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  • 1
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    From drug response profiling to target addiction scoring in cancer cell models
    The Company of Biologists ; 2015
    In:  Disease Models & Mechanisms Vol. 8, No. 10 ( 2015-10-01), p. 1255-1264
    Details
    In: Disease Models & Mechanisms, The Company of Biologists, Vol. 8, No. 10 ( 2015-10-01), p. 1255-1264
    Abstract: Deconvoluting the molecular target signals behind observed drug response phenotypes is an important part of phenotype-based drug discovery and repurposing efforts. We demonstrate here how our network-based deconvolution approach, named target addiction score (TAS), provides insights into the functional importance of druggable protein targets in cell-based drug sensitivity testing experiments. Using cancer cell line profiling data sets, we constructed a functional classification across 107 cancer cell models, based on their common and unique target addiction signatures. The pan-cancer addiction correlations could not be explained by the tissue of origin, and only correlated in part with molecular and genomic signatures of the heterogeneous cancer cells. The TAS-based cancer cell classification was also shown to be robust to drug response data resampling, as well as predictive of the transcriptomic patterns in an independent set of cancer cells that shared similar addiction signatures with the 107 cancers. The critical protein targets identified by the integrated approach were also shown to have clinically relevant mutation frequencies in patients with various cancer subtypes, including not only well-established pan-cancer genes, such as PTEN tumor suppressor, but also a number of targets that are less frequently mutated in specific cancer types, including ABL1 oncoprotein in acute myeloid leukemia. An application to leukemia patient primary cell models demonstrated how the target deconvolution approach offers functional insights into patient-specific addiction patterns, such as those indicative of their receptor-type tyrosine-protein kinase FLT3 internal tandem duplication (FLT3-ITD) status and co-addiction partners, which may lead to clinically actionable, personalized drug treatment developments. To promote its application to the future drug testing studies, we have made available an open-source implementation of the TAS calculation in the form of a stand-alone R package.
    Type of Medium: Online Resource
    ISSN: 1754-8411 , 1754-8403
    URL: Article
    DOI: 10.1242/dmm.021105
    Language: English
    Publisher: The Company of Biologists
    Publication Date: 2015
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  • 2
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    Abstract 676: Axitinib targets gatekeeper-mutant BCR-ABL1(T315I)-driven leukemia in a distinct and selective fashion
    American Association for Cancer Research (AACR) ; 2015
    In:  Cancer Research Vol. 75, No. 15_Supplement ( 2015-08-01), p. 676-676
    Details
    In: Cancer Research, American Association for Cancer Research (AACR), Vol. 75, No. 15_Supplement ( 2015-08-01), p. 676-676
    Abstract: The T315I gatekeeper mutation confers resistance to majority of approved ABL1 inhibitors, with only ponatinib demonstrating efficacy in BCR-ABL1(T315I)-driven disease. However, vascular adverse events and selection of resistant compound mutations limit its clinical utility. Hence, there is an unmet need for novel therapies for patients with gatekeeper-mutated Ph+ leukemia. In this study we integrated comprehensive drug sensitivity testing with structural analysis to characterize the tyrosine kinase inhibitor axitinib as a putative novel therapy for BCR-ABL1(T315I)-driven leukemias. To address this we profiled BCR-ABL1(T315I)-driven CML/Ph+ ALL patient samples against 300 anti-cancer compounds (approved and investigational drugs). Ex vivo drug sensitivity testing of primary cells derived from a Ph+ ALL patient revealed a marked and cancer-selective response to the VEGFR inhibitor axitinib. Strikingly, axitinib exhibited higher sensitivity in T315I positive Ph+ patient samples in comparison to T315I negative CML and ALL patient samples. In line with the ex vivo drug response data, axitinib inhibited the kinase activity of ABL1(T315I) with similar potency as its primary target VEGFR2, while the potency to non-mutated ABL1 was 30-fold lower. Analogously, in engineered Ba/F3 cells, axitinib showed a 10-fold higher inhibition of T315I than non-mutated ABL1-driven autophosphorylation and cell growth. To better understand the molecular mechanisms of the BCR-ABL1(T315I) selectivity, we solved the crystal structure of axitinib in complex with ABL1(T315). The structure