scholarly journals Integrative Drug and Genomic Profiling Identify Therapeutic Vulnerabilities and Inform Precision Medicine for Pediatric Acute Myeloid Leukemia

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 2297-2297
Author(s):  
Han Wang ◽  
Kathy Chan ◽  
Po Yi Lee ◽  
Alex WK Leung ◽  
Chi Kong Li ◽  
...  
Keyword(s):  
Myeloid Leukemia ◽  
Drug Testing ◽  
Drug Response ◽  
Drug Sensitivity ◽  
Genomic Profiling ◽  
Targeted Agents ◽  
New Agents ◽  
Pediatric Aml ◽  
Refractory Aml

Abstract Background/Aims: Despite advances in chemotherapy-based treatment protocols, the outcomes of children with acute myeloid leukemia (AML) remain suboptimal. Implementation of targeted therapy based solely on genomics is challenging due to the complex mutational patterns and scarcity of pharmacologic agents for most lesions. In addition, pediatric and adult AML are genetically and biologically distinct, which poses a major hurdle for extrapolation of new agents approved for adult AML to the pediatric population. This study aims to adopt a functional approach that directly measure the response of patient-derived leukemic cells to targeted agents, and to establish the drug sensitivity pattern and identify candidates of immediate clinical relevance for precision usage in high-risk pediatric AML. Methods: A high-throughput drug screening, comprising 39 targeted agents (2 in Phase I, 10 in Phase II, 5 in Phase III, 22 FDA-approved) and 6 conventional chemotherapeutics, was performed on 30 pediatric AML samples collected at diagnosis or relapse using a serum-free, cytokine-supported culture system. A counter-screen of active drugs on cord blood hematopoietic stem cells was accomplished to reveal leukemia-selective activities. The robustness of the drug testing platform for predicting in vivo activities was validated in xenograft models. Genomic profiling was complementarily performed to identify the genetic markers and underlying mechanisms of drug sensitivity. Patients with refractory AML were treated with targeted agents based on drug profiling results, and assessed for clinical responses. Results: Unsupervised clustering revealed 5 distinct clusters of drug response: highly active compounds (IC50 <15 nM, 5 drugs); generally active compounds (IC50 <250 nM, 11 drugs); compounds with bimodal activities (wide IC50 ranges, 3 drugs); generally inactive compounds (16 drugs); and inactive compounds (IC50>2000 nM, 10 drugs). Targeted agents, including Bcl-2, HDAC, proteasome, HSP and survivin inhibitors, had substantially higher potency and selectivity over standard chemotherapeutic agents. New agents approved for adult AML were essentially inactive in pediatric AML. Drug sensitivity ex vivo accurately predicted in vivo single-agent and combinatorial activities with cytarabine in cell line- and patient-derived xenografts. Targeted resequencing of a 141-gene panel revealed novel mutations of prognostic relevance, such as KMT2C, in pediatric AML and their vulnerability to targeted agents. Whole-genome RNA-sequencing identified distinct gene expression signatures shaping the response to individual drugs. Administration of venetoclax to a child with refractory AML resulted in rapid blast clearance and achieved long-term remission. Complementary genomic profiling on serial specimens dictated the dynamic drug responses during disease evolution. Conclusions: Our study establishes a reliable drug testing platform and a pediatric-specific drug response profile of AML, which enables an evidence-based selection of targeted agents for patients without treatment options and endows therapies increasingly precise and personalized. The study also generates a valuable gene-drug-clinical dataset that could be leveraged to address the fundamental and translational biology of pediatric AML. It will ultimately impact the future design of clinical trials and protocols for managing this life-threatening malignancy. Disclosures No relevant conflicts of interest to declare.

Stratification of Multiple Myeloma Patients Based on Ex Vivo Drug Sensitivity and Identification of New Treatments for Patients with High-Risk Relapsed/Refractory Disease

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3006-3006
Author(s):  
Muntasir Mamun Majumder ◽  
Raija Silvennoinen ◽  
Pekka Anttila ◽  
David Tamborero ◽  
Samuli Eldfors ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Research Funding ◽  
Drug Testing ◽  
Drug Response ◽  
Drug Sensitivity ◽  
Plasma Cells ◽  
Ex Vivo ◽  
Resistant Group ◽  
Sensitive Group

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.

Large Interindividual Differences in In Vitro Calicheamicin Sensitivity May Underly Gemtuzumab Ozogamicin Resistance in Acute Myeloid Leukemia (AML).

