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 Custom High Throughput Drug Sensitivity Assay Reveals Therapeutic Options for Chronic Myeloid Leukemia Patients Resistant to or Intolerant of Tyrosine Kinase Inhibitors

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 268-268
Author(s):  
Vivian G. Oehler ◽  
Sylvia Chien ◽  
Jin Dai ◽  
Carrie L. Cummings ◽  
James Annis ◽  
...  
Keyword(s):  
High Throughput ◽  
Myeloid Leukemia ◽  
Research Funding ◽  
Drug Sensitivity ◽  
Kinase Inhibitors ◽  
Blast Phase ◽  
Drug Sensitivity Assay ◽  
Ic50 Values ◽  
Investigational Agents ◽  
New Diagnosis

Abstract Introduction. Tyrosine kinase inhibitors (TKIs) have revolutionized chronic phase (CP) chronic myeloid leukemia (CML) care with many patients achieving major and deeper molecular responses. However, for those who are resistant to or do not tolerate the approved TKIs, there are few alternatives. We therefore developed a custom high throughput drug screen comprised of both FDA approved and investigational agents. Methods. Fifty-six samples (50 individual patients) have undergone testing in the drug sensitivity assay, for which a large fraction exhibited resistance to approved agents. The Quellos High Throughput Core Laboratory's Cancer Drug Sensitivity has been CLIA approved for leukemia since 2015. Blood and bone marrow samples were obtained from CML patients with written informed consent. Mononuclear cells were isolated by density depletion. The myeloid population was obtained by lineage depletion of non-myeloid cells using magnetic beads and antibodies to erythroid lineage (CD235a), T (CD3) and B (CD19) lymphocytes, and NK (CD56) cells. Flow cytometry confirmed successful enrichment of the myeloid cell population. Cells were plated on extracellular matrix coated 384 well plates to test under conditions of adhesion mediated chemotherapy resistance. Initially, the assay was comprised of 32 drugs (11 patients) selected based on published activity in CML and resistant CML. The assay was then expanded to 64 drugs. Compounds are added (ranging from 5 pM to 100 μM) to patient samples using the CyBio CyBi-Well Vario and incubated at 37°C, 5% CO2 for 72 hours, then viability is assessed by CellTiterGlo. IC50s and AUCs are calculated for each drug using XLFit (IDBS) and a standard 4 parameter logistical model. Transcriptome analysis is planned for these samples. Results. Clinical characteristics are shown in Table 1. Mean and median BCR-ABL1 transcripts were 69% and 70% in diagnosis samples and 63% and 55% in resistant samples, respectively (P=0.607). ABL mutations were present in 5 patients (M244V, T315I, F359I). Additional myeloid mutations were present in 5 of 6 evaluable advanced phase samples, 4 of 17 evaluable diagnostic samples, and 3 of 10 evaluable resistant samples and included ASXL1, DNMT3A, IDH1, JAK2V617F, NRAS, RUNX1, and TET2. Figure 1 illustrates the breadth of sensitivity to agents in the assay. Figure 2 is a heat map of area under the curve (AUCs) illustrating the unique drug sensitivity patterns for all patients, with unsupervised clustering. For new diagnosis patients, the TKIs imatinib, dasatinib, nilotinib, bosutinib, and ponatinib ranked in the top 8 drugs. For primary resistant patients, the IC50 values for imatinib, nilotinib, bosutinib, and ponatinib were higher than the new diagnosis patients. For example, for ponatinib, the mean IC50 was 402.6 ± 354.7 X 10 E-9 M for primary resistant samples vs. 1.65 ± 0.45 X 10 E-9 M for diagnosis group, p=0.015 (Welch t test), or about 250-fold higher (less sensitive). In accelerated and blast phase samples drugs with IC50 values lower than 0.1 µM, a range that could correlate with in vivo drug response, were identified for all patients (range, 3-20 drugs per patient). Top candidates included proteasome and kinase inhibitors. In 2 patients harboring NRAS mutations, IC50 for trametinib was less than 0.1 µM as compared to patients without NRAS mutations, where the IC50s were higher. Clinical outcomes are available for nearly all patients. Although the study was not designed to select next-line TKI therapy in resistant patients, drug profiling was informative in many cases. Data for 7 resistant patients are shown in Table 2. For example, CML-012 had the lowest IC50 value (indicating most sensitive) for dasatinib, 4.1 X 10E-8 M and responded to dasatinib after failing imatinib (IC50 8.4 X 10E-7M). CML-003 did not respond to bosutinib (IC50 1.2 X 10E-6M) and did respond to dasatinib (IC50 1.2 X 10E-7M). CML-056 did not respond to nilotinib (IC50 1.4 X 10E-6M), dasatinib (IC50 6.9 X 10E-4M), or ponatinib (IC50 1.0 X 10E-6M). Notably, in all resistant patient samples we identified drugs with IC50 values lower than 0.1 µM. These therapeutics can be prioritized for further evaluation, either alone or in combination with TKIs, in resistant CML patients. Conclusion. In vitro drug sensitivity testing provides data for potential agents for patients with resistance or intolerance to FDA approved TKIs, or those that have entered accelerated phase or blast phase. Figure 1 Figure 1. Disclosures Oehler: Blueprint Medicines: Consultancy; Takeda: Consultancy; Pfizer: Research Funding; OncLive: Honoraria; BMS: Consultancy. Becker: Abbie: Research Funding; SecuraBio: Research Funding; Cardiff Oncology: Research Funding; Pfizer: Research Funding; BMS: Research Funding; CVS Caremark: Consultancy; Glycomimetics: Research Funding. OffLabel Disclosure: We developed a custom high throughput drug screen comprised of both FDA approved and investigational agents

