Hematopoietic stem cell destruction is the hallmark of BMF (1). Symptoms of BMF, such as anemia and infections, can be reduced if the BM niche is repopulated with new HSCs. However, most patients lack compatible donors for HSCT, but cells derived from cord blood tend to be a more universal match (29, 30). Using HSCs derived from cord blood for transplant can help repopulate the BM, reduce the risk of rejection, and decrease disease relapse (11). However, HSCT therapy using cord blood cells is restricted by limited availability of donors (15), as well as by the absolute numbers of stem cells present in each collected sample. Polymer-based expansion methods provide innovative means to overcome these limitations by expanding stem cell populations from cord blood, or other hematopoietic stem cell sources, to potentially increase HSCT access.

3D polymer-based cell matrices have been gaining in popularity for stem cell culture and expansion (31–35). Increasing attention focuses on the numerous tunable variables that can be incorporated in a polymer matrix, such as chemical identities, mechanical properties, and bulk matrix architecture ( Figure 1 ). The diverse structures allow for discrete environmental manipulation and can tolerate cytokine and growth factor loading, which in turn affects cell growth, function, and differentiation (34).

A recent study explored the expansion of CD34+ cells isolated from cord blood in a 3D polymer matrix (15). Here the authors use a zwitterionic hydrogel composed of poly(carboxybetaine) and crosslinked using click chemistry and degradable crosslinkers. Their designed zwitterionic gel (ZG), which incorporates both negative and positive charges in a single monomer unit, significantly outperformed gels made from poly(ethylene glycol) (PEG), a common non-charged polymer used for many biological applications. In additional grafting studies, cells cultured in the ZGs showed similar engraftment levels than the non-cultured controls, but with 100-fold fewer cells. The authors ultimately show clinically meaningful expansion both of CD34+ cells isolated from cord blood and bone marrow derived HSPCs in their 3D zwitterionic hydrogel.

Modulating matrix mechanical properties, such as stiffness and elasticity, also plays a significant role in stem cell differentiation and expansion (36). One study demonstrated how polyacrylamide gels with differing Young’s moduli (E, stiffness) affected mesenchymal stem cell differentiation (MSC). For example, softer gels (E = 0.1-1 kPa) promoted neuronal type expression, while stiffer gels (E = 25-40 kPa) promoted osteogenic differentiation (37). Gels can also be synthesized to have dynamic moduli, where the stiffness changes when exposed to certain stimuli, such as light, pH, or temperature. These gels allow for cellular adaptation, being able to differentiate into various cell types by simply changing the matrix modulus (38).

Tuning the bulk matrix architecture offers another variable for control. The woodpile structure ( Figure 1B , architecture) is commonly employed in 3D matrices as it can be easily manipulated to include diverse pore sizes and surface areas, both of which are important for cell growth and migration (25). A recent study investigated how gap sizes in a woodpile 3D matrix significantly affected bone marrow derived MSC (BM-MSC) migration (39). Matrices with larger gap sizes (100 μm) promoted the highest BM-MSC migration and increased the number of viable cells. Additionally, 3D cell matrices with submillimeter pore sizes were found to enhance CD34+ cell proliferation more than the nonporous matrices (33).

Polymers can also be used as intracellular delivery vehicles for drugs and biomacromolecules to aid in cellular manipulation ( Figure 2A ) (28). They are typically employed to help therapeutic cargo traverse the cell membrane, either by direct conjugation of the vehicle to the cargo, or by non-covalent complexation (40–42). Polymer-based delivery vehicles can also be used to protect the cargo from premature consumption or degradation. This aids in cargo pharmacokinetics, typically by increasing the half-life of the cargo or by increasing the toxicity threshold (27, 43).

