Systems and compositions for diagnosing Barrett's esophagus and methods of using the same

• US 10,018,631 B2
• Filed: 03/15/2012
• Issued: 07/10/2018
• Est. Priority Date: 03/17/2011
• Status: Active Grant

• Current Assignee: Cernostics, Inc. (Castle Biosciences, Inc.)
• Original Assignee: Cernostics, Inc. (Castle Biosciences, Inc.)
• Inventors: Thorne, Rebecca Jane, Campbell, Bruce B.
• Primary Examiner(s): Shafer, Shulamith H
• Application Number:
  US14/005,409
• Publication Number:
  US 20140141988A1
• Time in Patent Office: 2,308 Days
• Field of Search: None
• CPC Class Codes: C12Q 1/6886 for cancer immunoassay for cancer G01N33/574
• CPC Class Codes: G01N 2800/14 Disorders of ear, nose or throat
• CPC Class Codes: G01N 2800/56 Staging of a disease; Further complications associated with the disease
• CPC Class Codes: G01N 33/57407 Specifically defined cancers

Abstract

The invention provides a system, composition, and methods of using the systems and compositions for the analysis of a sample from a subject to accurately diagnose, prognose, or classify the subject with certain grades of or susceptibility to Barrett'"'"'s esophagus. In some embodiments, the system of the present invention comprises a means of detecting and/or quantifying morphological features, the expression of protein, or the expression of nucleic acids in a plurality of cells and correlating that data with a subject'"'"'s medical history to predict clinical outcome, treatment plans, preventive medicine plans, or effective therapies. In some embodiments, the invention relates to a method of classifying and compiling data taken from a cell sample from a subject analyzing the data, and converting the data from the system into a score by which a pathologist may calculate the likelihood that the subject develops cancer.

Claims

1. A method of assigning the risk of progression of Barrett'"'"'s esophagus to a subject, comprising:

a) obtaining an upper gastrointestinal sample from the subject, wherein the upper gastrointestinal sample comprises one or more of the following tissues;

surface epithelium, glandular epithelium, lamina propria, and stroma;

b) labeling a plurality of biomarkers using fluorescent probes, stains, or antibodies in the upper gastrointestinal sample from the subject, wherein the plurality of biomarkers are selected from the group consisting of p53, HIF-1alpha, beta-catenin, COX-2, and any combination thereof;

c) detecting the labeled biomarkers and nuclei with an optical scanner;

d) generating digital image data from the detected labeled biomarkers and nuclei;

e) storing the generated digital image data in a computer-readable storage medium;

f) analyzing the digital image data with a computer processor implementing computer-executable program code to produce pixel-based segmentation and object-based classification of subcellular compartments and tissues;

g) quantifying one or more descriptive features of each biomarker and nuclei, wherein the descriptive features are selected from the group consisting of mean intensity in a cell-based object including cell, cytoplasm, plasma membrane, and nucleus; and

ratio of intensity between cell-based objects including cell, cytoplasm, plasma membrane, and nucleus;

h) converting the analyzed and quantified digital image data to generate a score using a predictive statistical model developed in a set that comprises disease cases and unaffected controls,

wherein the score is computed by linear combination of descriptive features weighted by coefficients obtained via linear regression model, and the score is correlated to a risk of progression to high grade dysplasia or esophageal adenocarcinoma; and

i) using the score to identify which subjects to treat;

wherein the subject with a high risk score is treated using a clinical treatment selected from the group consisting of endoscopic surveillance, endoscopic mucosal resection, radiofrequency ablation, and any combination thereof; and

wherein the subject with a low risk score is not treated and avoids unnecessary invasive procedures and continues endoscopic surveillance at reduced frequency or discontinues endoscopic surveillance.

2. The method of claim 1, wherein the subject is identified as having a high risk score and has an increased risk of progression to low grade dysplasia, high grade dysplasia or esophageal adenocarcinoma.

3. The method of claim 1, wherein the subject is identified as having a low risk score and has no dysplasia, reactive atypia, indefinite for dysplasia, or low grade dysplasia.

5. The method of claim 1, wherein the upper gastrointestinal sample comprises a brushing, biopsy, or surgical resection of cells or tissue from the subject.

6. The method of claim 1, wherein the upper gastrointestinal sample is at room temperature or frozen.

7. The method of claim 1, wherein the upper gastrointestinal sample is freshly obtained, formalin fixed, alcohol fixed, or paraffin embedded.

