Novel Methods for Detecting Cell Interactions in the Tumor Microenvironment

PI Name: CONDEELIS, JOHN S  CONDEELI@AECOM.YU.EDU
Institute: Albert Einstein College of Medicine

Co-PI: James Castracane  jcastracane@uamail.albany.edu

We have developed methods to observe tumor cell invasion and intravasation directly in rat and mouse mammary tumors (intravital multiphoton imaging), and correlate these cell behaviors with metastatic potential and gene expression patterns of the invasive subpopulation of tumor cells in the primary tumor (in vivo invasion assay using catheter collection coupled to expression profiling). Furthermore, these methods can be used to characterize microenvironments that cause autocrine and paracrine-mediated cell migration resulting in the accumulation of cells around an initiating chemotactic signal. The current catheter method is limited to 4 hours of collection which makes it less suitable for looking at longer term events that require infiltration of cells from outside the tumor and angiogenesis. The extension of this technology to other ligands and longer times of cell collection will allow us to explore the variety of tumor cell-stromal cell interactions, for example involving immune cells and migrating endothelial cells, that are involved in invasion and metastasis of tumor cells. In addition, gaining access to these longer term cell interactions will allow exploration of gene expression changes that occur in tumor cells in response to different stromal cell interactions, and determine if there exists a common invasion signature of invasive tumor cells that may have prognostic and therapeutic value. Therefore, using nanotechnology, we will miniaturize and expand the versatility of the catheter collection protocol used in the in vivo invasion assay to create a NANo IntraVital Device (NANIVID).

The specific aims of the proposal are:
1. Preparation of human heterotransplant and human cell line-derived breast tumors for use with the NANIVID
2. Fabrication of the NANIVID for use in mouse and human mammary tumors
3. Test NANIVID prototypes:
3a. that can be inserted into a breast tumor
3b. that can report remotely the status of cell collection while in vivo
3c. that can be retrieved for identification of cell types and gene expression profiling
4.1. Define paracrine and autocrine loops involved in invasion of the NANIVID
4.2. Identify the cell types involved in invasion of the NANIVID
4.3. Determine gene expression profiles of invasive cells collected using the NANIVID
5. Extend the use of the NANIVID to human tumors

Therefore, the NANIVID will be an extension of a well documented and validated catheter based method but will extend its use to a wider variety of stromal cell types and to longer durations of collection. The NANIVID will be composed of a chemoattractant source (dendromer-EGF), capsule (cell trap), counter (nanochip for electrically sensing cell mass), remote reporter (passive response scanner allow reading of the chip output), retraction tool (mechanical retraction protocol to pull the NANIVID out of the tissue without disrupting its contents) and micro-fluidic pore (used to expel living intact cells from the NANIVID with a gentle flow of buffer). This technology will be capable of mapping cell-to-cell interactions in the human breast cancer microenvironment. The experiments using the NANIVID will define chemotactic responses and visualize them in real-time through several days. Experiments will utilize both mouse and human tissue allowing the identification of the tumor microenvironment for metastasis (TMEM) in humans and its comparison to the TMEM previously identified in mouse models. Thus it will provide unique insights into the origins and maintenance of the TMEM as the tumor progresses through to its malignant state. It will generate novel data and gene expression signatures in particular cell types that can then be queried in human breast cancers and correlated to prognosis.