Abstract
BackgroundMesenchymal/stromal stem cells (MSCs) are favorably regarded in anti-cancer cytotherapies for their spontaneous chemotaxis toward inflammatory and tumor environments associated with an intrinsic cytotoxicity against tumor cells. Placenta-derived or TRAIL-engineered adipose MSCs have been shown to exert anti-tumor activity in both in-vitro and in-vivo models of multiple myeloma (MM) while TRAIL-transduced umbilical cord (UC)-MSCs appear efficient inducers of apoptosis in a few solid tumors. However, apoptosis is not selective for cancer cells since specific TRAIL receptors are also expressed by a number of normal cells. To overcome this drawback, we propose to transduce UC-MSCs with a bicistronic vector including the TRAIL sequence under the control of IL-6 promoter (pIL6) whose transcriptional activation is promoted by the MM milieu.MethodsUC-MSCs were transduced with a bicistronic retroviral vector (pMIGR1) encoding for green fluorescent protein (GFP) and modified to include the pIL6 sequence upstream of the full-length human TRAIL cDNA. TRAIL expression after stimulation with U-266 cell conditioned medium, or IL-1α/IL-1β, was evaluated by flow cytometry, confocal microscopy, real-time PCR, western blot analysis, and ELISA. Apoptosis in MM cells was assayed by Annexin V staining and by caspase-8 activation. The cytotoxic effect of pIL6-TRAIL+-GFP+-UC-MSCs on MM growth was evaluated in SCID mice by bioluminescence and ex vivo by caspase-3 activation and X-ray imaging. Statistical analyses were performed by Student’s t test, ANOVA, and logrank test for survival curves.ResultspIL6-TRAIL+-GFP+-UC-MSCs significantly expressed TRAIL after stimulation by either conditioned medium or by IL-1α/IL-1β, and induced apoptosis in U-266 cells. Moreover, when systemically injected in SCID mice intratibially xenografted with U-266, those cells underwent within MM tibia lesions and significantly reduced the tumor burden by specific induction of apoptosis in MM cells as revealed by caspase-3 activation.ConclusionsOur tumor microenvironment-sensitive model of anti-MM cytotherapy is regulated by the axis pIL6/IL-1α/IL-1β and appears suitable for further preclinical investigation not only in myeloma bone disease in which UC-MSCs would even participate to bone healing as described, but also in other osteotropic tumors whose milieu is enriched of cytokines triggering the pIL6.
Highlights
Mesenchymal/stromal stem cells (MSCs) are favorably regarded in anti-cancer cytotherapies for their spontaneous chemotaxis toward inflammatory and tumor environments associated with an intrinsic cytotoxicity against tumor cells
Results promoter of IL-6 (pIL6)-tumor necrosis factor related apoptosis inducing ligand (TRAIL)+-green fluorescent protein (GFP)+-pMIGR1 vector construction and umbilical cord (UC)-MSC transduction pMIGR1 bicistronic retroviral vector containing poliovirus internal ribosome entry site (PIRES) and GFP sequences was modified to express full-length TRAIL under the control of pIL6 (Fig. 1a). pIL6-TRAIL construct was obtained by ligation of the relative polymerase chain reaction (PCR) products in the XhoI restriction site and subsequently cloned between the BglII and EcoRI sites on pMIGR1
Modified pMIGR1 construct and viral packaging plasmids were transfected in HEK293T competent cells and the culture supernatant containing viral particles bearing pIL6-TRAIL+-GFP+-pMIGR1 vector was used to infect UC-MSCs (Fig. 1b). pMIGR1 wildtype construct was used as control empty vector
Summary
Mesenchymal/stromal stem cells (MSCs) are favorably regarded in anti-cancer cytotherapies for their spontaneous chemotaxis toward inflammatory and tumor environments associated with an intrinsic cytotoxicity against tumor cells. Consistent work by different groups of investigators showed that both BM-derived and AT-derived MSCs are capable of inducing the tumor shrinkage in xenografted human glioma [11], gastric [12] or pancreatic [13] cancers, as well as in melanoma [14], and that this native anti-tumor cell growth activity may be definitely enhanced with MSCs transduced to express the tumor necrosis factor related apoptosis inducing ligand (TRAIL), namely a proapoptogen molecule linking the death receptors (DR) 4 and 5 on cancer cells [15] In this context, fetal MSCs from umbilical cord (UC) engineered to produce a membrane-TRAIL protein have been reported to efficiently restrain the intracranial glioblastoma growth in mice as an effect of their innate chemotactic tendency to migrate toward the tumor microenvironment while exposing the death ligand to the tumor cells [16]. In addition to the anti-myeloma activity, MSCs were found capable of repairing the bone loss within the bone lytic lesions in relation to their bone-regenerating constitutive capability [20]
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