Single-cell DNA sequencing reveals complex mechanisms of resistance to quizartinib.
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ABSTRACT:
SUBMITTER: Peretz CAC
PROVIDER: S-EPMC7948271 | biostudies-literature | 2021 Mar
REPOSITORIES: biostudies-literature
Publications
Single-cell DNA sequencing reveals complex mechanisms of resistance to quizartinib.
Peretz Cheryl A C CAC McGary Lisa H F LHF Kumar Tanya T Jackson Hunter H Jacob Jose J Durruthy-Durruthy Robert R Levis Mark J MJ Perl Alexander A Huang Benjamin J BJ Smith Catherine C CC
Blood advances 20210301 5
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Project description:Single cell DNA sequencing reveals complex mechanisms of resistance to quizartinib
| PRJNA687071 | ENA
Project description:Single cell DNA sequencing reveals complex mechanisms of resistance to quizartinib
| PRJNA687070 | ENA
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Project description:BackgroundBRAF 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.MethodsWe 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.ResultsWe 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.ConclusionsOur 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.
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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.
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Project description:Viruses deploy numerous strategies to infect plants, typically by forming complexes with another virus, leading to more efficient infection. However, the detailed plant responses to viral infection and the underlying mechanisms of co-infection remain unclear. Previously, we found that tomato spotted wilt orthotospovirus (TSWV) and Hippeastrum chlorotic ringspot orthotospovirus (HCRV) could infect plants in the field by forming a complex. In this study, we found that TSWV infected tobacco (Nicotiana benthamiana) plants in cooperation with HCRV, leading to a more efficient infection rate of both viruses. We then used the in-depth full-length transcriptome to analyze the responses of N. benthamiana to complex infection by TSWV-HCRV (TH). We found that infection with individual TSWV and HCRV triggered plant defense responses, including the jasmonic acid signaling pathway, autophagy, and secondary metabolism. However, TH co-infection could not trigger and even suppress some genes that are involved in these basal resistance responses, suggesting that co-infection is advantageous for the virus and not for the plants. Typically, the TH complex inhibits NbPR1 expression to suppress tobacco resistance. Moreover, the TH complex could alter the expression of microRNAs (miRNAs), especially novel-m0782-3p and miR1992-3p, which directly interact with NbSAM and NbWRKY6 and suppress their expression in tobacco, leading to downregulation of NbPR1 and loss of resistance in tobacco to TSWV and HCRV viruses. Overall, our results elucidated the co-infection mechanisms of TH in tobacco by deploying the miRNA of plants to suppress plant basal resistance and contributed to developing a novel strategy to control crop disease caused by this virus complex.
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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.
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