revealed that axitinib bound to a mutation induced active conformation of ABL1(T315I), different than the binding mode in non-mutated ABL1, likely explaining the increased potency towards ABL1(T315I). Moreover, axitinib occupied a distinct binding space than all approved ABL1 inhibitors, signifying that axitinib will have a unique mutation vulnerability profile. Compassionate two week treatment of a CML patient harboring the T315I mutation with axitinib, resulted in a 5-fold reduction of T315I transcript levels in the bone marrow, further suggesting that axitinib can produce specific and effective responses in patients with BCR-ABL1(T315I)-driven leukemia. In conclusion, we demonstrate that axitinib potently inhibits BCR-ABL1(T315I) via a gatekeeper mutant-selective mechanism. Since axitinib is in clinical use for treatment of refractory renal cell carcinoma with a manageable safety profile, our data provide a sound basis for readily repurposing axitinib for BCR-ABL1(T315I)-driven leukemia. Finally, the distinct mechanism of inhibition by axitinib serves as an exemplar for development of even more effective gatekeeper-mutant selective inhibitors targeting ABL1 as well as other clinically important kinases, such as EGFR and KIT. Citation Format: Tea Pemovska, Eric Johnson, Mika Kontro, Gretchen A. Repasky, Jeffrey Chen, Peter Wells, Ciarán N. Cronin, Michele McTigue, Olli Kallioniemi, Kimmo Porkka, Brion W. Murray, Krister Wennerberg. Axitinib targets gatekeeper-mutant BCR-ABL1(T315I)-driven leukemia in a distinct and selective fashion. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 676. doi:10.1158/1538-7445.AM2015-676
    Type of Medium: Online Resource
    ISSN: 0008-5472 , 1538-7445
    URL: Article
    DOI: 10.1158/1538-7445.AM2015-676
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    Language: English
    Publisher: American Association for Cancer Research (AACR)
    Publication Date: 2015
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  • 3
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    8-Chloro-Adenosine Inhibits Molecular Poor-Risk Acute Myeloid Leukemia (AML) and Leukemic Stem Cells (LSC) Growth Via Novel RNA- and ATP-Directed Mechanisms: A Novel Therapeutic Approach for AML
    American Society of Hematology ; 2015
    In:  Blood Vol. 126, No. 23 ( 2015-12-03), p. 792-792
    Details
    In: Blood, American Society of Hematology, Vol. 126, No. 23 ( 2015-12-03), p. 792-792
    Abstract: Only a minority of patients with AML are cured with currently available chemotherapy regimens. Thus, new drugs with novel mechanisms of action are urgently needed. In previous proof-of-concept studies, the halogenated ATP analog 8-chloro-adenosine (8-Cl-Ado) has shown activity against a variety of solid tumors and hematologic malignancies in vitro, favorable pharmacokinetic and pharmacodynamic profiles in preclinical animal studies and feasibility in a first-in-man phase I clinical trial in chronic lymphocytic leukemia (CLL), where plasma concentrations up to 1 uM could be achieved; since maximum tolerated dose was not yet reached in the CLL clinical trial, higher plasma concentration of 8-Cl-Ado can be anticipated. However, little is known regarding the activity of this compound in AML. In contrast to other nucleoside analogs used for treatment of AML (e.g., cytarabine, azanucleosides), 8-Cl-Ado is metabolized by adenosine kinase and its triphosphate derivative is incorporated into RNA and significantly inhibited RNA synthesis in AML cells (KG1a and MV-4-11) in a dose dependent manner (300 nM to 1 uM) after 24 h exposure as compared to vehicle-treated controls (p 〈 0.05 at 300 nM 8-Cl-Ado). Similarly, 20-80% inhibition of total RNA synthesis was observed in AML patient blasts, regardless of the harbored mutational or cytogenetic aberrations. This resulted in changes in expression levels of both coding and non-coding RNA (e.g. miR-126) in immunophenotypically distinct AML cell populations including leukemia-stem cell (LSC)-enriched CD34+CD38-fractions. In contrast, 24 h exposure to 8-Cl-Ado did not significantly inhibit DNA synthesis compared to vehicle at any concentration tested (p 〉 0.25 at all concentrations tested). 8-Cl-Ado was metabolized into its cytotoxic triphosphate, 8-Cl-ATP, which accumulated to intracellular concentrations of more than 600 uM after 12 h exposure to 10 uM 8-Cl-Ado. Accumulation of 8-Cl-ATP was associated with 〉 20% reduction of endogenous ATP compared to control-treated cells (p 〈 0.05). In regard to antileukemia activity, 8-Cl-Ado inhibited growth of AML cell lines (i.e., Molm-13, Molm-14, KG1a, MV-4-11, OCI-AML3) with IC50s ranging from 0.2 uM to 1.4 uM after 72 h of treatment. 