TABLE 2.
Studies using minicircles for the development of novel therapies.
| Sequence encoded | Vector length | Transfection method | Outcomes | Ref. |
|---|---|---|---|---|
| 3rd generation anti-CD44 CAR |
NR | Electroporation | Minicircle-generated anti-CD44-CAR T cells expressed CAR molecules with strong hepatocellular carcinoma tumor suppression activity in vitro and overcame tumor microenvironment barriers in mice. | [21] |
| 3rd generation anti-PSCA CAR |
4,575 | Electroporation | Unlike normal T cells, minicircle-generated PSCA CAR T cells had high cytokine secretion, strong antitumor effects, infiltrated tumor tissue, and persisted up to 28 days in mice. | [22] |
| 3rd generation NKG2D CAR |
NR | Electroporation | Minicircle-generated NKG2D CAR T cells demonstrated efficient and specific cytotoxic activity against human colorectal cancer in vitro and in vivo. | [23] |
| TIPE2 | NR | Hydrodynamic tail vein injection | Minicircle-mediated TIPE2 expression inhibited breast cancer cell proliferation and promoted in vivo anti-tumor immune responses by boosting CD8+ T cell and NK cell function. | [25] |
| Anti-EpCAM/CD3 | NR | Calcium phosphate nanoneedle-mediated cell perforation | Minicircle-mediated expression of an anti-EpCAM/CD3 bispecific antibody showed significant anti-cancer effects in vivo and increased survival of a xenograft mouse model of human ascites ovarian cancer by simultaneously conjugating immune cells and cancer cells. | [26] |
| miR-31 5p | NR | Cationic lipid | Minicircle transfection resulted in miRNA expression levels comparable to that of a lentiviral vector system used to generate cell lines stably expressing miR-31; This study validated WDR5 inhibition as a novel therapeutic option for nasopharyngeal carcinoma. | [27] |
| KLF4 | NR | IV injection | Minicircle-mediated KLF4 overexpression validated the role of KLF4 in the development and pathogenesis of inflammatory arthritis because it led to severe autoimmune arthritis in mice. KLF4 inhibition regulates the apoptosis of FLS and their expression of matrix metalloproteinases and proinflammatory cytokines. | [30] |
| Anti-alpha-synuclein shRNA | NR | RVG exosomes | Delivery of an anti-alpha-synuclein shRNA minicircle provided stable and prolonged gene downregulation and decreased aggregation of alpha-synuclein in the brain of a mouse model of Parkinson’s disease, improving clinical symptoms. | [31] |
| CBS | 2,336 | Hydrodynamic tail vein injection | Delivery of naked minicircle encoding CBS partially corrected metabolic and phenotypic defects in a mouse model of CBS deficiency. | [32] |
| sTNFR2-Fc | 3,000 | Electroporation | Minicircle-transfected MSCs produced the biologic TNFα inhibitor etanercept in vitro and had anti-inflammatory effects when injected into a collagen-induced rheumatoid arthritis mouse model. | [33] |
| IFNα IFNλ.3 |
1,656 1,677 |
Cationic lipid | Minicircles encoding liver-specific cytokine, IFNλ3, exhibited strong anti-HBV activity in transfected HBV-infected hepatocytes in vitro and suppressed viral antigen expression and viral DNA replication. | [34] |
| Bcl-2/GFP | NR | Electroporation/magnetofection | Minicircles encoding Bcl-2 attached to magnetic nanoparticles for in vivo transfection stimulated bone regeneration through the transient expression of Bcl-2, which prevented apoptosis of cell implants and promoted cell survival. | [36] |
| Sox9/Sox6/shANGPTL4 | NR | Cationic polymer | PEI minicircle particles encoding Sox9, Sox6, and shRNA against ANGPTL4 promoted chondrogenesis in vitro and suppressed osteoarthritis in mice. | [37] |
| GFP | 1,552 | Magnetofection | Neural stem cells engineered with minicircles in conjunction with magnetic nanoparticles were successfully grown and propagated on a novel neurosurgical-grade biomaterial scaffold with no adverse effects on key regenerative parameters. | [39] |
| BMP2/RFP TGFβ3/RFP |
7,300* 7,500* |
Cationic lipid | MSC-like, human iPSC-derived outgrowth cells transfected with two minicircles encoding TGFβ3 and BMP2, respectively, differentiated into the chondrogenic lineage and rescued osteochondral defects in rat models. | [58] |
| Sox2/Oct4/NanogLin28/GFP | NR | Cationic lipid | Transfection of Oct4, Sox2, Lin28, and Nanog-encoding minicircles to reprogram B16F10 murine melanoma cells resulted in incomplete reprogramming of cancer cells that did not form teratomas (an indicator of complete reprogramming). These cells, however, still displayed the characteristics of cancer stem cells and formed smaller, less aggressive tumors than the parental cell line. | [59] |
Italicized values are estimated lengths provided by the authors of those studies.
Bcl-2: B-cell lymphoma 2; BMP2: Bone morphogenic protein 2; CAR: Chimeric antigen receptor; CBS: Cystathionine β synthase; CD3: Cluster of differentiation 3; CD44: Cluster of differentiation 44; CD8: Cluster of differentiation 8; EpCAM: Epithelial cell adhesion molecule; FLS: Fibroblast-like synoviocytes; GFP: Green fluorescent protein; HBV: Hepatitis B virus; IFN: Interferon; IV: Intravenous; IPSCs: Induced pluripotent stem cells; KLF4: Kruppel-like factor 4; Lin28: Abnormal cell lineage 28; miRNA or miR: microRNA; MSCs: Mesenchymal stem cells; NK: Natural killer; NKG2D: Natural killer group 2 member D; NR: Not reported; Oct4: Octamer binding transcription factor 4; PEI: Polyethyleneimine; PSCA: Prostate stem cell antigen; Ref.: Reference; RFP: Red fluorescent protein; RVG: Rabies virus glycoprotein peptide; shRNA: Short hairpin RNA; shANGPLT4: Short hairpin angiopoietin-like protein 4; Sox: Sex-determining region Y-box transcription factor; sTNFR2-Fc: Soluble tumor necrosis factor receptor 2; TGFβ3: Transforming growth factor beta 3; TIPE2: Tumor necrosis factor alpha induced protein 8 family 2; TNF: Tumor necrosis factor; WDR5: WD repeat domain 5.