34 results on '"Minna Suvela"'
Search Results
2. Supplementary Table S1 from Individualized Systems Medicine Strategy to Tailor Treatments for Patients with Chemorefractory Acute Myeloid Leukemia
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Krister Wennerberg, Olli Kallioniemi, Kimmo Porkka, Caroline A. Heckman, Tero Aittokallio, Jonathan Knowles, Maija Wolf, Imre Västrik, Laura Turunen, Minna Suvela, Maria E. Rämet, Tero Pirttinen, Alun Parsons, Aino Palva, Satu Mustjoki, Astrid Murumägi, Pirkko Mattila, Jesus M. Lopez Marti, Muntasir Mamun Majumder, Tuija Lundán, Maija Lepistö, Anna Lehto, Sonja Lagström, Evgeny Kulesskiy, Riikka Karjalainen, Bjørn T. Gjertsen, Erkki Elonen, Pekka Ellonen, Maxim M. Bespalov, Henrikki Almusa, Agnieszka Szwajda, Samuli Eldfors, Henrik Edgren, Bhagwan Yadav, Mika Kontro, and Tea Pemovska
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XLSX file 452K, DSRT Oncology drug collection
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- 2023
3. Supplementary Table S3 from Individualized Systems Medicine Strategy to Tailor Treatments for Patients with Chemorefractory Acute Myeloid Leukemia
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Krister Wennerberg, Olli Kallioniemi, Kimmo Porkka, Caroline A. Heckman, Tero Aittokallio, Jonathan Knowles, Maija Wolf, Imre Västrik, Laura Turunen, Minna Suvela, Maria E. Rämet, Tero Pirttinen, Alun Parsons, Aino Palva, Satu Mustjoki, Astrid Murumägi, Pirkko Mattila, Jesus M. Lopez Marti, Muntasir Mamun Majumder, Tuija Lundán, Maija Lepistö, Anna Lehto, Sonja Lagström, Evgeny Kulesskiy, Riikka Karjalainen, Bjørn T. Gjertsen, Erkki Elonen, Pekka Ellonen, Maxim M. Bespalov, Henrikki Almusa, Agnieszka Szwajda, Samuli Eldfors, Henrik Edgren, Bhagwan Yadav, Mika Kontro, and Tea Pemovska
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XLSX file 36K, Clinical characteristics of patients at the time of diagnosis and DSRT, and responses of previous therapies
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- 2023
4. Supplementary Table S2 from Individualized Systems Medicine Strategy to Tailor Treatments for Patients with Chemorefractory Acute Myeloid Leukemia
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Krister Wennerberg, Olli Kallioniemi, Kimmo Porkka, Caroline A. Heckman, Tero Aittokallio, Jonathan Knowles, Maija Wolf, Imre Västrik, Laura Turunen, Minna Suvela, Maria E. Rämet, Tero Pirttinen, Alun Parsons, Aino Palva, Satu Mustjoki, Astrid Murumägi, Pirkko Mattila, Jesus M. Lopez Marti, Muntasir Mamun Majumder, Tuija Lundán, Maija Lepistö, Anna Lehto, Sonja Lagström, Evgeny Kulesskiy, Riikka Karjalainen, Bjørn T. Gjertsen, Erkki Elonen, Pekka Ellonen, Maxim M. Bespalov, Henrikki Almusa, Agnieszka Szwajda, Samuli Eldfors, Henrik Edgren, Bhagwan Yadav, Mika Kontro, and Tea Pemovska
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XLSX file 977K, Drug sensitivity profiles of AML and control samples
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- 2023
5. Supplementary Figures, Tables and Methods from Individualized Systems Medicine Strategy to Tailor Treatments for Patients with Chemorefractory Acute Myeloid Leukemia
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Krister Wennerberg, Olli Kallioniemi, Kimmo Porkka, Caroline A. Heckman, Tero Aittokallio, Jonathan Knowles, Maija Wolf, Imre Västrik, Laura Turunen, Minna Suvela, Maria E. Rämet, Tero Pirttinen, Alun Parsons, Aino Palva, Satu Mustjoki, Astrid Murumägi, Pirkko Mattila, Jesus M. Lopez Marti, Muntasir Mamun Majumder, Tuija Lundán, Maija Lepistö, Anna Lehto, Sonja Lagström, Evgeny Kulesskiy, Riikka Karjalainen, Bjørn T. Gjertsen, Erkki Elonen, Pekka Ellonen, Maxim M. Bespalov, Henrikki Almusa, Agnieszka Szwajda, Samuli Eldfors, Henrik Edgren, Bhagwan Yadav, Mika Kontro, and Tea Pemovska
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PDF file 400K, Supplementary Figures 1-9; Supplementary Tables 4-9 and Supplementary Methods
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- 2023
6. Ex vivo venetoclax sensitivity testing predicts treatment response in acute myeloid leukemia
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Heikki Kuusanmäki, Sari Kytölä, Ida Vänttinen, Tanja Ruokoranta, Amanda Ranta, Jani Huuhtanen, Minna Suvela, Alun Parsons, Annasofia Holopainen, Anu Partanen, Milla E.L. Kuusisto, Sirpa Koskela, Riikka Räty, Maija Itälä-Remes, Imre Västrik, Olli Dufva, Sanna Siitonen, Kimmo Porkka, Krister Wennerberg, Caroline A. Heckman, Pia Ettala, Marja Pyörälä, Johanna Rimpiläinen, Timo Siitonen, and Mika Kontro
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Hematology - Abstract
The BCL2 inhibitor venetoclax has revolutionized the treatment of acute myeloid leukemia (AML) patients not benefitting from intensive chemotherapy. Nevertheless, treatment failure remains a challenge, and predictive markers are needed, particularly for relapsed or refractory (R/R) AML. Ex vivo drug sensitivity testing may correlate with outcomes, but its prospective predictive value remains unexplored. Here we report the results of the first stage of the prospective Phase 2 VenEx trial evaluating the utility and predictiveness of venetoclax sensitivity testing using different cell culture conditions and cell viability assays in patients receiving venetoclax-azacitidine (NCT04267081). Participants with de novo AML ineligible for intensive chemotherapy, R/R AML, or secondary AML were included. The primary endpoint was the treatment response in ex vivo sensitive participants and the key secondary endpoints were the correlation of sensitivity with responses and survival. Venetoclax sensitivity testing was successful in 38/39 participants. Experimental conditions significantly influenced predictive accuracy. Blast-specific venetoclax sensitivity measured in conditioned medium most accurately correlated with treatment outcomes; 88% of sensitive participants achieved treatment response. Median survival was significantly longer for ex vivo sensitive participants (14. 6 months for s ensitive, 3. 5 for insensitive, p < 0 . 001). T his analysis illustrates the feasibility of integrating drug-response profiling into clinical practice and demonstrates excellent predictivity.
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- 2022
7. S100 Calcium Binding Protein Family Members Associate With Poor Patient Outcome and Response to Proteasome Inhibition in Multiple Myeloma
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Minxia Liu, Yinyin Wang, Juho J. Miettinen, Romika Kumari, Muntasir Mamun Majumder, Ciara Tierney, Despina Bazou, Alun Parsons, Minna Suvela, Juha Lievonen, Raija Silvennoinen, Pekka Anttila, Paul Dowling, Peter O’Gorman, Jing Tang, and Caroline A. Heckman
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multiple myeloma ,panobinostat ,drug resistance ,integumentary system ,QH301-705.5 ,S100 protein family ,proteasome inhibitors ,Biology (General) - Abstract
Despite several new therapeutic options, multiple myeloma (MM) patients experience multiple relapses and inevitably become refractory to treatment. Insights into drug resistance mechanisms may lead to the development of novel treatment strategies. The S100 family is comprised of 21 calcium binding protein members with 17 S100 genes located in the 1q21 region, which is commonly amplified in MM. Dysregulated expression of S100 family members is associated with tumor initiation, progression and inflammation. However, the relationship between the S100 family and MM pathogenesis and drug response is unknown. In this study, the roles of S100 members were systematically studied at the copy number, transcriptional and protein level with patients’ survival and drug response. Copy number analysis revealed a predominant pattern of gains occurring in S100 genes clustering in the 1q21 locus. In general, gains of genes encoding S100 family members associated with worse patient survival. However, S100 gene copy number and S100 gene expression did not necessarily correlate, and high expression of S100A4 associated with poor patient survival. Furthermore, integrated analysis of S100 gene expression and ex vivo drug sensitivity data showed significant negative correlation between expression of S100 family members (S100A8, S100A9, and S100A12) and sensitivity to some drugs used in current MM treatment, including proteasome inhibitors (bortezomib, carfilzomib, and ixazomib) and histone deacetylase inhibitor panobinostat. Combined proteomic and pharmacological data exhibited significant negative association of S100 members (S100A4, S100A8, and S100A9) with proteasome inhibitors and panobinostat. Clinically, the higher expression of S100A4 and S100A10 were significantly linked to shorter progression free survival in patients receiving carfilzomib-based therapy. The results indicate an association and highlight the potential functional importance of S100 members on chromosome 1q21 in the development of MM and resistance to established myeloma drugs, including proteasome inhibitors.
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- 2021
8. S100 Calcium Binding Protein Family Members Associate With Poor Patient Outcome and Response to Proteasome Inhibition in Multiple Myeloma
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Minxia, Liu, Yinyin, Wang, Juho J, Miettinen, Romika, Kumari, Muntasir Mamun, Majumder, Ciara, Tierney, Despina, Bazou, Alun, Parsons, Minna, Suvela, Juha, Lievonen, Raija, Silvennoinen, Pekka, Anttila, Paul, Dowling, Peter, O'Gorman, Jing, Tang, and Caroline A, Heckman
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multiple myeloma ,Cell and Developmental Biology ,panobinostat ,drug resistance ,integumentary system ,S100 protein family ,proteasome inhibitors ,Original Research - Abstract
Despite several new therapeutic options, multiple myeloma (MM) patients experience multiple relapses and inevitably become refractory to treatment. Insights into drug resistance mechanisms may lead to the development of novel treatment strategies. The S100 family is comprised of 21 calcium binding protein members with 17 S100 genes located in the 1q21 region, which is commonly amplified in MM. Dysregulated expression of S100 family members is associated with tumor initiation, progression and inflammation. However, the relationship between the S100 family and MM pathogenesis and drug response is unknown. In this study, the roles of S100 members were systematically studied at the copy number, transcriptional and protein level with patients’ survival and drug response. Copy number analysis revealed a predominant pattern of gains occurring in S100 genes clustering in the 1q21 locus. In general, gains of genes encoding S100 family members associated with worse patient survival. However, S100 gene copy number and S100 gene expression did not necessarily correlate, and high expression of S100A4 associated with poor patient survival. Furthermore, integrated analysis of S100 gene expression and ex vivo drug sensitivity data showed significant negative correlation between expression of S100 family members (S100A8, S100A9, and S100A12) and sensitivity to some drugs used in current MM treatment, including proteasome inhibitors (bortezomib, carfilzomib, and ixazomib) and histone deacetylase inhibitor panobinostat. Combined proteomic and pharmacological data exhibited significant negative association of S100 members (S100A4, S100A8, and S100A9) with proteasome inhibitors and panobinostat. Clinically, the higher expression of S100A4 and S100A10 were significantly linked to shorter progression free survival in patients receiving carfilzomib-based therapy. The results indicate an association and highlight the potential functional importance of S100 members on chromosome 1q21 in the development of MM and resistance to established myeloma drugs, including proteasome inhibitors.
