Industrialized, Artificial Intelligence-guided Laser Microdissection for Microscaled Proteomic Analysis of the Tumor Microenvironment

Mitchell, D. L.; Hunt, Allison L.; Conrads, Kelly A.; Hood, Brian L.; Makohon-Moore, Sasha C.; Rojas, Christine; Maxwell, G. Larry; Bateman, Nicholas W.; Conrads, Thomas P.

doi:10.3791/64171

Cited by 3 publications

(1 citation statement)

References 1 publication

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“…Regions of tissue annotated using HALO were exported for LMD (LMD7, Leica Microsystems) as previously described. 34 LMD harvested tissue for protein digestion was collected in MircoTubes (Pressure Biosciences, Inc., South Easton, MA, USA) containing 20 μL of 100 mM triethylammonium bicarbonate (TEAB, pH 8.0)/10% acetonitrile, capped and stored at -80°C until digestion. Tissue for RNA isolation was collected in 300 μL of Buffer RLT with 10% β-mercaptoethanol (QIAGEN Sciences, LLC, Germantown, MD, USA) and stored at -80°C until isolation.…”

Section: Methodsmentioning

confidence: 99%

ProteoMixture: A cell type deconvolution tool for bulk tissue proteomic data

Teng,

Schaaf,

Abulez

et al. 2024

iScience

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Section: Methodsmentioning

confidence: 99%

ProteoMixture: A cell type deconvolution tool for bulk tissue proteomic data

Teng,

Schaaf,

Abulez

et al. 2024

iScience

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Integration of Multi-omic Data in a Molecular Tumor Board Reveals EGFR-Associated ALK-Inhibitor Resistance in a Patient With Inflammatory Myofibroblastic Cancer

et al. 2023

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Inflammatory myofibroblastic tumors (IMTs) are intermediate-grade mesenchymal neoplasms commonly characterized by chromosomal rearrangements causing constitutive activation of anaplastic lymphoma kinase (ALK) and/or ALK mutations causing reduced sensitivity to ALK tyrosine kinase inhibitors (TKI). We present a patient with an IMT who initially responded to first-line alectinib, but who later suffered disease relapse and presently survives with moderate residual disease after receiving second-line lorlatinib. Biopsy specimens were analyzed using next generation sequencing (DNA-seq and RNA-seq) and reverse phase protein microarray (RPPA) as part of an institutional Molecular Tumor Board (MTB) study. An EML4-ALK rearrangement and EGFR activation (pEGFRY1068) were present in both the primary and recurrent tumors, while a secondary ALK I1171N mutation was exclusive to the latter. EGFR signaling in the background of a secondary ALK mutation is correlated with reduced ALK TKI sensitivity in vitro, implicating an important mechanism of drug resistance development in this patient. The RPPA results also critically demonstrate that ALK signaling (ALKY1604) was not activated in the recurrent tumor, thereby indicating that standard-of-care use of third- or fourth-line ALK TKI would not likely be efficacious or durable. These results underscore the importance of real-time clinical integration of functional protein drug target activation data with NGS in the MTB setting for improving selection of patient-tailored therapy.

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dia-PASEF Proteomics of Tumor and Stroma LMD Enriched from Archived HNSCC Samples

Panigrahi,

Hunt,

Assis

et al. 2024

Preprint

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We employed laser microdissection to selectively harvest tumor cells and stroma from the microenvironment of formalin-fixed, paraffin-embedded head and neck squamous cell carcinoma (HNSCC) tissues. The captured HNSCC tissue fractions were analyzed by quantitative mass spectrometry-based proteomics using a data independent analysis approach. In paired samples, we achieved excellent proteome coverage having quantified 6,668 proteins with a median quantitative coefficient of variation under 10%. We observed significant differences in relevant functional pathways between the spatially resolved tumor and stroma regions. Our results identified extracellular matrix (ECM) as a major component enriched in the stroma, including many cancer associated fibroblast signature proteins in this compartment. We demonstrate the potential for comparative deep proteome analysis from very low starting input in a scalable format that is useful to decipher the alterations in tumor and the stromal microenvironment. Correlating such results with clinical features or disease progression will likely enable identification of novel targets for disease classification and interventions.

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Industrialized, Artificial Intelligence-guided Laser Microdissection for Microscaled Proteomic Analysis of the Tumor Microenvironment

Cited by 3 publications

References 1 publication

ProteoMixture: A cell type deconvolution tool for bulk tissue proteomic data

ProteoMixture: A cell type deconvolution tool for bulk tissue proteomic data

Integration of Multi-omic Data in a Molecular Tumor Board Reveals EGFR-Associated ALK-Inhibitor Resistance in a Patient With Inflammatory Myofibroblastic Cancer

dia-PASEF Proteomics of Tumor and Stroma LMD Enriched from Archived HNSCC Samples

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