8. The method of claim 1, wherein the fluorescent probes comprise a fluorescent tag, preferably wherein each fluorescent probe is labeled with a different fluorophore.

9. The method of claim 1, wherein the detection of 2 or more, or 3 or more biomarkers are determined simultaneously.

10. The method of claim 1, wherein the subject is a human.

11. The method of claim 1, wherein the descriptive feature further comprises a plurality of clinical factors selected from the group consisting of the condition of a subject in the presence of a disease or in the absence of a disease, the health status of a subject, age, gender, chest pain type, neutrophil count, ethnicity, disease duration, diastolic blood pressure, systolic blood pressure, a family history parameter, a medical history parameter, a medical symptom parameter, height, weight, a body-mass index, resting heart rate, smoker/non-smoker status, whether the subject has stable chest pain, whether the subject has been diagnosed with a hiatal hernia, whether the subject has GERD, whether the subject has an gastritis, whether the subject has been previously diagnosed with Barrett'"'"'s esophagus, whether the subject has had a gastrointestinal procedure, whether the subject has diabetes, whether the subject has an inflammatory condition, whether the subject has an infectious condition, whether the subject is taking a steroid, whether the subject is taking an immunosuppressive agent, whether the subject is taking a chemotherapeutic agent, and a combination thereof.

4. A method of assigning the classification of Barrett'"'"'s esophagus to a subject, comprising:

a) obtaining an upper gastrointestinal sample from the subject, wherein the upper gastrointestinal sample comprises one or more of the following tissues;

surface epithelium, glandular epithelium, lamina propria, and stroma;

b) labeling a plurality of biomarkers using fluorescent probes, stains, or antibodies in ft the upper gastrointestinal sample from the subject, wherein the plurality of biomarkers are selected from the group consisting of HIF-1alpha, p53, COX-2, beta-catenin, and any combination thereof;

c) detecting the labeled biomarkers and nuclei with an optical scanner;

d) generating digital image data from the detected labeled biomarkers and nuclei;

e) storing the generated digital image data in a computer-readable storage medium;

f) analyzing the digital image with a computer processor implementing computer-executable program code to produce pixel-based segmentation and object-based classification of subcellular compartments and tissues;

g) quantifying one or more descriptive features of each biomarker and nuclei, wherein the descriptive features are selected from the group consisting of mean intensity in a cell-based object including cell, cytoplasm, plasma membrane, and nucleus; and

ratio of intensity between cell-based objects including cell, cytoplasm, plasma membrane, and nucleus;

h) converting the analyzed and quantified digital image data to generate a score using a predictive statistical model developed in a set that comprises disease cases and unaffected controls,

wherein the score is computed by linear combination of descriptive features weighted by coefficients obtained via linear regression model, and the score is correlated to a classification of Barrett'"'"'s esophagus, wherein the classification of Barrett'"'"'s esophagus is selected from the group consisting of no dysplasia, reactive atypia, indefinite for dysplasia, low grade dysplasia, high grade dysplasia, and esophageal adenocarcinoma; and

i) using the classification of Barrett'"'"'s esophagus to identify which subjects to treat;

wherein the subject with low grade dysplasia high grade dysplasia or esophageal adenocarcinoma are treated using a clinical treatment selected from the group consisting of endoscopic surveillance, endoscopic mucosal resection, radiofrequency ablation, and any combination thereof; and

wherein the subject with no dysplasia, reactive atypia, or indefinite for dysplasia are not treated and avoid unnecessary invasive procedures and continue endoscopic surveillance at reduced frequency or discontinues endoscopic surveillance.

12. The method of claim 4, wherein the upper gastrointestinal sample comprises a brushing, biopsy, or surgical resection of cells or tissue from the subject.

13. The method of claim 4, wherein the upper gastrointestinal sample is at room temperature or frozen.

14. The method of claim 4, wherein the upper gastrointestinal sample is freshly obtained, formalin fixed, alcohol fixed, or paraffin embedded.

15. The method of claim 4, wherein the fluorescent probes comprise a fluorescent tag, preferably wherein each fluorescent probe is labeled with a different fluorophore.

16. The method of claim 4, wherein the detection of 2 or more, or 3 or more biomarkers are determined simultaneously.

17. The method of claim 4, wherein the subject is a human.