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.

Response rates in patients with relapsed/refractory acute myeloid leukemia with FLT3-ITD mutation using 5-azacitadine plus sorafenib.

2012 ◽  
Vol 30 (30_suppl) ◽  
pp. 24-24 ◽  
Author(s):  
Mona Lisa Alattar ◽  
Hagop Kantarjian ◽  
Mark J. Levis ◽  
Mary Ann Richie ◽  
Naval Guastad Daver ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Intensive Therapy ◽  
Response Assessment ◽  
Flt3 Ligand ◽  
Refractory Acute Myeloid Leukemia ◽  
Putative Mechanism ◽  
Acute Myeloid

24 Background: Outcome of patients with relapsed acute myeloid leukemia (AML) with FLT3-ITD mutation is poor. Elevated FLT3 ligand levels (FL) secondary to cytotoxic chemotherapy is a putative mechanism of resistance to the FLT3 kinase inhibitors. We hypothesized that FL levels will be lower when combining sorafenib with less intensive therapy such as 5-Azacytidine (5-Aza). This study was conducted to evaluate the efficacy and tolerability of the combination of sorafenib and 5-Aza in patients with refractory or relapsed AML with FLT3-ITD mutation. Methods: Patients were eligible if they had relapsed or refractory AML and were 18 years of age or older. They received 5-Aza 75 mg/m2 IV daily x 7 days and sorafenib 400 mg PO BID continuously; cycles were repeated in approximately one month intervals. Results: 38 patients with AML with a median age of 60 years (range, 24-87) were enrolled. 4 were inevaluable as they never received therapy or discontinued it before response assessment. FLT-3-ITD was detected in 30 (88%) patients. They had received a median of 2 prior treatments (range, 0-6); 13 (38%) had received ≥ 3 prior regimens and 8 had failed prior FLT3 kinase inhibitor. Response rate was 41%, including 6 (17%) with CRi, 4 (12) with CRp, 3 (9%) with CR, and 1(3%) PR. The median time to achieving CR/CRi was 2 cycles (Range, 1-6) and the median duration of CR/CRi was 3 months (Range, 1–12). Conclusions: Combination of 5-Aza and sorafenib is effective for patients with relapsed AML with FLT-3-ITD mutation.

