Recently, the interest in using nucleic acids for therapeutic applications has been increasing. DNA molecules can be manipulated to express a gene of interest for gene therapy applications or vaccine development. Plasmid DNA can be developed to treat different diseases, such as infections and cancer. In most cancers, the immune system is limited or suppressed, allowing cancer cells to grow. DNA vaccination has demonstrated its capacity to stimulate the immune system to fight against cancer cells. Furthermore, plasmids for cancer gene therapy can direct the expression of proteins with different functions, such as enzymes, toxins, and cytotoxic or proapoptotic proteins, to directly kill cancer cells. The progress and promising results reported in animal models in recent years have led to interesting clinical results. These DNA strategies are expected to be approved for cancer treatment in the near future. This review discusses the main strategies, challenges, and future perspectives of using plasmid DNA for cancer treatment.

The ability of tumor cells to evade the immune system is one of the main challenges we confront in the fight against cancer. Multiple strategies have been developed to counteract this situation, including the use of immunostimulant molecules that play a key role in the anti-tumor immune response. Such a response needs to be tumor-specific to cause as little damage as possible to healthy cells and also to track and eliminate disseminated tumor cells. Therefore, the combination of immunostimulant molecules and tumor-associated antigens has been implemented as an anti-tumor therapy strategy to eliminate the main obstacles confronted in conventional therapies. The immunostimulant 4-1BBL belongs to the tumor necrosis factor (TNF) family and it has been widely reported as the most effective member for activating lymphocytes. Hence, we will review the molecular, pre-clinical, and clinical applications in conjunction with tumor-associated antigens in antitumor immunotherapy, as well as the main molecular pathways involved in this association.

Human papillomaviruses (HPVs) are responsible for about 25% of cancer cases worldwide. HPV-16 E7 antigen is a tumor-associated antigen (TAA) commonly expressed in HPV-induced tumors; however, it has low immunogenicity. The interaction of 4-1BBL with its receptor induces pleiotropic effects on innate, adaptive, and regulatory immunity and, if fused to TAAs in DNA vaccines, can improve the antitumor response; however, there is low transfection and antitumor efficiency. Oncolytic virotherapy is promising for antitumor gene therapy as it can be selectively replicated in tumor cells, inducing cell lysis, and furthermore, tumor cell debris can be taken in by immune cells to potentiate antitumor responses. In this study, we expressed the immunomodulatory molecule SA-4-1BBL fused to E7 on an oncolytic adenovirus (OAd) system. In vitro infection of TC-1 tumor cells and NIH-3T3 non-tumor cells with SA/E7/4-1BBL OAd demonstrated that only tumor cells are selectively destroyed. Moreover, protein expression is targeted to the endoplasmic reticulum in both cell lines when a signal peptide (SP) is added. Finally, in an HPV-induced cancer murine model, the therapeutic oncolytic activity of OAd can be detected, and this can be improved when fused to E7 and SP.

Caloric restriction (CR) and its related alternatives have been shown to be the only interventions capable of extending lifespan and decreasing the risk of cancer, along with a reduction in burden in pre-clinical trials. Nevertheless, the results from clinical trials have not been as conclusive as the pre-clinical results. Recognizing the challenges associated with long-term fasting, the application of caloric restriction mimetics (CRMs), pharmacological agents that mimic the molecular effects of CR, to harness the potential benefits while overcoming the practical limitations of fasting has resulted in an interesting alternative. This review synthesizes the findings of diverse clinical trials evaluating the safety and efficacy of CR and CRMs. In dietary interventions, a fast-mimicking diet was the most tolerated to reduce tumoral growth markers and chemotherapy side effects. CRMs were well tolerated, and metformin and aspirin showed the most promising effect in reducing cancer risk in a selected group of patients. The application of CR and/or CRMs shows promising effects in anti-cancer therapy; however, there is a need for more evidence to safely include these interventions in standard-of-care therapies.

The endoplasmic reticulum (ER) is a key organelle in cell homeostasis and cell health through antigen presentation to immune cells. Thus, the ER has become a therapeutic target to induce cellular immune responses. We previously reported the antitumor effect of a DNA vaccine that expresses the E7 antigen fused to the cyclooxygenase-2 (COX-2) protein. This inflammation-related enzyme contains a degradation cassette associated with the endoplasmic reticulum-associated degradation (ERAD) pathway. To avoid the use of full-length COX-2 and any risk of adverse effects due to the activity of its catalytic site, we designed new versions of the fusion protein. These new constructs encode the E7 antigen fused to the signal peptide and the ERAD sequence of COX-2 with or without the membrane-binding domain (MBD) as well as deletion of the catalytic site. We evaluated the antigen-specific antitumor effect of these DNA constructs in murine prophylactic and therapeutic cancer models. These assays showed that the ERAD cassette is the minimum sequence in the COX-2 protein that induces an antitumor effect when fused to the E7 antigen with the advantage of eliminating any potential adverse effects from the use of full-length COX-2.

Hair graying, a prototypical sign of human aging, is a progressive loss of pigmentation from growing hair shafts caused by disease and as a side effect of medications. Cerebrolysin is a neuropeptide preparation that mimics the effect of endogenous neurotrophic factors. Cerebrolysin has been widely used in neurologic conditions, such as cerebral stroke, Alzheimer’s disease, and dementia, among others. Cerebrolysin treatment has achieved to regain or maintain the cognitive ability of affected patients; however, up to date, there are no reports about the reactivation of hair pigmentation. We describe a previously not described effect occurring on patients receiving Cerebrolysin treatment for neurologic diseases and whether this effect is associated in reactivation of melanocytes and melanin expression. Here, we report five patients (mean age, 70.6 years), who also had age-related hair graying and scalp hair repigmentation during Cerebrolysin treatment. Macroscopic analysis revealed hair repigmentation consisted in diffuse darkening of the scalp hair. Impregnation and immunostaining analysis were performed on scalp biopsies taken before and after Cerebrolysin treatment; the results showed greater melanin and melanocyte marker MART-1/Melan-A staining following Cerebrolysin treatment. We present, to our knowledge, the first report on hair repigmentation is a previously not described effect occurring following Cerebrolysin treatment.