No abstract

Background: Nowadays, the prevention of parasitic diseases including leishmaniasis is one of the health concerns in the world, and cutaneous leishmaniasis is the most common type of these diseases. So far, no drug or vaccine has been approved for definitive treatment of this disease.Methods: In this study, the recombinant lentiviral vaccine containing a new multi-epitope of KMP11 and HASPB of the Leishmania major (L. major) was synthesized that had previously been designed in-silico. The designed multi-epitope was subcloned into the pCDH513 lentiviral vector, and the recombinant lentiviral multi-epitope vaccine (rLV-multi-epitope) was synthesized in the HEK293T cell by the packaging vectors. Also, the Western Blotting method was used to confirm the gene expression. Then, the rLV-multi-epitope vaccine was injected twice, along with two control groups, PBS, and rLV-empty to immunize the BALB /c mice. Twenty-one days after the second injection, the splenocytes of the mice were is­­­­olated and stimulated with the Leishmania lysate.Results: The results of the enzyme-linked immunoassay (ELISA) test not only showed the titer of IFN-γ and IL-4 was increased in the immunized group compared to the controls, but also indicated that the ratio of IFN-γ to IL-4 cytokines in the main group was increased significantly. As a result, the Th1 response was generated in the main group. Moreover, the humoral immune response was assessed and the results showed that the ratio of IgG2a to IgG1 antibody in the sera of the immunized mice was increased compared to the control groups. Also, the ratio of IgG2a to IgG1 was increased in the main group. Therefore, the humoral immune response was increased, which can also have a positive effect on increasing the Th1 response.Conclusions: Our results showed that immunization by the new rLV-multi-epitope vaccine could stimulate the immune system towards the Th1 through increasing the IFN-γ production.

BackgroundSince the incidence of various types of leishmaniasis, no definitive treatment has been considered for the disease, and due to its high prevalence worldwide, this issue has caused many concerns. Cutaneous leishmaniasis is the most common form of the disease which, can cause malignant lesions on the skin. Vaccination for the prevention and treatment of leishmaniasis can be the most effective way to combat this disease. In this study, we designed a new multi-epitope vaccine using immunoinformatics tools, which confirmed its effectiveness in the in silico.MethodsSequences Leish-111f protein (TSA, Leif, and LMSTI1) of Leishmania major (L. major) were downloaded from GenBank and with the help of immunoinformatic tools, was designed a new multi-epitope vaccine antigen of L. major.ResultTh and Tc epitopes of the leish-111f protein were predicted using bioinformatics tools. The final multi epitope was consisted of 18 CTL epitopes that joined by AAY linker. There are also 9 HTL epitopes in the structure of the final vaccine that were joined by GPGPG linker. The profilin adjuvant was also added into the construct by AAY Linker. There were 613 residues in the structure of the final construct. The multi epitope was stable and non-allergic. the data obtained from the binding of final multi-epitope vaccine-TLR11 residues (band lengths and weighted scores) showed that the ligand and the receptor have a high affinity to bind to each other. Moreover, in silico cloning approach, was improved the expression of proposed vaccine in E. coli host. Codon adaptation index and GC percent were calculated 1.0 and 53.35, respectivelyConclusionBased on these results, we hope that the multi-epitope vaccine, which contains the most appropriate epitopes of a strong Leishmania major immunogen, along with an adjuvant capable of binding to TLR11, will further stimulate the immune system against the L.major.

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.