8-Cl-Ado was also active against primary AML blasts including those harboring the poor-risk FLT3 -ITD mutation (IC50: 800 nM for FLT3-ITD-positive blasts). Furthermore, the effect of 8-Cl-Ado on tumor cells has been demonstrated to not be affected by p53 status. It is important to point out that loss of p53 and mutations in p53 are associated with drug resistance and adverse outcomes in AML. In an orthotopic mouse model where FLT3 -ITD-positive Molm-14 AML cells were xenografted, animals treated with 50 mg/kg/day through an implanted osmotic pump had 〉 70% reduction in tumor mass compared to vehicle-treated controls after 16 days of treatment (p 〈 0.05). Importantly, 24 h pre-treatment of human LSC-enriched CD34+CD38- blasts with 10 uM 8-Cl-Ado resulted in a significant inhibition of their colony forming ability compared to vehicle-treated controls ( 〉 50% reduction of colonies after 14 days, p 〈 0.05). To study the effect of 8-Cl-Ado on AML LSCs in vivo, sub-lethally irradiated Rag-2/gamma(c) double-knockout immunodeficient mice were i.v.-injected with ten million human LSC-enriched CD34+CD38- blasts pre-treated with 10 uM 8-Cl-Ado or vehicle control 24 h prior to i.v. injection. Significant longer survival was observed of mice engrafted with drug-treated cells compared to those engrafted with vehicle-treated controls (p 〈 0.0004). Taken together, 8-Cl-Ado is a promising agent with a unique RNA and ATP-targeting mechanism of action, excellent toxicity profile and encouraging preclinical anti-leukemia activity in AML. Based on these results, we have initiated a phase I/II clinical trial in patients with relapsed/refractory AML. Figure 1. Figure 1. Disclosures No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
    URL: Article
    DOI: 10.1182/blood.V126.23.792.792
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    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2015
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  • 4
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    Rationale for the combination of venetoclax and ibrutinib in T-prolymphocytic leukemia
    Ferrata Storti Foundation (Haematologica) ; 2021
    In:  Haematologica Vol. 106, No. 8 ( 2021-02-25), p. 2251-2256
    Details
    In: Haematologica, Ferrata Storti Foundation (Haematologica), Vol. 106, No. 8 ( 2021-02-25), p. 2251-2256
    Type of Medium: Online Resource
    ISSN: 1592-8721 , 0390-6078
    URL: Article
    DOI: 10.3324/haematol.2020.271304
    Language: Unknown
    Publisher: Ferrata Storti Foundation (Haematologica)
    Publication Date: 2021
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    detail.hit.zdb_id: 2030158-3
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  • 5
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    Axitinib effectively inhibits BCR-ABL1(T315I) with a distinct binding conformation
    Springer Science and Business Media LLC ; 2015
    In:  Nature Vol. 519, No. 7541 ( 2015-03-05), p. 102-105
    Details
    In: Nature, Springer Science and Business Media LLC, Vol. 519, No. 7541 ( 2015-03-05), p. 102-105
    Type of Medium: Online Resource
    ISSN: 0028-0836 , 1476-4687
    URL: Article
    DOI: 10.1038/nature14119
    RVK:
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    Language: English
    Publisher: Springer Science and Business Media LLC
    Publication Date: 2015
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    SSG: 11
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  • 6
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    Antileukemic Activity of 8-Chloro-Adenosine (8-Cl-Ado) Is Mediated By Mir-155 Degradation and ErbB3 Binding Protein (Ebp1)-Dependent p53 Activation: A Novel Therapeutic Approach for FLT3-ITD Acute Myeloid Leukemia (AML)
    American Society of Hematology ; 2018
    In:  Blood Vol. 132, No. Supplement 1 ( 2018-11-29), p. 3938-3938
    Details
    In: Blood, American Society of Hematology, Vol. 132, No. Supplement 1 ( 2018-11-29), p. 3938-3938
    Abstract: AML patients (pts) carrying the fms-related tyrosine kinase 3 (FLT3)-internal tandem duplication (ITD) have a poor prognosis. Combination of chemotherapy with tyrosine kinase inhibitors (TKIs) has improved survival of these pts, but a large proportion of them still die of their disease. Nucleoside analogs (NAs) are the backbone of several upfront and salvage chemotherapy regimens for AML, including FLT3-ITD. Although these agents have significant antileukemic activity, they are not effective in eradicating leukemia stem cells (LSCs), the likely reason for treatment failures in AML. Consequently, new strategies are needed to improve the outcome for this and other molecular subsets of AML pts. 8-Cl-Ado is a novel, ribose-containing, RNA-directed nucleoside analog which, different