Abstract C32: The critical roles of tumor-initiating cell and lymph node stromal microenvironment in human colorectal cancer extranodal metastasis
Authors: Myers Tamara , Obokhare Izi , Grimes Chelsea , Zhang Xin , Silinsky Jennifer , Cordova Jose , Green Heather , Margolin David , Li Li 
Published: August 2014

In the U.S., colorectal cancer (CRC) is the third most common cancer and the second most common cause of cancer-related mortality, with an estimated incidence of 143,000 cases and 51,000 deaths per year. The outcome of CRC patients depends on multiple factors, such as, staging (depth of tumor invasion into colon or rectum wall), lymph node (LN) involvement, local recurrence, and extra-nodal metastasis. Despite optimal oncologic treatment including surgery, chemotherapy and/or radiation therapy, up to 50% of the American Joint Committee on Cancer stage II and III patients will develop extra-nodal metastasis. The most common metastatic sites are the liver or lung. These metastatic cancer cells have acquired a chemoresistant phenotype and developed genetic alterations. Recent studies from our group and others have proposed that metastasis or recurrence is closely associated with the presence of CRC-cancer stem cells or -tumor initiating cells (Co-TIC) expressing CD133 and the chemokine (C-X-C motif) receptor 4 (CXCR4), and their interaction with the LN microenvironment.

Our hypothesis is that LN stromal cells prime, support, educate, and stimulate Co-TIC through soluble factors such as the chemokine (C-X-C motif) ligand 12 (CXCL12, ligand to CXCR4), and CD133+CXCR4+ Co-TIC are responsible for drug resistance and extra-nodal metastasis. In this study, we investigated the critical roles of Co-TIC and LN stromal microenvironment in human CRC extra-nodal metastasis using our unique humanized orthortopic xenograft model. The luciferase-tagged colon cancer cell lines and CRC cancer cells isolated from consented patients were injected intra-rectal (IR) to NOD/SCID mice. The LN stromal cell lines HK and CXCL12 knockdown HK (HK-KD-A3) cells were co-inoculated with CRC cells. In addition, Co-TIC were FACS sorted based on positivity of cell surface markers CD326, CD133 and CXCR4. Tumor growth and extra-nodal metastasis were followed by luciferase activity as detected by the IVIS imaging system and quantitative PCR for human DNA in mice livers and lungs.

The IR model closely mimicked the expected pattern of CRC spread by showing progressive CRC tumor growth under rectum mucosa with liver and lung metastasis in NOD/SCID mice both injected with CRC cell lines and patient cancer cells. The presence of HK cells or pre-incubation of cancer cells with HK cell conditioned media promoted CRC cell distant metastasis. CXCL12 knockdown HK-KD-A3 cells showed significantly impaired HK cell support of CRC primary tumor formation and more importantly distant organ metastasis. In the presence of HK cells, CD133+CXCR4+ Co-TIC showed increased tumor formation and extra-nodal metastasis in comparison to un-separated and double negative cancer cell populations.

Our results indicate both Co-TIC and LN stromal factors play crucial roles in CRC metastasis through CXCL12/CXCR4 axis. Blocking the signaling in Co-TIC/LN stromal interaction may lead to an effective therapy to prevent CRC extra-nodal metastasis, a major cause of cancer morbidity and mortality. In addition, the IR model would serve as a launching pad for evaluating the mechanism of CRC metastasis and testing novel targeted therapies.

Citation Format: Tamara Myers, Izi Obokhare, Chelsea Grimes, Xin Zhang, Jennifer Silinsky, Jose Cordova, Heather Green, David Margolin, Li Li. The critical roles of tumor-initiating cell and lymph node stromal microenvironment in human colorectal cancer extranodal metastasis. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr C32.