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- 2021
9. The Peptide–Drug Conjugate Melflufen Modulates the Unfolded Protein Response of Multiple Myeloma and Amyloidogenic Plasma Cells and Induces Cell Death
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Ken, Flanagan, Romika, Kumari, Juho J, Miettinen, Staci L, Haney, Michelle L, Varney, Jacob T, Williams, Muntasir M, Majumder, Minna, Suvela, Ana, Slipicevic, Fredrik, Lehmann, Nina N, Nupponen, Sarah A, Holstein, Caroline A, Heckman, Institute for Molecular Medicine Finland, Helsinki Institute of Life Science HiLIFE, Digital Precision Cancer Medicine (iCAN), and Research Programs Unit
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POTENTIATED ALKYLATOR MELFLUFEN ,MANAGEMENT ,LINES ,IN-VITRO ,3111 Biomedicine ,Hematology ,DEGRADATION ,PROTEASOME INHIBITORS ,AGENT ,STRESS PATHWAY - Abstract
Immunoglobulin light-chain (AL) amyloidosis is a rare disease caused by clonal plasma cell secretion of misfolded light chains that assemble as toxic amyloid fibrils, depositing in vital organs including the heart and kidneys, causing organ dysfunction. Plasma cell–directed therapeutics are expected to reduce production of toxic light chain by eliminating amyloidogenic cells in bone marrow, thereby diminishing amyloid fibril deposition and providing the potential for organ recovery. Melphalan flufenamide (melflufen) is a first-in-class peptide–drug conjugate that targets aminopeptidases and rapidly releases alkylating agents inside tumor cells. Melflufen is highly lipophilic, permitting rapid uptake by cells, where it is enzymatically hydrolyzed by aminopeptidases, resulting in intracellular accumulation of the alkylating agents, including melphalan. Previous data demonstrating sensitivity of myeloma cells to melflufen suggest that the drug might be useful in AL amyloidosis. We describe the effects of melflufen on amyloidogenic plasma cells in vitro and ex vivo, demonstrating enhanced cytotoxic effects in comparison to melphalan, as well as novel mechanisms of action through the unfolded protein response (UPR) pathway. These findings provide evidence that melflufen-mediated cytotoxicity extends to amyloidogenic plasma cells, and support the rationale for the evaluation of melflufen in patients with AL amyloidosis.
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- 2022
10. Single Cell RNA Sequencing Identifies Potential Molecular Indicators of Response to Melflufen in Multiple Myeloma
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Ana Slipicevic, Caroline A. Heckman, Sadiksha Adhikari, Nina N. Nupponen, Philipp Sergeev, Minna Suvela, Fredrik Lehmann, Maiju-Emilia Huppunen, and Juho J. Miettinen
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0303 health sciences ,Immunology ,Cell ,RNA ,Cell Biology ,Hematology ,Computational biology ,Biology ,medicine.disease ,Biochemistry ,3. Good health ,03 medical and health sciences ,0302 clinical medicine ,medicine.anatomical_structure ,medicine ,Multiple myeloma ,030304 developmental biology ,030215 immunology - Abstract
Introduction Melphalan flufenamide (melflufen), is a novel peptide-drug conjugate that targets aminopeptidases and selectively delivers alkylating agents in tumors. Melflufen was recently FDA approved for the treatment of relapsed/refractory multiple myeloma (MM) patients. Considering the challenges in treating this group of patients, and the availability of several new drugs for MM, information that can support treatment selection is urgently needed. To identify potential indicators of response and mechanism of resistance to melflufen, we applied a multiparametric drug sensitivity assay to MM patient samples ex vivo and analyzed the samples by single cell RNA sequencing (scRNAseq). Ex vivo drug testing identified MM samples that were distinctly sensitive or resistant to melflufen, while differential gene expression analysis revealed pathways associated with response. Methods Bone marrow (BM) aspirates from 24 MM patients were obtained after written informed consent following approved protocols in compliance with the Declaration of Helsinki. BM mononuclear cells from 12 newly diagnosed (ND) and 12 relapsed/refractory (RR) patients were used for multi-parametric flow cytometry-based drug sensitivity and resistance testing (DSRT) evaluation to melflufen and melphalan, and for scRNAseq. Based on the results from the DSRT tests and drug sensitivity scores (DSS), we divided the samples into three groups - high sensitivity (HS, DSS > 40 (melflufen) or DSS > 16 (melphalan)), intermediate sensitivity (IS, 31 ≤ DSS ≤ 40 (melflufen) or 10 ≤ DSS ≤ 16 (melphalan)), and low sensitivity (LS, DSS < 31 (melflufen) or DSS < 10 (melphalan)). To identify genes, responsible for the general sensitivity to melphalan-based drugs we conducted differential gene expression (DGE) analyses separately for melphalan and melflufen focusing on the plasma cell populations, comparing gene expression between HS and LS samples for both drugs ("HS vs. LS melphalan" and "HS vs. LS for melflufen", respectively). In addition, to explain the increased sensitivity of RR samples, we conducted the DGE analysis for ND vs. RR samples and searched for similarities between these three datasets. Results DSRT data indicated that samples from RRMM patients were significantly more sensitive to melflufen compared to samples from NDMM (Fig. 1A). In addition, we observed that samples with a gain of 1q (+1q) were more sensitive to melflufen while those with deletion of 13q (del13q) appeared to be less sensitive, although these results lacked significance (Fig. 1A). After separating the samples into different drug sensitivity groups (HS, IS, LS), DGE analysis showed significant downregulation of the drug efflux and multidrug resistance protein family member ABCB9 in the melflufen HS group opposed to the LS group (2.2-fold, p < 0.001). A similar pattern was detected for the melphalan HS vs. LS comparison suggesting that this alteration might be a common indicator of sensitivity to melphalan-based drugs. Furthermore, in the melflufen HS group we observed downregulation of the matrix metallopeptidase inhibitors TIMP1 and TIMP2 (3-fold and 1.6-fold, p < 0.001, respectively), and cathepsin inhibitors CST3 and CSTB (3.2-fold and 1.3-fold, p < 0.001, respectively) (Fig. 1B). This effect was observed in both "ND vs. RR" and "HS vs. LS for melflufen" comparisons, but not for melphalan, suggesting that these changes are associated with disease progression and specific indicators of sensitivity to melflufen. Moreover, gene set enrichment analysis (GSEA) showed activation of pathways related to protein synthesis, as well as amino acid starvation for malignant and normal cell populations in the HS group. Conclusion In summary, our results indicate that melflufen is more active in RRMM compared to NDMM. In addition, samples from MM patients with +1q, which is considered an indicator of high-risk disease, tended to be more sensitive to melflufen. Based on differential GSEA and pathway enrichment, several synergizing mechanisms could potentially explain the higher sensitivity to melflufen, such as decreased drug efflux and increased drug uptake. Although these results indicate potential indicators of response and mechanisms of drug efficacy, further validation of these findings is required using data from melflufen treated patients. Figure 1 Figure 1. Disclosures Slipicevic: Oncopeptides AB: Current Employment. Nupponen: Oncopeptides AB: Consultancy. Lehmann: Oncopeptides AB: Current Employment. Heckman: Orion Pharma: Research Funding; Oncopeptides: Consultancy, Research Funding; Novartis: Research Funding; Celgene/BMS: Research Funding; Kronos Bio, Inc.: Research Funding.
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- 2021
11. In Vitro and inVivo Activity of Melflufen in Amyloidosis
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Caroline A. Heckman, Minna Suvela, Muntasir Mamun Majumder, Fredrik Lehmann, Sarah A. Holstein, Ana Slipicevic, Ken Flanagan, Juho J. Miettinen, Romika Kumari, Michelle L. Varney, and Nina N. Nupponen
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0301 basic medicine ,Melphalan ,Immunology ,Plasma cell ,Immunoglobulin light chain ,Biochemistry ,03 medical and health sciences ,0302 clinical medicine ,medicine ,AL amyloidosis ,Multiple myeloma ,biology ,Chemistry ,Amyloidosis ,Cell Biology ,Hematology ,medicine.disease ,3. Good health ,030104 developmental biology ,medicine.anatomical_structure ,biology.protein ,Cancer research ,Bone marrow ,Antibody ,030215 immunology ,medicine.drug - Abstract
Background: Immunoglobulin light-chain (AL) amyloidosis is a rare disease caused by plasma cell secretion of misfolded light chains that assemble as amyloid fibrils and deposit on vital organs including the heart and kidneys, causing organ dysfunction. Plasma cell directed therapeutics, aimed at preferentially eliminating the clonal population of amyloidogenic cells in bone marrow are expected to reduce production of toxic light chain and alleviate deposition of amyloid thereby restoring healthy organ function. Melphalan flufenamide ethyl ester, melflufen, is a peptidase potentiated alkylating agent with potent toxicity in myeloma cells. Melflufen is highly lipophilic, permitting rapid cellular uptake, and is subsequently enzymatically cleaved by aminopeptidases within cells resulting in augmented intracellular concentrations of toxic molecules, providing a more targeted and localized treatment. Previous data demonstrating multiple myeloma plasma cell sensitivity for melflufen suggests that the drug might be useful to directly eliminate amyloidogenic plasma cells, thereby reducing the amyloid load in patients. Furthermore, the increased intracellular concentrations of melflufen in myeloma cells indicates a potential reduction in systemic toxicity in patients, an important factor in the fragile amyloidosis patient population. To assess potential efficacy in amyloidosis patients and to explore the mechanism of action, we examined effects of melflufen on amyloidogenic plasma cells invitro and invivo. Methods: Cellular toxicity and apoptosis were measured in response to either melflufen or melphalan in multiple malignant human plasma cell lines, including the amyloidosis patient derived light chain secreting ALMC-1 and ALMC-2 cells, as well as primary bone marrow cells from AL amyloidosis patients, using annexin V and live/dead cell staining by multicolor flow cytometry, and measurement of cleaved caspases. Lambda light chain was measured in supernatant by ELISA, and intracellular levels were detected by flow cytometry. To assess efficacy of melflufen in vivo, the light chain secreting human myeloma cell line, JJN3, was transduced with luciferase and adoptively transferred into NSG mice. Cell death in response to melflufen or melphalan was measured by in vivo bioluminescence, and serum light chain was monitored. Results: Melflufen demonstrated increased potency against multiple myeloma cell lines compared to melphalan, inducing malignant plasma cell death at lower doses on established light chain secreting plasma cell lines. While ALMC-1 cells were sensitive to both melphalan and melflufen, the IC50 for melphalan at 960 nM was approximately 3-fold higher than melflufen (334 nM). However, ALMC-2 cells were relatively insensitive to melphalan (12600 nM), but maintained a 100-fold increase in sensitivity to melflufen (121 nM). Furthermore, while 40% of primary CD138+ plasma cells from patients with diagnosed AL amyloidosis responded to melflufen treatment in vitro, only 20% responded to melphalan with consistently superior IC50 values for melflufen (Figure 1). Light chain secreting cell lines and AL amyloidosis patient samples were further analyzed by single cell sequencing. We further examined differential effects on apoptosis and the unfolded protein response in vitro in response to either melflufen or melphalan. This is of particular interest in amyloidosis, where malignant antibody producing plasma cells possess an increased requirement for mechanisms to cope with the amplified load of unfolded protein and associated ER stress. As AL amyloidosis is ultimately a disease mediated by secretion of toxic immunoglobulin, we assessed the effects of melflufen on the production of light chain invitro, measuring a decrease in production of light chain in response to melflufen treatment. Finally, we took advantage of a recently described adoptive transfer mouse model of amyloidosis to assess the efficacy of melflufen and melphalan in eliminating amyloidogenic clones and reducing the levels of toxic serum light chain in vivo. Conclusions: These findings provide evidence that melflufen mediated toxicity, previously described in myeloma cells, extends to amyloidogenic plasma cells and further affects the ability of these cells to produce and secrete toxic light chain. This data supports the rationale for the evaluation of melflufen in patients with AL amyloidosis. Figure 1 Disclosures Flanagan: Oncopeptides AB: Employment. Slipicevic:Oncopeptides AB: Employment. Holstein:Celgene: Consultancy; Takeda: Membership on an entity's Board of Directors or advisory committees; Adaptive Biotechnologies: Membership on an entity's Board of Directors or advisory committees; GSK: Consultancy; Genentech: Membership on an entity's Board of Directors or advisory committees; Sorrento: Consultancy. Lehmann:Oncopeptides AB: Employment. Nupponen:Oncopeptides AB: Employment. Heckman:Celgene: Research Funding; Novartis: Research Funding; Oncopeptides: Research Funding; Orion Pharma: Research Funding.
