Double inv(3)(q21q26.2) in acute myeloid leukemia is resulted from an acquired copy neutral loss of heterozygosity of chromosome 3q and associated with disease progression

AbstractLineage-defining transcription factors (TFs) are compelling targets for leukemia therapy, yet they are among the most challenging proteins to modulate directly with small molecules. We previously used CRISPR screening to identify a Salt-Inducible Kinase 3 (SIK3) requirement for the growth of acute myeloid leukemia (AML) cell lines that overexpress the lineage TF MEF2C. In this context, SIK3 maintains MEF2C function by directly phosphorylating histone deacetylase 4 (HDAC4), a repressive cofactor of MEF2C. Here, we evaluated whether inhibition of SIK3 with the tool compound YKL-05-099 can suppress MEF2C function and attenuate disease progression in animal models of AML. Genetic targeting of SIK3 or MEF2C selectively suppressed the growth of transformed hematopoietic cells underin vitroandin vivoconditions. Similar phenotypes were obtained when exposing cells to YKL-05-099, which caused cell cycle arrest and apoptosis in MEF2C-expressing AML cell lines. An epigenomic analysis revealed that YKL-05-099 rapidly suppressed MEF2C function by altering the phosphorylation state and nuclear localization of HDAC4. Using a gatekeeper allele ofSIK3, we found that the anti-proliferative effects of YKL-05-099 occurred through on-target inhibition of SIK3 kinase activity. Based on these findings, we treated two different mouse models of MLL-AF9 AML with YKL-05-099, which attenuated disease progressionin vivoand extended animal survival at well-tolerated doses. These findings validate SIK3 as a therapeutic target in MEF2C-positive AML and provide a rationale for developing drug-like inhibitors of SIK3 for definitive pre-clinical investigation and for studies in human patients with leukemia.Key PointsAML cells are uniquely sensitive to genetic or chemical inhibition of Salt-Inducible Kinase 3in vitroandin vivo.A SIK inhibitor YKL-05-099 suppresses MEF2C function and AMLin vivo.

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Update of a Phase I Study of Sorafenib in Patients with Refractory/Relapsed Acute Myeloid Leukemia or High-Risk Myelodysplastic Syndrome.

Blood ◽

10.1182/blood.v110.11.893.893 ◽

2007 ◽

Vol 110 (11) ◽

pp. 893-893 ◽

Cited By ~ 7

Author(s):

