, a crucial and initial step in the process of immunity, and lead to an upregulation of programmed death-ligand 1 PD–L1 on DCs further suppressing T cell function ^[26][81]. Leaky blood vessels increase the intra-tumoral pressure and together with the down-regulation of cell-adhesion molecules complicates extravasation of tumor infiltrating lymphocytes (TILs). Hypoxia itself lead to upregulation of PD–L1 and the simultaneous up-regulation of VEGF-A which furthermore impairs the function of the antigen presenting cells. Finally, the balance of TILs shifts towards increased infiltration of regulatory T cells (Treg) on behalf of cytotoxic effector cells (CD8⁺) due to regulatory changes in the ECs ^[27][82]. This is primarily due to an increased expression of FAS ligand on tumor-associated ECs that prevents effector T cells from crossing the EC barrier by inducing apoptosis. Treg are resistant to FAS ligand creating a relative overexpression of Treg in the tumor compared to the effector cells. Treg furthermore inhibit the antigen presenting cells in the tumor enhancing the immune suppressive environment. Tumor vasculature normalization thus creates an immune friendly microenvironment and may turn a “cold” tumor into a “warm”. Added to this is the recent discovery of how stimulated immune cells themselves lead to vascular normalization, partly through CD8⁺ T–cells and interferon gamma (IFN–Ƴ), creating a beneficial immune-vasculature crosstalk and a rationale for combining these two classes of therapeutics with a potential synergistic benefit ^[28][83]. The optimal dosing and timing of these treatment combinations will likely differ between individual tumor types and represent an essential key for unlocking their full potential.

2.2. Anti-angiogenetic Therapy and Immunotherapy in Colorectal Cancer

This potential synergism between anti-angiogenic and immune checkpoint inhibitor drugs has resulted in numerous clinical trials testing the combination of PD–1/PD–L1 antibodies with anti–VEGF drugs. By now, a number of randomized trials have shown remarkable results which have led to approvals by the FDA and/or EMA in renal cell carcinoma (axitinib plus pembrolizumab, and axitinib plus avelumab), endometrial carcinoma (pembrolizumab plus lenvatinib), non-squamous NSCLC (bevacizumab and atezolizumab), and hepatocellular carcinoma (bevacizumab plus atezolizumab). This broad clinical activity suggest that a combination strategy may also be of benefit in colorectal cancer ^[29][84].

The first clinical data in CRC were presented by Bendell et al. at the ASCO–GI conference in 2015 ^[30][85]. Among 13 patients with treatment refractory disease, they demonstrated one objective tumor response by adding bevacizumab to MPDL3280A (atezolizumab). The following year, at the AACR 107th annual meeting, Wallin et al. presented the results from 23 patients with mCRC treated with first-line FOLFOX, bevacizumab, and atezolizumab ^[31][86]. They demonstrated promising efficacy data, with a median PFS of 14.1 months and parallel translational research argued for immune-related activity by this combination. The corresponding papers to these two initial abstracts are so far not published.

In 2017, Yoshida et al. published the results of their pilot study (COMVI study) in anticancer research ^[32][87]. Six patients with previously untreated mCRC were included in a prospective single arm study. All patients received, as standard therapy, oxaliplatin (130 mg/m²) on day one, capecitabine (1000 mg/m²) twice daily on the days 1–14, and bevacizumab (7.5 mg/kg) on day one. To this backbone, they added cultured αβ T–lymphocytes (>5 × 10⁹) combined with interleukin–2 and anti–CD3 on day 18. Two patients achieved a complete response, three a partial response, and one demonstrated stable disease as the best outcome. The median progression free and overall survival was 567 and 966 days, respectively. Adverse events were mild to moderate. Although a small pilot study, these published results, for the first time in patients with CRC, demonstrated that combining chemotherapy and anti-angiogenesis with immune-modulating therapies were feasible and efficacy data were promising.

An additional two abstracts were presented the following years but they did not quite meet the initial expectations. In 2018, at the ESMO congress, Grothey et al. presented a late-breaking abstract from cohort 2 of the MODUL trial ^[33][88]. After induction therapy with the FOLFOX + bevacizumab regimen, 445 patients were randomized to maintenance fluoropyrimidine and bevacizumab ± atezolizumab. The updated analyses revealed no difference in median PFS and OS between the two strategies. Mettu et al. presented an abstract at the poster discussion session at the same congress the following year ^[34][89]. In this study, 133 patients with mCRC, with treatment resistant disease, were randomized to receive capecitabine and bevacizumab plus placebo or atezolizumab as last line treatment. The study reached its primary endpoint demonstrating a significant improvement of PFS by the addition of atezolizumab, although the numerical difference was only one month. The corresponding manuscripts have not been published either.

The first publication based on a commercially available immuno-therapeutic, within this specific field in CRC, was published earlier this year, in April 2020, and revealed the results from the dose expansion phase Ib trial REGONIVO (EPOC1603) ^[35][90]. Fukuoka et al. included patients with gastric or CRC, 25 of each, who had progressed on a minimum of two previous lines of palliative treatment. All the patients with CRC had previously received anti-angiogenetic treatment, the cancer in one patient had deficient mismatch repair (dMMR) but the remaining 24 were all proficient (microsatellite stable), and six had RAS mutations. Patients were treated with nivolumab 3 mg/kg every two weeks and regorafenib once daily, day 1–21, in a four-week cycle. During the dosing-finding, part of the study regorafenib was reduced from initially 160 mg to the recommended 80 mg at which no patients experienced dose-limiting toxicity. Among the patients with CRC 9 (36%) achieved an objective tumor response and median PFS was 7.9 months. A trend towards better outcome for the patients with lung metastases, compared to liver metastases was presented, which could be due to a more immunosuppressive environment in the liver compared to the lung. This study provided real clinical evidence of synergy between the investigated drugs. Neither of the two would be expected to provide meaningful benefit as singe agents in this group (except the one patient with a dMMR tumor) but combined, one out of three responded.

These results, together with similar findings in other cancer types, have paved the way for multiple trials assessing the clinical benefit from combining immunotherapy and anti-angiogenetic treatment in CRC. An example is the recently published study protocol AtezoTRIBE by Antoniotti et al. ^[36][91]. In this randomized phase II trial untreated patients with unresectable mCRC will receive FOLFOXIRI and bevacizumab ± atezolizumab in four months followed by maintenance 5–fluoruracil, leucovorin, bevacizumab ± atezolizumab. The study is estimated to be completed in April 2021. A supplementary search at clinical.trials.gov for trials in CRC combining immunotherapy with an anti-angiogenetic drug revealed more than 20 ongoing clinical trials (Table 2Table 6). With a specific focus on CRC only, this level of clinical activity underlines the potential impact gained by combining these two classes of therapeutics.

2.1. Tumor-Microenvironment

Tumor-associated blood vessels have a unique architecture and physiologic properties as previously addressed. These properties lead to the generation of an immune suppressive tumor microenvironment ^[24][79]. Malignant tumors have the ability to evade immune suppression through changes in the recruitment, trafficking and infiltration of effector T cells and their final recognition and killing of cancer cells. This is the consequence of several processes related to tumor-initiated angiogenesis. The initial up-regulation of VEGF-A inhibit the maturation of dendritic cells (DCs) ^[25][80]