Encapsulin Based Self-Assembling Iron-Containing Protein Nanoparticles for Stem Cells MRI Visualization

Abstract

Over the past decade, cell therapy has found many applications in the treatment of different diseases. Some of the cells already used in clinical practice include stem cells and CAR-T cells. Compared with traditional drugs, living cells are much more complicated systems that must be strictly controlled to avoid undesirable migration, differentiation, or proliferation. One of the approaches used to prevent such side effects involves monitoring cell distribution in the human body by any noninvasive technique, such as magnetic resonance imaging (MRI). Long-term tracking of stem cells with artificial magnetic labels, such as magnetic nanoparticles, is quite problematic because such labels can affect the metabolic process and cell viability. Additionally, the concentration of exogenous labels will decrease during cell division, leading to a corresponding decrease in signal intensity. In the current work, we present a new type of genetically encoded label based on encapsulin from Myxococcus xanthus bacteria, stably expressed in human mesenchymal stem cells (MSCs) and coexpressed with ferroxidase as a cargo protein for nanoparticles’ synthesis inside encapsulin shells. mZip14 protein was expressed for the enhancement of iron transport into the cell. Together, these three proteins led to the synthesis of iron-containing nanoparticles in mesenchymal stem cells—without affecting cell viability—and increased contrast properties of MSCs in MRI.

New Approach to Non-Invasive Tumor Model Monitoring via Self-Assemble Iron Containing Protein Nanocompartments

“…We hope that the encapsulin-based label will allow us to study the growth and metastasis of 4T1-Qt tumors in vivo via MRI. Our previous study [ 13 ] has shown that the stable heterologous expression of Myxococcus xanthus encapsulin genes can be achieved in human mesenchymal stem cells (MSCs). The presence of transgenic sequences in cells did not reduce the rate of cell proliferation, and the T2 relaxation time of MSCs containing nanoparticles in encapsulins was significantly lower than the T2 relaxation time of intact MSCs.…”

“…Western blot analysis was carried out as described earlier [ 13 ]. Briefly, 4T1-Qt cells were lysed using RIPA buffer, and the resulting lysate was precipitated (15 min, 14,000× g ).…”

Engineered Protein Nanocages for Concurrent RNA and Protein Packaging In Vivo

“…Engineered encapsulins have shown potential as nanoreactors, , drug delivery systems, imaging agents, − and immunotherapies. ,, Straightforward genetic and chemical shell modification allows small-molecule conjugation, peptide loop insertion, pore modification, ,, and fusion of protein domains to the N- and C-terminus of the encapsulin protomer. ,,, Recently, engineered encapsulins capable of triggered reversible disassembly, enabling in vitro cargo loading and stimulus-responsive cargo release, have also been reported …”

Engineered protein nanocages for concurrent RNA and protein packagingin vivo

“…Engineered encapsulins have shown potential as nanoreactors, [50,51] drug delivery systems, [52] imaging agents, [53–55] and immunotherapies. [30,56,57] Straightforward genetic and chemical shell modification allows small molecule conjugation, [33] peptide loop insertion, [58] pore modification, [50,59,60] and fusion of protein domains to the N- and C-terminus of the encapsulin protomer.…”

“…[46,47] This feature -a dedicated and modular protein loading mechanism -has been widely utilized to package non-native cargo proteins into the encapsulin shell via simple genetic fusion of TPs to proteins of interest. [48][49][50] Engineered encapsulins have shown potential as nanoreactors, [50,51] drug delivery systems, [52] imaging agents, [53][54][55] and immunotherapies. [30,56,57] Straightforward genetic and chemical shell modification allows small molecule conjugation, [33] peptide loop insertion, [58] pore modification, [50,59,60] and fusion of protein domains to the N-and C-terminus of the encapsulin protomer.…”