UC-RAPID | SBIR

OUSD (R&E) critical technology area(s): Biomanufacturing, Biotechnology

Objective: Develop and demonstrate a universal, species-agnostic cell culture platform that (1) enables rapid establishment of viable, reproducible cell lines - including induced pluripotent stem cells (iPSCs) where feasible - from diverse non-model species (vertebrate and invertebrate); (2) leverages autonomous, closed-loop culture automation to compress protocol development timelines and ensure cross-laboratory reproducibility; and (3) provides the cellular foundation required to develop and validate in vitro gene-drive and related genetic-biocontrol technologies in non-model species. The platform shall address the long-standing bottleneck in which establishing cell culture for a new, non-model species requires years of manual, trial-and-error optimization - an obstacle that today blocks gene-drive development in non-model invasive species - thereby unlocking multiple applications across biotechnology, agriculture, conservation, and public-health markets.

Description: Cell culture is foundational to modern biotechnology, biomanufacturing, drug discovery, vaccine production, regenerative medicine, and genetic engineering. However, most established cell culture protocols, media formulations, and immortalized or pluripotent cell lines exist only for well-characterized "model" organisms (e.g., human, mouse, rat). For many vertebrate species, including those of emerging interest to defense, biosecurity, agricultural, conservation, and public-health stakeholders, no validated cell culture baseline exists.

Filling this gap is critical for gene-drive and genetic-biocontrol research. Gene drives, genetic elements engineered to propagate a trait through a target population faster than standard inheritance, hold significant promise for controlling disease vectors, suppressing invasive or pest species, and protecting agriculture, ecosystems, and public health. Yet engineering, optimizing, validating and characterizing the safety and efficacy of gene-drive constructs requires robust, species-specific cell lines for the target organism that can enable rapid in vitro assessment of drive activity (e.g., homing/conversion efficiency, off-target activity, and stability) that would otherwise require slow, whole-organism breeding studies.

Establishing cell culture for a novel, non-model species today is a slow and irreproducible process. It requires years of manual optimization across an enormous parameter space (media composition, growth factors, substrate, oxygen tension, temperature, passaging cadence, reprogramming-factor delivery, and differentiation cues), depends heavily on individual operator skill, and frequently fails to transfer between laboratories. This bottleneck limits the speed for standing up new biological capabilities, constrains the breadth and resilience of the bioeconomy, and directly impedes the responsible development of gene-drive and biocontrol countermeasures.

This Phase II SBIR topic seeks a generalizable platform that fundamentally changes this paradigm by combining (1) species-agnostic biological methods for deriving and maintaining cells from arbitrary vertebrate and invertebrate species, (2) autonomous, closed-loop automation that systematically explores culture parameter space and produces reproducible, transferable protocols and cell lines, and (3) the cellular substrate and engineering workflows needed to build and validate in vitro testbeds for gene-drive technologies in those cell lines.

Proposers must address Component 1 and Component 2 below and are highly encouraged to address Component 3. A proposer may submit only one proposal to this topic.

Component 1: Cross-Species Universality (Species-Agnostic Cell Culture & iPSC Derivation) Develop generalizable biological methods that allow viable, stable cell lines to be established from diverse, non-model vertebrate and invertebrate species without bespoke, multi-year, per-species protocol development.

Areas of interest include, but are not limited to:

• Universal or rapidly tunable media, substrate, and supplement formulations applicable across taxonomically diverse species (e.g., across multiple vertebrate classes such as mammals and reptiles).
• Species-agnostic strategies for primary cell isolation, immortalization, and/or induced pluripotency (iPSC reprogramming), including reprogramming-factor delivery and selection approaches that generalize across species.
• Methods for establishing, characterizing, and validating pluripotency, viability, genomic stability, and lineage differentiation potential in species lacking established reference reagents or markers.
• A principled, design-of-experiments (DOE) or model-driven framework for rapidly converging on viable culture conditions for a previously uncharacterized species.
• Cryopreservation, banking, and quality-control approaches that ensure cell line stability and reproducibility across time and locations.