Combination of Sorafenib and 5-Azacytidine Has Significant Activity in Patients with Relapsed/Refractory or Untreated Acute Myeloid Leukemia and FLT3-ITD mutation

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1519-1519
Author(s):  
Farhad Ravandi ◽  
Mona Lisa Alattar ◽  
Mark J. Levis ◽  
Guillermo Garcia-Manero ◽  
Mary A Richie ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Research Funding ◽  
Kinase Inhibitors ◽  
Response Assessment ◽  
Cytotoxic Chemotherapy ◽  
Off Label Use ◽  
Off Label ◽  
Acute Myeloid ◽  
Refractory Aml

Abstract Abstract 1519 Background: The outcome of patients (pts) with acute myeloid leukemia (AML) and FLT3-ITD mutation is poor, particularly in the relapse setting. Sorafenib is a potent inhibitor of FLT3 kinase with reported clinical activity as a single agent (Metzelder S, Blood, 2009), and in combination with chemotherapy (Ravandi F, JCO, 2010). A potential mechanism of resistance to FLT3 kinase inhibitors is high levels of FLT3 ligand (FL) as seen after myelosuppressive chemotherapy. We hypothesized that combining sorafenib with a less myelosuppressive agent, such as 5-azacytidine (AZA), may lead to higher and more durable responses than cytotoxic chemotherapy. Furthermore, both drugs have demonstrated a potential for inducing differentiation in AML cells, thereby providing further rationale for the combination. Methods: Pts were eligible if they had relapsed or refractory AML, were 18 years of age or older, and had adequate performance status (ECOG ≤ 2) and organ function. Older pts without prior therapy were also eligible, if they were deemed unsuitable to receive chemotherapy. Presence of FLT3-ITD was not a requirement but these pts were targeted for enrollment. Treatment regimen included AZA 75 mg/m2 daily for 7 days together with sorafenib 400 mg twice daily for 28 days; cycles were repeated in approximately 4 to 5-week intervals. Overall responses were assessed after the completion of at least one cycle of therapy and at the time of the best peripheral blood and bone marrow response. Plasma samples were collected on approximately Day 1 and Day 10 of each cycle. To assess the degree of FLT3 inhibition, the plasma inhibitory activity (PIA) assay was performed using the Molm-14 cell line (Levis M, Blood, 2006). Plasma FL concentrations were measured using an ELISA kit (R&D Systems). Results: 43 pts with AML with a median age of 64 years (range, 24–87) were enrolled. They included 19 (44%) pts with diploid cytogenetics, 11 (26%) with chromosome 5/7 or complex cytogenetic abnormalities, and 13 (30%) with miscellaneous abnormalities. Prior to the initiation of treatment, FLT3-ITD was detected in 40/43 (93%) pts with a median allelic ratio of 0.28 (range, 0 – 0.93). They had received a median of 2 prior treatments (range, 0–7). 16 (37%) pts had received ≥3 prior regimens and 9 had failed therapy with FLT3 kinase inhibitors (5 with AC220, 1 with PKC412, and 6 with sorafenib, either as monotherapy or with chemotherapy or plerixafor); 3 had failed 2 prior FLT3 inhibitors. 6 pts were inevaluable as they discontinued therapy before response assessment at one month and 3 are too early for response assessment. The overall CR/CRi/PR rate among the 34 evaluable pts is 44%, including 10 (29%) with CRi and 4 (12%) with CR and 1 (3%) with PR (in this pt, bone marrow blast declined from 51% to 6% with normalization of blood counts). Overall, pts have received a median of 3 (range, 1–9) treatment cycles with the median number of cycles to response among the responders being 2 (range, 1 – 4) and the median time to achieving response, 2.1 months (range, 0.9 – 4.6 months). The median duration of CR/CRi Is 2.3 months (range, 1 – 12.2+ months). Six pts have proceeded to allogeneic stem cell transplant. The most common study drug-related adverse events were rash and fatigue with no deaths attributable to study medications. One pt developed grade 3 cardiomyopathy suspected to be related to the study regimen. Of the 34 pts included in the clinical analysis, there were 22 pts from whom plasma samples spanning at least one cycle of therapy were available. Among them, 64% achieved FLT3 inhibition to a targeted level of less than 15% of baseline during their first cycle of therapy. Median survival in pts who achieved this degree of inhibition was 238 days, while median survival in pts who did not reach this level was 154 days (p=0.13). Mean FL levels at cycle 1, day 1 and cycle 1, day 10 were 9 pg/mL and 17 pg/mL, respectively. Mean FL levels at cycle 2, day 0 and cycle 2, day 10 were 27 pg/mL and 54 pg/mL, respectively. Conclusions: Combination of AZA and Sorafenib is effective for the treatment of older pts and pts with relapsed and refractory AML and FLT3-ITD mutation. While not statistically significant, there was a trend toward improved survival in pts with adequate FLT3 inhibition during cycle 1. FL levels did not rise to the levels seen in pts receiving cytotoxic chemotherapy. Disclosures: Ravandi: Bayer/Onyx: Honoraria, Research Funding; Celgene: Honoraria, Research Funding. Off Label Use: Off-label use of sorafenib and 5-azacytidine in patients with acute myeloid leukemia. Levis:Astellas Pharma: Consultancy; Plexxikon: Consultancy; Symphogen: Consultancy. Garcia-Manero:Celgene: Research Funding. Andreeff:Hoffmann-La Roche: Research Funding; Karyopharm Therapeutics: Unrestricted gift, Unrestricted gift Other. Cortes:Celgene: Research Funding.