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- 2019
12. Associations between Microrna Expression, Disease Progression and Ex Vivo Drug Response in Multiple Myeloma
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Romika Kumari, Alun Parsons, Juha Lievonen, Minna Suvela, Caroline A. Heckman, Ashwini Kumar, Pekka Anttila, and Raija Silvennoinen
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0303 health sciences ,business.industry ,Immunology ,Disease progression ,Treatment outcome ,Buparlisib ,Cancer ,Cell Biology ,Hematology ,medicine.disease ,Biochemistry ,3. Good health ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,chemistry ,microRNA ,Cancer research ,Drug response ,Medicine ,business ,Ex vivo ,Multiple myeloma ,030304 developmental biology ,030215 immunology - Abstract
Targeted drug treatment strategies have significantly prolonged the overall survival rate among multiple myeloma (MM) patients. However, high relapse rates and multiple drug resistance still pose major challenges. Although, the underlying molecular features of the disease have been explored both at the genomic and transcriptomic levels, the functional role of microRNAs (miRNA) in MM disease progression and prognosis is yet to be investigated at a personalized level. In earlier studies, microRNAs have been implicated to regulate gene expression and were determined to play crucial roles in the biology of MM by acting as oncogenes or tumor suppressors. Nevertheless, considering the heterogeneity of MM, little is known about the roles of miRNAs in controlling MM disease progression and drug response at an individualized systems level. We collected bone marrow aspirates from MM patients at diagnosis (n=20) and relapse (n=25) after informed consent and following approved protocols in accordance with the Declaration of Helsinki. CD138+ plasma cells were enriched from the bone marrow samples and used for miRNA-sequencing and drug sensitivity and resistance testing (DSRT). The miRNA was prepared from the CD138+ cells and subjected to sequencing using Illumina compatible technologies. DSRT was performed and responses to 83 clinically approved drugs and investigational compounds were measured as drug sensitivity scores (DSS) as described previously (Majumder et al., Oncotarget 2017). The pairwise comparative analysis of miRNA expression and drug responses was performed using Spearman's rank-order correlations, to elucidate significant associations of miRNA expression with drug sensitivity and resistance. Additionally, using DEseq2 the differential miRNA expression was determined for the newly diagnosed and relapse samples to deconvolute the role of miRNAs in MM disease progression. The comparative analysis of the miRNA expression and drug sensitivity scores revealed statistically significant associations between miRNA expression and drug sensitivity measures with the Spearman coefficient (r) ranging from -0.71 to 0.64 (adjusted p-value ≤ 0.05) (Figure 1A). Negative associations were more prevalent, with 40 miRNAs negatively associated with ≥1 drug response from the total of 30 predicted drugs. miR-486, which is known to be an effective biomarker in diagnosis and prognosis of multiple cancer types (Jiang et al., Oncotarget 2018), was found to have significant negative correlation (r= -0.71 to -0.52, p-value ≤ 0.01) with the responses of 14 drugs. Similarly, negative correlation was observed for miR-144 with 12 drugs and miR-584 with 9 drugs. We observed that PI3K/mTOR inhibitors and HDAC inhibitors were common amongst all the significant negative correlations predicted. Specifically, the PI3K/mTOR inhibitors apitosilib, omipalisib and buparlisib were found to be negatively associated with the expression of 18, 14 and 7 miRNAs respectively. These observations can lead to the understanding of miRNA mediated regulation of molecular pathways involved in drug resistance. Differential miRNA expression analysis between newly diagnosed and relapse MM samples revealed the involvement of miRNAs in disease progression. The analysis resulted in total of 31 significant differentially expressed miRNAs with fold change ≥2 and adjusted p-value ≤ 0.1 (Figure 1B). Several miRNAs known to play crucial roles in cancer diagnosis and prognosis were found to be significantly upregulated in the relapse samples. In particular, 25 miRNAs were upregulated, including following miR-17/92 cluster members: miR-18b, miR-20a, miR-92b and miR-106a, which are known to have an oncogenic role in various cancer types (Mogilyansky & Rigoutsos, Cell Death and Differentiation 2013). Interestingly, 12/31 differentially regulated miRNAs were located on chromosome X. Although cytogenetic alteration data predicted that chromosome 1q gain is significantly prominent in the relapse samples (p-value = 0.009), only 3/31 differentially regulated miRNAs were located on chromosome 1. These results demonstrate the role of miRNAs in regulating drug response and disease progression in multiple myeloma. Monitoring miRNA expression profiles in MM patients can facilitate the assessment of treatment outcome and prognosis, and miRNAs could potentially be useful prognostic and treatment biomarkers for MM. Disclosures Silvennoinen: Amgen: Research Funding; Bristol-Myers Squibb (BMS): Research Funding; Takeda: Research Funding; Celgene: Research Funding. Heckman:Celgene: Research Funding; Novartis: Research Funding; Oncopeptides: Research Funding; Orion Pharma: Research Funding.
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- 2019
13. Identification of precision treatment strategies for relapsed/refractory multiple myeloma by functional drug sensitivity testing
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Bhagwan Yadav, Esa Jantunen, Pekka Anttila, Juha Lievonen, Samuli Eldfors, Muntasir Mamun Majumder, Raija Silvennoinen, Caroline A. Heckman, Riikka Karjalainen, Kimmo Porkka, Heikki Kuusanmäki, Alun Parsons, David Tamborero, Minna Suvela, Institute for Molecular Medicine Finland, University of Helsinki, Helsinki Institute of Life Science HiLIFE, Department of Oncology, Clinicum, Hematologian yksikkö, HUS Head and Neck Center, and HUS Comprehensive Cancer Center
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0301 basic medicine ,Drug ,LOW-DOSE DEXAMETHASONE ,media_common.quotation_subject ,medicine.medical_treatment ,precision medicine ,3122 Cancers ,Pharmacology ,drug sensitivity and resistance testing ,SINGLE-AGENT ,Targeted therapy ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,TARGETED THERAPY ,Refractory ,Panobinostat ,medicine ,COMBINATION ,Multiple myeloma ,media_common ,PANOBINOSTAT ,business.industry ,WORKING GROUP ,Precision medicine ,medicine.disease ,3. Good health ,GLUCOCORTICOID-RECEPTOR ,multiple myeloma ,functional screening ,030104 developmental biology ,Oncology ,chemistry ,030220 oncology & carcinogenesis ,CELLS ,PHASE-II ,high-risk myeloma ,3111 Biomedicine ,business ,RISK-STRATIFICATION ,Progressive disease ,Ex vivo ,Research Paper - Abstract
Novel agents have increased survival of multiple myeloma (MM) patients, however high-risk and relapsed/refractory patients remain challenging to treat and their outcome is poor. To identify novel therapies and aid treatment selection for MM, we assessed the ex vivo sensitivity of 50 MM patient samples to 308 approved and investigational drugs. With the results we i) classified patients based on their ex vivo drug response profile; ii) identified and matched potential drug candidates to recurrent cytogenetic alterations; and iii) correlated ex vivo drug sensitivity to patient outcome. Based on their drug sensitivity profiles, MM patients were stratified into four distinct subgroups with varied survival outcomes. Patients with progressive disease and poor survival clustered in a drug response group exhibiting high sensitivity to signal transduction inhibitors. Del(17p) positive samples were resistant to most drugs tested with the exception of histone deacetylase and BCL2 inhibitors. Samples positive for t(4; 14) were highly sensitive to immunomodulatory drugs, proteasome inhibitors and several targeted drugs. Three patients treated based on the ex vivo results showed good response to the selected treatments. Our results demonstrate that ex vivo drug testing may potentially be applied to optimize treatment selection and achieve therapeutic benefit for relapsed/refractory MM.
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- 2017
14. Abstract 458: Precision systems medicine in acute myeloid leukemia: real-time translation of tailored therapeutic opportunities arising from ex-vivo drug sensitivity testing and molecular profiling
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Disha Malani, Olli Kallioniemi, Alun Parson, Satu Mustjoki, Minna Suvela, Caroline A. Heckman, Katja Suomi, Karoliina Laamanen, Laura Turunen, Kimmo Porkka, Ashwni Kumar, Bjørn Tore Gjertsen, Imre Vastrik, Muntasir Mamun Majumder, Pekka Ellonen, Evgeny Kulesskiy, Maija Wolf, Maria Nurmi, Oscar Brück, Astrid Muruimägi, Swapnil Potdar, Sari Kytölä, Krister Wennerberg, Samuli Eldfors, Tero Aittokallio, Simon Anders, Riikka Karjalainen, Bhagwan Yadav, Jani Saarela, Siv Knappila, Matti Kankainen, Aino Palva, Elina Lehtinen, Mika Kontro, and Pirkko Mattila
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Oncology ,Drug ,Cancer Research ,medicine.medical_specialty ,NPM1 ,business.industry ,media_common.quotation_subject ,Myeloid leukemia ,Transcriptome ,Systems medicine ,Internal medicine ,Sensitivity testing ,medicine ,Prospective cohort study ,business ,Ex vivo ,media_common - Abstract
Acute myeloid leukemia (AML) is an aggressive disease of clonal hematopoietic progenitor cells. Here, we applied ex-vivo drug sensitivity and resistance testing on AML patient cells with 362 emerging and 153 approved cancer drugs together with genomic and transcriptomic profiling to identify and tailor therapies for patients with advanced disease. Ex-vivo testing with freshly isolated patient cells revealed cancer-specific efficacies of approved drugs in 97% of the 164 patient cases, including 47% of the cases with no actionable driver mutations. We identified 142 statistically significant associations between drug responses and somatic mutations, including increased sensitivity to JAK inhibitors in patients with NPM1 mutations. Transcriptomic profiles predicted drug responses better than genomics and helped to identify additional response markers, especially beyond mutations. For example, overexpression of HOX family genes was associated with sensitivity to JAK inhibitors in patients with NPM1 mutation. In a prospective study, we translated the functional drug response and molecular profile data to the clinic and suggested tailored therapy with targeted drugs for 26 relapsed or refractory AML patients. In an observational intervention study, acting on these recommendations resulted in a temporary complete clinical remission or leukemia-free state in 39% of the cases. In summary, we conclude that ex-vivo testing of drugs on patient AML cells i) revealed clinically actionable drug efficacies in almost all AML patients, including patients with no actionable mutations, ii) predicted cases with actionable driver mutations with no pharmacological dependency on the target, and iii) enabled real-time tailoring of therapy with 39% clinical response rate in chemorefractory advanced AML. Taken together, we believe this real-time systems medicine approach could become a powerful strategy for tailoring therapies for individual patients in the future. Citation Format: Disha Malani, Ashwni Kumar, Bhagwan Yadav, Mika Kontro, Swapnil Potdar, Oscar Bruck, Säri Kytölä, Jani Saarela, Samuli Eldfors, Riikka Karjalainen, Muntasir M. Majumder, Imre Västrik, Pekka Ellonen, Matti Kankainen, Minna Suvela, Siv Knappila, Alun Parson, Aino Palva, Pirkko Mattila, Evgeny Kulesskiy, Laura Turunen, Karoliina Laamanen, Elina Lehtinen, Maria Nurmi, Katja Suomi, Astrid Muruimägi, Bjorn T. Gjertsen, Satu Mustjoki, Simon Anders, Maija Wolf, Tero Aittokallio, Krister Wennerberg, Caroline Heckman, Kimmo Porkka, Olli Kallioniemi. Precision systems medicine in acute myeloid leukemia: real-time translation of tailored therapeutic opportunities arising from ex-vivo drug sensitivity testing and molecular profiling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 458.
