Drug repurposing: Misconceptions, challenges, and opportunities for academic researchers.

Drug repurposing is promoted as a cost- and time-effective mechanism for providing new medicines. Often, however, there is insufficient consideration by academic researchers of the processes required to ensure that a repurposed drug can be used for a new indication. This may explain the inability of drug repurposing to fulfill its promise. Important aspects, often overlooked, include financial and intellectual property considerations, the clinical and regulatory path, and clinical equipoise, which provides ethical justification for randomized controlled trials. The goal of drug repurposing is to obtain a new regulator-approved label for an existing drug, and so, the trajectory for drug repurposing and traditional drug development is similar. Here, we discuss factors critical for a successful repurposed medicine to help academic investigators better identify drug repurposing opportunities.

Surface proteins of SARS-CoV-2 drive airway epithelial cells to induce interferon-dependent inflammation

SARS-CoV-2, the virus that has caused the COVID-19 pandemic, robustly activates the host immune system in critically ill patients. Understanding how the virus engages the immune system will facilitate the development of needed therapeutic strategies. Here we demonstrate both in vitro and in vivo that the SARS-CoV-2 surface proteins Spike (S) and Envelope (E) activate the key immune signaling interferon (IFN) pathway in both immune and epithelial cells independent of viral infection and replication. These proteins induce reactive oxidative species generation and increases in human and murine specific IFN-responsive cytokines and chemokines, similar to their upregulation in critically ill COVID-19 patients. Induction of IFN signaling is dependent on canonical but discrepant inflammatory signaling mediators as the activation induced by S is dependent on IRF3, TBK1, and MYD88 while that of E is largely MYD88 independent. Furthermore, these viral surface proteins, specifically E, induced peribronchial inflammation and pulmonary vasculitis in a mouse model. Finally we show that the organized inflammatory infiltrates are dependent on type I IFN signaling, specifically in lung epithelial cells. These findings underscore the role of SARS-CoV-2 surface proteins, particularly the understudied E protein, in driving cell specific inflammation and their potential for therapeutic intervention. Author SummarySARS-CoV-2 robustly activates widespread inflammation, but we do not understand mechanistically how the virus engages the immune system. This knowledge will facilitate the development of critically needed therapeutic strategies to promote beneficial immune responses will dampening harmful inflammation. Here we demonstrate that SARS-CoV-2 surface proteins spike and envelope alone activated innate cell function and the interferon signaling pathway. This activation occurred in both immune and epithelial cells, and mechanistic studies demonstrated dependence on known key inflammatory signaling mediators, IRF3, TBK1, and MYD88. In animal studies, we showed that these viral surface proteins induce epithelial cell IFN-dependent lung pathology, reminiscent to acute COVID-19 pulmonary infection. These findings underscore the need for further investigation into the role of SARS-CoV-2 surface proteins, particularly the understudied E protein, in driving cell specific inflammation.

Induction of T cell-mediated immunity using a c-Myb DNA vaccine in a mouse model of colon cancer.

Overexpression of the proto-oncogene c-Myb occurs in more than 80% of colorectal cancer (CRC) and is associated with aggressive disease and poor prognosis. To test c-Myb as a therapeutic target in CRC we devised a DNA fusion vaccine to generate an anti-CRC immune response. c-Myb, like many tumor antigens, is weakly immunogenic as it is a "self" antigen and subject to tolerance. To break tolerance, a DNA fusion vaccine was generated comprising wild-type c-Myb cDNA flanked by two potent Th epitopes derived from tetanus toxin. Vaccination was performed targeting a highly aggressive, weakly immunogenic, subcutaneous, syngeneic, colon adenocarcinoma cell line MC38 which highly expresses c-Myb. Prophylactic intravenous vaccination significantly suppressed tumor growth, through the induction of anti-tumor immunity for which the tetanus epitopes were essential. Vaccination generated anti-tumor immunity mediated by both CD4+ and CD8+ T cells and increased infiltration of immune effector cells at the tumor site. Importantly, no evidence of autoimmune pathology in endogenous c-Myb expressing tissues was detected as a consequence of breaking tolerance. In summary, these results establish c-Myb as a potential antigen for immune targeting in CRC and serve to provide proof of principle for the continuing development of DNA vaccines targeting c-Myb to bring this approach to the clinic.

PI-88 and novel heparan sulfate mimetics inhibit angiogenesis.

