Hepatocellular carcinoma is a prevalent malignant tumor of the digestive system and is one of the leading causes of cancer-related deaths. According to global cancer statistics for 2020, it ranks 6th in incidence and 3rd in tumor-related mortality worldwide, and the situation in China is even worse (1, 2). Less than 10% of people with hepatocellular carcinoma survive the disease for five years. The disease develops quietly and around half of the patients are already in a middle to late stage when they are first diagnosed. Macrovascular invasion is seen on imaging, and the prognosis is exceedingly dismal, with an expected survival of about 2 to 5 months (3, 4).

Currently, multi-target tyrosine kinase inhibitors (TKIs) such as sorafenib, Lenvatinib, and Donafenib as first-line targeted therapeutic agents for advanced hepatocellular carcinoma have been shown to slow tumor progression and extend survival time. However, the ORR of sorafenib is only 2%, and Lenvatinib was non-inferior to sorafenib in terms of OS (13.6 vs. 12.3 months) in REFLECT clinical trial. Among them, Lenvatinib had the highest ORR of 24.1% (5–7). The primary study endpoint of survival time was not met with Nivolumab in phase III clinical trial CheckMate-459 (8). Similarly, KEYNOTE-240, a phase III study of Pembrolizumab for second-line treatment of advanced hepatocellular carcinoma, failed to demonstrate superiority of anti-PD-1 therapy in comparison with routine therapy (9). Though monotherapy of Immune checkpoint inhibitors (ICIs) obtains higher ORR compared to Sorafenib, but the clinical efficacy and survival outcomes of patients with uHCC are still unsatisfied.

At present, ICIs in combination with anti-angiogenic therapies are currently a popular direction in the systemic therapy of intermediate-advanced uHCC. In the IMBrave 150 phase III clinical trial, Atezolizumab combined with Bevacizumab outperformed the sorafenib group in uHCC patients in terms of OS and PFS (10). It was recommended by the FDA as a first-line treatment for untreated advanced hepatocellular carcinoma. Several clinical studies such as Pembrolizumab plus Lenvatinib, Sintilimab plus a bevacizumab biosimilar (IBI305), Camrelizumab plus Apatinib, have shown longer OS and PFS as well as higher ORR (11–13). Transcatheter artery chemoembolization (TACE) is one of the preferred locoregional treatments which is recommended by the guidelines for patients with uHCC (14). Necrosis of tumor tissue induced by TACE can release tumor antigens, which may promote tumor-specific immune responses (15). The synergistic anti-tumor effect of TACE combined with PD - (L) 1 inhibitor and TKI has been confirmed that the combinational therapy can significantly improve the survival and oncological benefit of Chinese patients compared with TACE monotherapy. However, TACE-induced embolic syndrome increases patient suffering and abnormal liver function are often seen in patients with intermediate to advanced HCC, which trigger us to think it over and seek for other effective local treatment options. The high-level radiobiologic effect of high-dose irradiation by stereotactic body radiotherapy (SBRT), which is significantly superior to conventional fractionated radiotherapy, can induce reactive oxygen species-related DNA damage, cause immunogenic death of tumor cells (16–18). Tumor cell death exposes a large number of tumor-specific antigens, which activate cross-presentation of antigen-presenting cells to effector T cells, evoking local immune responses and also inducing adaptive immune responses outside the radiation area and distant metastases to produce a “distant effect” (19). Meanwhile, it is now preferred that anti-VEGF plays a facilitative and synergistic role when patients receive ICIs.

The combination of multiple regimens has shown superior therapeutic effects and will be a popular research direction. We believe that after exposing the specific antigens of tumor cells via SBRT may lead to better efficacy of immunotherapy. The purpose of this article is to summarize the effectiveness and safety of the combination treatment regimen of Toripalimab combined with Anlotinib after SBRT in patients with unresectable hepatocellular carcinoma.