from other NAs, is incorporated into newly transcribed RNA rather than in DNA, causing inhibition of RNA transcription. Among the AML molecular subsets, we identified FLT3-ITD blasts as one of the most sensitive to 8-Cl-Ado; however the mechanism of this differential effect remains unknown. Ex-vivo treatment with 8-Cl-Ado induced dose-dependent growth inhibition and apoptosis in FLT3-ITD AML cell lines and primary blasts including the LSC-enriched CD34+CD38- immunophenotypic subpopulation, with IC50s ranging from 200 nM to 1400 nM and a negligible effect on normal hematopoietic stem cells. In an orthotopic murine model, mice xenografted with FLT3-ITD-positive MV4-11 cells and treated (upon engraftment) with 75 mg 8-Cl-Ado/kg/day through an implanted osmotic pump survived significantly longer than vehicle-treated controls (median survival 45 days vs. 27.3 days, p=0.002). MicroRNA-155 (miR-155) is the most over-expressed miRNA in FLT3-ITD and reportedly plays a key role in FLT3-ITD blast hyper-proliferation [PMID 20656931, PMID 25971362]. Thus, silencing of miR-155 has been proposed as a novel therapeutic approach for FLT3-ITD AML [PMID 25971362] . As 8-Cl-Ado is incorporated mainly into RNA, we reasoned that it could also be incorporated into miR-155 (and other miRNAs). Consistent with our hypothesis, we detected co-localization of 8-Cl-Ado and miR-155 in FLT3-ITD primary blast cells and MV4-11, using fluorescence-labeled 8-Cl-Ado (8-Cl-Ado-FAM) and miR-155 staining (SmartFlare probes), suggesting that 8-Cl-Ado interacts directly with miR-155. Using quantitative RT-PCR we demonstrated ~50% miR-155 down-regulation in 8-Cl-Ado-FAM or 8-Cl-Ado-treated MV4-11 cells and FLT3-ITD primary blast cells, compared to vehicle-treated controls. On a molecular level, 8-Cl-Ado-dependent miR-155 down-regulation was accompanied by up-regulation of SHIP1, a bona fide miR-155 target phosphatase that decreased p-AKT levels thereby negatively regulating FLT3-ITD-dependent AKT signaling required for leukemia cell growth and survival. This effect also disrupted the interaction of AKT and ErbB3 binding protein (Ebp1), a highly expressed protein that regulates p53 expression and prevents DNA fragmentation and apoptosis in normal and leukemic cells. Thus, we hypothesized by disrupting the AKT/Ebp1 interplay via miR-155 down-regulation, 8-Cl-Ado induces pro-apoptotic Ebp1-dependent p53 expression and activation and leukemia cell death. Indeed, we showed that 8-Cl-Ado treatment caused AKT/Ebp1 dissociation and p53 activation in primary FLT3-ITD AML blasts. Conversely, overexpression of miR-155 reversed 8-Cl-Ado-induced apoptosis. By combining 8-Cl-Ado with the TKI quizartinib we elicited a synergistic anti-leukemic effect in primary AML blasts [combination index (CI) values 〈 1 at doses required to inhibit 50, 75, and 90% of cell growth (ED50, ED75, ED90)]. In an orthotopic mouse model where FLT3-ITD-positive MV4-11 cells were xenografted, mice treated with 75 mg 8-Cl-Ado/kg/day plus quizartinib (0.5 mg/kg/day) survived significantly longer than mice treated with the individual agents or vehicle alone (median survival 54.2 days vs. 27.3 days for controls, p=0.016). Taken together, these results support 8-Cl-Ado as a promising novel agent via a unique mechanism of action for FLT3-ITD AML, mediated by miR-155 degradation. 8-Cl-Ado also synergizes with TKIs both in vitro and in vivo thereby representing a potentially novel clinical approach for FLT3-ITD AML. A single-agent phase I/II clinical trial in relapsed/refractory AML is already ongoing at our institution. Figure. Figure. Disclosures Wennerberg: Novartis: Research Funding. Gandhi:Pharmacyclics: Research Funding.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
    URL: Article
    DOI: 10.1182/blood-2018-99-119880
    RVK:
    XA 33000
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    XA 10000
    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2018
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  • 7
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    Primary T-Prolymphocytic Leukemia (T-PLL) Cells Are Sensitive To BCL-2 and HDAC Inhibitors: Results From High-Throughput Ex Vivo Drug Testing
    American Society of Hematology ; 2013
    In:  Blood Vol. 122, No. 21 ( 2013-11-15), p. 3828-3828
    Details
    In: Blood, American Society of Hematology, Vol. 122, No. 21 ( 2013-11-15), p. 3828-3828