John Delmonte ◽

Hagop M. Kantarjian ◽

Michael Andreeff ◽

Stefan Faderl ◽

John J. Wright ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

High Risk ◽

Myelodysplastic Syndrome ◽

Phase I ◽

Disease Progression ◽

Myeloid Leukemia ◽

Maximal Response ◽

Daily Dose ◽

Grade 3 ◽

Acute Myeloid

The critical importance of the Ras, VEGF, and FLT3 pathways in the pathogenesis of myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) has been well established. FLT3 abnormalities, internal tandem duplication (ITD) and point mutations, occur in about 30% of pts with AML and the FLT3-ITD mutation independently confers poor prognosis. Sorafenib is an oral multikinase inhibitor targeting the above pathways and is highly potent against FLT3-ITD mutants (IC50 1–3 nM) (ASH abstract, 2006). We are conducting a phase I trial to evaluate the safety and efficacy of two different schedules of sorafenib. To date, 21 patients (pts) with refractory/relapsed AML (n=20) and high risk MDS (n=1) have been enrolled. Pts were randomized to sorafenib for 5 days per week for 21 days (arm A; n=11) or for 14 days every 21 days (arm B; n=10). In both arms the starting dose level (DL) is 200 mg twice daily. Successive dose levels are 600, 800, and 1200 mg daily in a standard 3+3 design. Peripheral blood (PB) and bone marrow (BM) samples were obtained for evaluation of FLT3 status and phosphorylated and total FLT3 and ERK expression. Median age is 62 years (range, 33–82), number of prior therapies 2 (range, 1–5), time from diagnosis to sorafenib treatment 9 months (range, 2–46), and median duration on study was 1.2 months (range, 0.1–3.4). Twenty pts are evaluable. 9/20 (45%) pts received ≤ 1 cycle of sorafenib because of disease progression (n=6), self-discontinuation (n=2), or no benefit (n=1), of whom 5 (56%) were FLT3-ITD negative, 3 (33%) were FLT3-ITD positive, and 1 (11%) was not tested. In contrast, 11/20 (55%) pts received > 1 cycle of sorafenib, of whom 8 (73%) were FLT3-ITD positive and 3 (27%) were FLT3-ITD negative; reasons for discontinuation were disease progression (n=5), self-discontinuation (n=2), stem cell transplant (n=2), or no benefit (n=2). Sorafenib has been well tolerated with 1 pt achieving a DLT of grade 3 hyperbilirubinemia at the 800 mg daily dose in arm B, but the MTD has not been reached; this cohort has been expanded. The only other grade 3 toxicity has been pleural effusion at the 600 mg daily dose in arm A, not considered a DLT because it occurred during cycle 2. A ≥ 50% reduction in PB or BM blasts was obtained in 11/20 (55%) pts. 9/11 (82%) pts harbored the FLT3-ITD mutation and had a median duration of response of 42 days (range, 15–87). In these 9 pts, the median PB absolute blast count at baseline and after maximal response to sorafenib was 10.3 (range, 0.2–18.7) and 0 (range, 0–1)(p=0.008). Median BM blast percentage at baseline and after maximal response to sorafenib was 72% (range, 14–96) and 42% (range, 12–58) (p=0.002), with 1 pt achieving a morphologic complete remission in the BM. Serial determinations of phosphorylation status following sorafenib (at 0, 2, 24,120 hours) in pts with the FLT3-ITD mutation demonstrated inhibition of phospho-FLT3 in 3/3 and phospho-ERK in 5/5 pts. In conclusion, sorafenib administration is safe in AML and appears to preferentially target the FLT3-ITD mutation. This study continues to accrue pts to define the MTD and it will be followed by combination studies of standard chemotherapy with sorafenib, with an emphasis on targeting pts with AML expressing the FLT3-ITD mutation.

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Functional and Phenotypic Characterization Of Acute Myeloid Leukemia By Analysis Of Diagnostic/Relapse Paired Samples

Blood ◽

10.1182/blood.v122.21.2595.2595 ◽

2013 ◽

Vol 122 (21) ◽

pp. 2595-2595

Author(s):

