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US-20260126452-A1 - SYSTEM AND METHOD FOR QUANTITATIVE PATHOLOGY USING MASS SPECTROMETRY IMAGING

US20260126452A1US 20260126452 A1US20260126452 A1US 20260126452A1US-20260126452-A1

Abstract

An apparatus for facilitating quantitative anatomic pathology using mass spectrometry imaging includes a solid support having at least one flat surface, a tissue homogenate having a thickness mounted to the at least one flat surface of the solid support, and a quantitative array having a thickness and comprising a tissue microarray having a plurality of wells and a series of varying concentrations of an isotopically labeled metabolite deposited in the plurality of wells. The quantitative array is mounted over the tissue homogenate on the solid support.

Inventors

  • Nathalie Agar
  • Sankha S. Basu
  • Sylwia A. Stopka
  • Sandro Santagata
  • Marcia C. Haigis
  • Michael Regan
  • Gerard Baquer

Assignees

  • THE BRIGHAM AND WOMEN’S HOSPITAL, INC.
  • PRESIDENT AND FELLOWS OF HARVARD COLLEGE

Dates

Publication Date
20260507
Application Date
20221031

Claims (20)

  1. 1 . An apparatus for facilitating quantitative anatomic pathology using mass spectrometry imaging, the apparatus comprising: a solid support having at least one flat surface; a tissue homogenate having a thickness mounted to the at least one flat surface of the solid support; and a quantitative array having a thickness and comprising a tissue microarray having a plurality of wells and a series of varying concentrations of an isotopically labeled metabolite deposited in the plurality of wells, wherein the quantitative array is mounted over the tissue homogenate on the solid support.
  2. 2 . The apparatus according to claim 1 , wherein the solid support is a slide.
  3. 3 . The apparatus according to claim 1 , wherein the tissue homogenate thickness is the same as the quantitative array thickness.
  4. 4 . The apparatus according to claim 1 , further comprising: a tissue sample having a thickness and mounted to the at least one flat surface of the solid support and adjacent to the quantitative array.
  5. 5 . The apparatus according to claim 4 , further comprising: a matrix assisted laser desorption ionization matrix deposited on the quantitative array and the tissue sample.
  6. 6 . The apparatus according to claim 4 , wherein the tissue sample thickness is the same as the quantitative array thickness.
  7. 7 . A method for making an apparatus for facilitating quantitative anatomic pathology using mass spectrometry imaging, the method comprising: applying a material solution to a tissue microarray (TMA) to form an imprint mold having a plurality of wells; dispensing a series of varying concentrations of an isotopically labeled metabolite onto the plurality of wells to form a quantitative array; sectioning the quantitative array to produce a section with a predetermined thickness; sectioning a tissue homogenate to produce a section with a predetermined thickness; mounting the section of the tissue homogenate to a solid support having at least one flat surface; and mounting the section of the quantitative array over the tissue homogenate on the solid support.
  8. 8 . The method according to claim 7 , wherein the material solution is formed with a material configured to minimize diffusion of the series of isotopically labeled metabolites.
  9. 9 . The method according to claim 7 , wherein the predetermined thickness of the tissue homogenate is the same as the predetermined thickness of the quantitative array.
  10. 10 . The method according to claim 7 , further comprising: sectioning a tissue sample to produce a section with a predetermined thickness; and mounting the section of the tissue sample to the solid support adjacent to the quantitative array.
  11. 11 . The method according to claim 10 , further comprising: applying a matrix assisted laser desorption ionization matrix on the section of the quantitative array and the section of the tissue sample.
  12. 12 . The method according to claim 10 , wherein the predetermine thickness of the tissue sample is the same as the predetermined thickness of the quantitative array.
  13. 13 . The method according to claim 11 , wherein the matrix includes an isotopically labeled internal standard.
  14. 14 . A method for quantitative anatomic pathology using mass spectrometry imaging, the method comprising: preparing a test slide comprising a tissue homogenate, a quantitative array mounted over the tissue homogenate and comprising a plurality of wells and a series of varying concentrations of an isotopically labeled metabolite deposited in the plurality of wells, and a tissue sample mounted on the test slide adjacent to the quantitative array; performing mass spectrometry imaging on the test slide to generate a set of mass spectrometry data and a set of calibration curves for the isotopically labeled metabolites; generating quantitative data for a plurality of metabolites based on the mass spectrometry data and the set of calibration curves; generating a report comprising at least the quantitative data for the plurality of metabolites.
  15. 15 . The method according to claim 14 , wherein the mass spectrometry imaging is matrix assisted laser desorption ionization mass spectrometry imaging and the test slide further comprises a matrix assisted laser desorption ionization matrix applied to the quantitative array and the tissue sample
  16. 16 . The method according to claim 15 , wherein the matrix assisted laser desorption ionization mass spectrometry imaging comprises performing a Fourier Transform Ion Cyclotron Resonance (FT-ICR) mass spectrometry imaging technique.
