## 4 Discussion
The different affected pathways and targets are represented in Figure 3.  
Treatment based on the synergistic combination of drug and gene silencing via siRNA offered a novel and promising therapeutic strategy for gastrointestinal cancers. This systematic review has elucidated its great potential, observing a wide variety of drug-siRNA and nanoplatform combinations with very promising results both in vitro and in vivo. These results were seen in the reduction in cell viability and proliferation, increased rate of apoptosis, decreased cell migration and increased chemosensitivity compared to the administration of the free drugs. Also, these therapies achieved a considerable reduction of tumor xenograft volume and weight in murine models, a reduction in angiogenesis in the introduced tumor tissue and an increase in survival. In the different studies, the authors used a wide range of different cancer cell lines obtained from different animal models, as many of them naturally generate an insensitivity to certain drugs or artificially acquire resistance to further studies, such as HepG2-SI cells insensitive to sorafenibib [45]. Also, a vast variety of in vitro and in vivo techniques were used to analyze different parameters and characteristics, e.g., immunohistochemistry and hematoxylin-eosin staining, western blot, qPCR, cell viability (calculating the IC\({}_{50}\) cytotoxicity), apoptosis detection by flow cytometer and TUNEL staining, among others. In the in vivo assays, the authors analyzed in vivo biodistribution and the antitumor effect, i.e., the accumulation of the nanoparticles in the tumor. They achieved good in vitro and in vivo results and were represented in most studies. This also showed its great applicability. All the analyzed studies were focused on a preclinical level of treatment, using murine models as in vivo elements. The next step for the development of these promising strategies is to reach the clinical phase to ensure their maximum applicability in humans and to have a real therapeutic use, not only as an antitumor element but also as a translational medical tool for a multitude of diseases. Figure 3: Different siRNA strategies and affected cellular pathways within their cancerous effects were employed among all the studies analyzed. This approach should be applicable at all levels and customizable, without patient rejection and with few side effects. Currently, there are many preclinical trials focused on the combination of gene therapy, chemotherapeutic, and nanoparticles, but they have not yet reached the level of clinical trials in humans in most cases. Therefore, few therapies containing these three elements have been approved by any regulatory agency. However, gene therapy or chemotherapeutic alone in nanoparticles, have reached the clinical phase and in many cases have been approved by the FDA or EMA. These treatments aim to overcome potential low tumor localization, low stability, and rapid elimination from the bloodstream. They show promising antitumor activities and adequate safety profiles when used individually. For example, notable siRNA anticancer naturenotherapeutics include CALAA-01 for solid tumors targeting RRM2, selectively targeting transferrin receptors on the surface of tumor cells, with a completed phase I clinical trial in 2012 (NCT00689065); or DCR-MYC against hepatocellular carcinoma targeting c-Myc to inhibit cell proliferation and growth, with completed phase II (NCT02314052), among others. And as an exceptional case, we could find the use of Atu027, a novel liposomal RNA interference therapeutic targeting PKN3, to inhibit angiogenesis and tumor invasion, with completed phase Ib/IIa in 2016 combined with gemcitabine (NCT01808638) against advanced or metastatic pancreatic adenocarcinoma. The study employed two cohorts, with non-pancreatic cancer and with advanced pancreatic cancer, as one of the few studies that combined gene therapy and chemotherapy that reached the clinical phase [13; 71; 72].