21) (RUNX1-ETO”) secondaria a sclerosi multipla trattata con mitoxantrone. Ancora una volta abbiamo identificato una regione di 9 paia di basi (ATGCCCCAG) che mostrava una omologia del 90% con la sequenza ATGCCCTAG presente nell’introne 6 di PML evidenziata nei pazienti con t-APL secondarie a trattamento con mitoxantrone. In questo contesto è da notare che l’Accademia Americana di Neurologia nel 2000 ha approvato il mitoxantrone per il trattamento della sclerosi multipla. Following the response to post-marketing findings such as potential risk of therapy related leukemia, decreased systolic function and heart failure, the US Food and Drug administration (FDA) has added a “black box” warning to the prescribing information for the mitoxantrone.Therapy-related acute promyelocytic leukemia (t-APL) with the t(15;17) translocation is a well-recognized complication of cancer treatment with agents targeting topoisomerase II. However, cases are emerging following mitoxantrone therapy for multiple sclerosis (MS). Analysis of 12 cases of mitoxantrone-related t-APL in MS patients revealed an altered distribution of chromosome 15 breakpoints compared to de novo APL, biased towards disruption within PML intron 6 (11/12, 92% vs 622/1022, 61%: p=0.035). Despite this intron spanning ~1kb, the breakpoint in five mitoxantrone-treated patients fell within an 8bp region (1482-9) corresponding to the “hotspot” previously reported in t-APL complicating mitoxantrone-containing breast cancer therapy. Another shared breakpoint was identified within the ~17kb RARA intron 2 involving two t-APL cases arising after mitoxantrone treatment for MS and breast cancer, respectively. Analysis of PML and RARA genomic breakpoints in functional assays in 4 cases, including the shared RARA intron 2 breakpoint at 14446-49, confirmed each to be preferential sites of topoisomerase IIa-mediated DNA cleavage in the presence of mitoxantrone. This study further supports the presence of preferential sites of DNA damage induced by mitoxantrone in PML and RARA genes that may underlie the propensity to develop this particular subtype of leukemia following exposure to this agent. On extending our genomic analysis on therapy-related acute myeloid leukemia associated with t(16;21) (RUNX1-ETO2) arising after treatment of multiple sclerosis with mitoxantrone t-AML. We identified that genomic breakpoint region of RUNX1 contained a ATGCCCCAG nucleotide sequence showing ~90% homology to a “hotspot” DNA region ATGCCCTAG present in intron 6 of PML which was identified in therapy-related acute promyelocytic leukemia cases arising following treatment with mitoxantrone. Of note, in year 2000 Report of the Therapeutics and Technology Assessment Subcommittee of the American Academy of Neurology had approved mitoxantrone for progressive multiple sclerosis. Following the response to post-marketing findings such as potential risk of therapy related leukemia, decreased systolic function and heart failure, the US Food and Drug administration (FDA) has added a “black box” warning to the prescribing information for the mitoxantrone

Investigation on the mechanisms underlying the chromosomal translocations in therapy-related acute myeloid leukemias

