# US 2014/0193811 A1 · Culture Based Screening Assay and Methods of Use Thereof to Identify Agents Which Modulate Tumor Development, Invasion and Differentiation > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 2014/0193811 A1 - Document ID: US20140193811A1 - Status: Published application - Kind: A1 - Application number: 14/086,495 - Filed: 2013-11-21 - Published: 2014-07-10 - Earliest claimed: 2007-10-31 - Assignees: The Trustees of the University of Pennsylvania - Applicants: The Trustees of the University of Pennsylvania - Inventors: Anil K. Rustgi; Carmen Z. Michaylira; Douglad B. Stairs - Web page: https://patentgrep.com/patent/US20140193811A1 ## CPC - C12Q 1/6876: Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes - C12N 5/0679: {Cells of the gastro-intestinal tract} - C12N 5/0697: {Artificial constructs associating cells of different lineages, e.g. tissue equivalents (blood vessels C12N5/0691)} - G01N 33/5044: {involving specific cell types} - G01N 33/54306: {Solid-phase reaction mechanisms} - C12N 2500/25: Insulin-transferrin; Insulin-transferrin-selenium - C12N 2500/32: Amino acids - C12N 2500/40: Nucleotides, nucleosides or bases - C12N 2501/39: Steroid hormones - C12N 2501/392: Sexual steroids - C12N 2501/395: Thyroid hormones - C12N 2502/1323: Adult fibroblasts - C12N 2502/23: Gastro-intestinal tract cells - C12N 2502/30: tumour cells - C12N 2503/04: Screening or testing on artificial tissues - C12N 2533/54: Collagen; Gelatin - C12Q 2600/112: Disease subtyping, staging or classification - C12Q 2600/158: Expression markers - C12Q 2600/16: Primer sets for multiplex assays - G01N 2333/4706: stimulating, promoting or activating activity - 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 - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) - 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 A 3D organotypic culture which phenocopies aggressive, invasive cancer and methods of use thereof are provided. ## Claims ### Claim at position 1 1-19. (canceled) ### Claim 20 A method of in vitro screening of a compound comprising a) incubating an organotypic culture comprising an artificial stroma including a mixture of collagen and human fibroblasts isolated from a tissue of interest, said stroma being overlayed with primary keratinocytes, said keratinocytes comprising nucleic acids encoding at least one activated oncogene and at least one inactivated tumor suppressor gene, said culture exhibiting enhanced cell migration and invasion in an artificial extracellular matrix relative to control keratinocytes which lack said nucleic acids in the presence and absence of said compound, and observing the effect of said compound upon the migration and invasive characteristics of said keratinocytes relative to cells which are not incubated with said compound. ### Claim 21 (depends on 20) The method of claim 20, further comprising observing the effect of said compound on cultures which do not contain said nucleic acids. ### Claim 22 (depends on 20) The method according to claim 20, further comprising seeding a malignant cell on an artificial stroma to create a positive control model of tumor formation, tumor invasion or tumor-stroma interaction. ### Claim 23 (depends on 20) The method of claim 20, wherein said method comprises contacting said culture with a cellular migration and invasive process altering agent and assessing the effect of said compound on said culture in the presence of said agent. ### Claim 24 (depends on 23) The method of claim 23, wherein said agent is an MMP-9 inhibitor. ### Claim 25 (depends on 23) The method of claim 23, wherein the agent in an AKT inhibitor. ### Claim 26 (depends on 20) The method of claim 20, further comprising determining whether said compound modulates the expression level of at least one cell adhesion molecule selected from the group of N-cadherin, melanoma cell adhesion molecule, periostin and cadherin11. ### Claim 27 (depends on 20) The method of claim 20, further comprising determining whether said compound modulates the expression level of fibroblast hepatocyte growth factor. ### Claim 28 (depends on 20) The method according to claim 20, wherein said compound is screened for toxicity to human tissue, and wherein said observing step comprises observing the effects of said compound on the morphology and life span of said keratinocytes thereby identifying a compound which reduces the life span of the cells or has a negative impact on the morphology of said cells. ### Claim 29 (depends on 20) A method of in vitro screening of a compound comprising contacting an organotypic culture as claimed in claim 20, with said compound wherein said cells of interest are obtained from a tumor isolated from a human patient and observing the effect of said compound upon said migration and invasive characteristics of said cells relative to tumor derived organotypic cultures which are not contacted with said compound. ### Claim 30 (depends on 29) The method of claim 29, wherein said method comprises contacting said culture with a cellular migration and invasive process altering agent and assessing the effect of said compound on said culture in the presence of said agent. ### Claim 31 A genetic signature associated with cells exhibiting increased invasiveness, comprising nucleic acids encoding N-cadherin, melanoma cell adhesion molecule, periostin and cadherin 11, said nucleic acids exhibiting differential expression in invading versus non-invading cells as set forth in Table 1. ### Claim 32 An assay kit for determining the presence a genetic signature encoding proteins associated with increased tumor cell migration and invasion in a biopsy sample of esophageal cells, comprising antibodies immunologically specific for at least two proteins selected from the group consisting of cadherin 2, type 1, N cadherin, melanoma cell adhesion molecule, periostin, osteoblast specific factor, cadherin 11, type 2, OB-cadherin, claudin 1, claudin 4 claudin 7, occludin, one of said at least one protein or antibody being detectably labeled. ### Claim 33 (depends on 32) The kit of claim 32 comprising antibodies immunologically specific for N-cadherin, melanoma cell adhesion molecule, periostin and cadherin11. ### Claim 34 (depends on 32) The assay kit according to claim 32, wherein said antibodies are immobilized on a solid support. ### Claim 35 (depends on 32) The assay kit according to claim 32 further comprising said proteins obtained from normal tissue for use as a control sample. ### Claim 36 (depends on 31) A method for identifying patients at increased risk for metastatic esophageal cancer comprising obtaining a biological sample from said patient and assessing said patient for the presence of at least one of the molecules present in the genetic signature of claim 31. ### Claim 37 (depends on 36) The method of claim 36, wherein said biological sample is sera from said patient and said molecule is periostin. ### Claim 38 (depends on 37) The method of claim 37, wherein said periostin is detected using antibodies immunologically specific therefor and an immunocomplex formed between said periostin and said antibody is detected. ### Claim 39 (depends on 37) The method of claim 37, wherein said periostin is detected using nucleic acids that hybridize to nucleic acids encoding said periostin, said nucleic acids comprising a detectable label. ### Claim 40 (depends on 37) The method of claim 37, wherein all of the proteins encoded by the nucleic acids of the genetic signature are detected.