# US 2014/0155332 A1 · Selective inhibitors of tumor-initiating cells > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 2014/0155332 A1 - Document ID: US20140155332A1 - Status: Published application - Kind: A1 - Application number: 14/003,021 - Filed: 2012-03-02 - Published: 2014-06-05 - Earliest claimed: 2011-03-04 - Assignees: Children's Medical Center Corporation - Inventors: Judy Lieberman; Fabio Petrocca - Web page: https://patentgrep.com/patent/US20140155332A1 ## CPC - C12Q 1/6886: for cancer (immunoassay for cancer G01N33/575) - A61K 31/165: having aromatic rings, e.g. colchicine, atenolol, progabide - A61K 31/167: having the nitrogen of a carboxamide group directly attached to the aromatic ring, e.g. lidocaine, paracetamol - A61K 31/713: Double-stranded nucleic acids or oligonucleotides - A61K 38/05: Dipeptides - G01N 33/57515: of the breast - C12Q 2600/158: Expression markers - G01N 2500/10: involving cells - G01N 2800/52: Predicting or monitoring the response to treatment, e.g. for selection of therapy based on assay results in personalised medicine; Prognosis - G01N 2800/56: Staging of a disease; Further complications associated with the disease - 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 - A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D) - G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q) ## Abstract Described herein are novel malignancy associated gene signature biomarkers, and assays and methods thereof, to classify prognosis or malignant potential of a cancer and identify cancer-initiating cells. The malignancy associated gene signature biomarkers, assays and methods described herein provide, in part, new methodologies to screen for novel drugs for treating cancers and tumors, such as, for example, triple-negative breast tumors. Using the assays and methods described herein proteasome inhibitors, histone deacetylase inhibitors, and glycolysis inhibitors, were identified as being highly effective in altering gene expression signatures specifically in malignant or cancer-initiating cells. ## Claims ### Claim at position 1 -23. (canceled) ### Claim 24 A method of classifying a cancer in a subject in need thereof, the method comprising: a. assaying expression of ten or more of the 154 malignancy associated response signature biomarkers of SEQ ID NOs: 1-154 in a biological sample obtained from the subject having a cancer, and b. comparing the expression of the ten or more of the 154 malignancy associated response signature biomarkers of SEQ ID NOs: 1-154 in the biological sample obtained from the subject having a cancer with a reference value, wherein increased expression of 1.8-fold or greater of at least ten of the biomarkers in the biological sample obtained from the subject relative to the reference value indicates that the cancer is classified as having a poor prognosis or being a malignant cancer, and absence of increased expression of 1.8-fold or greater of at least ten of the biomarkers relative to the reference value indicates that the cancer does not have poor prognosis or is not a malignant cancer. ### Claim 25 (depends on 24) The method of claim 24, wherein the cancer is selected from a breast cancer, a triple-negative breast cancer, Luminal B breast cancer, and epithelial breast cancer. ### Claim at position 4 26-28. (canceled) ### Claim 29 (depends on 27) The method of claim 27, wherein if the cancer is classified as having a poor prognosis or being a malignant cancer, the method further comprises the step of administering at least one proteasome inhibitor to the subject. ### Claim 30 (depends on 24) The method of claim 24, wherein if the cancer is classified as having a poor prognosis or being a malignant cancer and wherein if at least two of the ten or more genes having increased expression is a proteasomal malignancy associated response signature biomarker of SEQ ID NOs: 1, 2, 4, 7, 15, 16, 21, 29, 33, 34, 38, 60, 74, 76, and 149, the method further comprises the step of administering at least one proteasome inhibitor to the subject. ### Claim 31 (depends on 29) The method of claim 29, wherein the at least one proteasome inhibitor specifically inhibits any of the proteasomal malignancy associated response signature biomarkers set forth in SEQ ID NOs: 1, 2, 4, 7, 15, 16, 21, 29, 33, 34, 38, 60, 74, 76, and 149. ### Claim 32 (depends on 29) The method of claim 29, wherein the at least one proteasome inhibitor specifically inhibits the malignancy associated response signature biomarker set forth in SEQ ID NOs: 134 (MCL-1). ### Claim 33 (depends on 29) The method of claim 29, wherein the at least one proteasome inhibitor is an siRNA or