br OshF m and shF m
3.6. OshF(m) and shF(m) have similar anti-tumor activity, but OshF (m) has a stronger ability to alter the tumor microenvironment
To facilitate future clinical research and improve vaccine expres-sion in vivo, we optimized the nucleotide sequence of shF(m) and reduced the sequence homology for the monitoring of the vaccine distribution in vivo (Fig. 6A). The newly optimized shF(m) was called OshF(m). The Fosfomycin calcium of FAPa in the OshF(m) group was signifi-cantly higher than that of shF(m), as verified by western blotting (Fig. 6B). The results of prophylactic experiment showed that OshF
(m) had better effect than shF(m) in inhibiting the growth of tumors (Fig. S1C, P < 0.05). To compare OshF(m) and shF(m) with respect to the anti-tumor effect and regulatory ability in the TME, in a thera-peutic setting, we set up three groups of mice (n = 8), i.e., a Vec
group, OshF(m) group, and shF (m) group. Mice were immunized once a week on the seventh day after tumor challenge (2 104 4T1 cells) and were immunized three times (Fig. 6C). Unlike the results of prophylactic experiment, there was no significant differ-ence between OshF(m) and shF(m) in the inhibition of tumor growth and induction of the anti-tumor immune response (Fig. 6D, F, and G). The same three groups of mice (n = 12) were vaccinated according to the therapeutic settings to evaluate survival time. OshF(m) and shF
(m) prolonged the survival of mice to a certain extent compared with that of the control group, but there was still no significant difference between the two groups. However, the relative expression levels of FAPa, SDF-1, and collagen I, which are indicators of the level of CAFs, in the OshF(m) group were significantly lower than those in the shF
(m) group (Fig. 6H, I, and J, P < 0.05). Flow cytometry showed that the infiltrated MDSCs in the OshF(m) and shF(m) groups were dramati-cally reduced (Fig. 6K, P < 0.01). In addition, the application of the vaccines did not affect the weight growth of mice (Fig. S2). Thus, OshF(m) and shF(m) had similar effects on tumor suppression and survival in mice in a therapeutic setting, but OshF(m) had stronger ability to inhibit tumor growth in a prophylactic setting.
Recent advances in cancer immunotherapy, such as the devel-opment of immune checkpoint inhibitors and chimeric antigen
receptor T cell therapies, have pushed cancer therapy into a new era [38–41]. However, a relatively small fraction of patients benefit from these approaches due to an immunosuppressed tumor microenvironment, which provides nutrients for tumor growth and suppresses the immune system [1–3,7]. As a major part of the TME, CAFs have a prominent role in the growth, progression, and metastasis of tumors by producing soluble factors that modu-late the ECM [9–11]. In addition, their genetic stability makes CAFs an attractive target for cancer immunotherapy [17,18].
Several studies, including our previous work, have verified that cancer vaccines targeting FAPa, which is expressed on CAFs, atten-uate tumor growth by inducing CD8+ T cell infiltration, which ulti-mately reduce the number of CAFs and eliminates immunosuppressive components in the TME [10,26,32,35,36,42]. In addition, more than 90% of epitheliomas, such as breast cancer, colorectal cancer, pancreatic cancer, and lung cancer, express FAPa [21,22], and high expression levels of FAPa in tumor stroma are associated with aggressive progression, metastasis, and recurrence in many different cancer types [43–45], suggesting that FAPa is a promising treatment target for various cancers. In the studies of immunotherapies targeting FAPa, the anti-tumor effects induced by cancer vaccines were detected in prophylactic settings or ther-apeutic settings, in which vaccines were administered to mice only a few days after tumor inoculation [10,26,32,35,36,42]. To verify