Jenny M. Ho ◽

Liran I. Shlush ◽

Amanda Mitchell ◽

Rene Marke ◽

Jessica McLeod ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Stem Cell ◽

Disease Progression ◽

Myeloid Leukemia ◽

Phenotypic Characterization ◽

Stem Cell Markers ◽

Surface Markers ◽

Paired Samples ◽

Flow Cytometric ◽

Acute Myeloid

Abstract Treatment of relapsed acute myeloid leukemia (AML) results in lower complete remission rates compared to treatment of AML at diagnosis. Immunophenotypic changes in leukemic blasts are common from diagnosis to relapse (Baer et al. Blood. 2001), suggesting that the underlying biology of AML changes with disease progression. A better understanding of the biologic properties of AML cells at different stages of disease will facilitate the development of biomarker tools and more effective therapies. We therefore studied paired diagnostic/relapse samples obtained from AML patients. Cells from 11 paired samples were transplanted into immune deficient mice (NOD.SCID IL2Rg null) over a range of cell doses. In 10 of 11 patients, a leukemic graft could be generated after transplantation of lower cell doses from the relapse sample compared to the paired diagnostic sample. By limiting dilution analysis, leukemia stem cell (LSC) frequency was higher in relapse samples (1 in 5.8×102 to 1 in 2.4×106, median 1 in 2.0×103) compared to diagnostic samples (1 in 5.0×103 to 1 in 6.1×106, median 1 in 5.5×104); the fold increase in LSC frequency ranged from 2.2 to 745 (median 8.6). Multiparameter flow cytometric analysis carried out on 13 paired diagnostic/relapse samples demonstrated an increase in 2 known stem cell markers, CD34 and CD117, from diagnosis to relapse: CD34 was gained or increased at relapse in 7/13 (54%) of paired samples, while CD117 was gained or increased at relapse in 9/13 (69%) of paired samples. We plan to take a comprehensive approach to examine the surface marker expression of paired diagnostic/relapse samples by a high throughput flow cytometric screen (HTS) of both bulk and LSC-containing AML populations to identify markers that are altered at relapse. As a first step, we have performed HTS of 373 surface markers on 10 AML patient samples, including a relapse sample from a diagnostic/relapse pair. A significant proportion of markers (155/373, 42%) were expressed on less that 5% of cells in all 10 AML samples analyzed. We will therefore focus on the remaining markers in our HTS analysis of paired samples. Surface markers that are differentially expressed from diagnosis to relapse will be further characterized in order to gain insight into disease progression and identify potential therapeutic targets. Disclosures: No relevant conflicts of interest to declare.

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Safety, Pharmacokinetics, and Efficacy of BP-100-1.01 (Liposomal Grb-2 Antisense Oligonucleotide) in Patients with Refractory or Relapsed Acute Myeloid Leukemia (AML), Philadelphia Chromosome Positive Chronic Myelogenous Leukemia (CML), Acute Lymphoblastic Leukemia (ALL), and Myelodysplastic Syndrome (MDS)

Blood ◽

10.1182/blood.v126.23.3801.3801 ◽

2015 ◽

Vol 126 (23) ◽

pp. 3801-3801 ◽

Cited By ~ 3

Author(s):

Maro Ohanian ◽

Hagop M. Kantarjian ◽

Farhad Ravandi ◽

Gautam Borthakur ◽

Guillermo Garcia-Manero ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Acute Lymphoblastic Leukemia ◽

