ENAapplication/xmlftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/016/SRR21844816/SRR21844816_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/014/SRR21844814/SRR21844814_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/012/SRR21844812/SRR21844812_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/018/SRR21844818/SRR21844818_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/019/SRR21844819/SRR21844819_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/016/SRR21844816/SRR21844816_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/013/SRR21844813/SRR21844813_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/014/SRR21844814/SRR21844814_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/017/SRR21844817/SRR21844817_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/011/SRR21844811/SRR21844811_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/017/SRR21844817/SRR21844817_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/015/SRR21844815/SRR21844815_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/011/SRR21844811/SRR21844811_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/015/SRR21844815/SRR21844815_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/018/SRR21844818/SRR21844818_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/013/SRR21844813/SRR21844813_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/019/SRR21844819/SRR21844819_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR218/012/SRR21844812/SRR21844812_1.fastq.gzprimaryOK200GenomicsBader, Biomedical Engineering, Johns Hopkins Universityhttps://www.ebi.ac.uk/ena/browser/view/PRJNA888356Mus musculusxref:PubMed:36604566Inter-patient and intra-tumoral heterogeneity complicate the identification of predictive biomarkers and effective treatments for basal triple negative breast cancer (b-TNBC). Invasion is the initiating event in metastasis and can occur by both collective and single-cell mechanisms. We cultured primary organoids from a b-TNBC genetically engineered mouse model in 3D collagen gels to characterize their invasive behavior. We observed that organoids from the same tumor presented different phenotypes that we classified as non-invasive, collective and disseminative. To identify molecular regulators driving these invasive phenotypes, we developed a workflow to isolate individual organoids from the collagen gels based on invasive morphology and perform RNA sequencing. We next tested the requirement of differentially regulated genes for invasion using shRNA knock-down. Strikingly, KRAS was required for both collective and disseminative phenotypes. We then performed a drug screen targeting signaling nodes upstream and downstream of KRAS. We found that inhibition of EGFR, MAPK/ERK, or PI3K/AKT signaling reduced invasion. Of these, ERK inhibition was striking for its ability to potently inhibit collective invasion and dissemination. We conclude that different cancer cells in the same b-TNBC tumor can express different metastatic molecular programs and identified KRAS and ERK as essential regulators of collective and single cell dissemination. Overall design: C3(1)-TAg organoids were plated in 3D fibrillar collagen I for 3 days. We classified organoids based on their invasive phenotype (non-invasive, invasive (collective) and disseminative. Organoids were then removed from the gels with sharp forceps and were kept on dry ice. Trizol (Invitrogen) was then added and RNA extracted. Three independent experiments were conducted.ENAKI-RAS, primary breast cancer, malignant tumor of the breast, Breast Neoplasms, Neoplasms, Kras2, NS3, Benign Neoplasm, mammary neoplasm, Mammary Cancers, Human Mammary Neoplasms, Tumor, Malignant, Human, Breast Malignant Tumor, ras, tumor of the breast, "neoplasm of breast (disorder)" EXACT [SNOMEDCT_2005_07_31:126926005], Breast Tumor, BREAST NEOPL, Kras-2, "mammary tumor" EXACT [CSP2005:2016-0671], breast tumor, cancer of the breast, neoplasm, C-K-RAS, k-ras, KRAS2, KRAS1, rask2, p21B, BC, Mammary Neoplasms, malignant neoplasm of breast, Human Mammary, Malignancy, Mammary Neoplasm, Mammary Carcinoma, K-RAS4B, K-RAS4A, Human Mammary Carcinomas, Cancer of the Breast, Neoplasias, neoplasm of breast, neoplasm of the breast, breast neoplasm, cancer of breast, malignant neoplasm, Malignant Tumor of Breast, Mammary, Mammary Cancer, Malignancies, Breast, mammary cancer, mammary tumor, Breast Malignant Neoplasms, Tumors, Cancer, Human Mammary Neoplasm, Breast Carcinoma, Carcinoma, breast tumour, Breast Carcinomas, "breast neoplasm" EXACT [MTH:120], Malignant Neoplasm, Collective, Ki-ras, K-RAS2B, K-RAS2A, K-ras, c-Ki-ras, "breast tumor" EXACT [NCI2004_11_17:C2910], Cell, Breast Malignant Neoplasm, MT, Benign, Mammary Carcinomas, p21, Neoplasm, Human Mammary Carcinoma, NOS, AI929937, Malignant Neoplasm of Breast, tumor of breast, Carcinomas, Breast Neoplasm, primary cancer, NS, tumour of the breast, NEOPL BREAST, Programs., Benign Neoplasms, Cancers, breast neoplasm (disease), "mammary neoplasm" RELATED [], malignant tumor, Malignant Neoplasms, tumour of breast, breast cancer, RASK2, kras, Breast Tumors, Breast Cancer, CFC2, Breast Malignant Tumors, Cancer of Breast, cancer, Neoplasia, breastKI-RAS, primary breast cancer, malignant tumor of the breast, Breast Neoplasms, Neoplasms, Kras2, NS3, Benign Neoplasm, mammary neoplasm, Mammary Cancers, Human Mammary Neoplasms, Tumor, Malignant, Human, Breast Malignant Tumor, ras, tumor of the breast, "neoplasm of breast (disorder)" EXACT [SNOMEDCT_2005_07_31:126926005], Breast Tumor, BREAST NEOPL, Kras-2, "mammary tumor" EXACT [CSP2005:2016-0671], breast tumor, cancer of the breast, neoplasm, C-K-RAS, k-ras, KRAS2, KRAS1, rask2, p21B, BC, Mammary Neoplasms, malignant neoplasm of breast, Human Mammary, Malignancy, Mammary Neoplasm, Mammary Carcinoma, K-RAS4B, K-RAS4A, Human Mammary Carcinomas, Cancer of the Breast, Neoplasias, neoplasm of breast, neoplasm of the breast, breast neoplasm, cancer of breast, malignant neoplasm, Malignant Tumor of Breast, Mammary, Mammary Cancer, Malignancies, Breast, mammary cancer, mammary tumor, Breast Malignant Neoplasms, Tumors, Cancer, Human Mammary Neoplasm, Breast Carcinoma, Carcinoma, breast tumour, Breast Carcinomas, "breast neoplasm" EXACT [MTH:120], Malignant Neoplasm, Collective, Ki-ras, K-RAS2B, K-RAS2A, K-ras, c-Ki-ras, "breast tumor" EXACT [NCI2004_11_17:C2910], Cell, Breast Malignant Neoplasm, MT, Benign, Mammary Carcinomas, p21, Neoplasm, Human Mammary Carcinoma, NOS, AI929937, Malignant Neoplasm of Breast, tumor of breast, Carcinomas, Breast Neoplasm, primary cancer, NS, tumour of the breast, NEOPL BREAST, Programs., Benign Neoplasms, Cancers, breast neoplasm (disease), "mammary neoplasm" RELATED [], malignant tumor, Malignant Neoplasms, tumour of breast, breast cancer, RASK2, kras, Breast Tumors, Breast Cancer, CFC2, Breast Malignant Tumors, Cancer of Breast, cancer, Neoplasia, breastfalseTriple negative breast tumors contain heterogeneous cancer cells expressing distinct KRAS-dependent collective and disseminative invasion programsTriple negative breast tumors contain heterogeneous cancer cells expressing distinct KRAS-dependent collective and disseminative invasion programs2025-09-242023-01-23PRJNA888356GSE2150691009036604566