Convective Foam Drying for Room-Temperature Preservation of CAR-T Cell Therapies

A Proposal for the ARPA-H BioStabilization Systems (BoSS) Program

Technical Area 2: Scalable Cell Processing Systems

Notice ID: ARPA-H-SOL-26-136 | February 2026

Executive Summary

This proposal presents a novel approach to room-temperature preservation of chimeric antigen receptor T-cell (CAR-T) therapies through convective foam drying. Current CAR-T products require cryogenic storage below -150°C, creating significant logistical barriers, substantial costs, and limiting patient access to these transformative cancer treatments. Our approach leverages the unique physics of foam structures to achieve controlled desiccation of living cells, addressing the fundamental engineering challenge that has prevented room-temperature cell preservation at therapeutic scale.

By creating thin liquid films within a foam matrix, we enable uniform water removal from millions of cells simultaneously while maintaining the precise drying kinetics required for cellular survival. This proposal details the scientific rationale, technical approach, risk mitigation strategies, and development roadmap for bringing foam-dried CAR-T cells from concept to clinical reality.

1. The Controlled Drying Challenge

1.1 Why Desiccation is Fundamentally Difficult

Room-temperature preservation through desiccation requires removing approximately 95% of cellular water while maintaining the structural and functional integrity of living cells. This presents a paradox: water is essential for life, yet its presence enables the chemical reactions and physical processes that cause degradation during storage. The goal is to transition cells into a stable, glass-like (vitrified) state where molecular motion is arrested and decay processes are halted.

Unlike cryopreservation, which simply slows reactions by reducing temperature, desiccation fundamentally changes the physical state of the cell. This transition must be carefully controlled to prevent irreversible damage.

1.2 The Impact of Drying Rate

The rate at which water is removed from cells determines whether they survive the desiccation process. Both extremes are lethal:

• Too Fast: Rapid water loss causes osmotic shock, membrane rupture, and protein aggregation. Protective molecules cannot redistribute quickly enough to stabilize cellular structures before damage occurs.

• Too Slow: Extended drying times at ambient conditions allow ongoing metabolism, oxidative damage, and protein degradation. Cells may trigger apoptotic pathways before reaching the protective vitrified state.

The optimal drying profile exists in a narrow window: fast enough to reach vitrification before cumulative damage becomes lethal, yet slow enough to allow protective molecules to stabilize cellular components. This window varies by cell type, making process optimization essential.

1.3 Critical Thresholds in Cellular Desiccation

1.4 The Geometry Problem

Drying kinetics are governed by diffusion physics. Water must travel from the cell interior, across the membrane, through the extracellular medium, and finally to the evaporation surface. Diffusion time scales with the square of distance, creating an enormous challenge for bulk samples.

A therapeutic CAR-T dose contains 10⁸ to 10¹⁰ cells in approximately 1-10 mL volume. Drying this volume uniformly using conventional approaches would require either impractically long times (risking metabolic damage) or aggressive conditions (risking mechanical and thermal damage). The fundamental insight of our approach is that foam structures solve this geometry problem.

2. Foam Drying: A Physics-Based Solution

2.1 The Foam Advantage

Foam drying elegantly addresses the geometry problem by restructuring bulk liquid into a network of thin films. When a cell suspension is converted to foam, cells distribute throughout thin liquid lamellae (typically 10-100 μm thick) separated by gas bubbles. This transformation achieves thin-film drying physics at bulk volumes.

This 400-fold increase in surface area-to-volume ratio and corresponding reduction in diffusion distance fundamentally changes the drying dynamics, enabling rapid, uniform water removal that would be impossible with bulk liquid samples.

2.2 The Shear Stress Challenge

Shear stress represents the primary risk to cell viability during foam formation. When gas bubbles are introduced into a cell suspension, fluid mechanical forces act on cells in several ways:

• Bubble Formation Shear: As bubbles nucleate and grow, surrounding fluid accelerates, generating velocity gradients that stretch and deform cells.

• Interface Adsorption: Cells may be drawn to the gas-liquid interface where surface tension forces can cause membrane deformation or rupture.

• Bubble Coalescence: When bubbles merge, the rapid film rupture and fluid rearrangement generates localized high-shear events.

• Drainage Flow: Liquid draining from foam films creates shear within the thin lamellae where cells reside.

T cells, with their relatively soft cytoskeleton compared to other cell types, are particularly sensitive to mechanical stress. Studies have shown that shear rates above 1000-5000 s⁻¹ can cause significant viability loss in lymphocytes. Our foam generation approach must therefore maintain shear rates well below these thresholds.

2.3 Solutions for Shear Stress Mitigation

We propose a multi-pronged approach to minimize shear-induced cell damage:

2.3.1 Microfluidic Foam Generation

Rather than using conventional mechanical agitation (whisking, homogenization), we will employ microfluidic devices that generate bubbles through controlled flow focusing. In these systems, gas and liquid streams meet at precisely engineered junctions where bubbles form one at a time under laminar flow conditions. This approach offers several advantages: bubble size is highly uniform (reducing Laplace pressure variations), shear rates are predictable and controllable, and the gentle nature of the process is inherently compatible with sensitive biological materials. Our preliminary calculations indicate achievable shear rates of 100-500 s⁻¹, well within the safe range for T cells.

