PDL1 blockade produces remarkable clinical responses, thought to occur by T cell reactivation through prevention of PDL1-PD1 T cell inhibitory interactions. Here, we find that PDL1 cell-intrinsic ...signaling protects cancer cells from interferon (IFN) cytotoxicity and accelerates tumor progression. PDL1 inhibited IFN signal transduction through a conserved class of sequence motifs that mediate crosstalk with IFN signaling. Abrogation of PDL1 expression or antibody-mediated PDL1 blockade strongly sensitized cancer cells to IFN cytotoxicity through a STAT3/caspase-7-dependent pathway. Moreover, somatic mutations found in human carcinomas within these PDL1 sequence motifs disrupted motif regulation, resulting in PDL1 molecules with enhanced protective activities from type I and type II IFN cytotoxicity. Overall, our results reveal a mode of action of PDL1 in cancer cells as a first line of defense against IFN cytotoxicity.
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•PDL1 expression represents a direct defense barrier against IFN toxicity•PDL1 contains non-classical conserved signal transduction motifs•PDL1 signaling motifs as hotspots for hyperactivating somatic mutations in cancer•Interfering with PDL1 signaling motifs sensitizes cancer cells to IFN cytotoxicity
Gato-Cañas et al. find that PDL1 protects cancer cells from interferon toxicity by counteracting interferon signaling through the activities of two non-classical conserved motifs. Human cancers acquire somatic mutations within these motifs that enhance PDL1 anti-interferon activities, favoring tumor progression.
The role of long noncoding RNAs (lncRNAs) in viral infection is poorly studied. We have identified hepatitis C virus (HCV)‐Stimulated lncRNAs (CSRs) by transcriptome analysis. Interestingly, two of ...these CSRs (PVT1 and UCA1) play relevant roles in tumorigenesis, providing a novel link between HCV infection and development of liver tumors. Expression of some CSRs seems induced directly by HCV, while others are upregulated by the antiviral response against the virus. In fact, activation of pathogen sensors induces the expression of CSR32/EGOT. RIG‐I and the RNA‐activated kinase PKR sense HCV RNA, activate NF‐κB and upregulate EGOT. EGOT is increased in the liver of patients infected with HCV and after infection with influenza or Semliki Forest virus (SFV). Genome‐wide guilt‐by‐association studies predict that EGOT may function as a negative regulator of the antiviral pathway. Accordingly, EGOT depletion increases the expression of several interferon‐stimulated genes and leads to decreased replication of HCV and SFV. Our results suggest that EGOT is a lncRNA induced after infection that increases viral replication by antagonizing the antiviral response.
Synopsis
This study shows that the expression of several cellular lncRNAs is induced in HCV‐infected cells, either directly triggered by the viral infection or activated by the antiviral response. One of these is EGOT, which blocks the interferon response and favors viral replication.
HCV infection directly induces lncRNAs that partly play relevant roles in cell proliferation, providing a novel link between HCV infection and hepatocarcinogenesis.
Also the cellular antiviral response to HCV or other RNA viruses activates the expression of the lncRNAs (such as EGOT).
EGOT is induced by NF‐κB after PKR or RIG‐I activation and functions as a negative regulator of the type I IFN response.
Increased EGOT expression in infected cells favors viral replication.
This study shows that the expression of several cellular lncRNAs is induced in HCV‐infected cells, either directly triggered by the viral infection or activated by the antiviral response. One of these is EGOT, which blocks the interferon response and favors viral replication.
Progressive familial intrahepatic cholestasis type 3 (PFIC3) is a rare monogenic disease caused by mutations in the ABCB4 gene, resulting in a reduction in biliary phosphatidylcholine. Reduced ...biliary phosphatidylcholine cannot counteract the detergent effects of bile salts, leading to cholestasis, cholangitis, cirrhosis and ultimately liver failure. Here, we report results from treating two- or five-week-old Abcb4
mice with an AAV vector expressing human ABCB4, resulting in significant decreases of PFIC3 disease biomarkers. All male mice achieved a sustained therapeutic effect up through 12 weeks, but the effect was achieved in only 50% of females. However, two-week-old females receiving a second inoculation three weeks later maintained the therapeutic effect. Upon sacrifice, markers of PFIC3 disease such as, hepatosplenomegaly, biliary phosphatidylcholine and liver histology were significantly improved. Thus, AAV-mediated gene therapy successfully prevented PFIC3 symptoms in a clinically relevant mouse model, representing a step forward in improving potential therapy options for PFIC3 patients.