Personalized Approach To Treatment of Acute Myeloid Leukemia Using a High-Throughput Chemosensitivity Assay

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 483-483 ◽  
Author(s):  
Ka Yee Yeung ◽  
C. Anthony Blau ◽  
Vivian G. Oehler ◽  
Su-In Lee ◽  
Christopher P. Miller ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Complete Remission ◽  
High Throughput ◽  
Myeloid Leukemia ◽  
In Vitro Cytotoxicity ◽  
Personalized Approach ◽  
New Diagnosis ◽  
Acute Myeloid ◽  
Refractory Aml

Most patients with acute myeloid leukemia either relapse or fail to respond to initial therapy. Moreover, each patient’s AML contains multiple mutations that although varying from one population to another, are unique to that individual, prompting development of individualized approaches to AML therapy. We have devised a high-throughput sensitivity assay for 160 drugs; 45 are FDA approved and 115 are investigational, representing multiple mechanisms of action and signaling pathways. 30 primary patient blood marrow samples and 15 acute leukemia cell lines have been analyzed. Peripheral blood blasts from individual patients were thawed, viable cells isolated and purified to >80% using magnetic bead separations. In vitro chemotherapy cytotoxicity testing is performed in a 384 well high throughput format, with eight concentrations of each drug. The output is cell survival assessed via a luminescent detection method after a 4-day incubation with various drugs. Fitted curves (idbs XLfit) were derived from plots of survival versus drug concentrations used in the study. Leukemia cells are tested adherent to coated plates to mimic adhesion in the bone marrow microenvironment, a property that confers drug resistance. The assay exhibits excellent reproducibility from independently thawed samples from the same patients, with a Spearman correlation coefficient of 0.9 (p=1.6 X 10-141). Gene expression microarrays were also performed for the same 30 patients. Our assay of the 30 patients revealed that there were over 50 drugs that exhibited cytotoxicity in at least some patients. There was wide variation in the drug sensitivity patterns exhibited by the patient blasts samples, and each was unique. Notably, all patient samples were susceptible to several drugs with IC50s in the range that might predict clinical response. For patients who achieved complete remission, we showed statistically significant association with in vitro cytotoxicity in response to 7 drugs that are commonly employed to treat AML, including mitoxantrone (p=0.002 for 0.1 µM, p=0.01 for 0.3 µM), clofarabine (p=0.0009 for 0.1 µM, p=0.003 for 0.3 µM, p=0.007 for 1µM), daunorubicin (p=003 for 0.1 µM, p=0.005 for 0.3 µM), etoposide (p=0.02 at for 0.1 µM, p=0.01 for 0.3 µM, p=0.01 for 1 µM), and fludarabine (p=0.05 for 0.3 µM, p=0.002 for 1 µM). In addition, we used a multivariate statistical method to identify drug combinations that are effective in predicting the complete remission of AML patients. In