# Integration of _Salmonella_ into Combination Cancer Therapy
## 4 Clinical Application of _Salmonella_ in Combination Cancer Therapy  
### Clinical Trials  
Several genera of bacteria have been studied in pre-clinical settings, and a few bacterial platforms have been selected for testing in human patients, namely _Listeria monocytogenes_, _Clostridium novyi_, and _S. typhimurium_. These bacterial species have received the most attention due to the increased understanding of their physiology, pathogenicity, and genetics which, in turn, led to the development of attenuation strategies critical for safeadministration of live bacteria in humans. Among bacterial species tested in clinical settings, strains of _Listeria monocytogenes_ have shown encouraging and promising results [21]. Besides, clinical signs of tumor colonization and objective evidence of tumor responses have been observed following the intravenous and intratumoral administration of _Clostridium novoi_-NT spores [165; 166; 167]. The observed tumor destruction and absence of viable tumor cells were attributed to gas pockets produced by _Clostridium novoi_-NT. However, treatment with oncolytic bacteria alone failed to eradicate all cancer cells, and thereby led to tumor progression or recurrence [168; 166]. The first _Salmonella_ strain studied in human clinical trials was _S. typhimurium_ VNP20009, and was tested against metastatic melanoma and metastatic renal carcinoma. The study revealed that the maximum tolerated dose was 3.0 \(\times\) 10\({}^{8}\) CFU/m\({}^{2}\) injected through the intravenous route. Despite the observed tumor colonization and increased pro-inflammatory cytokines in some of the treated patients, no objective tumor regression was reported [169]. Another clinical trial utilized _S. typhimurium_ VNP20009 to treat metastatic melanoma patients. Consistent with previous findings, no tumor regression was documented among treated patients [170]. In order to improve therapeutic efficacy, VNP20009 was engineered to express the _E coli_ CD enzyme, which converts the non-toxic 5-FC into anti-tumor 5-FU, and was tested in three patients with head and neck squamous carcinoma and esophageal adenocarcinoma through intratumoral injection. Tumor colonization was observed in two patients for at least 15 days post administration, with a 3:1 tumor-to-plasma ratio of 5-FU; this ratio was <1.0 in the non-colonized patient. No adverse effects were reported following the treatment [171]. Later trials utilized the oral route for _Salmonella_ administration. _S. typhimurium_ Ty2 Ia that expresses VEGFR2 (VXM01 vaccine) has been used against advanced pancreatic cancer. Although the engineered bacteria succeeded in enhancing VEGFR2-specific T cell effector responses and reducing tumor perfusion, adverse events such as neutrophil, lymphoma, and diarrhea were observed in treated subjects [172]. SalpIL2 (_S. typhimurium_ that expresses human IL-2) showed no significant benefits in phase I clinical study against metastatic gastrointestinal cancer. On the other hand, an increase in circulating NK cells and NK-T cells was reported among treated patients [152], which provides insights into the possibility of combining _Salmonella_ strains with other approaches of cancer immunotherapy. **Table** 2 summarizes previous and ongoing clinical trials using _Salmonella_-based cancer therapy. The results of clinical trials highlighted the discrepancy in the therapeutic outcome between pre-clinical and clinical models. This could be related to differences in tumor contextures and growth rates that might alter bacterial invasiveness and proliferation within tumor tissues [173]. Others attributed the inefficiency of BMCT in clinical trials to the over-attenuation of the utilized bacteria which, in turn, compromises its anti-tumor effects [174].