Cancer therapy has experienced a breakthrough with the use of immune checkpoint inhibitors (ICIs) based on monoclonal antibodies (mAbs), which are able to unleash immune responses against tumors ...refractory to other therapies. Despite the great advancement that ICIs represent, most patients with gastrointestinal tumors have not benefited from this therapy. In addition, ICIs often induce adverse effects that are related to their systemic use. Local administration of ICIs in tumors could concentrate their effect in the malignant tissue and provide a higher safety profile. A new and attractive approach for local delivery of ICIs is the use of gene therapy vectors to express these blocking antibodies in tumor cells. Several vectors have been evaluated in preclinical models of gastrointestinal tumors to express ICIs against PD-1, PD-L1, and CTLA-4, among other immune checkpoints, with promising results. Vectors used in these settings include oncolytic viruses, self-replicating RNA vectors, and non-replicative viral and non-viral vectors. The use of viral vectors, especially when they have replication capacity, provides an additional adjuvant effect that has been shown to enhance antitumor responses. This review covers the most recent studies involving the use of gene therapy vectors to deliver ICIs to gastrointestinal tumors.
A promising therapy for patients with B-cell lymphoma is based on vaccination with idiotype monoclonal antibodies (mAbs). Since idiotypes are different in each tumor, a personalized vaccine has to be ...produced for each patient. Expression of immunoglobulins with appropriate post-translational modifications for human use often requires the use of stable mammalian cells that can be scaled-up to reach the desired level of production. We have used a noncytopathic self-amplifying RNA vector derived from Semliki Forest virus (ncSFV) to generate BHK cell lines expressing murine follicular lymphoma-derived idiotype A20 mAb. ncSFV/BHK cell lines expressed approximately 2 mg/L/24 h of A20 mAb with proper quaternary structure and a glycosylation pattern similar to that of A20 mAb produced by hybridoma cells. A20 mAb purified from the supernatant of a ncSFV cell line, or from the hybridoma, was conjugated to keyhole limpet hemocyanin and used to immunize Balb/c mice by administration of four weekly doses of 25 µg of mAb. Both idiotype mAbs were able to induce a similar antitumor protection and longer survival compared to non-immunized mice. These results indicate that the ncSFV RNA vector could represent a quick and efficient system to produce patient-specific idiotypes with potential application as lymphoma vaccines.
Nanobodies are camelid-derived single-domain antibodies that present some advantages versus conventional antibodies, such as a smaller size, and higher tissue penetrability, stability, and ...hydrophilicity. Although nanobodies can be delivered as proteins, in vivo expression from adeno-associated viral (AAV) vectors represents an attractive strategy. This is due to the fact that AAV vectors, that can provide long-term expression of recombinant genes, have shown an excellent safety profile, and can accommodate genes for one or several nanobodies. In fact, several studies showed that AAV vectors can provide sustained nanobody expression both locally or systemically in preclinical models of human diseases. Some of the pathologies addressed with this technology include cancer, neurological, cardiovascular, infectious, and genetic diseases. Depending on the indication, AAV-delivered nanobodies can be expressed extracellularly or inside cells. Intracellular nanobodies or “intrabodies” carry out their function by interacting with cell proteins involved in disease and have also been designed to help elucidate cellular mechanisms by interfering with normal cell processes. Finally, nanobodies can also be used to retarget AAV vectors, when tethered to viral capsid proteins. This review covers applications in which AAV vectors have been used to deliver nanobodies, with a focus on their therapeutic use.
