Single-cell DNA sequencing reveals complex mechanisms of resistance to quizartinib.
Ontology highlight
ABSTRACT:
SUBMITTER: Peretz CAC
PROVIDER: S-EPMC7948271 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Single cell DNA sequencing reveals complex mechanisms of resistance to quizartinib
| PRJNA687070 | ENA
Project description:Single cell DNA sequencing reveals complex mechanisms of resistance to quizartinib
| PRJNA687071 | ENA
Project description:BRAF is mutated in ?42% of human melanomas (COSMIC. http://www.sanger.ac.uk/genetics/CGP/cosmic/) and pharmacological BRAF inhibitors such as vemurafenib and dabrafenib achieve dramatic responses in patients whose tumours harbour BRAF(V600) mutations. Objective responses occur in ?50% of patients and disease stabilisation in a further ?30%, but ?20% of patients present primary or innate resistance and do not respond. Here, we investigated the underlying cause of treatment failure in a patient with BRAF mutant melanoma who presented primary resistance.We carried out whole-genome sequencing and single nucleotide polymorphism (SNP) array analysis of five metastatic tumours from the patient. We validated mechanisms of resistance in a cell line derived from the patient's tumour.We observed that the majority of the single-nucleotide variants identified were shared across all tumour sites, but also saw site-specific copy-number alterations in discrete cell populations at different sites. We found that two ubiquitous mutations mediated resistance to BRAF inhibition in these tumours. A mutation in GNAQ sustained mitogen-activated protein kinase (MAPK) signalling, whereas a mutation in PTEN activated the PI3 K/AKT pathway. Inhibition of both pathways synergised to block the growth of the cells.Our analyses show that the five metastases arose from a common progenitor and acquired additional alterations after disease dissemination. We demonstrate that a distinct combination of mutations mediated primary resistance to BRAF inhibition in this patient. These mutations were present in all five tumours and in a tumour sample taken before BRAF inhibitor treatment was administered. Inhibition of both pathways was required to block tumour cell growth, suggesting that combined targeting of these pathways could have been a valid therapeutic approach for this patient.
| S-EPMC3999800 | biostudies-other
Project description:A frightening resurgence of bed bug infestations has occurred over the last 10 years in the U.S. and current chemical methods have been inadequate for controlling this pest due to widespread insecticide resistance. Little is known about the mechanisms of resistance present in U.S. bed bug populations, making it extremely difficult to develop intelligent strategies for their control. We have identified bed bugs collected in Richmond, VA which exhibit both kdr-type (L925I) and metabolic resistance to pyrethroid insecticides. Using LD(50) bioassays, we determined that resistance ratios for Richmond strain bed bugs were ?5200-fold to the insecticide deltamethrin. To identify metabolic genes potentially involved in the detoxification of pyrethroids, we performed deep-sequencing of the adult bed bug transcriptome, obtaining more than 2.5 million reads on the 454 titanium platform. Following assembly, analysis of newly identified gene transcripts in both Harlan (susceptible) and Richmond (resistant) bed bugs revealed several candidate cytochrome P450 and carboxylesterase genes which were significantly over-expressed in the resistant strain, consistent with the idea of increased metabolic resistance. These data will accelerate efforts to understand the biochemical basis for insecticide resistance in bed bugs, and provide molecular markers to assist in the surveillance of metabolic resistance.
| S-EPMC3198472 | biostudies-literature
Project description:FLT3-ITD mutations occur in 20-30% of AML patients and are associated with aggressive disease. Patients with relapsed FLT3-mutated disease respond well to 2nd generation FLT3 TKIs but inevitably relapse within a short timeframe. In this setting, until overt relapse occurs, the bone marrow microenvironment facilitates leukemia cell survival despite continued on-target inhibition. We demonstrate that human bone marrow derived conditioned medium (CM) protects FLT3-ITD+ AML cells from the 2nd generation FLT3 TKI quizartinib and activates STAT3 and STAT5 in leukemia cells. Extrinsic activation of STAT5 by CM is the primary mediator of leukemia cell resistance to FLT3 inhibition. Combination treatment with quizartinib and dasatinib abolishes STAT5 activation and significantly reduces the IC50 of quizartinib in FLT3-ITD+ AML cells cultured in CM. We demonstrate that CM protects FLT3-ITD+ AML cells from the inhibitory effects of quizartinib on glycolysis and that this is partially reversed by treating cells with the combination of quizartinib and dasatinib. Using a doxycycline-inducible STAT5 knockdown in the FLT3-ITD+ MOLM-13 cell line, we show that dasatinib-mediated suppression of leukemia cell glycolytic activity is STAT5-independent and provide a preclinical rationale for combination treatment with quizartinib and dasatinib in FLT3-ITD+ AML.
| S-EPMC7606260 | biostudies-literature
Project description:Objective: Mantle cell lymphoma (MCL) is a rare subtype of non-Hodgkin lymphoma (NHL) with high heterogeneity and a high recurrence rate. How heterogenous cell populations contribute to relapse remains to be elucidated. Methods: We performed single cell RNA sequencing (scRNA-seq) on approximately 4,000 bone marrow cells sampled from one patient with multidrug resistant MCL. We identified 10 subpopulations comprising 4 malignant B cell subtypes, 3 T cell subtypes, 2 dendritic cell subtypes and 1 natural killer (NK) cell subtype. Subsequently, we identified cell markers, including a series of genes associated with immune escape and drug resistance. In addition, we explored the roles of these genes in the mechanism of immune escape and drug resistance, and we verified the expression imbalance and clinical prognostic potential by using GEO datasets including 211 MCL samples. Results: The major immune escape mechanisms of MCL included anti-perforin activity, decreased immunogenicity and direct inhibition of apoptosis and cell killing, as mediated by type I and II B cells. The drug resistance mechanisms of different cell clusters included drug metabolism, DNA damage repair, apoptosis and survival promotion. Type III B cells closely communicate with other cells. The key genes involved in the resistance mechanisms showed dysregulated expression and may have significant clinical prognostic value. Conclusion: This study investigated potential immune escape and drug resistance mechanisms in MCL. The results may guide individualized treatment and promote the development of therapeutic drugs.