- Published
- 2019
15. Individualized Systems Medicine Strategy to Tailor Treatments for Patients with Chemorefractory Acute Myeloid Leukemia
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Jonathan Knowles, Erkki Elonen, Laura Turunen, Tero Aittokallio, Maria E. Rämet, Kimmo Porkka, Satu Mustjoki, Aino Palva, Pekka Ellonen, Henrikki Almusa, Mika Kontro, Samuli Eldfors, Tuija Lundán, Caroline A. Heckman, Tea Pemovska, Agnieszka Szwajda, Bhagwan Yadav, Maija Lepistö, Tero Pirttinen, Anna Lehto, Krister Wennerberg, Maxim M. Bespalov, Olli Kallioniemi, Muntasir Mamun Majumder, Sonja Lagström, Jesus M. Lopez Marti, Minna Suvela, Imre Vastrik, Maija Wolf, Pirkko Mattila, Henrik Edgren, Astrid Murumägi, Evgeny Kulesskiy, Bjørn Tore Gjertsen, Alun Parsons, and Riikka Karjalainen
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0303 health sciences ,Myeloid ,business.industry ,Myeloid leukemia ,Drug resistance ,medicine.disease ,Bioinformatics ,Somatic evolution in cancer ,3. Good health ,Systems medicine ,03 medical and health sciences ,Drug repositioning ,0302 clinical medicine ,Breast cancer ,medicine.anatomical_structure ,Oncology ,030220 oncology & carcinogenesis ,medicine ,Personalized medicine ,business ,030304 developmental biology - Abstract
We present an individualized systems medicine (ISM) approach to optimize cancer drug therapies one patient at a time. ISM is based on (i) molecular profiling and ex vivo drug sensitivity and resistance testing (DSRT) of patients' cancer cells to 187 oncology drugs, (ii) clinical implementation of therapies predicted to be effective, and (iii) studying consecutive samples from the treated patients to understand the basis of resistance. Here, application of ISM to 28 samples from patients with acute myeloid leukemia (AML) uncovered five major taxonomic drug-response subtypes based on DSRT profiles, some with distinct genomic features (e.g., MLL gene fusions in subgroup IV and FLT3-ITD mutations in subgroup V). Therapy based on DSRT resulted in several clinical responses. After progression under DSRT-guided therapies, AML cells displayed significant clonal evolution and novel genomic changes potentially explaining resistance, whereas ex vivo DSRT data showed resistance to the clinically applied drugs and new vulnerabilities to previously ineffective drugs. Significance: Here, we demonstrate an ISM strategy to optimize safe and effective personalized cancer therapies for individual patients as well as to understand and predict disease evolution and the next line of therapy. This approach could facilitate systematic drug repositioning of approved targeted drugs as well as help to prioritize and de-risk emerging drugs for clinical testing. Cancer Discov; 3(12); 1416–29. ©2013 AACR. See related commentary by Hourigan and Karp, p. 1336 This article is highlighted in the In This Issue feature, p. 1317
- Published
- 2013
16. JAK1/2 and BCL2 inhibitors synergize to counteract bone marrow stromal cell-induced protection of AML
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Caroline A. Heckman, Bjørn Tore Gjertsen, Riikka Karjalainen, Alun Parsons, Jing Tang, Olli Kallioniemi, Tea Pemovska, Minna Suvela, Tero Aittokallio, Mihaela Popa, Mireia Mayoral Safont, David Tamborero, Komal Kumar Javarappa, Krister Wennerberg, Mika Kontro, Dmitrii Bychkov, Kimmo Porkka, Muntasir Mamun Majumder, Jonathan Knowles, Minxia Liu, Bhagwan Yadav, and Emmet McCormack
- Subjects
0301 basic medicine ,Myeloid ,Chronic lymphocytic leukemia ,Pharmacology ,Biochemistry ,Tyrosine-kinase inhibitor ,chemistry.chemical_compound ,Mice ,0302 clinical medicine ,hemic and lymphatic diseases ,Tumor Cells, Cultured ,Sulfonamides ,Drug Synergism ,Hematology ,3. Good health ,Leukemia ,Leukemia, Myeloid, Acute ,STAT Transcription Factors ,medicine.anatomical_structure ,Proto-Oncogene Proteins c-bcl-2 ,030220 oncology & carcinogenesis ,Female ,Signal Transduction ,Stromal cell ,medicine.drug_class ,Immunology ,Antineoplastic Agents ,Bone Marrow Cells ,Cell Line ,03 medical and health sciences ,Nitriles ,medicine ,Animals ,Humans ,Protein Kinase Inhibitors ,Venetoclax ,business.industry ,Cell Biology ,Janus Kinase 1 ,Janus Kinase 2 ,medicine.disease ,Bridged Bicyclo Compounds, Heterocyclic ,030104 developmental biology ,Pyrimidines ,chemistry ,Drug Resistance, Neoplasm ,Pyrazoles ,Bone marrow ,Stromal Cells ,business ,Ex vivo - Abstract
The bone marrow (BM) provides a protective microenvironment to support the survival of leukemic cells and influence their response to therapeutic agents. In acute myeloid leukemia (AML), the high rate of relapse may in part be a result of the inability of current treatment to effectively overcome the protective influence of the BM niche. To better understand the effect of the BM microenvironment on drug responses in AML, we conducted a comprehensive evaluation of 304 inhibitors, including approved and investigational agents, comparing ex vivo responses of primary AML cells in BM stroma-derived and standard culture conditions. In the stroma-based conditions, the AML patient cells exhibited significantly reduced sensitivity to 12% of the tested compounds, including topoisomerase II, B-cell chronic lymphocytic leukemia/lymphoma 2 (BCL2), and many tyrosine kinase inhibitors (TKIs). The loss of TKI sensitivity was most pronounced in patient samples harboring FLT3 or PDGFRB alterations. In contrast, the stroma-derived conditions enhanced sensitivity to Janus kinase (JAK) inhibitors. Increased cell viability and resistance to specific drug classes in the BM stroma-derived conditions was a result of activation of alternative signaling pathways mediated by factors secreted by BM stromal cells and involved a switch from BCL2 to BCLXL-dependent cell survival. Moreover, the JAK1/2 inhibitor ruxolitinib restored sensitivity to the BCL2 inhibitor venetoclax in AML patient cells ex vivo in different model systems and in vivo in an AML xenograft mouse model. These findings highlight the potential of JAK inhibitors to counteract stroma-induced resistance to BCL2 inhibitors in AML.
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- 2016
17. Abstract 3899: Discovery and clinical implementation of individualized therapies in acute myeloid leukemia based on ex vivo drug sensitivity testing and multi-omics profiling
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Siv Knappila, Matti Kankainen, Aino Palva, Piia Mikkonen, Maria Nurmi, Imre Vastrik, Mika Kontro, Karjalainen Riikka, Sanna Timonen, Satu Mustjoki, Krister Wennerberg, Disha Malani, Maija Wolf, Oscar Brück, Pekka Ellonen, Ashwini Kumar, Tero Aittokallio, Pirkko Mattila, Laura Turunen, Caroline A. Heckman, Bhagwan Yadav, Karoliina Laamanen, Kimmo Porkka, Sari Kytölä, Swapnil Potdar, Alun Parson, Poojitha Ojamies, Astrid Murumägi, Evgeny Kulesskiy, Bjorn Tore Gjersten, Olli Kallioniemi, Muntasir Mamun Majumder, Minna Suvela, Jani Saarela, Samuli Eldfors, Simon Anders, and Elina Lehtinen
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Trametinib ,Oncology ,Cobimetinib ,Drug ,Cancer Research ,medicine.medical_specialty ,business.industry ,media_common.quotation_subject ,Myeloid leukemia ,medicine.disease ,Drug repositioning ,chemistry.chemical_compound ,Leukemia ,chemistry ,Internal medicine ,medicine ,business ,Exome ,Ex vivo ,media_common - Abstract
Acute myeloid leukemia (AML) is a heterogeneous disease characterized by multiple molecular subtypes and lack of effective targeted therapies. Here, we performed extensive molecular profiling and ex vivo drug testing with 515 approved and emerging cancer drugs on 164 AML patient samples. The aim was to i) assign individualized treatment options to advanced AML patients in real time, ii) explore drug response patterns across the molecular subtypes of AML and iii) identify opportunities to repurpose existing and emerging cancer drugs. Bone marrow samples (n=164) from 129 consecutive AML patients and 17 healthy donors were studied from the Helsinki University Hospital and the Haukeland University Hospital, Bergen. Mononuclear cells were resuspended either in mononuclear cell medium (MCM) or stroma conditioned medium (CM) and tested for drug sensitivity and resistance as previously described (PMID: 24056683) and studied by exome and transcriptome sequencing. The study protocol allowed us to return data to the clinician for consideration of novel treatment options. For the meta-analysis of associations between drug responses and molecular and clinical parameters, Wilcoxon signed ranked test and logistic regression were applied. Clustering of all patient samples based on ex vivo drug response patterns in both media types identified 7 distinct functional groups of AML. For example, a subgroup of samples was highly resistant to chemotherapeutics and all targeted drugs except BCL-2 inhibitors. The differences in drug responses in the two media types highlighted the importance of assay conditions for ex vivo drug testing. Strong clustering of several drugs in the same drug classes was often observed as well as clustering across different classes, for example between BET (JQ1, I-BET151, birabresib) and MEK (trametinib, cobimetinib) inhibitors. About 24 percent of the FLT3 negative AML patients manifested strong ex vivo sensitivity to glucocorticoids, highlighting a potential drug repositioning opportunity in this subset of AML patients. Overall, we identified 320 significant associations between drugs and mutated driver genes including association between NPM1 mutation and sensitivity to JAK inhibitors. Altogether, targeted treatment opportunities were clinically tested in 25 occasions in chemorefractory AML patients. The tailored clinical therapy led to transient complete remission or leukemia free state in 36% (9/25) of these cases. In conclusion, we discovered and clinically implemented individualized therapeutic options for AML patients, which resulted in a 36% clinical responses in a non-randomized proof-of-concept study. The associations identified between ex-vivo drug response and driver mutations provided novel drug repositioning opportunities in specific molecular subtypes. Citation Format: Disha Malani, Ashwini Kumar, Bhagwan Yadav, Mika Kontro, Swapnil Potdar, Oscar Brück, Sari Kytölä, Jani Saarela, Samuli Eldfors, Poojitha Ojamies, Karjalainen Riikka, Muntasir Mamun Majumder, Imre Västrik, Pekka Ellonen, Matti Kankainen, Minna Suvela, Siv Knappila, Alun Parson, Aino Palva, Pirkko Mattila, Evgeny Kulesskiy1, Laura Turunen, Karoliina Laamanen, Elina Lehtinen, Piia Mikkonen, Maria Nurmi, Sanna Timonen, Astrid Murumägi, Bjorn Tore Gjersten, Satu Mustjoki, Tero Aittokallio, Krister Wennerberg, Simon Anders, Maija Wolf, Caroline Heckman, Kimmo Porkka, Olli Kallioniemi. Discovery and clinical implementation of individualized therapies in acute myeloid leukemia based on ex vivo drug sensitivity testing and multi-omics profiling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3899.