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 107-107
Author(s):  
Bianca F. Goemans ◽  
Gertjan J.L. Kaspers ◽  
Susanne J.H. Vijverberg ◽  
Anne H. Loonen ◽  
Ursula Creutzig ◽  
...  
Keyword(s):  
Myeloid Leukemia ◽  
Drug Sensitivity ◽  
Gemtuzumab Ozogamicin ◽  
Cross Resistance ◽  
Newly Diagnosed ◽  
Primary Resistance ◽  
In Vitro Sensitivity ◽  
Resistant Patient ◽  
Pediatric Aml

Abstract Gemtuzumab ozogamicin (GO or Mylotarg®) is increasingly being used in the treatment of AML. GO consists of a cytotoxic drug - calicheamicin which is conjugated to an anti-CD33 antibody. Most AML patients highly express CD33 on their blasts. Studies relating CD33 expression to response to GO have failed to show an association. Although 30–50% of patients respond to Mylotarg®, the causes of primary resistance to this drug remain unclear. Previous reports have studied P-gp status and CD33 expression as a cause of resistance to GO. Another factor that might determine response to GO is cellular drug resistance to calicheamicin. In this study we examined in vitro resistance to calicheamicin in 90 initially diagnosed and 32 relapsed pediatric AML samples using the 4 day MTT assay (concentration range 0.000004 – 0.4 μg/ml). The LC50 value, the drug concentration at which 50% of the cells is killed by the drug, is used as a measure of sensitivity. In addition to calicheamicin, some samples were also tested successfully for in vitro sensitivity to etoposide, cytarabine, daunorubicin, idarubicin, mitoxantrone, 6-thioguanine, vincristine and L-asparaginase. The characteristics of the 122 pediatric AML samples included are as follows: 62% boys, median age 9.6 years, median WBC 53.0*109/L, FAB types M0 n=9, M1 n=10, M2 n=18, M3 n=9, M4 n=33, M5 n=24, M7 n=3, unknown n=16. There was a more than 100,000 fold difference in calicheamicin sensitivity between the most sensitive and the most resistant patient samples. FAB M2 samples taken at initial diagnosis (n=13) were significantly more resistant to calicheamicin compared to the other FAB types (Resistance Ratio (RR)=2.5, median LC50 0.033 vs. 0.013 μg/ml, p=0.008). Newly diagnosed AML samples were significantly more sensitive to calicheamicin compared to relapsed AML samples (RR=0.68, median LC50 0.023 vs. 0.034 μg/ml, p=0.042) (although these patients had not been treated with calicheamicin). There was strong cross-resistance between calicheamicin and the anthracyclines idarubicin (Spearmans rho = 0.73, p<0.0001, n=23), daunorubicin (rho=0.61, p<0.0001, n=103) and mitoxantrone (rho=0.52, p=0.039, n=16). In addition, there was moderate cross-resistance with etoposide (rho=0.42, p<0.0001, n=101). No cross-resistance was observed between calicheamicin and cytarabine (rho=0.11, p=0.28, n=106), 6-thioguanine (rho=0.20, p=0.054, n=97), vincristine (rho=0.12, p=0.44, n=46) or L-asparaginase (rho=0.21, p=0.16, n=45). In conclusion, the interpatient differences in calicheamicin sensitivity are the largest differences in in vitro drug sensitivity we have ever observed in pediatric AML. FAB M2 samples are 2.5 fold more resistant to calicheamicin than samples with other FAB types. Initially diagnosed pediatric AML samples are 1.5 fold more sensitive to calicheamicin than relapsed AML samples. There is marked cross-resistance between calicheamicin and the related anthracyclin compounds. Given the large differences in sensitivity to calicheamicin in pediatric AML samples, it is likely that calicheamicin resistance plays a role in resistance to Mylotarg®.

scholarly journals A Biomaterial Screening Approach Reveals Microenvironmental Mechanisms of Drug Resistance

2017 ◽  
Author(s):  
Alyssa D. Schwartz ◽  
Lauren E. Barney ◽  
Lauren E. Jansen ◽  
Thuy V. Nguyen ◽  
Christopher L. Hall ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Systems Biology ◽  
Drug Screening ◽  
Drug Response ◽  
Drug Sensitivity ◽  
Mek Inhibitor ◽  
Tumor Burden ◽  
Adaptive Resistance