| S-EPMC7476085 | biostudies-literature
Project description:Metastasis is a complex biological process that has been difficult to delineate in human colorectal cancer (CRC) patients. A major obstacle in understanding metastatic lineages is the extensive intra-tumor heterogeneity at the primary and metastatic tumor sites. To address this problem, we developed a highly multiplexed single-cell DNA sequencing approach to trace the metastatic lineages of two CRC patients with matched liver metastases. Single-cell copy number or mutational profiling was performed, in addition to bulk exome and targeted deep-sequencing. In the first patient, we observed monoclonal seeding, in which a single clone evolved a large number of mutations prior to migrating to the liver to establish the metastatic tumor. In the second patient, we observed polyclonal seeding, in which two independent clones seeded the metastatic liver tumor after having diverged at different time points from the primary tumor lineage. The single-cell data also revealed an unexpected independent tumor lineage that did not metastasize, and early progenitor clones with the "first hit" mutation in APC that subsequently gave rise to both the primary and metastatic tumors. Collectively, these data reveal a late-dissemination model of metastasis in two CRC patients and provide an unprecedented view of metastasis at single-cell genomic resolution.
| S-EPMC5538546 | biostudies-literature
Project description:Measuring multiple omics profiles from the same single cell opens up the opportunity to decode molecular regulation that underlies intercellular heterogeneity in development and disease. Here, we present co-sequencing of microRNAs and mRNAs in the same single cell using a half-cell genomics approach. This method demonstrates good robustness (~95% success rate) and reproducibility (R2 = 0.93 for both microRNAs and mRNAs), yielding paired half-cell microRNA and mRNA profiles, which we can independently validate. By linking the level of microRNAs to the expression of predicted target mRNAs across 19 single cells that are phenotypically identical, we observe that the predicted targets are significantly anti-correlated with the variation of abundantly expressed microRNAs. This suggests that microRNA expression variability alone may lead to non-genetic cell-to-cell heterogeneity. Genome-scale analysis of paired microRNA-mRNA co-profiles further allows us to derive and validate regulatory relationships of cellular pathways controlling microRNA expression and intercellular variability.
| S-EPMC6327095 | biostudies-literature
Project description:The treatment of chronic myeloid leukemia (CML), a disease caused by t(9;22)(q34;q11) reciprocal translocation, has advanced largely through the use of targeted tyrosine kinase inhibitors (TKIs). To identify molecular differences that might distinguish TKI responders from non-responders, we performed single cell RNA sequencing on cells (n = 41,723 cells) obtained from the peripheral blood of four CML patients at different stages of treatment to generate single cell expression profiles. Analysis of our single cell expression profiles in conjunction with those previously obtained from the bone marrow of additional CML patients and healthy donors (total = 69,263 cells) demonstrated that imatinib treatment significantly altered leukocyte population compositions in both responders and non-responders, and affected the expression profiles of multiple cell populations, including non-neoplastic cell types. Notably, in imatinib poor-responders, patient-specific pre-treatment unique stem/progenitor cells became enriched in peripheral blood compared to the responders. These results indicate that resistance to TKIs might be intrinsic in some CML patients rather than acquired, and that non-neoplastic immune cell types may also play vital roles in dispersing the responsiveness of patients to TKIs. Furthermore, these results demonstrated the potential utility of peripheral blood as a diagnostic tool in the TKI sensitivity of CML patients.
| S-EPMC7803567 | biostudies-literature
Project description:T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive leukemia that is most frequent in children and is characterized by the presence of few chromosomal rearrangements and 10 to 20 somatic mutations in protein-coding regions at diagnosis. The majority of T-ALL cases harbor activating mutations in NOTCH1 together with mutations in genes implicated in kinase signaling, transcriptional regulation, or protein translation. To obtain more insight in the level of clonal heterogeneity at diagnosis and during treatment, we used single-cell targeted DNA sequencing with the Tapestri platform. We designed a custom ALL panel and obtained accurate single-nucleotide variant and small insertion-deletion mutation calling for 305 amplicons covering 110 genes in about 4400 cells per sample and time point. A total of 108 188 cells were analyzed for 25 samples of 8 T-ALL patients. We typically observed a major clone at diagnosis (>35% of the cells) accompanied by several minor clones of which some were less than 1% of the total number of cells. Four patients had >2 NOTCH1 mutations, some of which present in minor clones, indicating a strong pressure to acquire NOTCH1 mutations in developing T-ALL cells. By analyzing longitudinal samples, we detected the presence and clonal nature of residual leukemic cells and clones with a minor presence at diagnosis that evolved to clinically relevant major clones at later disease stages. Therefore, single-cell DNA amplicon sequencing is a sensitive assay to detect clonal architecture and evolution in T-ALL.
| S-EPMC7885827 | biostudies-literature