antisense RNA agent. ### Claim 34 (depends on 29) The method of claim 29, wherein the at least one proteasome inhibitor is bortezomib. ### Claim 35 (depends on 24) The method of claim 24, wherein if the cancer is classified as having a poor prognosis or being a malignant cancer, the method further comprises the step of administering at least one histone deacetylase inhibitor to the subject. ### Claim 36 (depends on 35) The method of claim 35, wherein the at least one histone deacetylase inhibitor is an siRNA or antisense RNA agent. ### Claim 37 (depends on 35) The method of claim 35, wherein the at least one histone deacetylase inhibitor is trichostatin A (TSA) or Vorinostat. ### Claim 38 (depends on 24) The method of claim 24, wherein when the cancer is classified as having a poor prognosis or being a malignant cancer, the method further comprises the step of administering at least one glygolytic inhibitor to the subject. ### Claim 39 (depends on 38) The method of claim 38, wherein the at least one glycolysis inhibitor is an siRNA or antisense RNA agent. ### Claim 40 (depends on 24) The method of claim 24, wherein if the cancer is classified as having a poor prognosis or being a malignant cancer, relative survival rate, relative risk of metastasis, treatment option, or any combination thereof is determined for the subject. ### Claim 41 A method of classifying a cancer in a subject in need thereof, the method comprising: a. assaying expression of at least five biomarkers set forth in SEQ ID NOs: 1-4, 6, 7, 13, 14, 23, 29, 33-35, 37, 38, 66, 72, 77, 78, 84, 91, 93, and 151 in a biological sample obtained from the subject having a cancer, and b. comparing the expression of the five or more of the 23 malignancy associated response signature biomarkers in the biological sample obtained from the subject having a cancer with a reference value, wherein increased expression of 1.8-fold or greater of at least ten of the biomarkers in the biological sample obtained from the subject relative to the reference value indicates that the cancer is classified as having a poor prognosis or being a malignant cancer, and absence of increased expression of 1.8-fold or greater of at least ten of the biomarkers relative to the reference value indicates that the cancer does not have poor prognosis or is not a malignant cancer. ### Claim 42 (depends on 41) The method of claim 41, wherein the cancer is selected from a breast cancer, a triple-negative breast cancer, Luminal B breast cancer, and epithelial breast cancer. ### Claim at position 19 43-46. (canceled) ### Claim 47 (depends on 41) The method of claim 41, wherein when the cancer is classified as having a poor prognosis or being a malignant cancer, the method further comprises the step of administering at least one proteasome inhibitor to the subject. ### Claim 48 (depends on 41) The method of claim 41, wherein when the cancer is classified as having a poor prognosis or being a malignant cancer and wherein at least one of the five or more genes having increased expression is a proteasomal malignancy associated response signature biomarker of SEQ ID NOs: 1, 2, 4, 7, 29, 33, 34, and 38, the method further comprises the step of administering at least one proteasome inhibitor to the subject. ### Claim 49 (depends on 47) The method of claim 47, wherein the at least one proteasome inhibitor specifically inhibits any of the proteasomal malignancy associated response signature biomarkers set forth in SEQ ID NOs: 1, 2, 4, 7, 15, 16, 21, 29, 33, 34, 38, 60, 74, 76, and 149. ### Claim 50 (depends on 46) The method of claim 46, wherein the at least one proteasome inhibitor specifically inhibits the malignancy associated response signature biomarker set forth in SEQ ID NOs: 134 (MCL-1). ### Claim 51 (depends on 46) The method of claim 46, wherein the at least one proteasome inhibitor is an siRNA or antisense RNA agent. ### Claim 52 (depends on 46) The method of claim 46, wherein the at least one proteasome inhibitor is bortezomib. ### Claim 53 A method of classifying a cancer in a subject in need thereof, the method comprising: a. assaying expression of at least three malignancy associated response signature biomarkers set forth in SEQ ID NOs: 1, 2, 4, 7, 15, 16, 21, 29, 33, 34, 38, 60, 74, 76, and 149 in a biological sample obtained from the subject having a cancer, and b. comparing the expression of the at least three malignancy associated response signature biomarkers in the biological sample obtained from the subject having a cancer with a reference value, wherein increased expression of 1.8-fold or greater of at least three of the biomarkers in the biological sample obtained