TOC FigureDrug response screening, gene expression, and kinome signaling were combined across biomaterial platforms to combat adaptive resistance to sorafenib.Insight BoxWe combined biomaterial platforms, drug screening, and systems biology to identify mechanisms of extracellular matrix-mediated adaptive resistance to RTK-targeted cancer therapies. Drug response was significantly varied across biomaterials with altered stiffness, dimensionality, and cell-cell contacts, and kinome reprogramming was responsible for these differences in drug sensitivity. Screening across many platforms and applying a systems biology analysis were necessary to identify MEK phosphorylation as the key factor associated with variation in drug response. This method uncovered the combination therapy of sorafenib with a MEK inhibitor, which decreased viability on and within biomaterials in vitro, but was not captured by screening on tissue culture plastic alone. This combination therapy also reduced tumor burden in vivo, and revealed a promising approach for combating adaptive drug resistance.AbstractTraditional drug screening methods lack features of the tumor microenvironment that contribute to resistance. Most studies examine cell response in a single biomaterial platform in depth, leaving a gap in understanding how extracellular signals such as stiffness, dimensionality, and cell-cell contacts act independently or are integrated within a cell to affect either drug sensitivity or resistance. This is critically important, as adaptive resistance is mediated, at least in part, by the extracellular matrix (ECM) of the tumor microenvironment. We developed an approach to screen drug responses in cells cultured on 2D and in 3D biomaterial environments to explore how key features of ECM mediate drug response. This approach uncovered that cells on 2D hydrogels and spheroids encapsulated in 3D hydrogels were less responsive to receptor tyrosine kinase (RTK)-targeting drugs sorafenib and lapatinib, but not cytotoxic drugs, compared to single cells in hydrogels and cells on plastic. We found that transcriptomic differences between these in vitro models and tumor xenografts did not reveal mechanisms of ECM-mediated resistance to sorafenib. However, a systems biology analysis of phospho-kinome data uncovered that variation in MEK phosphorylation was associated with RTK-targeted drug resistance. Using sorafenib as a model drug, we found that co-administration with a MEK inhibitor decreased ECM-mediated resistance in vitro and reduced in vivo tumor burden compared to sorafenib alone. In sum, we provide a novel strategy for identifying and overcoming ECM-mediated resistance mechanisms by performing drug screening, phospho-kinome analysis, and systems biology across multiple biomaterial environments.

scholarly journals Discovery of Novel Drug Sensitivities in T-Prolymphocytic Leukemia (T-PLL) By High-Throughput Ex Vivo Drug Testing and Genetic Profiling

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 917-917
Author(s):  
Emma I Andersson ◽  
Leopold Sellner ◽  
Malgorzata Oles ◽  
Tea Pemovska ◽  
Paavo Pietarinen ◽  
...  
Keyword(s):  
Drug Screening ◽  
Research Funding ◽  
Drug Testing ◽  
Drug Response ◽  
Drug Sensitivity ◽  
Response Pattern ◽  
Ex Vivo ◽  
Drug Responses ◽  
Ic50 Values ◽  
Stat Pathway

Abstract Introduction T-PLL is a mature post-thymic T-cell neoplasm with an aggressive clinical course (5-year overall survival 21%). Almost 75% of T-PLL cases harbor chromosome 14 translocations resulting in aberrant activation of the proto-oncogene TCL1A. Furthermore, in the majority of T-PLL cases the ATM gene is mutated or deleted, and recently it was reported that mutations in genes involved in the JAK-STAT pathway were found in 76% of T-PLL cases. Due to the rareness and aggressive nature of the disease, clinical trials are difficult to execute. By using a high-throughput ex vivo drug sensitivity and resistance testing (DSRT) platform covering 306 approved and investigational oncology drugs we systematically investigated the heterogeneity of drug responses in PLL-patients. As the impact of mutations on drug sensitivity is not well understood we aimed to identify relevant associations between the drug responses and genetic lesions in T-PLL patients. Methods Primary cells (MNCs) from seven T-PLL patients were obtained for drug screening. Samples were seeded in 384-well plates and 306 active substances were tested using a 10,000-fold concentration range resulting in a dose-response curve for each compound. Cell viability was measured after 72 h incubation and differential drug sensitivity scores (sDSS), representing leukemia-specific responses, were calculated by comparing patient samples to healthy donors. Hierarchical clustering of the drug responses was performed with Cluster 3.0 and Java Tree View. To assess the performance of the drug screening platform we also exchanged six samples with the German Cancer Research Center in Heidelberg for a comparison of results between two independent drug screening systems. To understand heterogeneous pathway dependencies, drug sensitivities were correlated with somatic genetic variants and recurrent chromosomal aberrations. Genetic characterization was performed by exome sequencing of tumor and matched healthy cells to profile known recurrent genetic variants (ATM, STAT5b, IL2RG, JAK1, JAK3) as well as CNVs (TCL1A translocations, ATM deletions, recurrent chromosomal aberrations). Results Four out of seven patient samples showed high sensitivity to small molecule BCL2 inhibitors navitoclax (IC50: 10-68nM) and ABT-199 (IC50: 14-45nM) and to HDAC inhibitors panobinostat and belinostat (IC50: 2-65nM). Intriguingly, the CDK inhibitor SNS-032 was effective in 6/7 patient samples (IC50: 7-95nM). SNS-032 inhibits Cdk2, Cdk7 and Cdk9, which control transcription of anti-apoptotic proteins including MCL1 and XIAP. As the AKT1/MTOR pathway is activated in many T-PLL patients due to expression of the TCL1A oncoprotein, it was interesting to observe that patient samples did not show any response to AKT inhibitors (MK-2206 and GDC-0068 IC50 values >1000 nM) nor to MTOR inhibitors (rapalogs temsirolimus and everolimus). Similarly, T-PLL cells were insensitive to JAK-inhibitors. Clustering of drug responses from T-PLL patients with primary AML and ALL patient samples revealed the drug response profiles to be specific for T-PLL patients (Figure). 6/7 patients clustered together while the only patient (PLL4) in our cohort with confirmed mutations in the JAK-STAT pathway genes STAT5b (P702S) and IL2RG (K315E) exhibited a non-sensitive response pattern when compared to other samples (Figure). Interestingly, exome sequencing did not reveal any JAK mutations in our PLL-cohort (n=5) nor additional STAT5b or IL2RG mutations in other patients except in this unresponsive patient. In the comparison between the platforms the correlation of the censored IC50 values from the 60 overlapping drugs was r=0.75. Similar fits of dose-response curves were seen for most drugs, although there were notable exceptions, which may be due to divergent culture conditions and day of read-out. Conclusions Ex vivo drug testing of primary patient cells has the potential to provide novel personalized drug candidates (such as BCL2, HDAC and CDK inhibitors) for T-PLL. The drug response pattern was T-PLL specific warranting further clinical testing. Drug screening, mutation analysis and RNA sequencing of additional patients is currently ongoing (n=20) to validate whether drug responses can be predicted based on the mutation profile or aberrant gene expression. Figure Clustering of T-PLL, AML and ALL patient samples based on DSRT results. Figure. Clustering of T-PLL, AML and ALL patient samples based on DSRT results. Disclosures 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. Mustjoki:Bristol-Myers Squibb: Honoraria, Research Funding; Novartis: Honoraria, Research Funding.