  17. 17 . The method according to claim 14 , further comprising excluding mass spectrometry data located outside of the quantitative array wells.
  18. 18 . The method according to claim 14 , wherein the matrix includes an isotopically labeled internal standard for normalization.
  19. 19 . The method according to claim 14 , wherein the set of calibration curves are generated using a weighted linear regression model.
  20. 20 . The method according to claim 14 , further comprising performing normalization to the matrix peak.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS This application is based on, claims priority to, and incorporates herein by reference in its entirety U.S. Ser. No. 63/273,863 filed Oct. 29, 2021, and entitled “MALDI MSI Quantitative Anatomic Pathology.” STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH This invention was made with government support under Grant Nos. CA201469, CA210180 and EB028741, awarded by National Institutes of Health. The government has certain rights in the invention FIELD The present disclosure relates generally to quantitative mass spectrometry and, more particularly, to systems and methods for quantitative anatomic pathology using mass spectrometry imaging, for example, simultaneous quantitation of multiple biomarkers and drugs from tissue sections. BACKGROUND Altered cellular metabolism is a hallmark of many different cancers and has proven to be a promising source of therapeutic targets. Exploiting these vulnerabilities, however, requires accurate metabolic characterization in heterogeneous specimens, which are often composed of neoplastic, stromal, and immune cells in a complex tumor microenvironment. Although liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) remains the gold standard for many analytical applications, most methods involve liquid matrices such as serum, plasma, or urine. Tissue-based quantification of small molecules using LC-MS/MS requires extensive sample preparation, making it challenging to implement in clinical laboratories. Furthermore, since these approaches involve homogenization or other tissue destructive processes, knowledge of spatial variability is lost, an important characteristic of endogenous metabolites in heterogeneous tumors. Matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI MSI) is a powerful analytical technique that provides spatially-preserved detection and quantification of analytes in tissue specimens. For example, MALDI-MSI can provide a powerful platform to map the spatial distribution of large biopolymers, proteins, peptides, lipids, small molecules, and drugs directly in tissue sections. Accurate and reliable quantification using MALDI-MSI, however, has faced several technical challenges. One challenge, common to most MS-based techniques, is variable ionization efficiency and ion suppression due to matrix effects. This has been addressed in LC-MS/MS methods by creating calibration curves spiked into comparable biological matrices as well as through normalization to spiked stable isotope internal standards (IS). However, matrix effect challenges are considerably more challenging when mapping metabolites in tissue specimens due to wide pixel-to-pixel variability in tissue composition and the lack of chromatographic separation. Several approaches to account for matrix effects and to allow for better accuracy and precision have been applied to MALDI-MSI. For example, to generate calibration curves, the tissue mimetics may be used for the absolute quantitation for MALDI-MSI. The first mimetic approach consisted of spiking a range of different drug concentrations into a set of tissue homogenates that were pre-weighed into microcentrifuge tubes and then transferred into a home-built mold. This was further refined by creating cylindrical molds consisting of layers of serially frozen spiked-tissue homogenates in increasing concentrations to create calibration curves. Despite these improvements, both methods are still arguably cumbersome to generate, and require considerable space on a relatively small MALDI slide, leaving minimal room for experimental samples. In addition to the use of calibration curves, stable isotope internal standards have also been applied, though most often to analyze drugs in tissue, and less commonly for endogenous metabolites. It would be desirable to provide a system and method for quantitative anatomic pathology using mass spectrometry imaging that can simultaneously quantitate a plurality of biomolecules and drugs with improved precision, accuracy and analytical characteristics. SUMMARY In accordance with an embodiment, an apparatus for facilitating quantitative anatomic pathology using mass spectrometry imaging includes a solid support having at least one flat surface, a tissue homogenate having a thickness mounted to the at least one flat surface of the solid support, and a quantitative array having a thickness and comprising a tissue microarray having a plurality of wells and a series of varying concentrations of an isotopically labeled metabolite deposited in the plurality of wells. The quantitative array is mounted over the tissue homogenate on the solid support In accordance with another embodiment, a method for making an apparatus for facilitating quantitative anatomic pathology using mass spectrometry imaging includes applying a material solution to a tissue microarray (TMA) to form an imprint mold having a plurality of wells, dispensing a series of varying concentratio