from the subject relative to the reference value indicates that the cancer is classified as having a poor prognosis or being a malignant cancer, and absence of increased expression of 1.8-fold or greater of at least three of the biomarkers relative to the reference value indicates that the cancer does not have poor prognosis or is not a malignant cancer. ### Claim 54 (depends on 52) The method of claim 52, wherein the cancer is selected from a breast cancer, a triple-negative breast cancer, a Luminal B breast cancer and an epithelial breast cancer. ### Claim at position 28 55-56. (canceled) ### Claim 57 (depends on 52) The method of claim 52, wherein if the cancer is classified as having a poor prognosis or being a malignant cancer, the method further comprises the step of administering at least one proteasome inhibitor to the subject. ### Claim 58 (depends on 57) The method of claim 57, wherein the at least one proteasome inhibitor specifically inhibits any of the proteasomal malignancy associated response signature biomarkers set forth in SEQ ID NOs: 1, 2, 4, 7, 15, 16, 21, 29, 33, 34, 38, 60, 74, 76, and 149. ### Claim 59 (depends on 57) The method of claim 57, wherein the at least one proteasome inhibitor specifically inhibits the malignancy associated response signature biomarker set forth in SEQ ID NOs: 134 (MCL-1). ### Claim 60 (depends on 57) The method of claim 57, wherein the at least one proteasome inhibitor is an siRNA or antisense RNA agent. ### Claim 61 (depends on 57) The method of claim 57, wherein the at least one proteasome inhibitor is bortezomib. ### Claim 62 An assay comprising the steps of: a. measuring expression of ten or more of the 154 malignancy associated response signature biomarkers set forth in SEQ ID NOs: 1-154 in a biological sample obtained from a subject having cancer, and b. comparing the expression of the ten or more of the 154 malignancy associated response signature biomarkers set forth in SEQ ID NOs: 1-154 in the biological sample obtained from the subject having a cancer with a reference value, wherein increased expression of at least ten of the measured biomarkers in the biological sample obtained from the subject relative to the reference value diagnoses the patient as having a poor prognosis or malignant cancer, and absence of increased expression of at least ten of the measured biomarkers relative to the reference value diagnoses the patient as not having a poor prognosis or malignant cancer, wherein the prognosis comprises relative survival rate, relative risk of metastasis, treatment option, or any combination thereof. ### Claim 63 (depends on 62) The assay of claim 62, wherein the increased expression of the ten or more of the 154 malignancy associated response signature biomarkers in the biological sample determines a prognosis for the subject having the cancer. ### Claim 64 (canceled) ### Claim 65 (depends on 62) The assay of claim 62, wherein the cancer is a breast cancer. ### Claim 66 (depends on 65) The assay of claim 65, wherein the breast cancer is selected from a triple-negative breast cancer, a Luminal B breast cancer, and an epithelial breast cancer. ### Claim at position 39 67-68. (canceled) ### Claim 69 (depends on 62) The assay of claim 62, wherein if the subject is diagnosed as having a poor prognosis or malignant cancer, the subject is administered at least one proteasome inhibitor. ### Claim 70 (depends on 62) The assay of claim 62, wherein if the subject is diagnosed as having a poor prognosis or malignant cancer and wherein at least two of the ten or more genes having increased expression is a proteasomal malignancy associated response signature biomarker of SEQ ID NOs: 1, 2, 4, 7, 15, 16, 21, 29, 33, 34, 38, 60, 74, 76, and 149, the subject is administered at least one proteasome inhibitor. ### Claim 71 (depends on 69) The assay of claim 69, wherein the at least one proteasome inhibitor specifically inhibits any of the proteasomal malignancy associated response signature biomarkers set forth in SEQ ID NOs: 1, 2, 4, 7, 15, 16, 21, 29, 33, 34, 38, 60, 74, 76, and 149. ### Claim 72 (depends on 69) The assay of claim 69, wherein the at least one proteasome inhibitor specifically inhibits the malignancy associated response signature biomarker set forth in SEQ ID NOs: 134 (MCL-1). ### Claim 73 (depends on 69) The assay of claim 69, wherein the at least one proteasome inhibitor is an siRNA or antisense RNA agent. ### Claim 74 (depends on 69) The assay of claim 69, wherein the at least one proteasome inhibitor is bortezomib. ### Claim 75 (depends on 69) The assay of claim 69, wherein if the subject is diagnosed as having a poor prognosis