Component 2: Autonomous, Closed-Loop Culture Automation Develop automation that performs cell culture operations with minimal human intervention and that systematically and reproducibly drives protocol optimization for new species.

Areas of interest include, but are not limited to:

• Per-vessel or per-well environmental independence (e.g., independent control of temperature, gas composition, and media conditions) to enable many simultaneous, independent optimization experiments.
• Automated execution of routine and complex culture operations, including feeding, passaging, reprogramming, differentiation, imaging, and sampling.
• Integrated, automated imaging, data capture, and analysis (including machine-learning–enabled morphology/health assessment) feeding a closed-loop optimization engine that proposes and executes the next experimental conditions.
• Software, data architecture, and electronic protocol capture that make optimized protocols portable and reproducible across instruments and laboratories.
• Throughput and parallelization sufficient to compress new-species protocol development from years to weeks or months.

Component 3: Gene-Drive and Genetic-Biocontrol Enablement Develop the cellular substrates, engineering workflows, and validation methods needed to characterize gene-drive and related genetic-biocontrol technologies in non-model target species.

Areas of interest include, but are not limited to:

• Establishment of species-specific cell lines (including germline-relevant or developmentally relevant cell types) suitable as a testbed for gene-drive construct development.
• In vitro assays to characterize gene-drive efficiency, homing/conversion rates, off-target effects, fitness costs, and genetic stability prior to any whole-organism work.
• Containment, reversibility, and safeguard strategies (e.g., molecular confinement, reversal/immunizing drives) evaluable at the cellular level..

Proposers addressing Component 3 must explicitly address responsible-research practices, applicable biosafety/biosecurity and regulatory considerations, and appropriate institutional oversight. All Component 3 work under this topic shall be limited to in vitro / cellular research; no environmental release, and no work resulting in a gene-drive-competent whole organism, is contemplated or permitted under this effort. [BR4.1]

Component Integration
Proposers should describe how species-agnostic biology, closed-loop automation, and (where applicable) gene-drive enablement combine into a single coherent platform - e.g., how the automation systematically searches biological parameter space to converge on a validated cell line for a new target species, and how that cell line then serves as a reproducible substrate for gene-drive engineering and in vitro validation.

PHASE I

This is a Direct-to-Phase II (DP2) topic. No Phase I award will be made under this topic. To be eligible, the proposer must demonstrate that scientific and technical merit and feasibility equivalent to a Phase I project has been achieved prior to submission, through prior work (e.g., internally funded research, other federal or non-federal funding, or other documented prior effort).

Proposers shall provide documentation establishing, at minimum, prior demonstration of feasibility for the component(s) addressed:

1. Cross-species feasibility (Component 1): Demonstrated establishment and/or maintenance of viable cell lines—and/or successful iPSC derivation—from at least two non-model vertebrate and/or invertebrate species, with supporting characterization data (viability, stability, and, where applicable, pluripotency or differentiation). Demonstration spanning more than one taxonomic class is viewed favorably.
2. Automation feasibility (Component 2): Demonstrated operation of automated or closed-loop cell culture functionality (e.g., automated feeding/passaging, independent per-vessel environmental control, automated imaging/analysis, or an operational optimization loop), with supporting performance data.
3. Gene-drive enablement feasibility (Component 3, if addressed): Demonstrated genome editing and/or construct delivery in a non-model cell line, and/or a relevant in vitro assay for genetic-element performance, with supporting data and documented biosafety/oversight practices.

Documentation may include prior data, peer-reviewed or technical reports, prototype descriptions, imaging/characterization results, and prior award or internal-funding history.

Proposals that do not adequately substantiate prior Phase I-equivalent feasibility for the component(s) addressed will be deemed non-responsive.