La leucemia acuta promielocitica (APL) è caratterizzata dalla traslocazione t(15;17) con la formazione del gene chimerico PML-RARa. La APL secondaria a trattamento con inibitori della topoisomerasi II (t-APL) rappresenta una nota complicanza del trattamento chemioterapico in pazienti affetti da cancro. Tuttavia negli ultimi anni sono stati descritti parecchi casi di t-APL in pazienti affetti da sclerosi multipla (SM) e trattati con mitoxantrone. In 12 pazienti affetti da MS con t-APL secondaria all’uso di mitoxantrone, l’analisi genomica ha mostrato un’alterata distribuzione all’interno dell’introne 6 del gene PML dei punti di rottura sul cromosoma 15 rispetto alle APL de novo (11/12, 92% vs 622/1022, 61%: p=0.035). Infatti, nonostante l’introne 6 abbia una dimenzione di circa 1kb, in 5 pazienti con t-apl secondaria a MS il punto di rottura nel cromosoma 15 cade in una regione di 8 paia di basi (hotspot) precedentemente desritta in pazienti affetti da t-APL secondaria a carcinoma della mammella (KM). Inoltre, all’interno del gene RARa , che si estende per circa 17 kb abbiamo identificato un altro punto di rottura comune a due pazienti con t-APL secondaria a trattamento con mitoxantrone per MS e KM. In 4 casi, l’utilizzo di saggi funzionali ci ha permesso di confermare sia in PML che in RARa la presenza di loci che sono risultati essere siti preferenziali di taglio da parte della topoisomerasi IIa in presenza di mitoxantrone. Questo studio conferma ulteriormente la presenza nei geni PML e RARa di porzioni di DNA particolarmente sensibili al danno indotto da mitoxantrone che possono spiegare la propensione a sviluppare questo la t-APL in pazienti trattati con questo chemioterapico. Abbiamo esteso l’analisi genomica anche in un caso con leucemia acuta mieloide con traslocazione t(16;21) (RUNX1-ETO”) secondaria a sclerosi multipla trattata con mitoxantrone. Ancora una volta abbiamo identificato una regione di 9 paia di basi (ATGCCCCAG) che mostrava una omologia del 90% con la sequenza ATGCCCTAG presente nell’introne 6 di PML evidenziata nei pazienti con t-APL secondarie a trattamento con mitoxantrone. In questo contesto è da notare che l’Accademia Americana di Neurologia nel 2000 ha approvato il mitoxantrone per il trattamento della sclerosi multipla. Following the response to post-marketing findings such as potential risk of therapy related leukemia, decreased systolic function and heart failure, the US Food and Drug administration (FDA) has added a “black box” warning to the prescribing information for the mitoxantrone.Therapy-related acute promyelocytic leukemia (t-APL) with the t(15;17) translocation is a well-recognized complication of cancer treatment with agents targeting topoisomerase II. However, cases are emerging following mitoxantrone therapy for multiple sclerosis (MS). Analysis of 12 cases of mitoxantrone-related t-APL in MS patients revealed an altered distribution of chromosome 15 breakpoints compared to de novo APL, biased towards disruption within PML intron 6 (11/12, 92% vs 622/1022, 61%: p=0.035). Despite this intron spanning ~1kb, the breakpoint in five mitoxantrone-treated patients fell within an 8bp region (1482-9) corresponding to the “hotspot” previously reported in t-APL complicating mitoxantrone-containing breast cancer therapy. Another shared breakpoint was identified within the ~17kb RARA intron 2 involving two t-APL cases arising after mitoxantrone treatment for MS and breast cancer, respectively. Analysis of PML and RARA genomic breakpoints in functional assays in 4 cases, including the shared RARA intron 2 breakpoint at 14446-49, confirmed each to be preferential sites of topoisomerase IIa-mediated DNA cleavage in the presence of mitoxantrone. This study further supports the presence of preferential sites of DNA damage induced by mitoxantrone in PML and RARA genes that may underlie the propensity to develop this particular subtype of leukemia following exposure to this agent. On extending our genomic analysis on therapy-related acute myeloid leukemia associated with t(16;21) (RUNX1-ETO2) arising after treatment of multiple sclerosis with mitoxantrone t-AML. We identified that genomic breakpoint region of RUNX1 contained a ATGCCCCAG nucleotide sequence showing ~90% homology to a “hotspot” DNA region ATGCCCTAG present in intron 6 of PML which was identified in therapy-related acute promyelocytic leukemia cases arising following treatment with mitoxantrone. Of note, in year 2000 Report of the Therapeutics and Technology Assessment Subcommittee of the American Academy of Neurology had approved mitoxantrone for progressive multiple sclerosis. Following the response to post-marketing findings such as potential risk of therapy related leukemia, decreased systolic function and heart failure, the US Food and Drug administration (FDA) has added a “black box” warning to the prescribing information for the mitoxantrone

Targeting transcriptional kinase of CDK7 halts proliferation of multiple myeloma cells by modulating the function of canonical NF-kB pathway and cell cycle regulatory proteins