Myelodysplastic Syndrome ◽

Disease Progression ◽

Myeloid Leukemia ◽

Research Funding ◽

Lymphoblastic Leukemia ◽

Myelogenous Leukemia ◽

Average Decrease ◽

Acute Myeloid

Abstract Background: Essential to cancer cell signaling, the growth receptor bound protein-2 (Grb-2) is evolutionarily conserved and utilized by oncogenic tyrosine kinases including Bcr-Abl to activate Ras, ERK, and AKT. BP-100-1.01is a neutrally-charged, liposome-incorporated antisense designed to inhibit Grb-2 expression. Aim: To define the safety, maximum tolerated dose (MTD), optimal biologically active dose, pharmacokinetics and anti-leukemia activity of BP-100-1.01 in patients (pts) with hematologic malignancies. Methods: This is a standard 3+3 phase I dose-finding study in pts with relapsed or refractory acute myeloid leukemia (AML), chronic myeloid leukemia in blast phase (CML-BP), acute lymphoblastic leukemia (ALL) and myelodysplastic syndrome (MDS). The starting dose was 5 mg/m2 twice weekly, IV over 2-3 hours for 28 days. Dose escalation proceeded through 5, 10, 20, 40, 60, and 90 mg/m2.Uponcompletion of single agent phase 1, combination of cytarabine 20 mg SubQ BID x 10 days + 60 mg/m2 of BP-100-1.01 was studied (Cohort 1B). Flow cytometric analysis was performed on peripheral blood samples from cohorts 3, 4, 5, 6 and 1B collected at baseline, on day 15 and at end-of-treatment (EOT). Fluorescent-labeled antibodies specific for Grb-2 or phosphorylated Erk (pErk) were utilized to determine Grb-2 protein levels and pErk levels in CD33-expressing cells. Results: A total of33 pts were included (13 in Cohort 1, 6 in Cohort 2, 3 each in Cohorts 3, 4, 5, and 4 in cohort 6). One patient has been treated in cohort 1B. The median age was 64 yrs (range, 32-89) and diagnoses were AML (n=24), CML-BP (n=5) and MDS (n=4). The median number of prior therapies was 4 (range, 1- 8). Of 33 pts, 21 were evaluable and 11 failed completion of a full 28-Day cycle due to disease progression (with no toxicity) and were replaced, per protocol. Only one pt (treated at 5 mg/m2) experienced dose limiting toxicity (DLT), grade 3 mucositis and hand-foot syndrome, while receiving concurrent hydroxyurea for proliferative CML-BP. The patient had a previous history of hydroxyurea-induced mucositis. Being the first patient to receive BP-100-1.01, these toxicities were considered possibly related to BP-100-1.01. The cohort was expanded to a total of 6 pts. No other DLTs have been noted in any pt. Among 21 evaluable pts, 11 experienced at least a 50% reduction in peripheral or bone marrow blasts from baseline. Additionally 2 pts with improvement in leukemia cutis lesions received 1 cycle each. Furthermore, 6 pts demonstrated transient improvement (n=3) and/or stable disease (n=3). Among the 21 evaluable pts, a median of 1 cycle was administered (1-5): Four pts received 2 cycles, 3 pts received 5 cycles, and all others received 1 cycle. Notably one pt (treated at 5 mg/m2)with CML-BP showed a significant reduction in blasts from 81% to 5%. Due to leptomeningeal disease progression therapy was discontinued before a full cycle. The 1st patient treated in cohort 1B achieved CR after 1 cycle. The patient did not experience any DLTs, but came off study due to failure to thrive in the context of dementia. The levels of Grb-2 and pErk proteins were indicated by their respective median fluorescent signals and are shown in the table. Median fluorescent signals of Grb-2 and pErk on days 15 and EOT were compared to baseline. On day 15 Grb-2 levels decreased by >25% in 7 out of 12 samples tested, and pErk levels by >25% in 6 out of 12 samples. The average decrease in Grb-2 levels was 61% (range: 47 to 85%) and in pErk levels 52% (range: 28 to 82%). On the last measured sample (EOT or day 22), BP-100-1.01 decreased >25% Grb-2 levels in 11 out of 13 samples, and >25% pErk levels in 7 out of 13 samples. The average decrease in Grb-2 levels was 49% (range: 28 to 91%) and in pErk levels was 52% (range: 27 to 91%). Table 1. Patient Number Grb-2 decrease (Day 15) pErk decrease (Day 15) Grb-2 decrease (Day 22 or EOT) pErk decrease (Day 22 or EOT) 022 0 0 57 0 023 0 3 28 45 024 56 28 47 35 025 63 82 54 91 026 47 0 0 0 027 NS NS 34 27 028 0 0 30 54 029 57 51 65a 0a 030 54 55 43 47 031 0 0 0 0 032 85 54 91 63 033 6 13 53 2 034 63 42 40 0 NS = no sample collected aFewer cells were used in the analysis of this sample than other samples, because this sample had less cells than other samples Conclusions: BP-100-1.01, at dose range 5 mg/m2 to 90 mg/m2 is well tolerated with no MTD yet identified. There is suggestion of Grb-2 target protein down-regulation, and possible anti-leukemia activity. Disclosures Konopleva: Novartis: Research Funding; AbbVie: Research Funding; Stemline: Research Funding; Calithera: Research Funding; Threshold: Research Funding. Tari:Bopath Holdings: Employment. Cortes:BerGenBio AS: Research Funding; Teva: Research Funding; Novartis: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Ariad: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Astellas: Consultancy, Research Funding; Ambit: Consultancy, Research Funding; Arog: Research Funding; Celator: Research Funding; Jenssen: Consultancy.

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