or malignant cancer, the subject is administered at least one histone deacetylase inhibitor. ### Claim 76 (depends on 75) The assay of claim 75, wherein the at least one histone deacetylase inhibitor is an siRNA or antisense RNA agent. ### Claim 77 (depends on 75) The assay of claim 75, wherein the at least one histone deacetylase inhibitor is trichostatin A (TSA) or Vorinostat. ### Claim 78 (depends on 69) The assay of claim 69, wherein if the subject is diagnosed as having a poor prognosis or malignant cancer, the subject is administered at least one glygolytic inhibitor. ### Claim 79 (depends on 78) The assay of claim 78, wherein the at least one glycolysis inhibitor is an siRNA or antisense RNA agent. ### Claim 80 An assay comprising the steps of: a. dividing a cell culture grown from a biopsy obtained from a subject having cancer into at least 5 separate cultures; b. exposing each of the at least 5 separate cultures to a different inhibitory agent, wherein each said different inhibitory agent specifically inhibits a different malignancy associated response signature biomarker set forth in SEQ ID NOs: 1-154 c. growing each of the at least 5 separate cell cultures of step (b) for at least 12 hours; d. measuring viability of the cells from each of the cultures of step (c), wherein if the total viability of the cells in at least 60% of the cultures is decreased by at least 25%, then the biopsy obtained from the subject comprises cancer-initiating cells. ### Claim 81 (depends on 80) The assay of claim 80, wherein the inhibitory agents are selected from siRNA agents, antisense RNA agents, or small molecules, that specifically inhibit any of the cancer gene signature biomarkers set forth in SEQ ID NOs: 1-154. ### Claim 82 (depends on 80) The assay of claim 80, wherein the inhibitory agents specifically inhibit any of the malignancy associated response signature biomarkers set forth in SEQ ID NOs: 1-4, 6, 7, 13, 14, 23, 29, 33-35, 37, 38, 66, 72, 77, 78, 84, 91, 93, and 151. ### Claim 83 (depends on 80) The assay of claim 80, wherein at least one inhibitory agent specifically inhibits any of the malignancy associated response signature biomarkers set forth in SEQ ID NOs: 1-4, 6, 7, 13, 14, 23, 29, 33-35, 37, 38, 66, 72, 77, 78, 84, 91, 93, and 151. ### Claim 84 (depends on 80) The assay of claim 80, wherein at least one inhibitory agent specifically inhibits any of the proteasomal malignancy associated response signature biomarkers set forth in SEQ ID NOs: 1, 2, 4, 7, 15, 16, 21, 29, 33, 34, 38, 60, 74, 76, and 149. ### Claim 85 (depends on 80) The assay of claim 80, wherein at least one inhibitory agent specifically inhibits any of the mitosis cancer gene signature biomarkers set forth in SEQ ID NOs: 9, 17, 22, 23, 31, 37, 35, 52, 68, 80, 101, 124, 130, and 137. ### Claim 86 (depends on 80) The assay of claim 80, wherein at least one inhibitory agent specifically inhibits any of the RNA splicing malignancy associated response signature biomarkers set forth in SEQ ID NOs: 5, 10, 11, 18, 20, 26, 36, 41, 57, 61, 123, 127, and 151. ### Claim 87 (depends on 80) The assay of claim 80, wherein at least one inhibitory agent specifically inhibits any of the molecular transport malignancy associated response signature biomarkers set forth in SEQ ID NOs: 12, 13, 32, 48, 51, 56, 64, 91, 96, 98, 108, 121, and 147. ### Claim 88 (depends on 80) The assay of claim 80, wherein at least one inhibitory agent specifically inhibits any of the metabolic malignancy associated response signature biomarkers set forth in SEQ ID NOs: 46, 49, 65, 69, 72, 78, 104, 107, 112, 126, 138, 141, and 153. ### Claim at position 60 89-101. (canceled) ### Claim 102 (depends on 1) A method of identifying a candidate therapeutic agent against a cancer initiating cell comprising the step of a. exposing a BPLER cell culture to a test agent, wherein said BPLER cell culture comprises human breast primary epithelial cells (BPE) transformed with a defined set of genetic elements; b. measuring expression of at least ten of the 154 malignancy associated response signature biomarkers of claim 1 in the culture, c. comparing the expression of the same at least 10 biomarkers of malignancy associated response signature biomarkers of claim 1 as was measured in step (b) to an expression signature reference from a BPLER cell culture that has not been exposed to the test agent, wherein a decrease of expression of at least 5 of the at least 10 genes in the test culture compared to the expression signature reference indicates that the test agent is a candidate therapeutic agent against a cancer initiating cell. ### Claim at position 62 103-105. (canceled)