particular, we found that the in vitro cytotoxicity data of a combination of three drugs (BAY 11-7085, TPCA-1, ON 01910.Na) are more predictive of the complete remission of patients compared to using each of these drugs individually (i.e. 91.2% accuracy compared to 84.8%, 84.2%, 79.1% respectively). We also performed linear regression analyses to study the relationship between in vitro cytotoxicity in the high throughput screen of each drug versus the patients’ clinical features (including complete remission, having received the tested drug, new diagnosis vs. relapse status, age, gender, bilirubin, albumin, lactate dehydrogenase, white blood count, platelet count, blast, absolute neutrophil count, % bone marrow blasts, hemoglobin, fibrinogen, cytogenetics risk category). The following patients’ clinical features showed significant correlations with the in vitro cytotoxicity of our assays in selected drugs: complete remission for mitoxantrone (p= 0.04 for 0.3 µM), flavoperidol (p= 0.02 0.1 µM), and fludarabine (p= 0.01 0.3 µM, p=0.02 for 1 µM); drug the patient received for clofarabine (p= 0.03 for 0.3 µM, p=0.007 for 1 µM); new diagnosis vs. relapse for clofarabine (p= p= 0.02 for 0.3 µM, p=0.006 for 1 µM). This assay serves as the basis for a new clinical trial (ClinicalTrials.gov Identifier: NCT01872819) now open to enrollment for “personalized” leukemia therapy, for which patients with refractory AML are assigned drugs based on the results of this test. This new personalized approach to individualized therapy for refractory AML may provide novel drugs to patients and new insights into leukemia drug resistance. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Azacitidine and lenalidomide as an alternative treatment for refractory acute myeloid leukemia: a case report

2014 ◽  
Vol 133 (3) ◽  
pp. 271-274 ◽  
Author(s):  
Juliana Todaro ◽  
Patrícia Weinschenker Bollmann ◽  
Edna Terezinha Rother ◽  
Auro del Giglio
Keyword(s):  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Therapeutic Option ◽  
Monosomy 7 ◽  
Refractory Acute Myeloid Leukemia ◽  
Third Line ◽  
Line Chemotherapy ◽  
Monosomal Karyotype ◽  
Acute Myeloid ◽  
Refractory Aml

CONTEXT:Refractory acute myeloid leukemia (AML) is a difficult disease to control with second or third-line chemotherapy regimens. In this report, we describe using azacitidine in combination with lenalidomide as salvage therapy.CASE REPORT:52-year-old female was diagnosed with refractory AML and high-risk cytogenetics: complex monosomal karyotype consisting of t (3, 3) in association with monosomy 7 and del 5q. Morphological remission associated with maintenance of the cytogenetic abnormality of chromosome 3 and disappearance of the abnormalities relating to chromosomes 5 and 7 was achieved after three cycles of combination therapy with azacitidine and lenalidomide.CONCLUSION:Azacitidine plus lenalidomide can be a therapeutic option for patients with refractory AML, as illustrated in this case.

