Biomarker Analysis Tool

In collaboration with the bioinformatics firm 4HF Biotech GmbH which is a bioinformatics firm specializing in cancer data mining to discover new anti-cancer drugs, we offer data mining for analysis of the genetic signature of tumor cell lines that are sensitive for a customer drug to reveal potential biomarkers.

The Biomarker Analysis is based on the sensitivity profile of the customer compound such as the results of the ProLiFiler - our Cell Line Panel Screening service.

The Biomarker Analysis tool identifies genes or genetic alterations which are indicative for the efficacy of a drug and which may serve as predictive biomarkers in disease models and patient cohorts.

Additional Information

Biomarker Analysis Details

Primary data set: We will perform the ProLiFiler assay for testing the anti-proliferative effects of your drug on a set of 140 human tumor cell lines for IC50 determination. This is called the sensitivity profile of your compound.

Database: Our collaborator 4HF Biotech has built a comprehensive database, including data of more than 1,800 preclinical samples comprising multiple cancer datasets that are integrated into a single platform for visualization and statistical analysis.
The data relevant for the Biomarker Analysis Tool are mutations (whole-exome sequencing), gene copy numbers, and gene expression data (Affymetrix).

Approach: The ProLiFiler results are correlated with the gene expression, mutation, and somatic copy number alteration data of the tumor cell lines that were used in the ProLiFiler study. A correlation analysis is performed for each cell line to determine the correlation of the drug sensitivity to each gene in regards to the expression level, mutation status, and/or somatic copy number.

Analysis: The Biomarker Analysis is performed with two statistical tests for each correlation; one uses the IC50 value of the test drug; the other compares the gene data of responding and non-responding cell lines.

Deliverables:

Mutation analysis

  • Excel tables with raw data
  • Graphical presentation of the correlation of mutation status for each gene and the drug sensitivity for each cell line
  • Graphical presentation of the correlation of the position of each mutation and the drug sensitivity for each cell line
  • Optional: Presentation of the amino acid position of relevant mutations

Gene copy analysis

  • Excel tables with raw data
  • Graphical presentation of the correlation of gene copy gain and the drug sensitivity for each cell line
  • Graphical presentation of correlation of gene copy loss and the drug sensitivity for each cell line
  • For selected genes, additional plots can be generated.

Gene expression analysis

  • Excel table with raw data
  • Graphical presentation of the correlation of the expression of each gene and drug sensitivity for every cell line
  • Graphical presentation of the cluster analysis to identify differential gene expression signatures and different response rates to the test drug
  • Optional: Literature search of gene function for most relevant genes

Additional analyses can be performed upon requests such as target gene analysis, pathway analysis, subtype analysis, hypothesis-driven analysis, predictive model development, and more.

Case Study Nutlin-3a - Gene Expression

This case study investigates nutlin-3a which is a small molecule that occupies the p53 binding pocket of MDM2 and effectively disrupts the p53–MDM2 interaction that leads to activation of the p53 pathway.

case study biomarker analysis nutlin-3

Example of nutlin-3a gene expression analysis. Graphical presentation of Limma test results show which genes are highly expressed in drug-resistant and drug-sensitive cell lines. MDM2 and other p53 pathway-related genes were highly expressed in tumor cell lines, which were sensitive to nutlin-3 treatment.

example biomarker analysis nutlin-3 table

In the table are listed 10 genes which show high expression in tumor cell lines sensitive to nutlin-3a treatment. The results show that MDM2 expression is a predictor of sensitivity to nutlin-3a treatment.

Case Study Nutlin-3a - Mutation Analysis

This case study investigates nutlin-3a which is a small molecule that occupies the p53 binding pocket of MDM2 and effectively disrupts the p53–MDM2 interaction that leads to activation of the p53 pathway.

example study nutlin-3 biomarker analysis mutation

Example of nutlin-3a mutation analysis. Nutlin-3 was tested for dose-response in a panel of tumor cell lines. These cell lines contain a total of 16,998 mutated genes. The volcano plot shows for each gene (one dot) the Wilcoxon test p-value for the correlation of the drug response in proportion to the existence of mutants. For example, p53 mutations were found mostly in tumor cell lines that did not respond to nutlin-3a. KIAA1522, on the other hand, was found the most probable mutation that exists in cell lines responding to nutlin-3a treatment.

example study nutlin-3 mutation table top 3

The top three mutated genes correlated with nutlin-3a efficacy. P53 was found mutated in 10 cell lines, which responded to nutlin-3a treatment, and in 152 cell lines, which did not respond to nutlin-3a treatment. The mean IC50 value of nutlin-3a in tumor cell lines with p53 wild type was about 11.6 µM compared to 27.5 µM in cell lines with p53 mutation resulting in a difference of 16 µM between mutated and non-mutated cell lines which was used as a measure for resistance.

In a mutation position analysis, amino acid Arg 273 in TP53 was shown to be the strongest marker of resistance.

Case Study Nutlin-3a - sensitivity data

This case study investigates nutlin-3a which is a small molecule that occupies the p53 binding pocket of MDM2 and effectively disrupts the p53–MDM2 interaction that leads to activation of the p53 pathway.

cell line panel screening with IC50 values performed on nutlin-3a

The IC50 values of nutlin-3a were determined on 274 tumor cell lines and ranked. The tumor cell lines could be grouped as resistant, intermediated sensitive, and highly sensitive. The highest testing concentration was 30 µM with semi-log increments.

 

example study nutlin-3 IC50 ranking

The IC50 values of nutlin-3a are shown in a scatter plot for each cell line. The tumor types were ranked for the highest median IC50. The median IC50 for all tumor types (red line) coincides at 30 µM testing concentration because of the high number of resistant tumor cell lines.

The most susceptible tumor types are leukemia AML, melanoma, leukemia ALL, Hodgkin's lymphoma.