scholarly journals Dasatinib Inhibits FLT3/ITD and PTPN11mutated Acute Myeloid Leukemia Cells Overexpressing SRC Tyrosine Kinases

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1451-1451
Author(s):  
Sigal Tavor ◽  
Tali Shalit ◽  
Noa Chapal Ilani ◽  
Yoni Moskovitz ◽  
Nir Livnat ◽  
...  
Keyword(s):  
Gene Expression ◽  
Acute Myeloid Leukemia ◽  
Board Of Directors ◽  
Myeloid Leukemia ◽  
Drug Sensitivity ◽  
Kinase Inhibitors ◽  
Advisory Committees ◽  
Acute Myeloid

Background: Recent advances in acute myeloid leukemia(AML) targeted therapy improve overall survival. While these targeted therapies can achieve prolonged remissions, most patients will eventually relapseunder therapy. Our recent studies suggest that relapse most often originates from several sub-clones of leukemic stem cells (LSCs), present before therapy initiation, and selected due to several resistance mechanisms. Eradication of these LSCs during treatment induction /remission could thus potentially prevent relapse. The overall goal of the current study was to identify drugs which can be safely administrated to patients at diagnosis and that will target LSCs. Since simultaneously testing multiple drugs in vivo is not feasible, we used an in vitrohigh throughput drug sensitivity assay to identify new targets in primary AML samples. Methods: Drug sensitivity and resistance testing (DSRT) was assessed in vitro (N=46 compounds) on primary AML samples from patients in complete remission (N=29). We performed whole exome sequencing and RNAseq on samples to identify correlations between molecular attributes and in vitro DSRT. Results:Unsupervised hierarchical clustering analysis of in vitro DSRT, measured by IC50, identified a subgroup of primary AML samples sensitive to various tyrosine kinase inhibitors (TKIs). In this subgroup, 52% (9/17) of AML samples displayed sensitivity to dasatinib (defined as a 10-fold decrease in IC50 compared to resistant samples). Dasatinib has broad TKI activity, and is safely administered in the treatment of leukemia. We therefore focused our analysis on predicting AML response to dasatinib, validating our results on the Beat AML cohort. Enrichment analysis of mutational variants in dasatinib-sensitive and resistant primary AML samples identified enrichment of FLT3/ITD (p=0.05) and PTPN11(p=0.05) mutations among dasatinib responders. Samples resistant to dasatinib were enriched with TP53 mutations (p=0.01). No global gene expression changes were observed between dasatinib-sensitive and resistant samples in our cohort, nor in the Beat AML cohort. Following this, we tested the differential expression of specific dasatinib-targeted genes between dasatinib-responding and resistant samples. No significant differences were identified. However, unsupervised hierarchical clustering of dasatinib targeted genes expression in our study and in the Beat AML cohort identified a subgroup of AML samples (enriched in dasatinib responders) that demonstrated overexpression of three SRC family tyrosine kinases:FGR, HCK and LYN as well as PTK6, CSK, GAK and EPHB2. Analysis of the PTPN11 mutant samples revealed that the IC50 for dasatinib in 23 carriers of the mutant PTPN11 was significantly lower compared to the IC50 of PTPN11 wild type samples (p=0.005). LYN was also upregulated (p<0.001) in the mutant samples. We therefore hypothesized that gene expression of dasatinib-targeted genes could be used as a predictive biomarker of dasatinib response among FLT3/ITD carriers. We found that among FLT3/ITD AML carriers in the Beat AML cohort LYN, HCK, CSK and EPHB2 were significantly over-expressed in the dasatinib responding samples (N=27) as compared to the dasatinib resistant samples (N=35). To predict response to dasatinib among FLT3/ITD carriers we used a decision tree classifier based on the expression levels of these four genes. Our prediction model yielded a sensitivity of 74% and specificity of 83% for differentiating dasatinib responders from non-responders with an AUC of 0.84. Based on our findings, we selected FLT3/ITD AML samples and injected them to NSG-SGM3 mice. We found that in a subset of these samples, dasatinib significantly inhibited LSCs engraftment. This subset of FLT3/ITD AML samples expressed higher levels of LYN, HCK,FGR and SRC as compared to the FLT3/ITD samples that were not sensitive to dasatinib therapy in vivo. In summary, we identified a subgroup of AML patients sensitive to dasatinib, based on mutational and expression profiles. Dasatinib has anti-leukemic effects on both blasts and LSCs. Further clinical studies are needed to demonstrate whether selection of tyrosine kinase inhibitors, based on specific biomarkers, could indeed prevent relapse. Disclosures Tavor: Novartis: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Astellas: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees; BMS companies: Membership on an entity's Board of Directors or advisory committees.

