Dataset Information

Phosphoproteomic profiling of D816V-KIT cells

ABSTRACT: Acute Myeloid Leukaemia (AML) carries a 5 year survival rate of just 24%. Toxic chemotherapy regimens remain the backbone of standard of care for AML. The KIT tyrosine kinase is a recognised AML oncogene, associated with poor outcome. We recently identified DNA-PK as a novel therapeutic target in FLT3 mutant AML. The similarity between KIT and FLT3 regulated signalling pathways led us to investigate DNA-PK in KIT-mutant AML.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Blood Cell, Cell Culture

DISEASE(S): Acute Leukemia

SUBMITTER: Nicole Verrills

LAB HEAD: Nicole Verrills

PROVIDER: PXD030005 | Pride | 2023-01-26

REPOSITORIES: Pride

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Publications

Synergistic Targeting of DNA-PK and KIT Signaling Pathways in KIT Mutant Acute Myeloid Leukemia.

Murray Heather C HC Miller Kasey K Brzozowski Joshua S JS Kahl Richard G S RGS Smith Nathan D ND Humphrey Sean J SJ Dun Matthew D MD Verrills Nicole M NM

Molecular & cellular proteomics : MCP 20230120 3

Acute myeloid leukemia (AML) is the most common and aggressive form of acute leukemia, with a 5-year survival rate of just 24%. Over a third of all AML patients harbor activating mutations in kinases, such as the receptor tyrosine kinases FLT3 (receptor-type tyrosine-protein kinase FLT3) and KIT (mast/stem cell growth factor receptor kit). FLT3 and KIT mutations are associated with poor clinical outcomes and lower remission rates in response to standard-of-care chemotherapy. We have recently ide ...[more]

PMID: 36682716

Similar Datasets

Project description:Global high-throughput phosphoproteomic profiling is increasingly being applied to cancer specimens as a means to identify the oncogenic signaling cascades responsible for promoting disease initiation and disease progression; pathways that are often invisible to genomics analysis. Hence, phosphoproteomic profiling has enormous potential to inform and improve individualized anti-cancer treatment strategies. However, to achieve the adequate phosphoproteomic depth and coverage necessary to identify the activated, and hence, targetable kinases responsible for driving oncogenic signaling pathways; affinity phosphopeptide enrichment techniques are required and often coupled with offline high-pressure liquid chromatographic (HPLC) separation prior to nanoflow liquid chromatography–tandem mass spectrometry (nLC-MS/MS). These complex and time-consuming procedures, limit the utility of phosphoproteomics for the analysis of individual cancer patient specimens in real-time, and restrict phosphoproteomics to specialized laboratories often outside of the clinical setting. To address these limitations, here we have optimized a new protocol, phospho-Heavy-labeled-spiketide FAIMS Stepped-CV DDA (pHASED), that employs online phosphoproteome deconvolution using high-field asymmetric waveform ion mobility spectrometry (FAIMS) and internal phosphopeptide standards to provide accurate label-free quantitation (LFQ) data in real-time. Compared with traditional LFQ using shotgun phosphoproteomics workflows, pHASED provided increased phosphoproteomic depth and coverage (phosphopeptides = 4,617 pHASED, 2,789 LFQ), whilst eliminating the variability associated with offline prefractionation. pHASED was optimized using tyrosine kinase inhibitor (sorafenib) resistant isogenic FLT3-mutant acute myeloid leukemia (AML) cell line models. Bioinformatic analysis identified differential activation of the Serine/threonine protein kinase ataxia-telangiectasia mutated (ATM) pathway, responsible for sensing and repairing DNA damage in sorafenib-resistant AML cell line models, thereby uncovering a potential therapeutic opportunity. Herein, we have optimized a rapid, reproducible, and flexible protocol for the characterization of complex cancer phosphoproteomes in real-time; a step towards the implementation of phosphoproteomics in the clinic to aid in the selection of anti-cancer therapies for patients.

			Action	DRS
	HB_816_Combo_R1.mzML	Mzml
	HB_816_Combo_R1.mzid.gz	Mzid
	HB_816_Combo_R1.pdResult	Other
	HB_816_Combo_R1.raw	Raw
	HB_816_Combo_R2.raw	Raw