2.3.2 Protective Surfactant Selection

The choice of surfactant is critical for both foam stability and cell protection. We will use human serum albumin (HSA) as the primary surface-active agent. HSA naturally adsorbs to air-water interfaces, creating a protective protein layer that shields cells from direct contact with the gas phase. As an endogenous human protein already approved for clinical use, HSA introduces no additional biocompatibility concerns. Furthermore, albumin has documented protective effects during desiccation, serving both as a surfactant and a stabilizing excipient.

2.3.3 Viscosity Optimization

Increasing medium viscosity through the addition of trehalose (the primary cryoprotectant) serves dual purposes: it provides the molecular machinery for vitrification while simultaneously reducing shear stress during foam formation. Higher viscosity dampens velocity fluctuations, reduces bubble oscillation, and slows drainage. We will optimize trehalose concentration (targeting 15-25% w/v) to balance viscosity enhancement against osmotic stress on cells prior to foam generation.

2.3.4 Bubble Size Optimization

Larger bubbles generate lower Laplace pressures (ΔP = 2γ/r) and create thicker, more stable films between them. We will target bubble diameters of 200-500 μm, which provide sufficient surface area enhancement while minimizing the mechanical stresses associated with very small bubbles. This size range also allows cells to distribute freely within the lamellae without geometric constraints.

3. Convective Foam Drying: Technical Approach

3.1 Method Overview

Convective drying uses controlled airflow to remove water from materials. Applied to foam, this method offers precise control over drying kinetics through manipulation of temperature, humidity, and airflow rate. Unlike vacuum drying (which risks boiling) or spray drying (which involves excessive heat and shear), convective foam drying operates under gentle, physiologically compatible conditions.

The process involves generating foam from a cell suspension containing protectants, spreading the foam into thin layers, and passing conditioned air over the foam surface. Water evaporates from the film surfaces and is carried away by the airflow. As drying proceeds, trehalose and other protective molecules concentrate, eventually forming a glassy matrix that entraps cells in a preserved state.

3.2 Process Design

Our convective foam drying process consists of four integrated stages:

Stage 1: Pre-Loading with Protectants (2-4 hours)

CAR-T cells are incubated with trehalose (200-400 mM) and tardigrade-derived intrinsically disordered proteins at 37°C. This allows intracellular accumulation of protectants that will form the vitrification matrix. Concurrent addition of HSA (2-5% w/v) provides interfacial protection for subsequent foam formation.

Stage 2: Microfluidic Foam Generation (30-60 minutes)

The protectant-loaded cell suspension is processed through parallelized microfluidic foam generators. Nitrogen gas (to prevent oxidation) is introduced at controlled flow rates to create uniform foam with target bubble diameter of 200-500 μm. The foam is collected and spread into layers approximately 5-10 mm thick in shallow trays within a closed, sterile drying chamber.

Stage 3: Humidity-Ramped Convective Drying (4-8 hours)

Conditioned air is passed over the foam layers following a programmed humidity profile:

• Phase A (0-2 hours): High humidity (70-80% RH), moderate airflow. Slow initial drying allows protectant redistribution and cellular adaptation to osmotic stress.

• Phase B (2-5 hours): Decreasing humidity ramp (70% → 20% RH). Progressive water removal concentrates protectants toward vitrification. This is the critical transition phase.

• Phase C (5-8 hours): Low humidity (10-20% RH), extended hold. Final water removal achieves target moisture content below 5%. Vitrification is complete.

Stage 4: Conditioning and Packaging (1-2 hours)

Dried foam is transferred to storage containers under nitrogen atmosphere with desiccant. Secondary drying under mild vacuum removes residual bound water. Final moisture content is verified by near-infrared spectroscopy. Sealed containers are suitable for room-temperature storage.

3.3 Addressing Key Challenges

Challenge 1: Foam Stability During Drying

Problem: Foams are thermodynamically unstable and tend to collapse through drainage, coalescence, and coarsening. If foam structure is lost before drying completes, the thin-film geometry advantage is negated.

Solution: We exploit the drying process itself as a stabilization mechanism. As water evaporates, the remaining liquid phase becomes increasingly viscous due to concentrating trehalose and proteins. This viscosity increase dramatically slows drainage and arrests foam evolution well before complete drying. Additionally, HSA forms robust interfacial films that resist coalescence. Our humidity ramping profile is specifically designed to reach sufficient viscosity before foam collapse occurs. Preliminary modeling suggests foam stability windows of 6-10 hours are achievable with optimized formulations, exceeding our required drying time.

Challenge 2: Non-Uniform Drying

Problem: Even in foam, the exposed surface dries faster than the interior, potentially creating gradients in protectant concentration, residual moisture, and cell survival.