The heparan sulfate (HS) mimetic PI-88 is a promising inhibitor of tumor growth and metastasis expected to commence phase III clinical evaluation in 2007 as an adjuvant therapy for postresection hepatocellular carcinoma. Its anticancer properties are attributed to inhibition of angiogenesis via antagonism of the interactions of angiogenic growth factors and their receptors with HS. It is also a potent inhibitor of heparanase, an enzyme that plays a key role in both metastasis and angiogenesis. A series of PI-88 analogs have been prepared with enhanced chemical and biological properties. The new compounds consist of single, defined oligosaccharides with specific modifications designed to improve their pharmacokinetic properties. These analogs all inhibit heparanase and bind to the angiogenic fibroblast growth factor 1 (FGF-1), FGF-2, and vascular endothelial growth factor with similar affinity to PI-88. However, compared with PI-88, some of the newly designed compounds are more potent inhibitors of growth factor-induced endothelial cell proliferation and of endothelial tube formation on Matrigel. Representative compounds were also tested for antiangiogenic activity in vivo and were found to reduce significantly blood vessel formation. Moreover, the pharmacokinetic profile of several analogs was also improved, as evidenced primarily by lower clearance in comparison with PI-88. The current data support the development of HS mimetics as potent antiangiogenic anticancer agents.

Linked foreign T-cell help activates self-reactive CTL and inhibits tumor growth.

Transgenic mice expressing membrane-bound OVA under the rat insulin promoter, RIP-mOVA, has previously been suggested to display deletional tolerance toward the dominant CTL epitope, SIINFEKL, and provide an elegant model system to test the hypothesis that the lack of T cell help contributes to the tolerance. To understand how the CD8 tolerance is maintained in these mice, a set of neo-self-Ags, OVA, modified to contain a foreign Th peptide, were constructed and tested for their ability to induce CTL responses in RIP-mOVA mice. Immunization with these Th peptide-modified OVA molecules and not with the wild-type OVA induced self-reactive CTLs recognizing dominant CTL peptide, SIINFEKL. Importantly, immunization with the modified OVA constructs also prevented the growth of OVA-expressing tumors in transgenic mice. Since endogenous OVA Th peptides did not contribute toward breaking self CTL tolerance, these results also highlighted a very robust CD4 T cell tolerance toward OVA in RIP-mOVA mice that has not been previously described. These results therefore provide direct evidence that it is the tolerance in the CD4 Th cell compartment that helps maintain the CTL tolerance against self-Ag in these mice. Since the CTL tolerance can be broken or bypassed by foreign Th peptides inserted into a self Ag, potential of using this approach in generating effective therapeutic cancer vaccines is discussed.

Induction of TNF-alpha autoantibody production by AutoVac TNF106: a novel therapeutic approach for the treatment of allergic diseases.

BACKGROUND: Cytokines play an integral role in the coordination and persistence of allergic inflammatory processes and therefore represent prime targets for novel therapies in diseases such as asthma. Multiple attempts to generate low-molecular-weight cytokine inhibitors have failed, and the main attention has focused on the development of biological agents such as neutralizing antibodies. The present work describes a simple and effective method to induce the production of therapeutic anti-cytokine autoantibodies by active immunization against a modified endogenous cytokine. METHODS: Balb/c mice were subcutaneously injected with AutoVac TNF106, a recombinant murine TNF-alpha molecule containing a foreign immunogenic T helper epitope, and the induction of neutralizing anti-TNF-alpha autoantibodies was analysed. These mice were then sensitized with ovalbumin (OVA), and the effect of neutralizing anti-TNF-alpha autoantibodies on the allergen-induced airway inflammation was analysed. RESULTS: AutoVac TNF106-immunized mice developed high titres of neutralizing anti-TNF-alpha autoantibodies, which were maintained for at least 4 weeks after the last booster injection. Mice vaccinated with AutoVac TNF106 and further immunized against OVA showed diminished TNF-alpha levels in the bronchoalveolar lavage (BAL) fluid after OVA challenge. Moreover, pretreatment with AutoVac TNF106 resulted in significantly reduced numbers of eosinophils and neutrophils in BAL fluid in response to single or multiple allergen exposure. CONCLUSION: The induction of anti-TNF-alpha autoantibody production by the AutoVac TNF106 technology not only confirmed the role of TNF-alpha in the induction of allergic inflammation but also offers a novel approach to block the activity of cytokines in order to treat allergic inflammatory conditions.

HER-2 DNA and protein vaccines containing potent Th cell epitopes induce distinct protective and therapeutic antitumor responses in HER-2 transgenic mice.

Overexpression of the growth factor receptor HER-2 (c-erbB-2, neu) has transforming potential and occurs in approximately 20-30% of breast and ovarian cancers. HER-2 is a self Ag, but Abs and T cells specific for HER-2 have been isolated from cancer patients, suggesting HER-2 may be a good target for active immunotherapy. We constructed rat HER-2 DNA and protein vaccines containing potent Th cell epitopes derived from tetanus toxin and studied their potency in two strains of mice transgenic for the rat HER-2 molecule. Vaccination with HER-2 DNA protected nontransgenic mice from tumor challenge, but induced only moderate protection in one of the tumor models. However, vaccination with the modified HER-2 protein resulted in almost complete protection from tumor challenge in both tumor models. This protection could be mediated by Abs alone. In addition, protein vaccination efficiently eliminated pre-established tumors in both models, even when vaccination occurred 9 days after tumor implantation. These data demonstrate the potential of HER-2-based vaccines as therapeutic agents for the treatment of cancers overexpressing HER-2.