Mayaro virus (MAYV) of the genus alphavirus is a mosquito-transmitted emerging infectious disease that causes an acute febrile illness, rash, headaches, and nausea that may turn into incapacitating, persistent arthralgias in some victims. Since its discovery in Trinidad in 1954, cases of MAYV infection have largely been confined there and to the northern countries of South America, but recently, MAYV cases have been reported in some island nations in the Caribbean Sea. Accompanying these reports is evidence that new vectors, including Aedes spp. mosquitos, recently implicated in the global spread of Zika and chikungunya viruses, are competent for MAYV transmission, which, if true, could facilitate the spread of MAYV beyond its current range. Despite its status as an emerging virus, there are no licensed vaccines to prevent MAYV infection nor therapeutics to treat it. Here, we describe the development and testing of a novel DNA vaccine, scMAYV-E, that encodes a synthetically-designed consensus MAYV envelope sequence. In vivo electroporation-enhanced immunization of mice with this vaccine induced potent humoral responses including neutralizing antibodies as well as robust T-cell responses to multiple epitopes in the MAYV envelope. Importantly, these scMAYV-E-induced immune responses protected susceptible mice from morbidity and mortality following a MAYV challenge. Mayaro virus (MAYV) is a mosquito-transmitted virus that causes fever, headache, chills, nausea and joint pain that can last for months after infection. The rising number of cases, due to increased mosquito circulation, and the threat of an epidemic emphasize its importance as an emerging virus, but there are no licensed vaccines to prevent Mayaro infection nor therapeutics to treat it. In this study, we gathered fundamental knowledge on how the immune system responds to MAYV infection, and we evaluated the efficacy of a novel, synthetic DNA envelope vaccine (scMAYV-E) in mice. Analysis of immune responses in mice demonstrated that this vaccine induces potent T cell immunity and antibodies. Mice who receive the vaccine and then are challenged with live MAYV are protected against Mayaro disease. This data provides evidence that the DNA-based MAYV vaccine may be able to prevent Mayaro disease. Thus, the scMAYV-E vaccine is a promising step forward for MAYV vaccine development. Future testing will evaluate whether this vaccine is a viable means to halt the spread of MAYV and protect individuals from Mayaro disease.

Specific antibody therapy, including mAbs and bispecific T cell engagers (BiTEs), are important new tools for cancer immunotherapy. However, these approaches are slow to develop and may be limited in their production, thus restricting the patients who can access these treatments. BiTEs exhibit a particularly short half-life and difficult production. The development of an approach allowing simplified development, delivery, and in vivo production would be an important advance. Here we describe the development of a designed synthetic DNA plasmid, which we optimized to permit high expression of an anti-HER2 antibody (HER2dMAb) and delivered it into animals through adaptive electroporation. HER2dMAb was efficiently expressed in vitro and in vivo, reaching levels of 50 μg/ml in mouse sera. Mechanistically, HER2dMAb blocked HER2 signaling and induced antibody-dependent cytotoxicity. HER2dMAb delayed tumor progression for HER2-expressing ovarian and breast cancer models. We next used the HER2dMAb single-chain variable fragment portion to engineer a DNA-encoded BiTE (DBiTE). This HER2DBiTE was expressed in vivo for approximately 4 months after a single administration. The HER2DBiTE was highly cytolytic and delayed cancer progression in mice. These studies illustrate an approach to generate DBiTEs in vivo, which represent promising immunotherapies for HER2 ⁺ tumors, including ovarian and potentially other cancers.

Elevated low-density lipoprotein cholesterol (LDL-C) is one of the major contributors to cardiovascular heart disease (CHD), the leading cause of death worldwide. Due to severe side effects of statins, alternative treatment strategies are required for statin-intolerant patients. Monoclonal antibodies (mAbs) targeting proprotein convertase subtilisin/kexin type 9 (PCSK9) have shown great efficacy in LDL-C reduction. Limitations for this approach include the need for multiple injections as well as increased costs associated with patient management. Here, we engineered a DNA-encoded mAb (DMAb) targeting PCSK9 (daPCSK9), as an alternative approach to protein-based lipid-lowering therapeutics, and we characterized its expression and activity. A single intramuscular administration of mouse daPCSK9 generated expression in vivo for over 42 days that corresponded with a substantial decrease of 28.6% in non-high-density lipoprotein cholesterol (non-HDL-C) and 10.3% in total cholesterol by day 7 in wild-type mice. Repeated administrations of the DMAb plasmid led to increasing expression, with DMAb levels of 7.5 μg/mL at day 62. daPCSK9 therapeutics may provide a novel, simple, less frequent, cost-effective approach to reducing LDL-C, either as a stand-alone therapy or in combination with other LDL-lowering therapeutics for synergistic effect.

Ovarian cancer presents in 80% of patients as a metastatic disease, which confers it with dismal prognosis despite surgery and chemotherapy. However, it is an immunogenic disease, and the presence of intratumoral T cells is a major prognostic factor for survival. We used a synthetic consensus (SynCon) approach to generate a novel DNA vaccine that breaks immune tolerance to follicle-stimulating hormone receptor (FSHR), present in 50% of ovarian cancers but confined to the ovary in healthy tissues. SynCon FSHR DNA vaccine generated robust CD8+ and CD4+ cellular immune responses and FSHR-redirected antibodies. The SynCon FSHR DNA vaccine delayed the progression of a highly aggressive ovarian cancer model with peritoneal carcinomatosis in immunocompetent mice, and it increased the infiltration of anti-tumor CD8+ T cells in the tumor microenvironment. Anti-tumor activity of this FSHR vaccine was confirmed in a syngeneic murine FSHR-expressing prostate cancer model. Furthermore, adoptive transfer of vaccine-primed CD8+ T cells after ex vivo expansion delayed ovarian cancer progression. In conclusion, the SynCon FSHR vaccine was able to break immune tolerance and elicit an effective anti-tumor response associated with an increase in tumor-infiltrating T cells. FSHR DNA vaccination could help current ovarian cancer therapy after first-line treatment of FSHR+ tumors to prevent tumor recurrence.

Ovarian cancer is frequently diagnosed as peritoneal carcinomatosis. Unlike other tumor locations, the peritoneal cavity is commonly exposed to gut-breaching and ascending genital microorganisms and has a unique immune environment. IL-33 is a local cytokine that can activate innate and adaptive immunity. We studied the effectiveness of local IL-33 delivery in the treatment of cancer that has metastasized to the peritoneal cavity. Direct peritoneal administration of IL-33 delayed the progression of metastatic peritoneal cancer. Prolongation in survival was not associated with a direct effect of IL-33 on tumor cells, but with major changes in the immune microenvironment of the tumor. IL-33 promoted a significant increase in the leukocyte compartment of the tumor immunoenvironment and an allergic cytokine profile. We observed a substantial increase in the number of activated CD4⁺ T-cells accompanied by peritoneal eosinophil infiltration, B-cell activation and activation of peritoneal macrophages which displayed tumoricidal capacity. Depletion of CD4⁺ cells, eosinophils or macrophages reduced the anti-tumor effects of IL-33 but none of these alone were sufficient to completely abrogate its positive benefit. In conclusion, local administration of IL-33 generates an allergic tumor environment resulting in a novel approach for treatment of metastatic peritoneal malignancies, such as advanced ovarian cancer.