    Abstract: T-PLL is a rare mature post-thymic T-cell neoplasm with an aggressive clinical course and median overall survival of less than one year. Almost 75% of T-PLL cases harbor chromosome 14 translocations involving the T-cell receptor A/D locus resulting in aberrant activation of the proto-oncogenes TCL1A or MTCP1. T-PLL patients are difficult to treat as the leukemic cells are often resistant to most available chemotherapeutic drugs. Due to the rareness and aggressive nature of the disease, large clinical trials are difficult to execute. We therefore aimed to discover novel potential therapeutic targets using a high-throughput ex vivo drug sensitivity and resistance testing (DSRT) platform covering 306 approved and investigational oncology drugs. Methods Primary T-PLL cells were available from two patients. The first patient had a double positive CD4+CD8+CD3+ Vβ.14+ T-cell phenotype (patient 1) and cells underwent DSRT twice during a 5-month time-period (no treatment during that time). The second patient had a CD4+CD3+ phenotype (patient 2) and the cells were assayed once by DSRT. Fresh blood mononuclear cells (MNCs) were separated by Ficoll centrifugation from the patient samples (over 85 % leukemic cells in the MNC fraction) and healthy controls. Cells were seeded in 384-well plates and 306 active substances were tested using a 10,000-fold concentration range resulting in a dose-response curve for each compound. Cell viability was measured after 72 h incubation and differential drug sensitivity scores (DSS), representing leukemia-specific responses, were calculated by comparing patient samples with those obtained from healthy donors. In addition, both exome and RNA sequencing was performed from T-PLL cells (patient 1). Results Both patient samples showed high sensitivity to small molecule BCL2-inhibitors navitoclax (EC50 values 44nM and 10nM) and ABT-199 (EC50 23nM and 20nM) (Fig. 1 and 2). HDAC-inhibitors (quisinostat, belinostat and panobinostat) also showed high sensitivity in both patients in low nM concentrations (EC50 values 1-80nM). As AKT1/mTOR pathway is activated in most T-PLL patients due to the TCL1 oncoprotein, it was interesting to observe that neither of the patient samples showed any response to an AKT1 inhibitor (MK-2206 EC50 values 〉 1000 nM) nor to mTOR inhibitors (temsirolimus and everolimus)(Fig. 1). Furthermore, T-PLL cells were resistant to corticosteroids such as prednisolone and methylprednisolone. To further elucidate the molecular mechanism behind the drug responses, exome and RNA sequencing was performed from T-PLL cells (patient 1). No deletion was found in the ATM gene, but instead a homozygous missense mutation K2413Q was detected. This particular mutation is in the region coding for the FAT domain and while it has not been described earlier in T-PLL, it is in a cancer mutation hotspot region of ATM, suggesting that it is inactivating. No mutations directly linked to the BCL2-family genes were observed. In the RNA sequencing analysis, TCL1A was overexpressed when compared to the healthy CD4+ cells as expected. Similarly, AKT1 was overexpressed. The expression of BCL-2 and BCL-XL did not differ from those observed in healthy CD4+ cells while pro-apoptotic BCL-2 family members BID and BAD were elevated compared to the healthy control. Conclusions Primary T-PLL cells showed sensitivity to BCL-2 and HDAC inhibitors in a systematic high-throughput ex vivo drug sensitivity testing across a range of clinical and investigational drugs. The BCL-2 inhibitor sensitivity was not related to increased BCL-2 expression or activating mutations in the BCL-2 family genes, and further studies are needed to clarify the mechanism of action. However, the results suggest that BCL-2 inhibitors could be a novel promising candidate drug for T-PLL-patients and warrant further clinical development in this group of patients. In contrast, inhibitors of AKT and mTOR, kinases known to be activated by TCL1, showed no efficacy ex vivo in this assay. Disclosures: Porkka: BMS: Consultancy, Research Funding, Speakers Bureau; Novartis: Consultancy, Research Funding, Speakers Bureau. Mustjoki:Novartis: Consultancy, Speakers Bureau; BMS: Consultancy, Speakers Bureau.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
    URL: Article
    DOI: 10.1182/blood.V122.21.3828.3828
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    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2013
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  • 8
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    Metabolic Drug Survey Highlights Cancer Cell Dependencies and Vulnerabilities
    American Society of Hematology ; 2020
    In:  Blood Vol. 136, No. Supplement 1 ( 2020-11-5), p. 26-27
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    In: Blood, American Society of Hematology, Vol. 136, No. Supplement 1 ( 2020-11-5), p. 26-27
