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335 Development of Advanced Culture Systems for Expansion of Cancer Stem Cells

Fast-Track proposals will be accepted.

Direct-to-Phase II will not be accepted.

Number of anticipated awards: 4–7

Budget (total costs, per award):
Phase I: $225,000 for 9 months;
Phase II: $1,500,000 for 2 years

It is strongly suggested that proposals adhere to the above budget amounts and project periods. Proposals with budgets exceeding the above amounts and project periods may not be funded.

The deadline for receipt of all contract proposals submitted in response to this solicitation has expired. It was: November 5, 2014 by 4:30 PM ET.

Summary

Tumors consist of heterogeneous cell populations in which only a small fraction, less than 1%, is able to seed new tumors by transplantation, functionally defined as cancer stem cells (CSCs). There is growing interest in identifying markers and therapeutically targeting the CSC population in tumors. Recent studies have shown that CSCs have different drug sensitivities compared to the bulk population and represent an attractive therapeutic target. Studying these cells, however, has been a challenge due to their low abundance in vivo and the phenotypic plasticity they exhibit during expansion. Using current methods, isolated CSCs lose the expression of CSC markers and tumor initiating capacity when cultured in vitro or in vivo in xenograft animal models. The proportion of CSCs tends to an equilibrium level of less than 1% over time, and the cell population derived from CSC cultures typically recapitulates the heterogeneous nature of the original population. Thus, the goal of this contract topic is to meet the critical need to develop cell culture systems that can specifically grow CSCs for basic and translational research.

Developments in stem cell engineering and tissue engineering have generated new culture systems to accelerate the expansion of embryonic, induced pluripotent, and adult stem cell populations in vitro. These systems include technologies such as three dimensional (3D) culture systems containing extracellular matrix components and topological features, or bioreactors for large scale culture of cell spheroids. Preliminary data suggest that these technologies or similar culture systems may be applicable for quick and reproducible expansion of CSCs. Thus, commercial development of these culture systems specifically for CSC culture may have a significant impact in basic research and drug screening applications.

Project Goals

The purpose of this topic is to develop cell culture systems that can effectively grow cancer stem cells in vitro without the loss of cancer stem cell markers or tumor initiating potential.

The specific goals are to demonstrate that the cell culture system can expand the population of cancer stem cells isolated from established cancer cell lines, derived from either human or animal model systems (Phase I), and primary tumors from appropriate animal models or patient biospecimens (Phase II) that can then be harvested for use in downstream assays.

To successfully meet this goal, applicants will need to demonstrate that cancer stem cells grown using their system: 1) maintain the same tumor initiating potential using an established methodology; 2) maintain the same expression characteristics of accepted cancer stem cell markers; 3) can be easily and effectively harvested by a protocol that maintains the viability and tumor initiating phenotype of cancer stem cells; 4) can be readily used in downstream assays including, but not limited to, molecular read-outs (such as genomic, proteomic, metabolomic, or epigenomic analyses) or cell based read-outs (such as proliferation, migration, invasion, or apoptosis).

It is anticipated that applicants will employ innovative 3D cell culture or bioreactor systems in combination with defined media and growth factor conditions. However, applicants are free to employ any approach that will generate the desired results and meet the criteria listed above. Applicants may utilize feeder cells or co-culture conditions, provided the cancer stem cells can be effectively separated from other cell types at the point of harvest. Applicants are not restricted to specific cancer types, but should justify the choice of cancer type to study from both a scientific and commercial perspective.

Systems of particular interest will be amenable to scale up, demonstrate reliability and robustness at a price point that is compatible with market success and widespread adoption by the research community. Further, systems that can demonstrate success in expanding cancer stem cells from patient derived samples will also be of particular interest.

The focus of this contract concept is not to develop a screening platform for agents that selectively kill or arrest cancer stem cells, though a screen for such agents could be used to demonstrate the proof of principle that cancer stem cells grown using this technology can be used in downstream applications. Additionally, this contract is not intended to develop systems to expand non-cancer derived stem cells, though applicants may propose the use of such systems provided they can demonstrate effectiveness with cancer stem cells.

Phase I Activities and Expected Deliverables:

  • Culture system should include physical (i.e. scaffold, hydrogel, or matrix) and chemical (i.e. media, oxygen tension, pH) components
  • Culture system should be able to expand population up to 107 cells
  • Live cells should be able to be extracted and re-seeded from the culture system
  • Systems of particular interest will incorporate methods for freezing and long-term storage of expanded cells.
  • Validate culture system and SOP using a cancer cell line known to contain a CSC population.
    • Using an appropriate cell line model system, isolate the CSC subpopulation using established protocols and markers. Applicants should clearly outline the evidence for CSC subpopulations in the chosen cell line(s).
    • Culture the isolated CSC population using the developed culture system. Examples of cancer cell lines include MCF-7 breast cancer cells or HCT116 colorectal cancer cells.
    • Identify the sustainability of CSC markers and tumor initiating phenotype after culture in system using established protocols (e.g. CSC-specific cell surface markers, Hoechst 33342 exclusion, colony formation, tumor transplantation).
  • Submit a statement to NCI that specifies metrics and criteria used to evaluate the CSC population and phenotype, and justification from both a scientific and commercial perspective for why the specific cancer type or cell line is being used.
    • Specify SOP and biomarkers (cell marker or assays) used to identify CSC population.
    • Specify SOP for assays used to define CSC phenotype.

Phase II Activities and Expected Deliverables

  • Demonstrate ability of developed culture system to expand CSCs isolated from in vivo samples for at least one cancer type.
    • Culture CSC population isolated from tumor biospecimens (human or mouse) to a minimum of 107 cells.
    • Compare genomic, proteomic, metabolomic, and epigenomic profile of original CSC population and expanded population.
    • Demonstrate reproducibility by expanding CSC populations from 10 different biospecimens.
  • Test tumor initiating capacity of expanded CSC population using appropriate established in vivo assays.
Updated: June 24, 2015