scholarly journals Computational analysis of ABL kinase mutations allows predicting drug sensitivity against selective kinase inhibitors

Tumor Biology ◽  
2017 ◽  
Vol 39 (5) ◽  
pp. 101042831770164 ◽  
Author(s):  
Swapna Kamasani ◽  
Sravani Akula ◽  
Sree Kanth Sivan ◽  
Vijjulatha Manga ◽  
Justus Duyster ◽  
...  
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Large Scale ◽  
Drug Sensitivity ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Computational Study ◽  
Significant Proportion ◽  
Point Mutations ◽  
Abl Kinase

The ABL kinase inhibitor imatinib has been used as front-line therapy for Philadelphia-positive chronic myeloid leukemia. However, a significant proportion of imatinib-treated patients relapse due to occurrence of mutations in the ABL kinase domain. Although inhibitor sensitivity for a set of mutations was reported, the role of less frequent ABL kinase mutations in drug sensitivity/resistance is not known. Moreover, recent reports indicate distinct resistance profiles for second-generation ABL inhibitors. We thus employed a computational approach to predict drug sensitivity of 234 point mutations that were reported in chronic myeloid leukemia patients. Initial validation analysis of our approach using a panel of previously studied frequent mutations indicated that the computational data generated in this study correlated well with the published experimental/clinical data. In addition, we present drug sensitivity profiles for remaining point mutations by computational docking analysis using imatinib as well as next generation ABL inhibitors nilotinib, dasatinib, bosutinib, axitinib, and ponatinib. Our results indicate distinct drug sensitivity profiles for ABL mutants toward kinase inhibitors. In addition, drug sensitivity profiles of a set of compound mutations in ABL kinase were also presented in this study. Thus, our large scale computational study provides comprehensive sensitivity/resistance profiles of ABL mutations toward specific kinase inhibitors.

Sorafenib as a Salvage Therapy in FLT3-ITD Negative Relapse/ Refractory Acute Myeloid Leukemia

2019 ◽  
Vol 18 (10) ◽  
pp. 1489-1494
Author(s):  
Nan Yang ◽  
Zhenyang Gu ◽  
Zhanxiang Liu ◽  
Wenrong Huang ◽  
Shuhong Wang ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Salvage Therapy ◽  
Myeloid Leukemia ◽  
Kinase Inhibitor ◽  
Residual Disease ◽  
Disease Status ◽  
Therapeutic Effects ◽  
Cns Relapse ◽  
Acute Myeloid ◽  
Refractory Aml

Background: Multi-kinase inhibitor sorafenib showed dramatic effects in acute myeloid leukemia (AML) cells harboring fms-related tyrosine kinase 3-internal tandem duplication (FLT3-ITD) mutation. However, FLT3-ITD mutation only occurs in 25% of AML cases. The therapeutic effects of sorafenib in AML patients without FLT3-ITD are still in need of further investigation. Methods: A young AML patient with central nervous system (CNS) relapse was treated with sorafenib combined with chemotherapy. Another patient with refractory AML arising form chronic myelomonocytic leukemia (CMML) was treated with sorafenib monotherapy. Spinal and cranial magnetic resonance imaging (MRI), minimal residual disease (MRD) and peripheral blood cell count were monitored to evaluate disease status. Results: The patient with CNS relapse exhibited significant shrink of tumor volume. The other patient with refractory AML achieved hematological improvements. Conclusion: These two cases suggested that sorafenib might be utilized as a potent salvage therapy for some refractory/relapsed AML patients without the FLT3-ITD mutation.