    Abstract: Here we assembled a novel metabolic drug library covering 243 compounds allowing to systematically identify metabolic dependencies in high-throughput phenotypic screens. The CeMM Library of Metabolic Drugs (CLIMET) was compiled in a stepwise fashion, starting from 8000 candidate compounds, after a survey of public drug-target databases, and ending with 243 highly-curated compounds including extensive crosschecking for approval status, structural information, compound's potency and selectivity for the intended target, pathway/target redundancy, and commercial availability. To assess the potency of the compounds in CLIMET, we screened the full collection against a panel of 15 diverse myeloid leukemia cell lines. Each compound was tested for its effect on cell growth and survival in a 10,000-fold concentration range (1nM to 10uM) enabling the generation of dose response curves and calculation of area under the curve values (AUC) for each drug. We, further, functionally grouped the cell lines and drugs based on metabolic drug efficacy patterns and associated them with distinct genomic and metabolic attributes. Analysis of the metabolic drug response profiles revealed that 77 compounds (32%) affected cell viability with the top effective compounds targeting nucleotide metabolism, oxidative stress, and the PI3K/mTOR pathway. Unsupervised hierarchical clustering of the drug sensitivity profiles stratified the cell lines in 5 functional taxonomic groups, with the activity of 19 compounds significantly contributing to the cell line grouping (e.g. PF-02545920, GW 4064, mTOR inhibitors, daporinad). Comparison of the oxygen consumption rate and extracellular acidification rate showed that the examined cell lines have analogous baseline metabolic phenotypes, suggesting that the mitochondrial function of the cells as assessed by Seahorse analysis did not significantly influence the clustering. Genotype to phenotype associations were identified between FLT3mutations and sensitivity to 5-FU, lestaurtinib, and PF-02545920. Moreover, RAS mutations negatively correlated to mTOR and mitochondrial respiration inhibitor sensitivity, whereas TP53 mutations conferred a resistance phenotype to PI3K pathway inhibitors and antineoplastic agents. Selective sensitivities were detected to the lactate transporter (SLC16A1) inhibitor AZD3965, the PI3K inhibitor pictilisib, and the fatty acid synthase inhibitor GSK2194069, which could be explained by varied gene expression in sensitive cell lines and target/process dependency. CLIMET allows for identification of metabolic susceptibilities, grouping of cancer cells based on metabolic dependencies, as well as understanding of context-dependent mechanism of action of drugs. Functional drug testing may provide a rapid and robust approach to identify metabolic vulnerabilities, responding patients, and prioritize compounds for clinical evaluation as illustrated with our study. Disclosures Staber: Janssen: Consultancy, Honoraria; AbbVie: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Astra Zeneca: Consultancy, Honoraria; Roche: Consultancy, Honoraria, Research Funding; Celgene/ BMS: Consultancy, Honoraria; Gilead: Consultancy, Honoraria; msd: Consultancy, Honoraria.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
    URL: Article
    DOI: 10.1182/blood-2020-134769
    RVK:
    XA 33000
    RVK:
    XA 10000
    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2020
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  • 9
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    Identification Of AML Subtype-Selective Drugs By Functional Ex Vivo Drug Sensitivity and Resistance Testing and Genomic Profiling
    American Society of Hematology ; 2013
    In:  Blood Vol. 122, No. 21 ( 2013-11-15), p. 482-482
    Details
    In: Blood, American Society of Hematology, Vol. 122, No. 21 ( 2013-11-15), p. 482-482
    Abstract: Adult acute myeloid leukemia (AML) exemplifies the challenges of modern cancer drug discovery and development in that molecularly targeted therapies are yet to be translated into clinical use. No effective second-line therapy exists once standard chemotherapy fails. While many genetic events have been linked with the onset and progression of AML, the fundamental disease mechanisms remain poorly understood. There is significant genomic and molecular heterogeneity among patients. Several targeted therapies have been investigated for improved second-line AML therapy but none has been approved for clinical use to date. It would be critically important to identify patient subgroups that would benefit from such therapies and to identify combinations of drugs that are likely to be effective. Methods To identify and optimize novel therapies for AML, we studied 28 samples from 18 AML patients with an individualized systems medicine (ISM) approach. The ISM platform includes functional profiling of