Hydrophilic acylated surface protein B (HASPB) is an immunogenic Leishmania-specific protein that antibodies are produced against it in the sera of Leishmania-infected individuals. Kinetoplastid membrane protein 11 (KMP11) is another Leishmania antigen and considered as the suitable candidate for vaccine development Leishmaniasis. It is a highly conserved surface protein expressed in both promastigotes and amastigotes. In this study, KMP11 and HASPB coding sequences were cloned into a pCDH-cGFP lentiviral vector as a fusion protein to be used as a DNA vaccine against L. major. The KMP11-HASPB fusion protein was successfully expressed as evidenced by RT-PCR and Western blot assays. The effect of the vaccine was determined by evaluating the level of IFN-γ, IL-10, IgG1, and IgG2a performed using ELISA as well as determining the parasite load after challenge with L. major in vaccinated mice. The results revealed that IFN-γ, IL-10, IgG1, and IgG2a significantly increased after vaccination using KMP11-HASPB-expressing lentiviruses in BALB/c mice. It is noteworthy that the level of IFN-γ and IgG2a was higher than that of IL-10 and IgG1, respectively, which indicates the activation Th1 cells, macrophages, and cellular immunity. Moreover, the parasite load in the spleen and lymph node of vaccinated mice after challenge was significantly lower than that of controls.
Background: Cancer-related anemia (CRA) negatively influences cancer patients’ survival, disease progression, treatment efficacy, and quality of life (QOL). Current treatments such as iron therapy, red cell transfusion, and erythropoietin-stimulating agents (ESAs) may cause severe adverse effects. Therefore, the development of long-lasting and curative therapies is urgently required. Objective: In this study, a cell and gene therapy strategy was developed for in-vivo delivery of EPO cDNA by way of the genetic engineering of human Wharton’s jelly mesenchymal stem cells (hWJMSCs) to produce and secrete human EPO protein for extended periods after transplantation into the mice model of CRA. Methods: To evaluate CRA’s treatment in cancer-free and cancerous conditions, first, a recombinant breast cancer cell line 4T1 which expressed herpes simplex virus type 1 thymidine kinase (HSV1-TK) by a lentiviral vector encoding HSV1- TK was developed and injected into mice. After three weeks, all mice developed metastatic breast cancer associated with acute anemia. Then, ganciclovir (GCV) was administered for ten days in half of the mice to clear cancer cells. Meanwhile, another lentiviral vector encoding EPO to transduce hWJMSCs was developed. Following implantation of rhWJMSCs-EPO in a second group of mice, peripheral blood samples were collected once a week for ten weeks from both groups. Results: Analysis of peripheral blood samples showed that plasma EPO, hemoglobin (Hb), and hematocrit (Hct) concentrations significantly increased and remained at therapeutic for >10 weeks in both treatment groups. Conclusions: Data indicated that rhWJMSCs-EPO increased the circulating level of EPO, Hb, and Hct in both mouse subject groups, and improved the anemia of cancer in both cancer-free and cancerous mice.
Cancer-related anemia (CRA) negatively influences cancer patients’ survival, disease progression, treatment efficacy, and quality of life (QOL). Current treatments such as iron therapy, red cell transfusion, and erythropoietin-stimulating agents (ESAs) may cause severe adverse effects including hemolytic transfusion reaction and the possibility of host immunity against rhEPO. Therefore, development of long-lasting and curative therapies is highly required. Combined cell and gene therapy platform can introduce a new route for permanent production of erythropoietin (EPO) in the body with various degrees of clinical benefits and avoiding the need for repeat treatments. In this study, we developed cell and gene therapy strategy for in-vivo delivery of EPO cDNA via genetic engineering human Wharton’s jelly mesenchymal stem cells (hWJMSCs) to long-term produce and secret human EPO protein after transplantation into the mice model of CRA. To evaluate CRA's treatment in cancer-free and cancerous conditions, at first, we designed recombinant breast cancer cell line 4T1 expressing herpes simplex virus type 1 thymidine kinase (HSV1-TK) by a lentiviral vector encoding HSV1-TK and injected into mice. After 3 weeks, all mice develop metastatic breast cancer associated with acute anemia. Then, we administrated ganciclovir (GCV) for 10 days in half of the mice to clear cancer cells. Meanwhile, we designed another lentiviral vector encoding EPO to transduce hWJMSCs. Following implantation of rhWJMSCs-EPO, the whole peripheral blood samples were collected from the tail vein once per week for 10 weeks which were immediately analyzed for the measurements of EPO, hemoglobin (Hb), and hematocrit (Hct) plasma levels. The blood analysis showed that plasma EPO, hemoglobin (Hb), and hematocrit (Hct) concentration significantly increased and remained at a therapeutic level for >10 weeks in both treatment groups which indicates that the rhWJMSCs-EPO could improve CRA in both cancer-free and cancerous mice model.