- Published
- 2018
18. MYO9B polymorphisms in multiple sclerosis
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Annette Bang Oturai, Hanne F. Harbo, Lars P. Ryder, Aarno Palotie, Elisabeth Gulowsen Celius, Mark J. Daly, Frida Lundmark, Pentti J. Tienari, Keijo Koivisto, Ilkka Rautakorpi, Minna Suvela, Mauri Reunanen, Leena Peltonen, Anu Kemppinen, Tuula Pirttilä, Janna Saarela, Jan Hillert, Irina Elovaara, Institute for Molecular Medicine Finland, Research Programs Unit, Research Programme for Molecular Neurology, Neurologian yksikkö, Finnish Genome Center (-2009), Genomics of Neurological and Neuropsychiatric Disorders, and Department of Medical and Clinical Genetics
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Multiple Sclerosis ,Genotype ,Short Report ,Single-nucleotide polymorphism ,312 Clinical medicine ,Myosins ,Biology ,medicine.disease_cause ,White People ,Autoimmunity ,03 medical and health sciences ,0302 clinical medicine ,Polymorphism (computer science) ,Genetic variation ,Genetics ,medicine ,Humans ,311 Basic medicine ,Family ,Gene ,Genetics (clinical) ,030304 developmental biology ,0303 health sciences ,Polymorphism, Genetic ,318 Medical biotechnology ,Multiple sclerosis ,Case-control study ,217 Medical engineering ,medicine.disease ,3. Good health ,Case-Control Studies ,Immunology ,030211 gastroenterology & hepatology ,118 Biological sciences - Abstract
"Single-nucleotide polymorphisms (SNPs) in the 30 region of myosin IXB (MYO9B) gene have recently been reported to associate with different inflammatory or autoimmune diseases. We monitored for the association of MYO9B variants to multiple sclerosis (MS) in four Northern European populations. First, 18 SNPs including 6 SNPs with previous evidence for association to immune disorders, were tested in 730 Finnish MS families, but no linkage or family-based association was observed. To ensure the power to detect variants with a modest effect size, we further analyzed 10 variants in 899 Finnish cases and 1325 controls, and in a total of 1521 cases and 1476 controls from Denmark, Norway and Sweden, but found no association. Our results thereby do not support a major function of the tested MYO9B variants in MS. European Journal of Human Genetics (2009) 17, 840-843; doi: 10.1038/ejhg.2008.251; published online 14 January 2009" "Single-nucleotide polymorphisms (SNPs) in the 30 region of myosin IXB (MYO9B) gene have recently been reported to associate with different inflammatory or autoimmune diseases. We monitored for the association of MYO9B variants to multiple sclerosis (MS) in four Northern European populations. First, 18 SNPs including 6 SNPs with previous evidence for association to immune disorders, were tested in 730 Finnish MS families, but no linkage or family-based association was observed. To ensure the power to detect variants with a modest effect size, we further analyzed 10 variants in 899 Finnish cases and 1325 controls, and in a total of 1521 cases and 1476 controls from Denmark, Norway and Sweden, but found no association. Our results thereby do not support a major function of the tested MYO9B variants in MS. European Journal of Human Genetics (2009) 17, 840-843; doi: 10.1038/ejhg.2008.251; published online 14 January 2009" "Single-nucleotide polymorphisms (SNPs) in the 30 region of myosin IXB (MYO9B) gene have recently been reported to associate with different inflammatory or autoimmune diseases. We monitored for the association of MYO9B variants to multiple sclerosis (MS) in four Northern European populations. First, 18 SNPs including 6 SNPs with previous evidence for association to immune disorders, were tested in 730 Finnish MS families, but no linkage or family-based association was observed. To ensure the power to detect variants with a modest effect size, we further analyzed 10 variants in 899 Finnish cases and 1325 controls, and in a total of 1521 cases and 1476 controls from Denmark, Norway and Sweden, but found no association. Our results thereby do not support a major function of the tested MYO9B variants in MS. European Journal of Human Genetics (2009) 17, 840-843; doi: 10.1038/ejhg.2008.251; published online 14 January 2009"
- Published
- 2009
19. Increased Mutational Burden and Alterations to DNA Damage Repair Genes are Associated With Poor Prognosis and Sensitivity to PI3K-mTOR Inhibitors in Multiple Myeloma
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Muntasir Mamun Majumder, Ashwini Kumar, Minna Suvela, Juha Lievonen, Pekka Anttila, Raija Silvennoinen, David Tamborero, Esa Jantunen, Caroline A. Heckman, Samuli Eldfors, Alun Parsons, and Kimmo Porkka
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Cancer Research ,Poor prognosis ,Pathology ,medicine.medical_specialty ,business.industry ,Hematology ,DNA Damage Repair ,medicine.disease ,Discovery and development of mTOR inhibitors ,Oncology ,Cancer research ,Medicine ,business ,Gene ,PI3K/AKT/mTOR pathway ,Multiple myeloma - Published
- 2017
20. DNA Damage Repair Pathway Alterations in Multiple Myeloma Predict Poor Prognosis, but Correlate with Sensitivity to IGF1R-PI3K-mTOR and HDAC Inhibitors
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Juha Lievonen, Pekka Anttila, Muntasir Mamun Majumder, Esa Jantunen, Caroline A. Heckman, Ashwini Kumar, Kimmo Porkka, David Tamborero, Samuli Eldfors, Raija Silvennoinen, Minna Suvela, and Alun Parsons
- Subjects
Neuroblastoma RAS viral oncogene homolog ,Immunology ,Drug resistance ,Signal transduction inhibitor ,Biology ,medicine.disease_cause ,Bioinformatics ,Biochemistry ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,medicine ,Multiple myeloma ,030304 developmental biology ,0303 health sciences ,Navitoclax ,MEK inhibitor ,Cell Biology ,Hematology ,medicine.disease ,3. Good health ,Multiple drug resistance ,chemistry ,Cancer research ,KRAS ,030215 immunology - Abstract
Introduction Although several novel drugs have recently been approved or are in development for multiple myeloma (MM), there are few molecular indicators to guide treatment selection. In addition, the impact of recurrent myeloma alterations on drug response is often unclear. To address these limitations and elucidate genotype to phenotype relationships in myeloma, we comprehensively analyzed 100 MM samples and compared genomic, transcriptomic, and cytogenetic information to ex vivo drug response profiles and clinical outcome of individual MM patients. Our results reveal novel insights on i) drug response and resistance mechanisms, ii) biomarkers for drug response, and iii) potential treatment combinations to overcome drug resistance. Methods Bone marrow aspirates were collected from MM patients (n=100; newly diagnosed n=34; relapsed/refractory n=66) and healthy individuals (n=14). CD138+ plasma cells were enriched from the mononuclear cell fraction by immunomagnetic bead selection. Cells were screened against 142 oncology drugs tested in a 10,000-fold concentration range and 12 different drug combinations Somatic alterations were identified by exome sequencing of DNA from CD138+ cells and skin biopsies from each patient (n=85). RNA sequencing derived read counts from CD138+ cells of MM samples (n=67) were used for differential gene expression. Karyotype was determined by fluorescence in situ hybridization. Results For most drugs tested, no significant difference in response was observed between samples from newly diagnosed and relapsed refractory patients except for signal transduction inhibitors targeting IGF1R-PI3K-mTOR, MAPK and HSP90. A positive correlation was observed between mutational burden and sensitivity to targeted therapies. The median number of somatic alterations was 118 in sensitive compared to 50 in resistant samples. 14% of the samples exhibited a multidrug resistant phenotype and were resistant to proteasome inhibitors, immunomodulatory drugs and glucocorticoids. 30% of the resistant samples were from del(17p) patients. In addition, gene expression analysis revealed elevated expression of cell adhesion and integrin signaling molecules including ITGB3, ITGA2B, VCL, TLN1, MMP8, MMP9, plus ABCC3, which encodes a transporter protein shown to be associated with multidrug resistance. A combination of the protein kinase C inhibitor bryostatin-1 and pan-BCL2 inhibitor navitoclax was highly effective against the resistant samples. 26% of the patient samples harbored mutations in genes involved in DNA damage repair signaling, namely TP53, TP73, ATM and BAX, in a mutually exclusive pattern. In addition, patients with these mutations had a high relapse rate and poor overall survival (HR=7.2,95%CI 3.2-16.08). Interestingly, CD138+ cells from these patients showed activation of IGF1R-PI3K-mTOR signaling and were highly susceptible to inhibitors targeting this signaling axis. These samples were also highly sensitive to HDAC inhibitors. While no strong correlation between RAS pathway mutations (NRAS, KRAS, NF1, BRAF) and MEK inhibitor sensitivity was observed, samples with clonal RAS mutations tended to be more sensitive to MEK inhibitors compared to samples with subclonal mutations. Summary Our results suggest that drug resistance in myeloma may occur either via accumulation of somatic alterations or via cell adhesion mediated cytoprotection. Driver alterations in DNA damage signaling pathways were found to contribute to poor prognosis, but samples with these mutations showed enhanced sensitivity to IGF1R-PI3K-mTOR and HDAC inhibitors. Using genomic and transcriptomic data we identified molecular events that may shape the drug response landscape and found drug combinations that can overcome resistance mechanisms. Our results demonstrate that molecular information and ex vivo drug profiling may be useful to develop tailored treatment strategies and guide treatment decision, especially for relapsed/refractory myeloma patients. Disclosures Silvennoinen: Sanofi: Honoraria, Other: Lecture fee; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Lecture fee; Janssen: Honoraria, Research Funding; Celgene: Honoraria, Research Funding. Porkka:Bristol-Myers Squibb: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Novartis: Honoraria, Research Funding. Heckman:Pfizer: Research Funding; Celgene: Research Funding.
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- 2016
21. Stromal-Derived Factors Modulate Ex Vivo Drug Responses of Primary Acute Myeloid Leukemia Cells
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Muntasir Mamun Majumder, Imre Vastrik, Tero Aittokallio, Bhagwan Yadav, Jing Tang, Jonathan Knowles, Olli Kallioniemi, Minna Suvela, David Tamborero, Riikka Karjalainen, Dmitrii Bychkov, Caroline A. Heckman, Bjørn Tore Gjertsen, Alun Parsons, Tea Pemovska, Kimmo Porkka, Mika Kontro, and Krister Wennerberg
- Subjects
Drug ,Cancer Research ,Primary (chemistry) ,Stromal cell ,business.industry ,media_common.quotation_subject ,Myeloid leukemia ,Hematology ,Oncology ,Cancer research ,Medicine ,business ,Ex vivo ,media_common - Published
- 2015
22. Stratification of Multiple Myeloma Patients Based on Ex Vivo Drug Sensitivity and Identification of New Treatments for Patients with High-Risk Relapsed/Refractory Disease
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Juha Lievonen, Riikka Karjalainen, Heikki Kuusanmäki, Samuli Eldfors, Alun Parsons, Muntasir Mamun Majumder, Pekka Anttila, Raija Silvennoinen, Caroline A. Heckman, David Tamborero, Kimmo Porkka, and Minna Suvela
- Subjects
Neuroblastoma RAS viral oncogene homolog ,Oncology ,medicine.medical_specialty ,Immunology ,Pharmacology ,medicine.disease_cause ,Biochemistry ,03 medical and health sciences ,0302 clinical medicine ,In vivo ,Internal medicine ,medicine ,Vemurafenib ,Multiple myeloma ,030304 developmental biology ,0303 health sciences ,business.industry ,Melanoma ,Cell Biology ,Hematology ,medicine.disease ,3. Good health ,Leukemia ,KRAS ,business ,Ex vivo ,030215 immunology ,medicine.drug - Abstract
Introduction Response to treatment for multiple myeloma (MM) patients is variable and often unpredictable, which may be attributed to the heterogeneous genomic landscape of the disease. However, the effect of recurrent molecular alterations on drug response is unclear. To address this, we systematically profiled 50 samples from 43 patients to assess ex vivo sensitivity to 308 anti-cancer drugs including standard of care and investigational drugs, with results correlated to genomic alterations. Our results reveal novel insights about patient stratification, therapies for high-risk (HR) patients, signaling pathway aberrations and ex-vivo-in-vivo correlation. Methods Bone marrow (BM) aspirates (n=50) were collected from MM patients (newly diagnosed n=17; relapsed/refractory n=33) and healthy individuals (n=8). CD138+ plasma cells were enriched by Ficoll separation followed by immunomagnetic bead selection. Cells were screened against 308 oncology drugs tested in a 10,000-fold concentration range. Drug sensitivity scores were calculated based on the normalized area under the dose response curve (Yadav et al, Sci Reports, 2014). MM selective responses were determined by comparing data from MM patients with those of healthy BM cells. Clustering of drug sensitivity profiles was performed using unsupervised hierarchical ward-linkage clustering with Spearman and Manhattan distance measures of drug and sample profiles. Somatic alterations were identified by exome sequencing of DNA from CD138+ cells and skin biopsies from each patient, while cytogenetics were determined by fluorescence in situ hybridization. Results Comparison of the ex vivo chemosensitive profiles of plasma cells resulted in stratification of patients into four distinct subgroups that were highly sensitive (Group I), sensitive (Group II), resistant (Group III) or highly resistant (Group IV) to the panel of drugs tested. Many of the drug responses were specific for CD138+ cells with little effect on CD138- cells from the same patient or healthy BM controls. We generated a drug activity profile for the individual drugs correlating sensitivity to recurrent alterations including mutations to KRAS, DIS3, NRAS, TP53, FAM46C, and cytogenetic alterations del(17p), t(4;14), t(14;16), t(11;14), t(14;20), +1q and -13. Cells from HR patients with del(17p) exhibited the most resistant profiles (enriched in Groups III and IV), but were sensitive to some drugs including HDAC and BCL2 inhibitors. Samples from patients with t(4;14) were primarily in Group II and very sensitive to IMiDs, proteasome inhibitors and several targeted drugs. Along with known recurrently mutated genes in myeloma, somatic mutations were identified in genes involved in several critical signaling pathways including DNA damage response, IGF1R-PI3K-AKT, MAPK, glucocorticoid receptor signaling and NF-κB signaling pathways. The predicted impact of these mutations on the activity of the pathways often corresponded to the drug response. For example, all samples bearing NF1 (DSS=21±7.9) and 67% with NRAS (DSS=15±4.35) mutations showed higher sensitivity to MEK inhibitors compared to healthy controls (DSS=5±.21). However, sensitivity was less predictable for KRAS mutants with modest response only in 47% samples (DSS=7±2.14) . One sample bearing the activating V600E mutation to BRAF showed no sensitivity to vemurafenib, which otherwise has good activity towards V600E mutated melanoma and hairy-cell leukemia. Comparison of the chemosensitive subgroups with survival showed patients in Groups I and IV had high relapse rate and poor overall survival. The ex vivo drug sensitivity results were used to decide treatment for three HR patients with results showing good ex vivo -in vivo correlation. Summary Our initial results suggest that ex vivo drug testing and molecular profiling of MM patients aids stratification. Grouping of patients based on their ex vivo chemosensitive profile proved extremely informative to predict clinical phenotype and identify responders from non-responders. While some molecular markers could be used to predict drug response, others were less predictive. Nevertheless, ex vivo drug testing identified active drugs, particularly for HR and relapsed/refractory patients, and is a powerful method to determine treatment for this group of patients. Disclosures Silvennoinen: Genzyme: Honoraria; Sanofi: Honoraria; Janssen: Research Funding; Celgene: Research Funding; Research Committee of the Kuopio University Hospital Catchment Area for State Research Funding, project 5101424, Kuopio, Finland: Research Funding; Amgen: Consultancy, Honoraria. Porkka:Bristol-Myers Squibb: Honoraria; Celgene: Honoraria; Novartis: Honoraria; Pfizer: Honoraria. Heckman:Celgene: Honoraria, Research Funding.