Toward Individualized Therapy: Correlation of Mutation Analysis with in Vitro high Throughput Drug Sensitivity Testing in New Diagnosis and Relapsed Acute Myeloid Leukemia

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3851-3851
Author(s):  
Pamela S. Becker ◽  
Michael W. Schmitt ◽  
Zhiyi Xie ◽  
Andrew R Carson ◽  
Bradley Patay ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
High Throughput ◽  
Myeloid Leukemia ◽  
Drug Sensitivity ◽  
Missense Mutations ◽  
Sensitivity Testing ◽  
New Diagnosis ◽  
Duplex Sequencing ◽  
Acute Myeloid

Abstract Introduction: Whole genome sequencing has demonstrated tremendous heterogeneity in the mutations and chromosomal translocations associated with acute myeloid leukemia (AML), and there are several correlates with prognosis, yet we remain quite limited in our ability to predict specific chemotherapy drug sensitivity based on genomics with the exception of a few selected mutations or translocations, such as FLT3 -ITD or PML-RARA. One third of new diagnosis patients and over half of relapsed patients will not respond to initial chemotherapy regimens that incur appreciable toxicity and result in prolonged hospitalization. We therefore seek to define molecular information that might better predict response to conventional or novel therapies. Methods: MyAML™ uses next generation sequencing (NGS) to analyze the 3' and 5' UTR and exonic regions of 194 genes and potential genomic breakpoints within known somatic gene fusion breakpoints known to be associated with AML. Fragmented genomic DNA (~3.4Mb) is captured with a customized probe design, and sequenced with 300bp paired end reads on an Illumina MiSeq instrument to an average depth of coverage >1000x. Using a custom bioinformatics pipeline, MyInformatics™, single nucleotide variants (SNVs), insertion/deletions (indels), inversions and translocations are identified, annotated, characterized, and allelic frequencies calculated. Commonly associated variants in dbSNP and 1000 genomes may be eliminated, as well as variants with allele frequencies less than 5%. High throughput drug sensitivity testing was performed against a panel of 160 drugs, of which 56 are FDA approved and 104 are investigational. De-identified samples from 12 patients with de novo AML and 12 patients with relapsed AML were analyzed. For 2 patient samples, Duplex Sequencing was also performed to detect sub-clonal mutations below the detection limit of conventional NGS. Pearson and Spearman correlations were performed between all possible pairs of genes containing missense or indel mutations and the in vitro cytotoxicity response across the same set of 24 patients. Results: From the 24 patient samples analyzed to date, an average of 129 missense mutations were identified in each sample with an allelic frequency >5%. Of these, an average of over 21 missense variants were observed in COSMIC and less than 3 were novel (not in dbSNP). These samples also contained an average of over 12 coding indels (~5 frameshift and 7 inframe indels per sample). In addition, MyAML™ identified 3 samples with inv(16) and 6 samples with translocations, including the cryptic NUP98-NSD1 t(5;11) that was not detected by karyotyping. For 2 of the samples, Duplex Sequencing was performed at a depth of at least 6000X, and an accuracy of 10-7, and showed concordance of some of the mutations, with each method identifying additional mutations not observed by the other, an expected finding, as each method targeted distinct regions, and Duplex Sequencing had a greater depth of coverage. Fourteen genes were observed to exhibit at least one indel with a frameshift at frequency greater than 5% in more than one patient. In order to identify significantly associated drugs and genes containing indel mutations, we computed Pearson and Spearman correlations between drugs and these 14 genes across 24 patients. The correlation analyses revealed significant associations (p= 0.006 to 0.04) between indel mutations in three genes and chemosensitivity to drugs commonly used in AML such as cladribine, clofarabine, cytarabine, daunorubicin, etoposide, fludarabine and mitoxantrone. Similarly, significant associations (p<0.05) were identified between missense mutations in 5 genes and chemosensitivity to these drugs. Conclusion: Personalized data derived from a targeted genomic assay and in vitro chemotherapy sensitivity testing of individual patient AML samples will likely lead to innovation in treatment, identification of novel targeted agents, and improved outcomes in AML. Disclosures Xie: Invivoscribe: Employment. Carson:Genection: Employment. Patay:Genection: Employment.

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