scholarly journals In Vivo Evaluation of Mesothelin As a Therapeutic Target in Pediatric Acute Myeloid Leukemia

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1370-1370 ◽  
Author(s):  
Anilkumar Gopalakrishnapillai ◽  
Allison Kaeding ◽  
Christoph Schatz ◽  
Anette Sommer ◽  
Soheil Meshinchi ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Combination Therapy ◽  
Therapeutic Target ◽  
Myeloid Leukemia ◽  
Antibody Drug Conjugate ◽  
Pediatric Acute Myeloid Leukemia ◽  
Pediatric Aml ◽  
Acute Myeloid ◽  
Antibody Drug

Pediatric acute myeloid leukemia (AML) continues to have a cure rate of only 50% despite the use of highly intensive cytotoxic chemotherapy. Transcriptome sequencing of several AML samples by the NCI/COG TARGET AML Initiative identified mesothelin (MSLN) to be highly overexpressed in about one-third of pediatric AML (Tarlock et al., Blood, 128:2873, 2016). Because MSLN is not expressed in normal bone marrow samples (Fan et al., Blood, 130:3792, 2017) and only to a low level in other human organs and tissues, MSLN is an attractive therapeutic target for pediatric AML (Kaeding et al., Blood, 130:2641, 2017). The anti-MSLN antibody-drug conjugate (ADC) anetumab ravtansine (BAY 94-9343) generated by conjugating MSLN-antibody with tubulin inhibitor DM4 (Meso-ADC), and isotype control antibody conjugated with the same drug (Iso-ADC) were used to evaluate the efficacy of MSLN targeting in vivo. MSLN-overexpressing K562 (K562-MSLN) CML cells and MV4;11 (MV4;11-MSLN) AML cells were generated by lentiviral transduction of MSLN cDNA. Cell line-derived xenografts (CDX) were created by injecting the MSLN-transduced or parental (MSLN-) cells into NSG-SGM3 mice via the tail vein. Mice were randomly assigned to treatment groups when the median percentage of human cells in mouse peripheral blood was greater than 0.5%. K562-MSLN CDX mice treated with Meso-ADC (5 mg/Kg Q3dx3, i.v.) survived a median of 46 days longer than those treated with Iso-ADC (P=0.0011) and significantly longer than comparison groups, including K562-MSLN CDX mice treated with daunorubicin and Ara-C (DA, P=0.0008) or untreated (P=0.0018) (Fig. 1A). Median survival of K562 CDX mice treated with Meso-ADC, Iso-ADC, or untreated was similar (Fig. 1B). MV4;11-MSLN CDX mice treated with Meso-ADC exhibited complete remission and remained disease-free at 1 year post cell injection, with AML cell burden remaining <0.1% throughout the study period (Fig. 1C). In contrast, MV;11-MSLN CDX mice treated with Iso-ADC or untreated succumbed to disease at 72 and 38 days, respectively. Taken together, these results indicate that Meso-ADC was efficacious in reducing leukemia burden, and this effect required MSLN expression in target cells. We have generated a panel of patient-derived xenograft (PDX) lines by transplanting and serially propagating primary pediatric AML samples into NSG-SGM3 mice. The efficacy of Meso-ADC was also evaluated in a systemic PDX model using a MSLN+ PDX line (NTPL-146). NTPL-146 PDX mice treated with Meso-ADC (5 mg/Kg, Q3dx3 -x2 cycles) survived a median of 50 days longer than those treated with Iso-ADC (P=0.0018, Fig. 1D, arrows indicate time when each treatment cycle was initiated). In an independent experiment with NTPL-146 PDX mice, a survival benefit of Meso-ADC treatment over no treatment was observed after 1 cycle of Meso-ADC treatment (5 mg/Kg, Q3dx3, P=0.0019, Fig. 1E). Additionally, a combination therapy strategy with daunorubicin and Ara-C followed by Meso-ADC (DA -> Meso-ADC) resulted in improved median survival over Meso-ADC (P=0.0027) or DA treatment alone (P=0.0018) (Fig. 1E). The disseminated MSLN+ leukemia mouse models described herein support MSLN-targeted antibody-drug conjugate as a potential treatment strategy in MSLN+ AML. Furthermore, we provide the first in vivo demonstration of synergy between MSLN-targeted therapy and conventional chemotherapy in MSLN+ AML, warranting additional investigation to validate and optimize novel strategies for combination therapy. Figure 1 Disclosures Kaeding: Celgene: Employment. Schatz:Bayer AG: Employment. Sommer:Bayer AG: Employment, Equity Ownership.