- Published
- 2015
23. JAK1/2 and BCL2 Inhibitors Synergize to Counter-Act Bone Marrow Stromal Cell-Induced Protection of AML
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Riikka Karjalalainen, Tea Pemovska, Muntasir Mamun Majumder, Bhagwan Yadav, Jing Tang, Mika Kontro, Bjørn Gjertsen, Alun Parsons, Minna Suvela, Kimmo Porkka, Tero Aittokallio, Olli Kallioniemi, Krister Wennerberg, Jonathan Knowles, and Caroline Heckman
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Immunology ,Cell Biology ,Hematology ,Biochemistry - Abstract
BACKGROUND AND OBJECTIVES: The bone marrow (BM) microenvironment supports the survival of leukemic cells and influences their response to therapeutic agents by promoting drug tolerance and resistance. Novel therapeutic strategies are therefore needed that can override the BM mediated protection of AML cells in patients undergoing drug treatment. To address this we used a high-throughput drug screening method to identify novel drug combinations to reverse stromal-induced cytoprotection against the BCL2 antagonist venetoclax in primary AML samples. METHODS: Sensitivity of mononuclear cells collected from 18 AML BM aspirates or peripheral blood samples to a range of BCL2 inhibitors and tyrosine kinase inhibitors (TKIs) was assessed either in mononuclear cell medium (MCM, Promocell) or in a 25% HS-5 stromal cell-conditioned medium plus 75% RPMI medium mix (CM) to mimic cytoprotective bone marrow conditions. Cell viability was measured after 72 h and dose response curves generated for each tested drug. Drug sensitivity scores were calculated based on the area under the dose response curve. For the drug combination studies single agents (venetoclax, WEHI-539, ruxolitinib) were added simultaneously at fixed concentrations to AML cells and incubated for 72 h either in the MCM or CM medium. Cell viability was measured using the CellTiter-Glo assay. The expression of BCL2 genes was measured by qPCR after incubating the AML patient cells in either MCM or CM for 48 h. RESULTS: Incubation of primary AML cells in the CM culture condition led to reduced sensitivity to BCL2 family inhibitors, suggesting that stromal-derived factors in the CM promote cytoprotection. This effect was particularly pronounced for the selective BCL2 inhibitor venetoclax, where the CM-induced loss of sensitivity coincided with decreased BCL2 expression and increased BCL2L1 expression. In contrast, JAK inhibitors showed improved efficacy in CM compared to MCM culture conditions. To determine if the protective effects of CM stromal-like conditions against venetoclax could be diminished, the drug was tested in combination with the JAK1/2 inhibitor ruxolitinib using AML cells cultured in MCM or CM. When tested on AML cells from 4 patients with the FLT3-ITD alteration, we found that ruxolitinib rescued the sensitivity of venetoclax in leukemic cells in the presence of CM and the combination of two drugs exhibited synergistic effects in this setting. The combinatorial activity, however, was not recapitulated in the MCM condition. Since CM was found to induce BCL2L1 expression, venetoclax was also tested in combination with a BCLXLspecific inhibitor WEHI-539. Analogously to the ruxolitinib-venetoclax combination, synergistic activity between venetoclax and WEHI-539 was observed towards leukemic cells in the presence of CM. CONCLUSIONS: By applying a functional, drug-based approach to understand microenvironment-induced mechanisms of drug resistance in AML, we found that the activity of the selective BCL2 inhibitor venetoclax towards AML cells is adversely affected in stromal-based conditions, while JAK inhibitors, in contrast, exhibit increased efficacy in these conditions. Our results suggest stroma-derived cytokines induce JAK-STAT signaling in AML cells, which results in increased BCL2L1 expression and drives resistance to venetoclax. However, blocking JAK1/2 with ruxolitinib restores the sensitivity of AML cells to venetoclax. We found that JAK1/2 inhibitors such as ruxolitinib can act synergistically with BCL2/BCLXL inhibitors, suggesting clinically useful combination treatments. Disclosures Gjertsen: BerGenBio AS: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Boehringer Ingelheim: Membership on an entity's Board of Directors or advisory committees; Kinn Therapeutics AS: Equity Ownership. Porkka:Pfizer: Honoraria, Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; Celgene: Honoraria, Research Funding; Novartis: Honoraria, Research Funding. Kallioniemi:Vysis-Abbot: Patents & Royalties; Medisapiens: Membership on an entity's Board of Directors or advisory committees; IMI-Project Predect: Research Funding; Roche: Research Funding; Pfizer: Research Funding. Wennerberg:Pfizer: Research Funding. Heckman:Celgene: Honoraria, Research Funding.
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- 2015
24. Use of a genetic isolate to identify rare disease variants: C7 on 5p associated with MS
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Hanne F. Harbo, Leena Peltonen, Seppo Meri, Denis Bronnikov, Markku Viander, Irina Elovaara, Shaun Purcell, Philip L. De Jager, Åslaug R. Lorentzen, Aarno Palotie, Eveliina Jakkula, Minna Suvela, David A. Hafler, Jan Hillert, Pentti J. Tienari, Mauri Reunanen, Tuula Pirttilä, Frida Lundmark, Keijo Koivisto, Janna Saarela, Suvi P. Kallio, and Mark J. Daly
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Multiple Sclerosis ,Population ,Genome-wide association study ,Locus (genetics) ,Single-nucleotide polymorphism ,Biology ,03 medical and health sciences ,0302 clinical medicine ,Genetics ,Humans ,Allele ,education ,Molecular Biology ,Genetics (clinical) ,Finland ,030304 developmental biology ,Genetic association ,0303 health sciences ,education.field_of_study ,Haplotype ,Association Studies Articles ,General Medicine ,Complement C7 ,Haplotypes ,Case-Control Studies ,Chromosomes, Human, Pair 5 ,030217 neurology & neurosurgery ,Founder effect ,Genome-Wide Association Study - Abstract
Large case–control genome-wide association studies primarily expose common variants contributing to disease pathogenesis with modest effects. Thus, alternative strategies are needed to tackle rare, possibly more penetrant alleles. One strategy is to use special populations with a founder effect and isolation, resulting in allelic enrichment. For multiple sclerosis such a unique setting is reported in Southern Ostrobothnia in Finland, where the prevalence and familial occurrence of multiple sclerosis (MS) are exceptionally high. Here, we have studied one of the best replicated MS loci, 5p, and monitored for haplotypes shared among 72 regional MS cases, the majority of which are genealogically distantly related. The haplotype analysis over the 45 Mb region, covering the linkage peak identified in Finnish MS families, revealed only modest association at IL7R (P = 0.04), recently implicated in MS, whereas most significant association was found with one haplotype covering the C7-FLJ40243 locus (P = 0.0001), 5.1 Mb centromeric of IL7R. The finding was validated in an independent sample from the isolate and resulted in an odds ratio of 2.73 (P = 0.000003) in the combined data set. The identified relatively rare risk haplotype contains C7 (complement component 7), an important player of the innate immune system. Suggestive association with alleles of the region was seen also in more heterogeneous populations. Interestingly, also the complement activity correlated with the identified risk haplotype. These results suggest that the MS predisposing locus on 5p is more complex than assumed and exemplify power of population isolates in the identification of rare disease alleles.
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- 2009
25. No evidence for shared etiology in two demyelinative disorders, MS and PLOSL
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Keijo Koivisto, Irina Elovaara, Anna-Maija Kristiina Sulonen, Pentti J. Tienari, Aarno Palotie, Leena Peltonen, Janna Saarela, Mauri Reunanen, Tuula Pirttilä, Minna Suvela, Pekka Ellonen, and Suvi P. Kallio
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Linkage disequilibrium ,Multiple Sclerosis ,Chromosomal Proteins, Non-Histone ,Immunology ,DNA-Directed DNA Polymerase ,Biology ,medicine.disease_cause ,Polymorphism, Single Nucleotide ,Article ,Linkage Disequilibrium ,03 medical and health sciences ,0302 clinical medicine ,Gene Frequency ,Demyelinating disease ,medicine ,Genetic predisposition ,Immunology and Allergy ,Humans ,Genetic Predisposition to Disease ,Allele ,Receptors, Immunologic ,Allele frequency ,Finland ,030304 developmental biology ,Adaptor Proteins, Signal Transducing ,Genetics ,0303 health sciences ,Mutation ,Brain Diseases ,Membrane Glycoproteins ,TREM2 ,Multiple sclerosis ,Chromosome Mapping ,Membrane Proteins ,medicine.disease ,Neurology ,Neurology (clinical) ,Sequence Analysis ,030217 neurology & neurosurgery ,Demyelinating Diseases - Abstract
Loss-of-function mutations of DAP12 and TREM2 cause a recessively inherited disease PLOSL, manifesting in brain white matter. The genes of the DAP12-TREM2 signaling receptor are located on 19q13.12 and 6p21.1, to which linkage has been observed also in families affected by another immune-mediated demyelinating disease, MS. We have tested if allelic variation in DAP12 or TREM2 predisposes also to MS by monitoring carrier frequency of the Finnish PLOSL mutation in Finnish MS cases and by studying DAP12 and TREM2 in MS by linkage and association. To conclude, the DAP12-TREM2 complex unlikely has a role in genetic susceptibility of MS.
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- 2008
26. Human herpesvirus-6 and -7 DNA in cerebrospinal fluid of facial palsy patients
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Mervi Kanerva, Antti Vaheri, Anne J Jääskeläinen, Minna Suvela, Anne Pitkäranta, and Heli Piiparinen
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Adult ,Male ,Adolescent ,viruses ,Herpesvirus 6, Human ,Facial Paralysis ,Roseolovirus Infections ,Herpesvirus 7, Human ,medicine.disease_cause ,Polymerase Chain Reaction ,Virus ,Herpesviridae ,law.invention ,Cerebrospinal fluid ,law ,medicine ,Humans ,Child ,Polymerase chain reaction ,Aged ,Retrospective Studies ,biology ,virus diseases ,Cytomegalovirus ,General Medicine ,Middle Aged ,biology.organism_classification ,medicine.disease ,Microarray Analysis ,Virology ,Facial paralysis ,Herpes simplex virus ,Otorhinolaryngology ,Child, Preschool ,DNA, Viral ,Human herpesvirus 6 ,Female ,Follow-Up Studies - Abstract
Finding human herpesvirus (HHV)-7 and dual HHV-6A and -6B DNA in cerebrospinal fluid (CSF) of two facial palsy (FP) patients is intriguing but does not allow etiologic conclusions as such. HHV-6 or -7 DNA was revealed in 10% of the CSF samples tested from 70 immunocompetent adolescents and adults; a highly unusual result. How these findings are associated with the diseases they accompany remains to be defined.To determine whether herpes simplex virus (HSV)-1 and -2, varicella-zoster virus (VZV), HHV-6A, -6B, and -7, Epstein-Barr virus (EBV), and cytomegalovirus (CMV) DNA could be found in CSF of FP patients or controls.In all, 33 peripheral FP patients (26 idiopathic, 5 with herpesvirus infection, 1 puerperal, 1 Melkersson-Rosenthal syndrome) (34 CSF samples) and 36 controls (16 nonidiopathic FP, 7 hearing loss, 6 vertigo, 5 headache, 2 other) previously tested for HSV-1, VZV, and HHV-6 DNA by polymerase chain reaction (PCR) were tested with highly sensitive multiplex-PCR and an oligonucleotide microarray method.One FP patient had HHV-7 DNA and another had HHV-6A and -6B DNA simultaneously. In the control group, one HHV-7, one HHV-6A, and three HHV-6B DNA-positive specimens were found.