BackgroundCancer-related anemia (CRA) negatively influences cancer patients’ survival, disease progression, treatment efficacy, and quality of life (QOL). Current treatments such as iron therapy, red cell transfusion, and erythropoietin-stimulating agents (ESAs) may cause severe adverse effects. Therefore, development of long-lasting and curative therapies is highly required. Combined cell and gene therapy platform can introduce a new route for permanent production of erythropoietin (EPO) in the body with various degrees of clinical benefits and avoiding the need for repeat treatments.MethodsIn this study, we developed cell and gene therapy strategy for in-vivo delivery of EPO cDNA via genetic engineering human Wharton’s jelly mesenchymal stem cells (hWJMSCs) to long-term produce and secret human EPO protein after transplantation into the mice model of CRA. To evaluate CRA's treatment in cancer-free and cancerous conditions, at first, we designed recombinant breast cancer cell line 4T1 expressing herpes simplex virus type 1 thymidine kinase (HSV1-TK) by a lentiviral vector encoding HSV1-TK and injected into mice. After 3 weeks, all mice develop the metastatic breast cancer associated to the acute anemia. Then, we administrated ganciclovir (GCV) for 10 days in half of the mice to clear cancer cells. Meanwhile, we designed another lentiviral vector encoding EPO to transduce hWJMSCs. Following implantation of rhWJMSCs-EPO, the whole peripheral blood samples were collected from the tail vein once per week which were immediately analyzed for the measurements of EPO, hemoglobin (Hb), and hematocrit (Hct) plasma levels. Results We found that after implantation of rhWJMSCs-EPO, plasma EPO, Hb, and Hct concentration significantly increased which rose to a peak in the fourth week and remained at a therapeutic level for >17 weeks in the cancer-free group and >10 weeks in the cancerous group.ConclusionOur data indicate that the EPO-transduced hWJMSCs could improve the anemia of cancer in both cancer-free and cancerous mice model. This significant difference in length of time that Hb and Hct are in therapeutic levels in both treatment groups indicates that developing a precise targeted-therapy to eliminate cancer cells along with an effective treatment for CRA, as we presented here, could bring important clinical benefits.
Cancer-related anemia (CRA) negatively influences cancer patients’ survival, disease progression, treatment efficacy, and quality of life. Therefore, development of long-lasting and curative therapies is highly required. In this study, we developed cell and gene therapy strategy for in-vivo delivery of EPO cDNA via genetic engineering human Wharton’s jelly mesenchymal stem cells (hWJMSCs) to long-term produce human EPO protein after transplantation into the mice model of CRA. To evaluate CRA's treatment in cancer-free and cancerous conditions, we designed recombinant breast cancer cell line 4T1 expressing herpes simplex virus type 1 thymidine kinase (HSV1-TK) by a lentiviral vector encoding HSV1-TK and injected into mice. After confirming CRA in mice by blood analysis, half of them received ganciclovir for 10 days to clear cancer cells. Meanwhile, we designed another lentiviral vector encoding EPO to transduce hWJMSCs. Following implantation of rhWJMSCs-EPO, the whole peripheral blood samples were collected once per week for 10 weeks. The blood analyzing showed that plasma EPO, hemoglobin (Hb), and hematocrit (Hct) concentration significantly increased and remained at a therapeutic level for >10 weeks in the both treatment groups which indicate that the rhWJMSCs-EPO could improve CRA in both cancer-free and cancerous mice model.
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