Aplastic anemia (AA) is a form of bone marrow failure that is typically inherited, but can also be caused by drugs, such as hepatitis-C (HCV) treatments, and by diseases, such as the human immunodeficiency virus (HIV) virus. It is characterized by the bone marrow being incapable of producing enough blood cells for normal bodily functions. Filgrastim is a granulocyte colony-stimulating factor (G-CSF) used for hematopoietic cell growth and is known to treat neutropenia, aplastic anemia, and aids in myelosuppression after bone marrow transplantation (44–46). However, G-CSF is administered by daily injections, which is painful for patients and increases the risk of infection at the injection site (44). One study has shown the oral delivery of G-CSF mediated by a diethylene triamine penta acetic acid conjugated chitosan and poly(γ-glutamic acid) (γPGA-DTPA) nanoparticle ( Figure 2B , chitosan+γPGA-DTPA) (44, 47). They observed increased availability of G-CSF when encapsulated in the nanoparticle and delayed the maximum concentration release by 6 h, indicative of a sustained release rather than a burst release. Additionally, neutropenia rat models were treated with a one-time oral dose of the nanoparticle encapsulated G-CSF (NP-G-CSF) or free-form G-CSF administered by daily injections. Both treatments increased the absolute neutrophil count in the rats, but the NP-G-CSF was only administered once, demonstrating the simplicity and effectiveness of the nanoparticle platform to deliver G-CSF.

Alternative ISTs must be utilized to ensure patients have safe and effective immune suppression without the occurrence of drug resistance and dose related toxicity (12). One strategy to achieve immunosuppression is to enhance regulatory T cell (Treg) function. Tregs are T cells that suppress effector cell growth and division, thereby limiting the immune response (48). FOXP3 is the hallmark protein for Treg cell function, whereby elevated levels correlate with enhanced suppressive function by Tregs (48–50).

It has been shown that protein kinase C theta (PKCθ) enhances effector T cell function, while limiting Treg differentiation. Many studies have sought to inhibit PKCθ function by using small molecule inhibitors or by using gene editing technology, such as siRNA. Inhibiting PKCθ yielded more suppressive Tregs and even restored impaired Treg function (51). Alternatively, polymer materials can be used as delivery vehicles for inhibitory biomacromolecules, such as antibodies, providing a simple pathway to achieve similar results. A recent study highlights a designed polymer vehicle to intracellularly deliver an antibody against PKCθ into naïve CD4+ T cells and inhibit its function (16). This polymer mimics the structure of the designed protein transduction domain, Pep-1 (Chariot^Ⓡ), by using a block copolymer architecture with both hydrophobic and cationic blocks ( Figure 2B , MePh₁₀-b-dG₅) (52). This polymer has also been shown to significantly outperform Pep-1 (53), an analog of the naturally occurring cell penetrating peptide HIV1-TAT (54), and the commercial delivery agent AbDeliverIN™ (55). The MePh₁₀-b-dG₅ delivery vehicle successfully delivered an anti-PKCθ antibody, and yielded Tregs with elevated FOXP3 expression and suppressive function (16). The exact mechanism by which these polymers carry cargo across the cell membrane to enhance the therapeutic agent delivery continues to be an area of active investigation.

Gene therapy is another avenue for protein manipulation, typically achieved by viral deliveries, such as lentivirus and extracellular vesicles, or nonviral deliveries, such as microinjection, electroporation, and polymeric vehicles (56). Viral delivery vectors face significant limitations as they pose mutagenetic risks and suffer from poor production yields (57). Microinjection and electroporation are popular nonviral delivery methods for genetic materials, but are harsh on cells, difficult to apply to large cell populations, and not feasible for in vivo work, making these techniques inadequate for clinical applications (56, 58, 59). Polymers such as poly(ethyleneimine), poly(amidoamine), and poly(amino acids), are promising as nonviral alternatives and have been successful at intracellularly delivering DNA, mRNA, and other gene editing technology (26, 60–62).

The CRISPR/Cas9 system has been widely adopted as an efficient and effective gene editing technology (63, 64). Polymeric delivery vehicles can be tailor-made for the desired cargo making them ideal vehicles for intracellular CRISPR/Cas9 delivery. Recently, poly(β-amino esters) (PBAEs) were synthesized to encapsulate the CRISPR/Cas9 system and transfect human CD34+ and CD14+ cells ( Figure 2B , PBAE) (17). The transfection efficiency of the CRISPR/Cas9/PBAE nanoparticles (NPs) were >90% and maintained >86% cell viability. The NPs achieved 85% gene editing efficiency, which was measured by CD33 knockout. The edited cells were then injected into mice and the human cell engraftment was measured. Overall, there was no difference in human cell engraftment from the untreated control, both in the peripheral blood and the bone marrow of the mice (17).