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- 2008
27. Identification of precision treatment strategies for high risk multiple myeloma by ex vivo drug sensitivity testing
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David Tamborero, Samuli Eldfors, Mamun Majumder, J. Lievonen, Raija Silvennoinen, Caroline A. Heckman, Minna Suvela, Riikka Karjalainen, Alun Parsons, Heikki Kuusanmäki, Pekka Anttila, and Kimmo Porkka
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Drug ,Oncology ,0303 health sciences ,Cancer Research ,medicine.medical_specialty ,business.industry ,media_common.quotation_subject ,Hematology ,Pharmacology ,medicine.disease ,03 medical and health sciences ,0302 clinical medicine ,Sensitivity testing ,Internal medicine ,medicine ,Treatment strategy ,Identification (biology) ,business ,030217 neurology & neurosurgery ,Ex vivo ,Multiple myeloma ,030304 developmental biology ,media_common - Published
- 2015
28. The enhancement of homogenous mass extension reaction: comparison of two enzymes
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Kari Tuomainen, Kaisa Silander, Päivi Tikka-Kleemola, Olev Kahre, Maija Wessman, Aarno Palotie, Eija Hämäläinen, Minna Suvela, and Agu Artma
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Quality Control ,DNA polymerase ,Cost-Benefit Analysis ,DNA Mutational Analysis ,Single-nucleotide polymorphism ,DNA-Directed DNA Polymerase ,Molecular Inversion Probe ,Polymerase Chain Reaction ,Polymorphism, Single Nucleotide ,Mass Spectrometry ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Genotype ,SNP ,Humans ,Molecular Biology ,Genotyping ,030304 developmental biology ,Genetics ,0303 health sciences ,biology ,Cell Biology ,SNP genotyping ,Molecular Weight ,chemistry ,030220 oncology & carcinogenesis ,biology.protein ,DNA - Abstract
Reliable and efficient PCR and extension reactions using standardized procedures are key elements for successful single nucleotide polymorphism (SNP) genotyping projects. To improve the cost efficiency and overall performance of SNP genotyping we evaluated two commercial thermostable DNA polymerases used for the extension reaction in the homogeneous mass extension MassARRAY genotyping system. The aim was to study whether the quality, accuracy, and expenses of a new TERMIPol® DNA polymerase are competitive to the commonly used ThermoSequenase® DNA polymerase. We compared the enzymes by testing 96 SNPs genotyped for DNA samples of 31 unrelated individuals and one water control. The success rates, congruence between the genotypes and completeness of extension reactions support the use of TERMIPol®, especially when the amplification of the higher mass allele is difficult. Further, using TERMIPol® enabled successful genotyping (>93%) of several SNPs that failed (
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- 2006
29. Identification of Dual PI3K/mTOR and BCL2 Inhibitors for the Treatment of High Risk Multiple Myeloma
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Pekka Anttila, Kimmo Porkka, Heikki Kuusanmäki, Muntasir Mamun Majumder, Raija Silvennoinen, Alun Parsons, Marjaana Säily, Samuli Eldfors, Caroline A. Heckman, Minna Suvela, David Tamborero, Riikka Karjalainen, and Juha Lievonen
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Oncology ,Monosomy ,medicine.medical_specialty ,Immunology ,Plasma cell ,Biochemistry ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Internal medicine ,medicine ,Multiple myeloma ,030304 developmental biology ,0303 health sciences ,Navitoclax ,business.industry ,Venetoclax ,Cell Biology ,Hematology ,medicine.disease ,3. Good health ,Leukemia ,medicine.anatomical_structure ,chemistry ,Bone marrow ,business ,Ex vivo ,030215 immunology - Abstract
Introduction Multiple myeloma (MM) is an incurable malignant plasma cell disease with the highest incidence occurring at 65-70 years of age while 10% of patients are diagnosed below 55 years of age. The International Myeloma Working Group recently proposed new risk stratification standards for MM patients: high-risk (HR), standard (SR) and low-risk (LR) groups (Leukemia 2014, 28, 269−77). Although a median overall survival of LR patients is > 10 years from the diagnosis, new drugs and therapeutic innovations are urgently needed for HR patients (20%) who have a median overall survival of only two years. To identify new treatment options for MM patients, we compared ex vivo drug sensitivity data from primary CD138+ cells to standard risk stratification markers. Ex vivo responses indicated a number of investigational drugs as potential novel options for HR MM patients with links to risk markers. Methods Bone marrow aspirates were collected from newly diagnosed (n=14) and relapsed/refractory (n=21) MM patients. Cytogenetics were determined by fluorescence in situ hybridization (FISH) and the patients stratified based on the presence or absence of adverse FISH markers (t(4;14) and 17p del). Plasma cells (CD138+) were enriched from freshly isolated bone marrow samples and exome sequencing performed using DNA extracted from the CD138+ cells and matched skin biopsies. Ex vivo drug sensitivity was assessed by measuring the viability of the cells after 3-day incubation with 306 different oncology drugs in a 10,000-fold concentration range. Drug sensitivity scores were calculated based on the normalized area under the dose response curve (Scientific Reports 2014, 4, 5193) and select sensitivities determined by comparing results to healthy bone marrow cells. Based on drug sensitivities, the patients were classified in four different groups (sensitive, moderately sensitive, resistant and highly resistant). Results Of the 35 patients included in this study, 11 were classified as HR (31%) and 24 as SR/LR (69%). In the HR group 6/11 (55%) had t(4;14) and 5/11 patients (45%) had 17p13 del. In the SR/LR group common abnormalities included 13 monosomy/13q del (10/24), 1q gain (10/24) and K/NRAS mutation (11/24). Within the HR group, other co-occurring abnormalities included 1q gain (9/11), 13 monosomy/13q del (6/11), K/NRAS mutation (5/11), and TP53 mutation (2/11). Based on overall ex vivo drug sensitivity profiles of all patients, the majority of HR patients were classified as moderately sensitive (8/11; 73%) while SR/LR patients had diverse responses from sensitive to highly resistant. In the HR group, the highest select sensitivities were to BH3 mimetics and PI3K/mTOR inhibitors. While the t(4;14) is predicted to lead to upregulation and increased activity of the FGFR3, which could be targeted by FGFR inhibitors, none of the t(4;14) samples showed sensitivity to these drugs. However, with the exception of one t(4;14) sample, the rest all showed good sensitivity to dual PI3K/mTOR inhibitors, but not to rapalogs, suggesting that inhibition of PI3K and the mTORC1/2 complexes is required to inhibit t(4;14) cell growth rather than mTORC1 alone. Of the 17p del patients, 3/5 were classified as moderately sensitive, 1/5 sensitive and 1/5 highly resistant based on ex vivo drug response of CD138+ cells. All showed select sensitivity to BH3 mimetics/BCL2 inhibitors (navitoclax/ABT-263 and venetoclax/ABT-199/GDC-0199), while response to other drugs varied. Therefore, blocking cell survival signaling is likely essential for this group of HR MM patients. Conclusion By assessing the ex vivo sensitivity of primary plasma cells to a large collection of oncology drugs and comparing these data to standard risk stratification markers for MM, we have been able to identify potential new treatment options for high risk MM patients including dual PI3K/mTOR and BCL2- inhibitors. Although a larger cohort of patients is required to support the correlation between specific drug sensitivities and risk markers, these preliminary data indicate that currently used risk markers may be useful to predict the use of novel treatments. Disclosures Silvennoinen: Janssen-Cilag: Research Funding; Celgene: Research Funding; Janssen-Cilag: Honoraria; Sanofi: Honoraria; Celgene: Honoraria. Porkka:BMS: Honoraria; BMS: Research Funding; Novartis: Honoraria; Novartis: Research Funding; Pfizer: Research Funding. Heckman:Celgene: Research Funding.
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- 2014
30. Integration of Ex Vivo Drug Testing and in-Depth Molecular Profiling Reveals Oncogenic Signaling Pathways and Novel Therapeutic Strategies for Multiple Myeloma
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Caroline A. Heckman, Pekka Antilla, Bhagwan Yadav, Muntasir Mamun Majumder, Riikka Karjalainen, Marjanna Säily, Samuli Eldfors, Juha Lievonen, Krister Wennerberg, Kimmo Porkka, Heikki Kuusanmäki, Alun Parsons, Raija Silvennoinen, Olli Kallioniemi, Minna Suvela, and David Tamborero
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Drug ,media_common.quotation_subject ,Immunology ,Pharmacology ,Biochemistry ,Transcriptome ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,In vivo ,Medicine ,Multiple myeloma ,030304 developmental biology ,media_common ,0303 health sciences ,Navitoclax ,business.industry ,MEK inhibitor ,Cell Biology ,Hematology ,medicine.disease ,3. Good health ,Gene expression profiling ,chemistry ,business ,Ex vivo ,030215 immunology - Abstract
Introduction New drugs have improved survival for multiple myeloma (MM) patients, however, patient outcome remains highly variable, unpredictable and often very poor. To identify novel treatments and potential biomarkers, we applied high throughput ex vivo drug sensitivity testing combined with exome and transcriptome sequencing to samples collected from newly diagnosed and relapsed MM patients. Integration of results from the different platforms indicated several oncogenic signaling pathways driving drug response and highlighted the importance of a multi-targeted approach for treatment. Methods Bone marrow (BM) aspirates (n=48) were collected from MM patients (newly diagnosed n=14; relapsed/refractory n=26) and healthy individuals (n=8). CD138+ plasma cells were enriched by Ficoll separation followed by immunomagnetic bead selection. Cells were screened against 306 oncology drugs with the drugs tested in a 10,000-fold concentration range. Drug sensitivity scores were calculated based on the normalized area under the dose response curve (Yadav et al, Sci Reports, 2014). Importantly, MM selective responses were determined by comparing data from MM patients with those of healthy BM cells. Clustering of drug sensitivity profiles was performed using unsupervised hierarchical ward-linkage clustering with Spearman and Manhattan distance measures of drug and sample profiles. Somatic mutations were identified by exome sequencing of DNA from CD138+ cells and skin biopies from each patient, while gene expression profiles were derived from RNA sequencing of CD138+ cells. Results Cluster analysis of drug response profiles segregated the samples into four MM specific groups (Figure). Group I patients (n=12) were highly sensitive to many drugs, including several signal transduction inhibitors such as those targeting PI3K-AKT, MAPK and IGF pathways, as well as HSP90 and BCL2 inhibitors plus epigenetic/chromatin modifiers such as BET and HDAC inhibitors. Group II (n=15) showed a more modest response profile and were moderately sensitive to signal transduction inhibitors and epigenetic modifiers. Group III (n=9) were largely insensitive to most drugs in the panel except for BCL2 and proteasome inhibitors, while group IV (n=3) were resistant to all drugs except BCL2 inhibitors. Many samples were selectively sensitive to navitoclax (55%), dual PI3K/mTOR inhibitors (45%) and aminopeptidase inhibitors (20%), which had little effect on healthy control or MM CD138- cells. Only 33% of the samples responded to glucocorticoids. The majority of samples including healthy BM controls were sensitive to proteasome and CDK inhibitors, suggesting low selective cytotoxicity. However, drug sensitivity profiles of healthy control and CD138- cell populations were distinct from MM CD138+ samples indicating that observed CD138+ drug responses were specific for malignant plasma cells. In addition, we observed that drugs with overlapping target profiles tended to cluster together, indicating sample responses were similar to related drugs. Diagnostic and relapse samples were spread across the different response groups. Samples with mutations to genes involved in PI3K and NF-κB signaling tended to cluster in group I, while most samples with t(4;14) fell in Group II. Samples with RAS mutations were present in all response groups and no correlation with MEK inhibitor sensitivity was observed. 17p deletion samples were also found in all response groups, however, those with additional TP53 mutation tended to have increased drug sensitivity. Summary Our results indicate that PI3K/mTOR, MAPK, IGF1R, NF-κB and cell survival (e.g. BCL2, BCLXL) signaling are important pathways mediating MM ex vivo drug response. This matched with genomic and transcriptomic data, which identified alterations of genes involved in these pathways. Although additional work is needed to correlate ex vivo drug sensitivity with in vivo treatment response, our initial results suggest the possibility that MM patients could be subjected to stratified treatment based on combined ex vivo drug testing and molecular profiling. In addition, these results highlight the multiple signaling pathways active in MM and emphasize the need for improved combination strategies for treatment. Figure: Subgrouping of MM patient samples (I-IV) based on selective drug response profiles. H/D/R denotes healthy, diagnostic and relapse, respectively. Figure:. Subgrouping of MM patient samples (I-IV) based on selective drug response profiles. H/D/R denotes healthy, diagnostic and relapse, respectively. Disclosures Silvennoinen: Research Funding of Finland Government, Research Funding from Janssen-cilag, research funding from Celgene: Research Funding; Janssen-Cilag, Sanofi, Celgene: Honoraria. Wennerberg:Pfizer: Research Funding. Kallioniemi:Medisapiens: Consultancy, Membership on an entity's Board of Directors or advisory committees. Porkka:Bristol-Myers Squibb: Honoraria, Research Funding; Novartis: Honoraria, Research Funding. Heckman:Celgene: Research Funding.