AML patient cells ex vivo with drug sensitivity and resistance testing (DSRT), comprehensive molecular profiling as well as clinical background information. Data integration was done to identify disease- and patient-specific molecular vulnerabilities for translation in the clinic. The DSRT platform comprises 306 anti-cancer agents, each tested in a dose response series. We calculated differential drug sensitivity scores by comparing AML responses to those of control cells in order to distinguish cancer-specific drug effects. Next generation RNA- and exome-sequencing was used to identify fusion transcripts and mutations that link to drug sensitivities. Results Individual AML patient samples had a distinct drug sensitivity pattern, but unsupervised hierarchical clustering of the drug sensitivity profiles of the 28 AML patient samples identified 5 functional AML drug response subtypes. Each subtype was characterized by distinct combinations of sensitivities: Bcl-2 inhibitors (e.g. navitoclax; Group 1), JAK inhibitors (e.g. ruxolitinib) (Group 2) and MEK inhibitors (e.g. trametinib) (Groups 2 and 4), PI3K/mTOR inhibitors (e.g. temsirolimus; Groups 4 and 5), broad spectrum receptor tyrosine kinase inhibitors (e.g. dasatinib) (Groups 3, 4 and 5) and FLT3 inhibitors (e.g. quizartinib, sunitinib) (Group 5). Correlation of overall drug responses with genomic profiles revealed that RAS and FLT3 mutations were significantly linked with the drug response subgroups 4 and 5, respectively. Activating FLT3 mutations contributed to sensitivity to FLT3 inhibitors, as expected, but also to tyrosine kinase inhibitors not targeting FLT3, such as dasatinib. Hence, these data point to the potential synergistic combinatorial effects of FLT3 inhibitors with dasatinib for improved therapy outcome (Figure). Early clinical translational results based on compassionate use support this hypothesis. Therefore, by combinations of drugs we expect to see synergistic drug responses that can be translated into efficacious and safe therapies for relapsed AML cases in the clinic. Clinical application of DSRT results in the treatment of eight recurrent chemorefractory patients led to objective responses in three cases according to ELN criteria, whereas four of the remaining five patients had meaningful responses not meeting ELN criteria. After disease progression, AML patient cells showed ex vivo resistance to the drugs administered to the patients, as well as significant changes in clonal architecture during treatment response. Furthermore, we saw genomic alterations potentially explaining drug resistance, such as appearance of novel fusion genes. Summary The ISM approach represents an opportunity for improving therapies for cancer patients, one patient at the time. We show that the platform can be used to identify functional groups of AML linking to vulnerabilities to single targeted drugs and, importantly, unexpected drug combinations. This information can in turn be used for personalized medicine strategies and for creating hypotheses to be explored in systematic clinical trials, both for approved and investigational drugs. Disclosures: Off Label Use: Many of the compounds included in our DSRT platform are not indicated for AML therapy. Mustjoki:BMS: Honoraria, Research Funding; Novartis: Honoraria. Porkka:Novartis: Honoraria, Research Funding; BMS: Honoraria, Research Funding. Kallioniemi:Medisapiens: Membership on an entity’s Board of Directors or advisory committees; Roche: Research Funding.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
    URL: Article
    DOI: 10.1182/blood.V122.21.482.482
    RVK:
    XA 33000
    RVK:
    XA 10000
    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2013
    detail.hit.zdb_id: 1468538-3
    detail.hit.zdb_id: 80069-7
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    BTU Cottbus
    Wissenschaftspark Albert Einstein
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  • 10
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    Online Resource
    High-Throughput Ex Vivo Drug Sensitivity and Resistance Testing (DSRT) Integrated with Deep Genomic and Molecular Profiling Reveal New Therapy Options with Targeted Drugs in Subgroups of Relapsed Chemorefractory AML
    American Society of Hematology ; 2012
    In:  Blood Vol. 120, No. 21 ( 2012-11-16), p. 288-288
    Details
    In: Blood, American Society of Hematology, Vol. 120, No. 21 ( 2012-11-16), p. 288-288