As far as we know, there are no many reports about SBRT followed by ICIs and TKIs in intermediate and advanced HCC. Both TACE and SBRT can directly kill tumor cells to reduce tumor burden. SBRT can trigger not only local immune responses but also adaptive immune responses outside the radiation area and distant metastases, which is called “distant effect”. A single dose of 8 Gy*3 irradiation regimens has been proven to have a better distant effect and improve the OS and PFS in the NSCLC patients (20). Meta-analysis (21) of 1580 Chinese patients with advanced hepatocellular carcinoma showed that in patients with advanced hepatocellular carcinoma combined with portal vein or inferior vena cava trunk thrombosis, TACE plus radiotherapy prolonged survival compared with TACE alone or TACE plus sorafenib, indicating that advanced hepatocellular carcinoma combined with thrombosis is sensitive to radiotherapy. Chiang et al. (22) reported 5cases with uHCC received SBRT then followed by Nivolumab, achieving 100% ORR, which demonstrated impressive tumor control from the combination of SBRT and checkpoint inhibitors. Studies have certified that SBRT or conventional radiotherapy combined with PD-1, either before or during treatment, could provide better survival outcomes than patients who do not (23). A retrospective analysis of a small sample showed that palliative radiotherapy combined with PD-1/PD-L1 and anti-angiogenic agents was safe and effective in patients with BCLC C with an ORR of 40% and prolonged survival time (24).

Compelling reports suggest that VEGF has immunosuppressive properties for tumors apart from activating angiogenesis-related processes. In the past, anti-VEGF antibodies were thought to act more favorably to chemotherapy, for example, by normalizing tumor vasculature, but nowadays their facilitative and synergistic role in immunotherapy is preferred. Strong correlation was found between the increase of tumor infiltrating lymphocytes (such as CD4+and CD8+T cells) and the normalization of blood vessels imposed by VEGF pathway inhibitors (25, 26). All above convince us that SBRT together with anti-angiogenic will Significantly enhance the efficacy of immunotherapy to achieve better anti-tumor effects.

Notably, patients were supplemented with sufficient doses of enteric probiotics to maintain the balance of gut microbiome which should be given adequate attention. Existing views suggest that immune responses triggered by microbial-related molecular patterns such as intestinal flora translocation and ecological dysbiosis with intestinal flora metabolism are the main mechanisms by which intestinal flora promote the development of hepatocellular carcinoma through the intestine-hepatic axis (27). Numerous studies have shown that regulating gut microbiome may be a safer, low-cost potential strategy for the prevention or treatment of liver cancer by means of improving the immunological response to ICIs and enhancing the anti-tumor effects (28, 29), and plays an important role in improving the prognosis of patients with intermediate-advanced HCC.

Numerous clinical trials have shown that combinational therapy with PD-1 inhibitor and TKI achieved remarkable effect, with higher tumor response and longer survival than monotherapy. Combination regimens of multiple treatments can complement the shortcomings of a single treatment and provide better outcomes. However, the side effects caused by the combinational therapies will increase the risk of treatment-related adverse events and the safety needs further investigation. In our cohort, all patients in our study showed good tolerance to our regimen except one patient died of acute upper gastrointestinal hemorrhage. Several randomized clinical trials suggested that patients treated with PD-1 inhibitor and TKI were able achieved PFS ranging from 4.6 to 8.6 months approximately. A majority of the cases included had a combination of vascular carcinoma thrombosis (80%) and intrahepatic metastases (75%) with advanced stage. Based on this premise, we achieved a PFS of 7.4 months, with a 2-year overall-survival rate and a progression-free survival rate of 59.9% and 29.7%, respectively.

The limitations of this study conclude as follows: small sample size, single-center, as well as the selection bias of cases during the epidemic of COVID-19. In addition, ICIs and TKIs are still expensive at present and patients need good financial support. Also, the follow-up period was short in some cases, and the different treatment doses due to different treatment cycles may influence the assessment of efficacy. All of the above may affect the scientific validity of our conclusion of this trial.

In conclusion, exposure of tumor antigens of HCC via SBRT may improve the efficacy of combinational therapy with Toripalimab and Anlotinib for uHCC with manageable adverse effects, which deserves further exploration.

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.

The studies involving human participants were reviewed and approved by China Ethics Committee of Registering Clinical Trials. The patients/participants provided their written informed consent to participate in this study.

Conception and design: YP, YC, XC, HH. (II) Provision of study materials or patients: YC, YP, WF, XZ, HL, WF. (III)Collection and assembly of data: HH, ZC, JY. (IV) Data analysis and interpretation: HH, YC, ZC, JY. (V) Manuscript writing: HH, YC. (VI) Final approval of manuscript: All authors. All authors contributed to the article and approved the submitted version.