Solution: Multiple strategies address drying uniformity. First, foam layer thickness is limited to <10 mm to minimize through-thickness gradients. Second, the initial high-humidity phase slows surface drying, allowing the interior to equilibrate. Third, the open-cell structure of foam permits vapor transport through the interior, unlike a solid sample where surface “skinning” traps moisture. Fourth, bidirectional airflow (alternating direction) can be implemented to reduce edge effects. Fifth, real-time monitoring via NIR spectroscopy enables feedback control to adjust drying conditions dynamically.

Challenge 3: Oxidative Damage

Problem: Room-temperature storage exposes cells to oxidative stress over months or years, damaging lipids, proteins, and DNA even in the vitrified state.

Solution: Oxidation is addressed at multiple levels. Foam is generated using nitrogen rather than air, minimizing dissolved oxygen. Antioxidants (α-tocopherol, ascorbic acid) are included in the protectant formulation. Final packaging under nitrogen or argon atmosphere with oxygen scavengers ensures long-term protection. The glassy matrix itself limits oxygen diffusion. Light-protective packaging prevents photo-oxidation.

Challenge 4: Functional Recovery After Rehydration

Problem: Cells that survive drying may suffer damage that only manifests as functional impairment — reduced proliferation, cytotoxicity, or persistence — despite appearing viable.

Solution: The porous structure of dried foam enables rapid, uniform rehydration that minimizes osmotic shock. We will develop controlled rehydration protocols that gradually restore water content over 15-30 minutes. Comprehensive functional assays beyond simple viability (CAR expression, target killing, cytokine production, proliferation) will be implemented throughout development. Correlation of process parameters with functional outcomes will guide optimization toward therapeutic equivalence with fresh or cryopreserved CAR-T products.

Challenge 5: Scalability and GMP Compatibility

Problem: Research-scale processes often fail to translate to manufacturing environments that require closed systems, validated processes, and regulatory compliance.

Solution: GMP compatibility is designed into the process from inception rather than retrofitted. Microfluidic foam generators are inherently closed systems compatible with sterile processing. Convective drying chambers can be designed as sealed units with HEPA-filtered conditioned air. All materials (HSA, trehalose) are pharmaceutical-grade with established regulatory acceptance. Process analytical technology (PAT) for real-time moisture and temperature monitoring enables validated, reproducible manufacturing. Our development plan includes specific milestones for process validation and scale-up to clinical manufacturing volumes.

4. Development Roadmap

5. Expected Impact

Successful development of room-temperature CAR-T preservation would transform the cell therapy landscape:

• Expanded Access: Any hospital with standard pharmacy capabilities could store and administer CAR-T therapies, extending life-saving treatments to patients in community settings and underserved regions.

• Reduced Costs: Elimination of cold chain infrastructure and specialized handling could reduce per-dose logistics costs by 80-90%, potentially saving billions annually across the cell therapy industry.

• Improved Supply Chain Resilience: Room-temperature products are immune to power outages, shipping delays, and freezer failures that currently result in costly product losses.

• Pandemic Preparedness: Stable stockpiles of cell-based therapeutics could be pre-positioned for rapid deployment in public health emergencies.

• Global Health Equity: Developing nations without cryogenic infrastructure could access advanced cell therapies for the first time.

6. Conclusion

Convective foam drying represents a scientifically grounded, technically innovative approach to the grand challenge of room-temperature cell preservation. By transforming bulk liquid into thin films, we overcome the fundamental physics barrier that has limited previous approaches. By carefully engineering each process step — from surfactant selection to humidity ramping — we address the specific biological sensitivities of therapeutic T cells.

This proposal aligns directly with BoSS Technical Area 2 objectives: developing scalable cell processing systems that deploy biostabilization strategies. Our approach integrates naturally with the protective chemistry being developed under Technical Area 1, and our team is prepared to collaborate closely with complementary efforts across the program.

The path from laboratory demonstration to clinical reality is challenging but achievable. With ARPA-H support, we are confident that foam-dried CAR-T cells can become a transformative technology — making these remarkable therapies as easy to ship and store as aspirin, and finally delivering on the promise of cell therapy for all patients who need it.

Prompts Below: can you read this https://arpa-h.gov/explore-funding/programs/boss
are there papers supporting this?
why is this a challenge? what makes CAR-T complex and what makes this high-risk
expand on controlled drying process, (this intreguied me the most and was easier for me to understand becasue of my undergradate in pharmacueticals and cosmetics.)
Explore foam drying techniquies (this intreguied me the most and was easier for me to understand foam formulations are used in topical pharmaceuticals and cosmetics)
write up proposal, explain that the approach focuses on exploring controlled drying process, explain the impacts of drying process and how Desiccation required removing 95% of cellular water. Then explain foam drying, explaining the significance of sheer stress and address solutions for the challenges, then explain using convective foam drying and address solutions from the challenged raised