SARS-CoV-2 virus, the causative agent of COVID-19, has produced the largest pandemic in the 21st century, becoming a very serious health problem worldwide. To prevent COVID-19 disease and infection, ...a large number of vaccines have been developed and approved in record time, including new vaccines based on mRNA encapsulated in lipid nanoparticles. While mRNA-based vaccines have proven to be safe and effective, they are more expensive to produce compared to conventional vaccines. A special type of mRNA vaccine is based on self-amplifying RNA (saRNA) derived from the genome of RNA viruses, mainly alphaviruses. These saRNAs encode a viral replicase in addition to the antigen, usually the SARS-CoV-2 spike protein. The replicase can amplify the saRNA in transfected cells, potentially reducing the amount of RNA needed for vaccination and promoting interferon I responses that can enhance adaptive immunity. Preclinical studies with saRNA-based COVID-19 vaccines in diverse animal models have demonstrated the induction of robust protective immune responses, similar to conventional mRNA but at lower doses. Initial clinical trials have confirmed the safety and immunogenicity of saRNA-based vaccines in individuals that had previously received authorized COVID-19 vaccines. These findings have led to the recent approval of two of these vaccines by the national drug agencies of India and Japan, underscoring the promising potential of this technology.
Alphavirus vectors based on self-amplifying RNA (saRNA) generate high and transient levels of transgene expression and induce innate immune responses, making them an interesting tool for antitumor ...therapy. These vectors are usually delivered as viral particles, but it is also possible to administer them as RNA. We evaluated this possibility by in vivo electroporation of Semliki Forest virus (SFV) saRNA for local treatment of murine colorectal MC38 subcutaneous tumors. Optimization of saRNA electroporation conditions in tumors was performed using an SFV vector coding for luciferase. Then we evaluated the therapeutic potential of this approach using an SFV saRNA coding for interleukin-12 (SFV-IL-12), a proinflammatory cytokine with potent antitumor effects. Delivery of SFV-IL-12 saRNA by electroporation led to improvement in tumor control and higher survival compared with mice treated with electroporation or with SFV-IL-12 saRNA alone. The antitumor efficacy of SFV-IL-12 saRNA electroporation increased by combination with systemic PD-1 blockade. This therapy, which was also validated in a hepatocellular carcinoma tumor model, suggests that local delivery of saRNA by electroporation could be an attractive strategy for cancer immunotherapy. This approach could have easy translation to the clinical practice, especially for percutaneously accessible tumors.
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Self-amplifying RNA (saRNA) can mediate high levels of transgene expression and induce innate immune responses. Here we demonstrate that it is possible to administer saRNA by electroporation in tumors, leading to potent therapeutic responses. This approach could have easy translation to the clinic, especially for percutaneously accessible tumors.
Antiviral agents with different mechanisms of action could induce synergistic effects against SARS-CoV-2 infection. Some reports suggest the therapeutic potential of the heme oxygenase-1 (HO-1) ...enzyme against virus infection. Given that hemin is a natural inducer of the HO-1 gene, the aim of this study was to develop an in vitro assay to analyze the antiviral potency of hemin against SARS-CoV-2 infection. A SARS-CoV-2 infectivity assay was conducted in Vero-E6 and Calu-3 epithelial cell lines. The antiviral effect of hemin, and chloroquine as a control, against SARS-CoV-2 virus infection was quantified by RT-qPCR using specific oligonucleotides for the N gene. Chloroquine induced a marked reduction of viral genome copies in kidney epithelial Vero-E6 cells but not in lung cancer Calu-3 cells. Hemin administration to the culture medium induced a high induction in the expression of the HO-1 gene that was stronger in Vero-E6 macaque-derived cells than in the human Calu-3 cell line. However, hemin treatment did not modify SARS-CoV-2 replication, as measured by viral genome quantification 48 h post-infection for Vero-E6 and 72 h post-infection for the Calu-3 lineages. In conclusion, although exposure to hemin induced strong HO-1 up-regulation, this effect was unable to inhibit or delay the progression of SARS-CoV-2 infection in two epithelial cell lines susceptible to infection.
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•Antiviral potency of hemin against SARS-CoV-2 infection was assayed in vitro.•Hemin induced Heme Oxygenase-1 expression in Vero-E6 and Calu-3 epithelial cells.•Anti-inflammatory/oxidant activities of hemin products may reduce viral replication.•Hemin showed no antiviral effect against SARS-CoV-2 infection in cells.
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