scholarly journals Precision Medicine Assays Detect Novel Targetable FLT3 Fusions Amenable to Therapeutic Intervention in a Patient with Refractory Acute Myeloid Leukemia

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 5170-5170
Author(s):  
Bradley A Patay ◽  
Andrew Carson ◽  
Timothy J Martins ◽  
Sylvia Chien ◽  
Mary-Elizabeth M. Percival ◽  
...  
Keyword(s):  
High Throughput ◽  
Myeloid Leukemia ◽  
Drug Sensitivity ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Monosomy 7 ◽  
Drug Sensitivity Assay ◽  
Generation Sequencing ◽  
Acute Myeloid ◽  
Refractory Aml

Abstract Context: The 5-year survival rate for acute myeloid leukemia (AML) is 26.6%. The prognosis of patients with adverse events such as older age or unfavorable risk cytogenetics remains poor, even for those who undergo allogeneic hematopoietic cell transplant. AML is a heterogeneous disease and novel N-of-1 clinical trial designs may offer benefit to individuals compared to conventional clinical trials by offering improved utilization of investigational chemotherapy regimens. Precision medicine based assays that reveal a deeper understanding of the cancer biology and potential for novel therapeutics may improve survival in the future. Objective: The goal of the clinical trial on which this patient was enrolled, Individualized Treatment for Relapsed/Refractory Acute Leukemia Based on Chemosensitivity and Genomics/Gene Expression Data (ClinicalTrials.gov Identifier:NCT01872819) was to determine feasibility of a study that utilized results of comprehensive mutation analysis and an in vitrohigh throughput functional assay to choose treatment for individual patients with refractory AML. Feasibility was defined as initiating chosen treatment within 21 days. A secondary objective was to achieve a response (cytoreduction or at least partial response) greater than that expected for comparable refractory patient populations with other salvage regimens. Design, Setting, and Patient: A single center enrolled individuals who had failed at least 2 inductions at initial diagnosis or >1 salvage regimen for relapsed AML. Patients could receive any FDA approved drug or combination regimen based on molecular analysis and high throughput drug sensitivity assay. A 58 year old female was enrolled into this protocol with MECOM (EVI1) rearranged, Monosomy 7 refractory AML. Methods and Main Outcome Measures: The patient had various assays performed on her samples, including next generation sequencing and a high throughput in vitro assay that analyzed enriched blast samples for sensitivity to 150 drugs or drug combinations. MyAML™, a next generation sequencing panel, analyzed 194 genes including breakpoint hotspot loci with long paired end sequencing and high depth that optimized detection of large insertion and deletions and other structural variants found in AML at low variant allele frequency. Results: The MyAML assay detected multiple variants including: NRAS:c.38G>A; p.Gly13Asp VAF = 100%, RUNX1:c.494_495ins; p.165_R166ins VAF = 42%, WT1:c.1149_1150ins; p.E384Pfs*5 VAF=38%. Fusions were also detected: t(13;17)(q12.2;q11.2), t(8;13)(q21.13;q12.2), and t(9;12)(q32;p13.2) which involved FLT3 novel fusions. FLT3 internal tandem duplications (ITD) or tyrosine kinase domain (TKD) variants were not detected. The assay also detected the Monosomy 7 and confirmed the t(2;3) as a THADA-MECOM (EVI1) fusion to the nucleotide breakpoint. These variants involved activated signaling, myeloid transcription factor and DNA demethylation pathways. The high throughput drug sensitivity assay identified sensitivity to kinase inhibitors such as the MEK inhibitor selumetinib (IC50 8.1 X 10e-9 M), Flt3 inhibitor staurosporine (IC50 1.4 X 10e-8 M) and Abl kinase inhibitor ponatinib (IC50 2.7 X 10e-8 M). Insurance coverage for these agents was not able to be obtained as this indication would be considered off label. However, based on the FLT3 fusion we identified using MyAML, we decided to treat the patient with the tyrosine kinase inhibitor sorafenib that we were able to obtain through a patient assistance program. This novel case is the first demonstration of a FLT3 fusion detection with a drug sensitivity assay suggesting that kinase inhibitors with multiple targets might be suitable for this type of variant. Conclusion: Specialized assays designed to identify clonal and subclonal architecture of genes associated with specific diseases can reveal variants that present therapeutic options not currently utilized for high risk patients, suggesting broader use of this approach could improve current clinical outcomes. Disclosures Patay: Invivoscribe, Inc: Consultancy. Carson:Invivoscribe, Inc: Employment. Becker:GlycoMimetics: Research Funding.