PHASE II

Mature the platform and demonstrate it on previously uncharacterized species under realistic conditions. Expected activities include:

• Stand up and acceptance-test the integrated platform – species-agnostic culture methods, autonomous closed-loop automation (per-vessel control, automated feed/passage/imaging), and the DOE/optimization engine – validated against reference species prior to launching new-species campaigns.
• Demonstrate establishment of viable, characterized cell lines (including iPSCs where feasible) for at least four new, non-model species not used in the feasibility demonstration, at least one of which must be vertebrate. Proposals focusing on a greater number of vertebrate and/or long-life-cycle species, where in vitro methods deliver the greatest time-compression benefit over conventional whole-organism approaches, will be reviewed favorably.
• Quantify platform performance against the core metrics: time-to-protocol (target: years compressed to weeks/months), reproducibility (cross-run, cross-operator, and where feasible cross-site success rate), and breadth of species coverage.
• Demonstrate autonomous, closed-loop optimization converging on validated protocols, with electronic capture of transferable protocols.
• For efforts including Component 3, demonstrate an in vitro gene-drive/biocontrol culture system and characterization in an established non-model cell line, including efficiency and safeguard/containment assays, under appropriate biosafety oversight.
• Deliver characterized cell lines, validated protocols, datasets, and a final report documenting performance against metrics. Establish a credible path to TRL 6–7 by program end.

Target outcome: Advance the platform from TRL 3–4 (entry) to TRL 6 (representative prototype demonstrated in a relevant environment) by the end of the Phase II base.

Program Metrics:
The following metrics define success and are intended to push performance beyond the current state of the art. Proposers should propose specific, measurable targets meeting or exceeding these goals; proposals that credibly exceed them will be reviewed favorably.

• Time-to-Protocol: Less than 90 days to a validated cell line (and/or iPSC where feasible) for a previously uncharacterized, non-model species.
• Reproducibility: Greater than 85% cross-run success, with demonstrated cross-operator (and, where feasible, cross-site) reproducibility using electronically captured, transferable protocols.
• Species Coverage: Greater than 4 species spanning greater than 2 taxonomic classes, including at least one vertebrate.
• Successful Culture: Establishment and sustained maintenance of viable, characterized cell lines meeting pre-defined acceptance criteria (viability, growth, and marker/characterization profile).
• Genomic Stability: Greater than 20 passages with demonstrated genomic stability; equivalent of at least 5 generations maintained in all demonstrated species.
• Component 3 (Gene-Drive Enablement, if addressed): Demonstrated in vitro construct delivery and gene-drive characterization (e.g., editing/delivery efficiency, homing/conversion, off-target activity, and genetic stability), with functioning in vitro safeguard/containment mechanism

DP2 Base milestones for this program should include:

• Month 2 – Requirements Analysis: Finalize platform requirements, target-species set (at least four species, at least one of which must be vertebrate, with preference for long-life-cycle species), pre-defined acceptance criteria for the Target Metrics, characterization assays, and data schema. Establish biosafety/IBC oversight baseline for Component 3.
• Month 6 – Design Review: Review integrated platform design, species-agnostic methods, automation configuration, and DOE/closed-loop optimization plan; confirm acceptance-tested automation and reference-species validation; complete risk assessment sufficient to launch new-species campaigns.
• Month 8 – Optimization Campaign Initiation: Begin autonomous, closed-loop optimization to establish viable culture for new, non-model target species; confirm primary isolation/iPSC entry conditions.
• Month 12 – Heading Check: Review establishment progress against the Target Metrics: time-to-protocol, cross-run/cross-operator reproducibility, species-coverage trajectory, and genomic-stability/passage status; assess readiness for characterization and (if applicable) gene-drive in vitro work. Update risks and demonstration plan.
• Month 16 – Transition Planning: Develop technology transfer, commercialization, and scaling plans; identify transition partners; complete technoeconomic analysis.
• Month 17 – Capability Demonstration: Demonstrate establishment and characterization of validated cell lines/iPSCs for at least four new species (at least one of which must be vertebrate) against the Target Metrics: time-to-protocol, reproducibility, species coverage, and genomic stability; (Component 3) demonstrate in vitro gene-drive construct delivery, characterization, and safeguard/containment.
• Month 18 – Final Reporting: Summarize platform performance against the Target Metrics, delivered cell lines/protocols/datasets, lessons learned, and transition recommendations.