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- 2014
31. Landscape of Driver Lesions in Multiple Myeloma and Consequences for Targeted Drug Response
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Samuli Eldfors, Marjanna Säily, David Tamborero, Caroline A. Heckman, Raija Silvennoinen, Juha Lievonen, Alun Parsons, Muntasir Mamun Majumder, Minna Suvela, Kimmo Porkka, and Pekka Anttila
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0303 health sciences ,medicine.diagnostic_test ,Immunology ,Cancer ,Cell Biology ,Hematology ,Biology ,medicine.disease_cause ,Bioinformatics ,medicine.disease ,Biochemistry ,3. Good health ,Molecular cytogenetics ,03 medical and health sciences ,0302 clinical medicine ,Gene duplication ,medicine ,Cancer research ,KRAS ,Carcinogenesis ,Exome sequencing ,Multiple myeloma ,030304 developmental biology ,030215 immunology ,Fluorescence in situ hybridization - Abstract
Introduction Multiple myeloma (MM) is a heterogeneous disease that eventually becomes resistant to therapy. Determining the genomic lesions driving each stage of the tumor and identifying actionable items for novel targeted drugs will improve and increase therapeutic options for the malignancy. The aim of the present work is to obtain a comprehensive catalog of driver genomic lesions for both newly diagnosed (NDMM) and refractory/relapsed MM (RRMM) patients by integrating multiple genomic data and linking these to the action of targeted therapeutic approaches. Methods Molecular cytogenetics was assessed by fluorescence in situ hybridization and somatic mutations and copy number changes were identified by performing exome sequencing of DNA from CD138+ cell and skin paired samples collected from 30 MM patients (NDMM n=12; RRMM n=18). In addition, gene expression profiles were obtained by transcriptome sequencing. The proportion of tumor clones bearing a specific mutation was inferred from the variant allele frequency. Genetic alterations involved in the tumorigenesis of each patient (named drivers) were identified by combining an in silico method aimed to score their potential for being malignant with the a priori knowledge retrieved from the identification of complementary signals of positive selection in available tumor cohorts (Tamborero et al. Nat Sci Rep 2013). Selective drug response was assessed by testing the ex vivo sensitivity of patient derived CD138+ cells to 306 oncology drugs and comparing results with responses derived from healthy bone marrow control cells. Results Overall, 0.5 translocations, 3±2.8 mutations and 4.9±2.7 copy number changes per patient were identified as putative drivers. The total number of driver alterations did not differ between NDMM and RRMM samples, and no gene reached statistical significance for being more frequently altered in the latter group. However, the only mutations in RAS genes that appeared at sub-clonal proportions occurred in diagnosed samples, pointing out their positive selection among relapsed patients in which they were present in all clones. Translocations involving IGH@ were observed in 11 (37%) patients, and interestingly 3 other samples exhibited driver alterations in the oncogenes involved in these fusions (i.e. activating mutations in FGFR3 or gene amplification plus peaked overexpression of WHSC1 and CCND1). Recurrent alterations were observed among genes previously associated with MM, including DIS3 (n=15), KRAS (n=11), CYLD (n=8), TRAF3 (n=6) and FAM46C (n=5). Other genes not previously associated with or less-known to be involved in MM pathogenesis were also identified, including the histone methyltransferase MLL, the tumor necrosis factor associated genes FAF1 and TNFRSF13B, the p53-suppressing protein phosphatase PPM1D, and several genes related with blood cell differentiation and B-lymphocyte development (e.g. SOX7, BLK and PRDM1). Overall, the pathways most frequently targeted by driver alterations were MAPK (23 (77%) samples, mostly by mutations), NF-κB (17(57%) samples, mostly by gene copy loss), cell-cycle (18 (60%) samples), and RNA-processing (17 (57%) samples). Comparison of driver lesions to drug response using data derived from ex vivo testing of the same patient samples to different targeted small molecule inhibitors (e.g. PI3K/mTOR and MEK inhibitors) indicated that alterations affecting PI3K and p53 pathways were associated with increased drug sensitivity, while alterations involving activation of FGFR3 and copy loss of TRAF3 were associated with a more resistant phenotype. Conclusions The integration of multiple genomic data by combining different predictive computational tools can comprehensively identify cancer events in individual patients. Applying these tools to genomic data from MM patients identified both known and novel driver lesions, and some of these alterations were associated with the ex vivo response to selective drugs. However, further data is required to confirm biomarkers of response to those novel therapeutics and test potential benefits in MM patients. Disclosures Silvennoinen: Janssen, Sanofi, Celgene: Honoraria; Research Funding of Government Finland, Research Funding from Janssen and Celgene: Research Funding. Porkka:Novartis: Honoraria, Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding. Heckman:Celgene: Research Funding.
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- 2014
32. Identification Of AML Subtype-Selective Drugs By Functional Ex Vivo Drug Sensitivity and Resistance Testing and Genomic Profiling
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Riikka Karjalainen, Maija Lepistö, Tero Pirttinen, Jonathan Knowles, Maxim M. Bespalov, Samuli Eldfors, Sonja Lagström, Erkki Elonen, Anna Lehto, Bjørn Tore Gjertsen, Jesus M. Lopez Marti, Aino Palva, Alun Parsons, Tea Pemovska, Satu Mustjoki, Maria E. Rämet, Kimmo Porkka, Agnieszka Szwajda, Laura Turunen, Mika Kontro, Krister Wennerberg, Astrid Murumägi, Evgeny Kulesskiy, Pekka Ellonen, Pirkko Mattila, Caroline A. Heckman, Olli Kallioniemi, Minna Suvela, Imre Vastrik, Tero Aittokallio, Maija Wolf, Tuija Lundán, Henrik Edgren, Muntasir Mamun Majumder, Bhagwan Yadav, and Henrikki Almusa
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Trametinib ,0303 health sciences ,Ruxolitinib ,Sunitinib ,business.industry ,Immunology ,Cell Biology ,Hematology ,Drug resistance ,Bioinformatics ,Biochemistry ,Temsirolimus ,3. Good health ,Dasatinib ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,chemistry ,medicine ,Personalized medicine ,business ,030304 developmental biology ,030215 immunology ,medicine.drug ,Quizartinib - Abstract
Introduction 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.
- Published
- 2013
33. Abstract 5067: Exome sequencing reveals both DNA sequence and copy number changes in AML: Potential driver changes and mechanisms of drug resistance revealed from serial samples from the same patients
- Author
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Sari Hannula, Maija Wolf, Satu Mustjoki, Maija Lepistö, Astrid Murumägi, Muntasir Mamun Majumder, Pekka Ellonen, Olli Kallioniemi, Sonja Lagström, Mika Kontro, Caroline A. Heckman, Minna Suvela, Henrik Edgren, Henrikki Almusa, Krister Wennerberg, Naga Poojitha Kota Venkata, Jonathan Knowles, Kimmo Porkka, Pirkko Mattila, Riikka Karjalainen, Samuli Eldfors, Janna Saarela, and Alun Parsons
- Subjects
Genetics ,0303 health sciences ,Cancer Research ,medicine.medical_specialty ,Cancer ,Drug resistance ,Biology ,medicine.disease ,Somatic evolution in cancer ,Minimal residual disease ,DNA sequencing ,3. Good health ,03 medical and health sciences ,0302 clinical medicine ,Oncology ,030220 oncology & carcinogenesis ,Molecular genetics ,medicine ,Exome ,Exome sequencing ,030304 developmental biology - Abstract
Despite significant advances in characterizing the molecular genetics of AML, the clonal evolution of leukemic cells and the dynamic impact of genomic changes on the development of the disease and progression to drug resistance are not well understood. Here, we applied next-generation sequencing to quantify aberrant tumor subclones carrying specific mutant alleles of key cancer genes and developed a method to extract quantitative high-resolution copy number changes across the genome using exome sequencing data from matching cancer and normal DNA. Serial bone marrow (BM) samples collected from diagnosis to relapse to post-treatment drug resistance in a patient-centric manner made it possible to trace the clonal evolution of AML and to identify variants potentially involved in drug resistance. Exome sequencing from AML blast cells and normal skin biopsies was performed as part of the Finnish Hematology Registry and Biobanking (FHRB) effort. Consecutive paired samples from different patients revealed unique genetic patterns of clonal evolution and cancer progression in each patient. In a pre-resistant sample of one AML M5 patient, we identified four closely spaced insertions in the Wilm's Tumor (WT1) suppressor gene, none of which appear on the same sequence reads. This suggests the presence of multiple distinct leukemic subclones even before treatment resistance and underscores the strong selective advantage conferred by WT1 mutations. After relapse, one of the subclones was lost, and another one significantly increased suggesting that the relapse arose from the expansion of a pre-existing resistant subclone. In this patient, recurrent clones otherwise featured similar copy number changes and the same fusion genes as the primary diagnostic sample. In another AML patient developing recurrence an opposite pattern was observed: The relapsed, drug-resistant cells displayed an enormous increase of small microdeletions compared to the diagnostic, pre-treatment sample, while almost all sequence-level alterations in potential cancer genes were the same between the two samples. This suggests that a distinct type of DNA repair deficiency may have contributed to the drug resistant clone in this patient. Conclusions: Exome sequencing from paired samples of AML cells before and after relapse makes it possible to trace the clonal evolution of the disease and study the impact of therapy both at the level of sequence alterations of key cancer genes and simultaneously at the level of copy number changes inferred from exome sequence data. This analysis has highlighted multiple genomic patterns by which resistance may evolve in vivo during cancer treatment. Refined bioinformatic analysis and interpretation of exome-seq data provides a rich resource to identify genetic biomarkers of drug response and minimal residual disease. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5067. doi:1538-7445.AM2012-5067
- Published
- 2012
34. 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
- Author
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Bhagwan Yadav, Henrikki Almusa, Satu Mustjoki, Naga Poojitha Kota Venkata, Petteri Hintsanen, Pekka Ellonen, Samuli Eldfors, Agnieszka Szwajda, Muntasir Mamun Majumder, Tero Aittokallio, Pirkko Mattila, Astrid Murumägi, Evgeny Kulesskiy, Sonja Lagström, Henrik Edgren, Mika Kontro, Jesus M. Lopez Marti, Janna Saarela, Caroline A. Heckman, Tea Pemovska, Olli Kallioniemi, Disha Malani, Minna Suvela, Kimmo Porkka, Krister Wennerberg, Maija Lepistö, Riikka Karjalainen, and Alun Parsons
- Subjects
Oncology ,medicine.medical_specialty ,Ruxolitinib ,Immunology ,Drug resistance ,Bioinformatics ,Biochemistry ,03 medical and health sciences ,chemistry.chemical_compound ,Internal medicine ,0502 economics and business ,medicine ,030304 developmental biology ,0303 health sciences ,business.industry ,05 social sciences ,Ponatinib ,Foretinib ,Cell Biology ,Hematology ,medicine.disease ,3. Good health ,Dasatinib ,Leukemia ,chemistry ,050211 marketing ,Personalized medicine ,business ,Ex vivo ,medicine.drug - 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.
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