A neovascularized organoid derived from retrovirally engineered bone marrow stroma leads to prolonged in vivo systemic delivery of erythropoietin in nonmyeloablated, immunocompetent mice

Abstract

Marrow stromal cells (MSCs) are postnatal progenitor cells that can be easily cultured ex vivo to large amounts. This feature is attractive for cell therapy applications where genetically engineered MSCs could serve as an autologous cellular vehicle for the delivery of therapeutic proteins. The usefulness of MSCs in transgenic cell therapy will rely upon their potential to engraft in nonmyeloablated, immunocompetent recipients. Further, the ability to deliver MSCs subcutaneously -as opposed to intravenous or intraperitoneal infusions -would enhance safety by providing an easily accessible, and retrievable, artificial subcutaneous implant in a clinical setting. To test this hypothesis, MSCs were retrovirally engineered to secrete mouse erythropoietin (Epo) and their effect was ascertained in nonmyeloablated syngeneic mice. Epo-secreting MSCs when administered as 'free' cells by subcutaneous or intraperitoneal injection, at the same cell dose, led to a significant -yet temporaryhematocrit increase to over 70% for 55713 days. In contrast, in mice implanted subcutaneously with Matrigeltembedded MSCs, the hematocrit persisted at levels 480% for over 110 days in four of six mice (Po0.05 logrank). Moreover, Epo-secreting MSCs mixed in Matrigel elicited and directly participated in blood vessel formation de novo reflecting their mesenchymal plasticity. MSCs embedded in human-compatible bovine collagen matrix also led to a hematocrit 470% for 7578.9 days. In conclusion, matrixembedded MSCs will spontaneously form a neovascularized organoid that supports the release of a soluble plasma protein directly into the bloodstream for a sustained pharmacological effect in nonmyeloablated recipients.

Nonviral gene delivery of erythropoietin by mesenchymal stromal cells

“…Plasma EPO levels were also found to be elevated for up to 7 weeks. 16 In comparison, subcutaneous transplantation of 10 7 EPO-engineered MSCs showed hematocrit levels that were increased to 70-80% and returned to normal after 40-50 days. 20,21 We i.p.…”

“…42 In conclusion, we find that nucleofection can be used to generate EPO-engineered MSCs, which secrete biologically active EPO in sufficient amounts to stimulate hematopoiesis in vivo. Future strategies aimed at the prevention of graft loss (for example, encapsulation of MSCs, exploitation of tissue tropism) 11,16,20 and inhibition of autoimmune responses are likely to extend the duration of action of this nonviral cell-based gene therapy where required.…”

“…11 A few recent studies have successfully used viral transfer of the epo gene into non-human primate, rodent and human MSCs for cell-based gene therapy. [14][15][16][17] Here, we wanted to establish nonviral techniques for MSC-based delivery of EPO. This study is the first to examine nucleofection of MSCs for epo gene delivery in vivo.…”

Non-hematopoietic stem cells as factories for in vivo therapeutic protein production

“…The s.c. delivery of MSCloaded scaffolds would provide an easily accessible implant that could be retrieved once the therapeutic effect is fulfilled or in the event of an unexpected adverse reaction. A seminal work by Eliopoulos et al 50 reported that erythropoietin (Epo)-secreting MSCs (MSC Epo ), when administered as 'free' cells by s.c or i.p. injection, led to a temporary hematocrit increase.…”

Therapeutic potential of bone marrow-derived mesenchymal stem cells producing pigment epithelium-derived factor in lung carcinoma

“…This differentiation plasticity makes MSCs the ultimate candidates for future utilization Therapeutic potential of bone marrow-derived mesenchymal stem cells producing pigment epithelium-derived factor in lung carcinoma in cell therapy and tissue regeneration (16,17). Furthermore, MSCs expressing transgenes maintained long-term expression (up to 6 months) in vivo because of their hypoimmunogenic properties and production of immunosuppressive molecules (18,19). The tumor microenvironment is known to provide a preferential niche for MSCs homing and survival.…”