Combination Of PORF-mCD40L Gene Therapy With Cisplatin Improves Therapeutic Efficacy Of Murine Carcinoma

Objectives:The ligand for CD40(CD40L or CD154) is a 39KDa typeⅡtrans-membrance protein of the TNF superfamily that is preferentiallyexpressed on activated CD4~+T cell. The receptor for CD40L, CD40 (40 kd), isexpressed on APCs, including DCs, B cells, activated macrophages, andfollicular DCs. CD40/CD40 ligand (CD40L) interaction plays an essential rolein cell-mediated immune responses.DCs are professional APCs specialized insampling antigen throughout the body, migrating to lymphoid organs, andpresenting antigen to naive T cells. Stimuli from CD4~+ T-helper cells via theCD40/CD40L interaction is essential in bringing the DCs to a state in whichthey can autonomously trigger antigen-specific T-cell responses, the CD40L onantigen-stimulated CD4~+ T-helper cells activates DCs, with up-regulation ofT-cell costimulatory molecules such as B7 and intercellular adhesion molecule(ICAM)-1, and consequent direct stimulation of CD8~+ T-killer cells. As part ofthe activation of DCs, the CD40L-CD40 interaction induces the production ofcytokines that favor the development of a T-helper 1 (Th1) response.Havebeen shown that in vivo genetic modification of tumors with an adenovirus (Ad)vector engineered to express CD40 ligand (AdmCD40L) inducestumor-specific CTLs and suppresses tumor growth in vivo. Recent studiessuggest that the expression of the gene encoding CD40 Ligand (CD40L) may be an effective strategy to enhance the host immune response,and CD40L alsohave direct efficency to tumor. Based on the capacity of CD40L-CD40 inimmune response system and it's effects on tumor, We hypothesized theadministration of CD40L with the vector of liposome might induce or enhancea specific antitumor response with cisplatin in vivo and in vitro..Methods:To test this concept, A combined treatment strategy was designedusing pORF-mCD40L gene therapy plus cisplatin chemotherapy to determinethe antitumor ability. And the antitumor activity was evaluated in twoimmunocompetent mice loaded with Lewis lung carcinomas (LL/2) and Coloncarcinomas (CT26). The treatment protocols were designed by administeringpORF-mCD40L intratumor at 50μg / mouse twice a week for 4 weeks, andcisplatin intraperitoneally at 5 mg/kg/mouse a week for 4 weeks. Tumorvolume and survival time were observed. CD40L immunohistochemistrystaining of tumor tissues. Assessment of apoptotic cells in tumor tissues andCTL activities of lymphocytes were also conducted. The combined treatmentimproved antitumor activity and strengthen tumor regression and significantlyprolonged the life span of tumor bearing mice compared with the control. Thesynergistic treatment with cisplatin enlarge induction of apoptosis and elevatedCTL activities. In addition, we also made another experiment models as lungmetastasis model with CT26 and ascites model with H22 (in these models,we all gave the same dose with pORF-mCD40L), These studies provided thebasis for of the gene therapy with Liposome.Results:We found the antitumor effect of pORF-mCD40L in combinationwith cisplatin demonstrates an augmented effect than the groups of cispltinand pORF-mCD40L alone and saline contral, and resulted in depression ofestablished tumor growth and beneficial survival in murine tumor models.CTL activity of lymphocytes was found in T cell isolated from spleens of micetreated with pORF-mCD40L and cisplatin. The antitumor activity could be partly abrogated by depletion of CD4~+ or CD8~+ T lymphocytes with theantibodies of CD4 or CD8, furthermore, the later should apparently abrogatethe antitumor activity in vivo. There were also found increased IFN-γsecretingin pORF-mCD40L treated mices. HE and Immunohistochemical stainingrevealed that combined treatment group augmented induction of necrosis andapoptosis within the tumor tissues compared with other groups.in metastasismice, there are only 1-2 nudus in the lung of the mice treated withporf-CD40L, but control group was 200~+ nudus in their lung; pORF-mCD40Lalso indicated that it had evident effectiveness in controlling formation of ascites.Conclusions:our study indicated that pORF-mCD40L encapsuled with Liposome andcombined with cisplatin had synergistric antitumor effects in treating transplantedtumor subcutaneouly and in malignant ascites and metastatic tumor. Moreover,it is possible for Liposome as the vector of gene for the treatment of tunor.