    Abstract: Abstract 288 Introduction: Recent genomic analyses of acute myeloid leukemia (AML) patients have provided new information on mutations contributing to the disease onset and progression. However, the genomic changes are often complex and highly diverse from one patient to another and often not actionable in clinical care. To rapidly identify novel patient-specific therapies, we developed a high-throughput drug sensitivity and resistance testing (DSRT) platform to experimentally validate therapeutic options for individual patients with relapsed AML. By integrating the results with exome and transcriptome sequencing plus proteomic analysis, we were able to define specific drug-sensitive subgroups of patients and explore predictive biomarkers. Methods: Ex vivo DSRT was implemented for 29 samples from 16 adult AML patients at the time of relapse and chemoresistance and from 5 healthy donors. Fresh mononuclear cells from bone marrow aspirates ( 〉 50% blast count) were screened against a comprehensive collection of cytotoxic chemotherapy agents (n=103) and targeted preclinical and clinical drugs (n=100, later 170). The drugs were tested over a 10,000-fold concentration range resulting in a dose-response curve for each compound and each leukemia sample. A leukemia-specific drug sensitivity score (sDSS) was derived from the area under each dose response curve in relation to the total area, and comparing leukemia samples with normal bone marrow results. The turnaround time for the DSRT assay was 4 days. All samples also underwent deep exome (40–100×) and transcriptome sequencing to identify somatic mutations and fusion transcripts, as well as phosphoproteomic array analysis to uncover active cell signaling pathways. Results: The drug sensitivity profiles of AML patient samples differed markedly from healthy bone marrow controls, with leukemia-specific responses mostly observed for molecularly targeted drugs. Individual AML patient samples clustered into distinct subgroups based on their chemoresponse profiles, thus suggesting that the subgroups were driven by distinct signaling pathways. Similarly, compounds clustered based on the response across the samples revealing functional groups of compounds of both expected and unexpected composition. Furthermore, subsets of patient samples stood out as highly sensitive to different compounds. Specifically, dasatinib, rapalogs, MEK inhibitors, ruxolitinib, sunitinib, sorafenib, ponatinib, foretinib and quizartinib were found to be selectively active in 5 (31%), 5 (31%), 4 (25%), 4 (25%), 3 (19%), 3 (19%), 2 (13%), 2 (13%), and 1 (6%) of the AML patients ex vivo, respectively. DSRT assays of serial samples from the same patient at different stages of leukemia progression revealed patterns of resistance to the clinically applied drugs, in conjunction with evidence of dynamic changes in the clonal genomic architecture. Emergence of vulnerabilities to novel pathway inhibitors was seen at the time of drug resistance, suggesting potential combinatorial or successive cycles of drugs to achieve remissions in an increasingly chemorefractory disease. Genomic and molecular profiling of the same patient samples not only highlighted potential biomarkers reflecting the ex vivo DSRT response patterns, but also made it possible to follow in parallel the drug sensitivities and the clonal progression of the disease in serial samples from the same patients. Summary: The comprehensive analysis of drug responses by DSRT in samples from human chemorefractory AML patients revealed a complex pattern of sensitivities to distinct inhibitors. Thus, these results suggest tremendous heterogeneity in drug response patterns and underline the relevance of individual ex vivo drug testing in selecting optimal therapies for patients (personalized medicine). Together with genomic and molecular profiling, the DSRT analysis resulted in a comprehensive view of the drug response landscape and the underlying molecular changes in relapsed AML. These data can readily be translated into the clinic via biomarker-driven stratified clinical trials. Disclosures: Mustjoki: Bristol-Myers Squibb: Honoraria, Research Funding; Novartis: Honoraria. Kallioniemi:Roche: Research Funding; Medisapiens: Membership on an entity's Board of Directors or advisory committees. Porkka:Bristol-Myers Squibb: Honoraria, Research Funding; Novartis: Honoraria, Research Funding.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
    URL: Article
    DOI: 10.1182/blood.V120.21.288.288
    RVK:
    XA 33000
    RVK:
    XA 10000
    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2012
    detail.hit.zdb_id: 1468538-3
    detail.hit.zdb_id: 80069-7
    Library Location Call Number Volume/Issue/Year Availability
    BibTip Others were also interested in ...
    Online Resource
    Open Access Request
    Availability (electronic / print)
    BTU Cottbus
    Wissenschaftspark Albert Einstein
    EUV Frankfurt
    TH Wildau
    FH Potsdam
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