scholarly journals Results of CoALL 07-03 study childhood ALL based on combined risk assessment by in vivo and in vitro pharmacosensitivity

2019 ◽  
Vol 3 (22) ◽  
pp. 3688-3699 ◽  
Author(s):  
Franziska Schramm ◽  
Udo zur Stadt ◽  
Martin Zimmermann ◽  
Norbert Jorch ◽  
Arnulf Pekrun ◽  
...  
Keyword(s):  
Drug Testing ◽  
Drug Response ◽  
Lymphoblastic Leukemia ◽  
Term Outcome ◽  
Childhood All ◽  
Long Term Outcome ◽  
Key Points

Key Points Report of the long-term outcome of children with acute lymphoblastic leukemia upon risk-adapted therapy accrued in trial CoALL 07-03. Lack of correlation between in vitro and in vivo drug response as well as a lower predictive value of in vitro drug testing.

A phase I study of GTI-2040 (G), an antisense to ribonucleotide reductase (RNR), in combination with high-dose AraC (HiDAC) in acute myeloid leukemia (AML)

2006 ◽  
Vol 24 (18_suppl) ◽  
pp. 6561-6561
Author(s):  
G. Marcucci ◽  
R. B. Klisovic ◽  
W. Wei ◽  
S. Liu ◽  
P. Paschka ◽  
...  
Keyword(s):  
Phase I ◽  
Ribonucleotide Reductase ◽  
Dose Escalation ◽  
Myeloid Leukemia ◽  
High Dose ◽  
Phase Ii Trials ◽  
Refractory Aml ◽  
Dna Incorporation ◽  
Count Recovery

6561 Background: RNR converts ribonucleotides to deoxyribonucleotides for DNA synthesis. AraC is converted into AraC triphosphate (AraCTP) and competes with deoxycytidine for DNA incorporation. We hypothesized that RNR downregulation by G leads to lower deoxycytidine levels, preferential AraCTP incorporation into DNA and increased cytotoxicity. A CTEP-sponsored Phase I dose escalation study of G +HiDAC in relapsed/refractory AML tested this hypothesis. Methods: Cohort I (18–59 yrs) received G (dose level (DL) 1: 3.5 mg/m2/d) by continuous IV infusion (CIVI) on d 1–6 + AraC IV q12 hrs on d 2, 4, 6 (DL1: 2500 mg/m2/dose). Cohort II (≥60 yrs) received G CIVI on d 1–6 +AraC IV on d 2 −6 (DL1: 1500 mg/m2/d). An ELISA-based assay measured plasma and intracellular concentration (IC) of G. Results: To date, cohort I included 9 pts with relapsed and 9 with refractory AML; 9 had intermediate and 9 adverse risk cytogenetics (CyG); 8 received prior HiDAC. Cohort II included 10 pts with relapsed and 6 with refractory AML; 8 pts had intermediate and 8 high risk CyG; 5 pts received prior HiDAC. Toxicities were comparable to HiDAC alone. The younger pts had higher AUC and longer t1/2. Of 16 pts evaluable in cohort I (median time to 1st relapse 6 mos), 6 had complete remission (CR) and 1 incomplete CR (no disease and incomplete blood count recovery). In cohort II, no responses were observed. At 120 hrs of antisense infusion, median G IC in marrow cells was higher (i.e., 175 vs75 nM) in younger than in older pts. A median 50% decrease in RNR protein was noted in 5/9 and 5/10 pts in cohort I and II, respectively. In cohort I, a median 50% decrease and 200% increase in RNR was noted in CR (n=4) and non-responder (NR; n=9) pts, respectively. In cohort II R2 downregulation did not predict response. In cohort I 62% of the ICs was in nucleus and 21.2% in cytoplasm in CR pts (n=3) vs. 20.3% and 53.5% in NR pts(n=5). Conclusions: G/HiDAC is feasible. Robust plasma and IC levels of G and target downregulation are achievable in vivo. Responses (41%) were observed only in the younger cohort, where CR pts had higher G nuclear IC and target downregulation than NR pts. Dose escalation continues in the younger cohort to establish a dose for Phase II trials. No significant financial relationships to disclose.

