# US 11,441,191 B2 · Methods and kits comprising gene signatures for stratifying breast cancer patients

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- Number: US 11,441,191 B2
- Document ID: US11441191B2
- Status: Granted
- Kind: B2
- Application number: 16/308,564
- Filed: 2017-06-19
- Published: 2019-05-30
- Granted: 2022-09-13
- Earliest claimed: 2016-06-20
- Assignees: Istituto Europeo di Oncologia S.r.l.; Fondazione Istituto Firc Di Oncologia Molecolare (Ifom); The University of Milan
- Applicants: Istituto Europeo di Oncologia (IEO); Fondazione Istituto Firc di Oncologia Molecolare (IFOM); Universita degli Studi di Milano (University of Milan)
- Inventors: Pier Paolo Di Fiore; Salvatore Pece; Manuela Vecchi; Stefano Confalonieri
- Web page: https://patentgrep.com/patent/US11441191B2

## CPC

- C12Q 1/6886: for cancer (immunoassay for cancer G01N33/575)
- C12Q 2600/106: Pharmacogenomics, i.e. genetic variability in individual responses to drugs and drug metabolism
- C12Q 2600/112: Disease subtyping, staging or classification
- C12Q 2600/118: Prognosis of disease development
- C12Q 2600/158: Expression markers
- C12Q 2600/16: Primer sets for multiplex assays
- C12Q 2600/166: Oligonucleotides used as internal standards, controls or normalisation probes
- G16B 5/00: ICT specially adapted for modelling or simulations in systems biology, e.g. gene-regulatory networks, protein interaction networks or metabolic networks
- G16B 40/00: ICT specially adapted for biostatistics; ICT specially adapted for bioinformatics-related machine learning or data mining, e.g. knowledge discovery or pattern finding
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- G16B: BIOINFORMATICS, i.e. INFORMATION AND COMMUNICATION TECHNOLOGY [ICT] SPECIALLY ADAPTED FOR GENETIC OR PROTEIN-RELATED DATA PROCESSING IN COMPUTATIONAL MOLECULAR BIOLOGY

## Abstract

The present invention relates to refined prognostic clinical tools, methods, and kits for the evaluation of risk and treatment of distant recurrence in ER+/HER2− breast cancer patients.

## Claims

### Claim 1

A method for treating a human subject having a ER+/HER2− breast cancer comprising steps of:
(a) determining, in a breast tissue or breast cell sample from the subject the expression of group of genes;
(b) calculating a risk score based upon the expression of the group of genes, wherein a higher risk score indicates an increased risk of breast cancer recurrence in the subject,
wherein the risk score is calculated according to the following formula:
Risk score=Σi(βi*Cqnormalized),
wherein Cqnormalized is calculated according to the following formula:
Cqnormalized=AVG Cq−SF,
wherein SF is the difference between the AVG Cq value of at least one reference gene for each subject and a constant reference value K, wherein K=25.012586069, which represents the mean of the AVG Cq of the at least one reference gene calculated across a plurality of training samples;
(c) stratifying the subject into a high or low risk group; and
(d) administering a breast cancer treatment to the subject, wherein a subject stratified in a high risk group is provided a cancer treatment that is more aggressive than the cancer treatment provided to a subject stratified in a low risk group;
wherein said group of genes consists of:
(i) EIF4EBP1, MRPS23, and TOP2A;
(ii) APOBEC3B, CENPW, EIF4EBP1, EXOSC4, LY6E, MMP1, MRPS23, NDUFB10, and TOP2A;
(iii) ALYREF, APOBEC3B, CDK1, CENPW, EIF4EBP1, EXOSC4, H2AFJ, LY6E, MIEN1, MMP1, MRPS23, NDUFB10, NOL3, RACGAP1, SFN, and TOP2A; or
(iv) H2AFZ, CDK1, EXOSC4, PHLDA2, APOBEC3B, EIF4EBP1, SFN, PHB, EPB41L5, RACGAP1, MRPS23, TOP2A, H2AFJ, NOL3, MIEN1, CENPW, LY6E, ALYREF, MMP1, and NDUFB10.

### Claim 2 (depends on 1)

The method of claim 1, wherein the at least one reference gene is selected from the group consisting of GAPDH, GUSB, HPRT1, and TBP.

### Claim 3 (depends on 1)

The method of claim 1, wherein the gene expression is determined using reverse transcription and real-time quantitative polymerase chain reaction (RT-qPCR) with primers and/or probes specific for each gene of the said group of genes.

### Claim 4 (depends on 1)

The method of claim 1, wherein the sample is a breast tumor obtained from the subject, a cancerous breast cell obtained from the subject, or a breast cancer stem cell obtained from the subject.

### Claim 5 (depends on 1)

The method of claim 1, wherein the breast cancer treatment comprises surgery, radiation, anthracycline agents, alkylating agents, nucleoside analogs, platinum agents, vinca agents, anti-estrogen drugs, aromatase inhibitors, ovarian suppression agents, endocrine/hormonal agents, bisphophonate therapy agents, targeted biological therapy agents, and antibodies, or combinations thereof.

### Claim 6 (depends on 5)

The method of claim 5, wherein the breast cancer treatment comprises cyclophosphamide, fluorouracil, 5-fluorouracil, methotrexate, thiotepa, carboplatin, cisplatin, gemcitabine, anthracycline, taxanes, paclitaxel, protein-bound paclitaxel, doxorubicin, docetaxel, vinorelbine, tamoxifen, raloxifene, toremifene, fulvestrant, irinotecan, ixabepilone, temozolmide, topotecan, vincristine, vinblastine, eribulin, mutamycin, capecitabine, capecitabine, anastrozole, exemestane, letrozole, leuprolide, abarelix, buserlin, goserelin, megestrol acetate, risedronate, pamidronate, ibandronate, alendronate, denosumab, zoledronate, trastuzumab, tykerb or bevacizumab, or combinations thereof.