Multiple myeloma (MM) is an incurable plasma cell neoplasm. Despite several effective frontline therapeutic regimens, including Bortezomib (BTZ), relapse is almost inevitable; therefore, better therapeutic modalities to improve the outcomes are needed. Cyclin-dependent kinases (CDKs) are an essential constituent of the cellular transcriptional machinery and tumors including MM are critically dependent on transcription to maintain their oncogenic state. In the present study, we explored the efficacy of THZ1, a covalent CDK7 inhibitor in MM treatment using Bortezomib resistant (H929BTZR) cells and zebrafish xenografts. THZ1 showed anti-myeloma activity in the models of MM but had no effect on healthy CD34+ cells. THZ1 suppresses phosphorylation of carboxy-terminal domain of RNA polymerase II and downregulates the transcription of BCL2 family of proteins both in H929BTZS and H929BTZR cells leading to G1/S arrest and apoptosis. THZ1 mediates inhibition of bone marrow stromal cells-induced proliferation and activation of NF-kB signaling. The data derived from zebrafish xenografts of MM demonstrate that THZ1 combined with BTZ synergistically reduces tumor growth in zebrafish embryos. Collectively, our results reveal that THZ1 alone as well as in combination with BTZ has effective anti-myeloma activity

Over-representation of bcr3 subtype of PML/RAR α fusion gene in APL in Indian patients

Thirty six patients with acute promyelocytic leukemia were studied by reverse transcriptase-polymerase chain reaction (RT-PCR) and real-time PCR. There was concordance between the results achieved by both the methods except in one case, which was negative by RT-PCR but positive by real-time PCR. The prevalence of bcr3 (short isoform) was found to be significantly higher than that of bcr1 (long isoform ) (64 vs. 36%, P=0.03). No correlation was found between age, sex, and white blood cell (WBC) count at diagnosis. Molecular remission was achieved in 66.6% of patients with bcr3 isoform. Median WBC count at presentation was found to be higher than that in the West

Presenting features and treatment outcome of acute promyelocytic leukemia arising after multiple sclerosis

We report the clinical features and treatment outcome of 33 patients with multiple sclerosis who developed acute promyelocytic leukemia. Thirty patients were previously exposed to mitoxantrone. The median latency period between treatment initiation and acute promyelocytic leukemia diagnosis was 32 months. The PML-RARA bcr1 iso-form was identified in 87% of cases. Twenty-nine (90%) patients achieved hematologic remission after all-trans retinoic acid and chemotherapy (n=31) or arsenic trioxide and all-trans retinoic acid. Consolidation included modified chemotherapy or arsenic trioxide. At a median follow up of 26 months, 23 patients are in complete remission, 4 relapsed and one developed secondary leukemia. The 5-year cumulative incidence of relapse and overall survival were 23% and 68%, respectively. Although treatment heterogeneity and suboptimal post-remission therapy must be taken into account, overall results and development of secondary leukemia in one patient suggest that effective and less toxic agents like arsenic trioxide warrants further investigation in this context

Identification of a Potential "Hotspot" DNA Region in the RUNXI Gene Targeted by Mitoxantrone in Therapy-Related Acute Myeloid Leukemia with t(16;21) Translocation

Genes Chromosomes Cancer. 2009 Mar;48(3):213-21

Analysis of t(15;17) Chromosomal Breakpoint Sequences in Therapy-Related Versus De Novo Acute Promyelocytic Leukemia: Association of DNA Breaks with Specific DNA Motifs at PML and RARA Loci

We compared genomic breakpoints at the PML and RARA loci in 23 patients with therapy-related acute promyelocytic leukemia (t-APL) and 25 de novo APL cases.Eighteen of 23 t-APL cases received the topoisomerase II poison mitoxantrone for their primary disorder. DNA breaks were clustered in a previously reported 8 bp "hot spot" region of PML corresponding to a preferred site of mitoxantrone-induced DNA topoisomerase II-mediated cleavage in 39% of t-APL occurring in patients exposed to this agent and in none of the cases arising de novo (P = 0.007). As to RARA breakpoints, clustering in a 3' region of intron 2 (region B) was found in 65% of t-APL and 28% of de novo APL patients, respectively. Scan statistics revealed significant clustering of RARA breakpoints in region B in t-APL cases (P = 0.001) as compared to de novo APL (P = 1). Furthermore, approximately 300 bp downstream of RARA region B contained a sequence highly homologous to a topoisomerase II consensus sequence. Biased distribution of DNA breakpoints at both PML and RARA loci suggest the existence of different pathogenetic mechanisms in t-APL as compared with de novo APL