DP2 Base deliverables should include:

• Month 2 – Requirements Specification: Target-species list, pre-defined Target Metric acceptance criteria (time, reproducibility, species breadth, genomic stability), characterization assay panel, data/protocol schema, and Biosafety/IBC Oversight Plan (Component 3).
• Month 6 – Design Review Package: Integrated design and species-agnostic methods package, automation configuration and acceptance-test/reference-species results, optimization plan, updated schedule, risk register, and presentation.
• Month 8 – Optimization Campaign Plan: Target species, entry conditions, DOE design, run cadence, characterization checkpoints, and (if applicable) gene-drive workflow prerequisites and biosafety logistics.
• Month 12 – Heading Check Package: Cell viability/stability/pluripotency data, time-to-protocol, reproducibility, species-coverage, and genomic-stability/passage results measured against the Target Metrics, automation throughput, updated schedule/risk register, demonstration readiness assessment, and slide deck presentation.
• Month 16 – Transition Plan Report/PowerPoint: Commercialization and scaling strategy, transition-partner identification, and Technoeconomic Analysis (TEA).
• Month 17 – Demonstration Data Package: Performance results measured against the Target Metrics (time-to-protocol, reproducibility, species coverage, genomic stability) for at least four new species (at least one of which must be vertebrate); (Component 3) in vitro gene-drive construct, characterization, and safeguard/containment results.
• Month 18 – Final Report: Performance Data Package (metrics achieved vs. Target Metrics), delivered/characterized cell lines, validated electronically captured transferable protocols, complete datasets, lessons learned, and transition recommendations.

Phase III dual use applications

Demonstrate scalability, transferability, and deployment:

Defense applications: Biosecurity and genetic-biocontrol countermeasures; protection of force health from disease vectors; defense against invasive or pest species threatening installations, ecosystems, and agriculture; resilient and expeditionary biomanufacturing; rapid stand-up of biological production and engineering capabilities for novel or mission-relevant species; biological supply-chain resilience.

Commercial applications: Public-health vector control; agricultural pest and invasive-species management; cellular agriculture and cultivated foods; biopharmaceutical and vaccine host-line development; regenerative medicine; species conservation and biobanking; veterinary biotechnology; contract research and bioprocess host development.

References

1. Esfahani, Sajedeh Nasr, Yi Zheng, Auriana Arabpour, et al. “Derivation of Human Primordial Germ Cell-like Cells in an Embryonic-like Culture.” Nature Communications 15, no. 1 (2024): 167. https://doi.org/10.1038/s41467-023-43871-2.
2. Golkar-Narenji, Afsaneh, Piotr Dziegiel, Bartosz Kempisty, James Petitte, Paul Edward Mozdziak, and Artur Bryja. “In Vitro Culture of Reptile PGCS to Preserve Endangered Species.” Cell Biology International 47, no. 8 (2023): 1314–26. https://doi.org/10.1002/cbin.12033.
3. Kong, Xiangyu, and Thorold W. Theunissen. “Primate Embryo Model Leaps across Developmental Boundaries.” Nature, ahead of print, December 3, 2025. https://doi.org/10.1038/d41586-025-03729-7.
4. Panagopoulos, Andreas, Merula Stout, Sinan Kilic, et al. “Multigenerational Cell Tracking of DNA Replication and Heritable DNA Damage.” Nature 642, no. 8068 (2025): 785–95. https://doi.org/10.1038/s41586-025-08986-0.
5. Takahashi, Kazutoshi, and Shinya Yamanaka. “Induction of Pluripotent Stem Cells from Mouse Embryonic and Adult Fibroblast Cultures by Defined Factors.” Cell 126, no. 4 (2006): 663–76. https://doi.org/10.1016/j.cell.2006.07.024.

Keywords

Cell culture; pluripotent stem cells; embryonic stem cells; ESC; iPSC; automation; species agnostic cell culture; universal cell culture

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