scholarly journals Molecular Mechanism and Optimal Treatment Strategy in Acute Myeloid Leukemia with Resistance to Drugs and Radiation By NVP-LED225

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3691-3691 ◽  
Author(s):  
Fanyi Meng ◽  
Xiaodong Li ◽  
Bingjie Ding ◽  
Kaikai Huang ◽  
Qiuhua Zhu ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Myeloid Leukemia ◽  
Hedgehog Signaling ◽  
Leukemia Cells ◽  
Hedgehog Signaling Pathway ◽  
Treatment Groups ◽  
Gli 1 ◽  
Refractory Aml

Abstract PURPOSE: Total body irradiation (TBI) combined with chemotherapy is currently the most effective procedure of traditional preparative myeloablative regimen. However, resistance to chemotherapy in refractory acute myeloid leukemia (AML) is associated with short-time recurrence after Allo-HSCT. To investigate the mechanism of Hedgehog signaling pathway resulting in resistance, we used primary AML cells originated in refractory patients and 3 cell lines including HL60, HL60/ADR (a adriamycin-resistant cells), and HL60/RX (a radiation-resistant cell line established from HL60) as cellular modes to examine the expression of p-IGF-1R, IRS-1, p-Akt, Gli-1, MRP1, Bcl-2, and explored the safety and efficacy of NVP-LDE225 (an inhibitor of Hedgehog pathway) for enhancing the sensitivity to treatment in refractory leukemia in vitro and in vivo. Methods: The expression of Hedgehog signaling pathway was measured in 3 leukemia cell lines, and primary leukemia cells originated in refractory AML patients and non-refractory AML patients using western blot technique. In in vitro experiments, HL60/ADR cells and HL60/RX cells were treated with DMSO (control) or NVP-LDE225 for 48 h, and then assigned in six groups respectively: (a) Control, (b) ADM alone, (c) Radiation alone (d) NVP-LDE225 alone, (e) NVP-LDE225+ADM, (f) NVP-LDE225+Radiation. Flow cytometry, MTT assay and Western-Blot were performed separately to detect apoptosis, adriamycin uptake rate, proliferation inhibit rate and the expression of MRP1, p-IGF-1R, IRS-1, p-Akt, Gli-1, and Bcl-2. Furthermore, based on the results in vitro, the serial tumor volumes, general condition of the mice, complete blood counts, multiple organs injury, and the expression levels of hedgehog signaling pathway were used to detect the antitumor efficacy and toxicity of NVP-LDE225 in HL60/ADR and HL/RX xenograft model. Results: Our result showed that, compared with leukemia cells in non-refractory AML patients and HL60 cells, the expression of hedgehog signaling pathway protein was significantly elevated in refractory AML, HL60/ADR and HL60/RX cells(P<0.01). In addition, we also observed that NVP-LDE225 (10µM) could reverse ADM, DNR, HHT, and Ara-c resistance in HL60/ADR cells. Combining ADM with NVP-LDE225 (20µM) could markedly increase apoptosis and ADM positive rate of intracellular fluorescence detection than other groups (P<0.05). Moreover, combination of NVP-LDE225 with radiation could significantly increase radiation-induced apoptosis and exhibit higher expression of phosphorylation of histone H2AX and BAK than any other single treatment groups both in HL60/RX and HL60/ADR cells(P<0.05). In vivo, both in HL60/RX and HL60/ADR mice model, combination of NVP-LDE225 simultaneously and post 2 days or 5 days with ADM or radiation all presented significantly antitumor effect and high survival rate compared with the control or single agent mice (P<0.01), but there were no significant differences between the three combined treatment groups. Within the combination- and ADM-treated groups, nadir of the white blood cell and platelet counts were reached in day 8 with a gradual recovery starting at day 11. Finally, decrease in IRS-1, Gli-1, p-AKT, and NF-kb expressions were observed after treating with NVP-LDE225 in vitro and in tumor tissue. None of the pathological lesion in the heart, liver, kidney, brain, and lung was observed among all groups. Conclusion: The findings from this study demonstrated that Gli-1/p-Akt/NF-kb pathway play a key role for resistance to both drug and irradiation, and suggested that the combination of hedgehog pathway inhibitor (NVP-LDE225) with chemotherapy or radiotherapy could significantly enhance the antitumor activity via overcoming chemoresistance or radioresistance. The myelosuppression and toxicity could be well tolerated Disclosures No relevant conflicts of interest to declare.

Sign in / Sign up

Export Citation Format

Share Document