Consecutive low doses of cyclophosphamide preferentially target Tregs and potentiate T cell responses induced by DNA PLG microparticle immunization

Dana-Farber Cancer Institute, Boston, MA, USA.
Cellular Immunology (Impact Factor: 1.92). 02/2010; 262(2):150-61. DOI: 10.1016/j.cellimm.2010.02.007
Source: PubMed


Cyclophosphamide in combination with immunotherapeutic approaches preferentially impinges on T(reg) activity and allows for robust generation of T cell effectors. Reduced dosages of cyclophosphamide are necessary to restrict its cytotoxic effects to the negative regulatory cell populations while sparing effector lymphocytes. We investigated cyclophosphamide dosing in combination with ZYC300, a PLG-encapsulated plasmid DNA vaccine which encodes the cytochrome P450 family member, CYP1B1, a known human tumor-associated antigen. In mice, three consecutive, low doses of cyclophosphamide comprised a superior regimen in enhancing the magnitude, diversity of epitopes, and avidity to individual epitopes of specific T cell responses when compared to regimens that used either a single low or a single high dose. Consecutive low doses of cyclophosphamide predominantly targeted T(regs) while sparing overall T lymphocyte counts. Thus, we report the synergistic activity of pharmacologic T(reg) depletion with cyclophosphamide on quantitatively and qualitatively increasing T cell responses to a known human tumor-associated antigen.

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Available from: Christine M Barbon, Jul 03, 2014
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    • "Cord blood Tregs have been shown to be predominantly of the CD45RA + CD45RO − naïve phenotype in several studies (Kanegane et al., 1991; Wing et al., 2002; Takahata et al., 2004; Ly et al., 2009; Flanagan et al., 2010). Other phenotypic differences between cord and adult Tregs include the observation that CB Tregs are mostly CD27 + and thus at an earlier differentiation state than their mothers; they have a lower apoptotic potential as evidenced by lower CD95/Fas expression than their mothers; and less CD62L suggesting less of a Treg CM lymph node homing phenotype (Flanagan et al., 2010). CB Tregs also express less CCR6 than their matched mothers, which is the chemokine receptor that characterizes the Th17-and Th22-like Tregs (Duhen et al., 2012). "
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    ABSTRACT: Human newborns and infants are bombarded with multiple pathogens on leaving the sterile intra-uterine environment, and yet have suboptimal innate immunity and limited immunological memory, thus leading to increased susceptibility to infections in early life. They are thus the target age group for a host of vaccines against common bacterial and viral pathogens. They are also the target group for many vaccines in development, including those against tuberculosis (TB), malaria, and HIV infection. However, neonatal and infant responses too many vaccines are suboptimal, and in the case of the polysaccharide vaccines, it has been necessary to develop the alternative conjugated formulations in order to induce immunity in early life. Immunoregulatory factors are an intrinsic component of natural immunity necessary to dampen or control immune responses, with the caveat that they may also decrease immunity to infections or lead to chronic infection. This review explores the key immunoregulatory factors at play in early life, with a particular emphasis on regulatory T cells (Tregs). It goes on to explore the role that Tregs play in limiting vaccine immunogenicity, and describes animal and human studies in which Tregs have been depleted in order to enhance vaccine responses. A deeper understanding of the role that Tregs play in limiting or controlling vaccine-induced immunity would provide strategies to improve vaccine immunogenicity in this critical age group. New adjuvants and drugs are being developed that can transiently suppress Treg function, and their use as part of human vaccination strategies against infections is becoming a real prospect for the future.
    Frontiers in Microbiology 09/2014; 5:477. DOI:10.3389/fmicb.2014.00477 · 3.99 Impact Factor
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    • "Higher, cytotoxic doses of CPA impact all of the T-lymphocyte populations, resulting not only in reduced Treg counts but also reduced CD4+, CD8+, and total splenocyte counts [29,33]. Thus, a cytotoxic dose of CPA can impair the immune response to tumor-associated antigens [29]. These observations are consistent with Motoyoshi et al. who showed that low-dose CPA produced an antitumor immune response by selectively depleting Tregs in immunocompetent mice, while the effects of high-dose CPA were attributable solely to its cytotoxic effects [33]. "
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    ABSTRACT: Background L-BLP25 antigen-specific cancer immunotherapeutic agent is currently in phase III clinical trials for non-small cell lung cancer. Using a novel human MUC1 transgenic (hMUC1.Tg) lung cancer mouse model, we evaluated effects of L-BLP25 combined with low-dose cyclophosphamide (CPA) pretreatment on Th1/Th2 cytokine production and antitumor activity. Methods A chemically-induced lung tumor model was developed in hMUC1.Tg C57BL/6 mice by administering 10 weekly 0.75-mg/g doses of the chemical carcinogen urethane by intraperitoneal injection. Serum cytokines associated with Th1/Th2 polarization and inflammation were measured by multiplex cytokine assay during tumorigenesis. Antitumor activity of L-BLP25 (10 μg) with CPA (100 mg/kg) pretreatment was evaluated following either one or two eight-week cycles of treatment by preparing lung whole mounts and counting tumor foci, and assessing IFN-γ production by ELISpot assay. Results During the carcinogenesis phase, no detectable Th1- or Th2-associated cytokine responses were observed, but levels of pro-inflammatory cytokines were increased with distinctive kinetics. A single cycle of L-BLP25 consisting of eight weekly doses was ineffective, whereas adding a second cycle given during tumor progression showed a significant reduction in the incidence of tumor foci. Administering two cycles of L-BLP25 induced Th1 cytokines IL-12, IL-2 and IFNγ at 24 h after the last dose, while Th2 and inflammatory cytokines were elevated to a lesser extent. Conclusions Urethane-induced lung tumors in hMUC1.Tg mice can be used as a model to assess the efficacy of the MUC1 antigen-specific cancer immunotherapeutic agent L-BLP25. The results indicate that the antitumor response to L-BLP25 requires at least two cycles and pre-treatment with CPA. In addition, monitoring pro-inflammatory serum cytokines may be useful as a biomarker of L-BLP25 response. Taken together, the preclinical lung tumor model can be utilized for determining effective combinations of L-BLP25 with chemotherapy and/or other immunotherapies.
    Journal of Translational Medicine 03/2013; 11(1):64. DOI:10.1186/1479-5876-11-64 · 3.93 Impact Factor
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