Colorectal cancer (CRC) is third most prevalent malignant tumor and second most deadly cancer worldwide.¹ Economic growth and the diagnosis of CRC in younger may be attributed to hereditary, lifestyle, food, and environmental variables.¹ Even if screening programmes successfully minimise the death and morbidity from CRC,² between 20 and 35 percent of individuals with CRC have distant metastases at the time of diagnosis.^3,4 Five years of survival is around 13% for patients with metastatic CRC (mCRC).⁵ The National Comprehensive Cancer Network (NCCN) Guidelines for colon cancer promote molecular analysis, including oncogenic alterations for KRAS/NRAS/BRAF and mismatch repair (MMR) status, for mCRC before beginning a treatment.⁶ Only 4%–8% of mCRC have BRAF mutation V600E⁷ which is a punctiform substitution, from valine to glutamine in codon 600.⁸ The median overall survival (mOS) of BRAF V600E mutant mCRC is fewer than 15 months.⁹ The milestone of treatment for CRC is chemotherapy with or without targeted agents. The standard first-line chemotherapy for patients with BRAF mutations mCRC is fluorouracil, leucovorin, oxaliplatin, and irinotecan (FOLFOXIRI) with or without bevacizumab in patients fit or doublet of chemotherapy in association with bevacizumab, even if it was observed a poor effectiveness with progression free survival (PFS) and overall survival (OS) very low.¹⁰ Opposite to melanoma, patients with BRAF mutant CRC have not shown therapeutic benefit from BRAF inhibitors alone⁸ because the activation of alternate signalling pathways—such as PI3K/Akt,^11–14 and Wnt/β¹⁵—plays a driving role. In the last years many clinical trials with multi-target combination have been conducted. In previously treated BRAF mutant mCRC, combination targeted therapies involving BRAF inhibitor and Epidermal Growht factor receptore (EGFR) inhibitor produced positive results.^16–18 Moreover, a combined targeted therapy (encorafenib, binimetinib, and plus cetuximab) is being studied in the first line BRAF V600E mut. mCRC.¹⁹ Immune checkpoint inhibitors (ICIs) have been evaluated in CRC.²⁰ ICIs generate a substantial immune system activation and an effective anticancer immune response.²¹ The most used are monoclonal antibodies against cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) and programmed cell death 1 (PD-1), that have been approved for many cancer types in clinical practice.²² Studies already demonstrated that in mCRC with high microsatellite instability (MSI-H) which have a higher tumor mutation burden, they respond better to ICIs.²³ Advanced CRC commonly exhibits BRAF mutations associated to MSI-H/DNA deficient MMR (dMMR).²⁴ Thirty percent of BRAF mutant mCRC, are MSI-H. Treatment with ICIs has recently shown to be very helpful for achieving a long response in MSI-H/dMMR mCRC.²⁵ Instead, it is believed that ICIs monotherapy is not very effective in microsatellite instability (MSS) mCRC with BRAF mutation.²⁶ For this group of patients, ICIs are therefore being investigated in combination with other anticancer drugs, including BRAF inhibitors.²⁷ We address in this study the immunological environment of BRAF mutant CRC.

Using the terms colon, cancer, mutations, immunotherapy, BRAF, we searched PubMed (www.ncbi.nlm.nih.gov/pubmed) for full-text papers from 2017 to May 31, 2023.

Each of the found full-text articles was thoroughly read. We also looked over every abstract that was presented at international conferences between January 2020 and January 2024.

Two of the multiple different mechanisms that lead to CRC are the serrated and traditional adenoma-carcinoma pathways.²⁸ Many serrated premalignant lesions result from BRAF mutations, which show as an early event in the serrated pathway.^29,30 BRAF is essential to cell proliferation, differentiation, angiogenesis, apoptosis, and metastasis.³¹ Because BRAF kinase is constitutively phosphorylated when mutations occur, tumor growth is caused by the prolonged stimulation of MAPK pathway signalling.^11,32

Molecular characteristics and signalling pathways allowed the differentiation of three BRAF mutant types.³² The serrated pathway is closely related to class 1, which has the highest incidence but the worst prognosis among these. By contrast, class 3 CRCs are more likely to have a similar prognosis to wild-type CRCs.³³

Chromosomal instability (CIN), the CpG pathway of methylation phenotype (CIMP), and microsatellite instability (MSI) are at least three separate molecular pathways that drive CRC.^34,35 Advanced CRC is frequently associated with BRAF mutations, particularly the BRAF V600E mutation,^32,36,37 which may be linked to hypermethylation of the MLH1 promoter and the CpG island methylator phenotype.³⁸ The microsatellites belong to a large group of DNA motifs which have a length of 1–6 bp, harbouring high levels of sequence variation, common to all genomes.³⁹ Mostly observed at microsatellites, the status of dMMR causes many mutations or gene silencing.^40,41 Less than 10% of MSS CRC, include BRAF mutations.^24,37,42 Classification of BRAF mutated mCRC would be useful in predicting which treatment regimens to choose to improve patient outcomes.⁴³ Consensus molecular subtypes (CMS) state that BRAF mutant CRC mostly (up to 70%) belongs to CMS1 (MSI Immune), which has higher immune infiltration and improved OS.^44,45 Moreover, based on gene expression in CRC, subtypes of BRAF V600E mutation were proposed, independent of MSI status, PI3K mutation, gender, and sidedness. Through pathways including stimulation of IL2/STAT5, tumor necrosis factorα signalling, IL6/JAK/STAT3, and allograft rejection, BRAF V600E Mutation 1 (BM1) had an overall stronger immunological profile than BRAF V600E Mutation 2 (BM2). Better response rates, median progression-free survival (mPFS), and mOS were shown for BM1 in patients receiving EGFR inhibitors (dabrafenib, trametinib, and panitumumab) in addition to BRAF plus mitogen-activated protein kinase kinase (MEK).^46,47

MSI-H/dMMR subgroup

Recently there has been considerable progress in the use of ICIs in MSI-H CRC. Pembrolizumab, a PD-1 inhibitor, was tested in the phase II Keynote 164 trial (NCT02460198) in MSI-H/dMMR mCRC patients who had previously received systemic therapy. Pembrolizumab monotherapy showed response in five (55%) and one (25%) BRAF mutant patients in cohorts A (patients pre-treated with at least two standard lines of chemotherapy) and B (patients pre-treated with one line of systemic chemotherapy).⁴⁸ To treat MSI-H/dMMR mCRC, nivolumab, a PD-1 inhibitor, is being investigated in the phase II, multicenter, open-label CheckMate 142 trial (NCT02060188) either alone or in combination with Ipilimumab. Comparing the investigator-assessed objective response rate (ORR) of 31.1% (95% confidence interval [CI], 20.8%–42.9%) in 74 patients with MSI-H mCRC who had undergone at least one prior line of therapy the ORR, in BRAF mutated subgroup, was 25%.⁴⁹ Also, was studied in CheckMate 142, nivolumab with low-dose ipilimumab, a CTLA-4 inhibitor. The treatment gave an ORR of 65% (95% CI, 55%–73%) and a disease control rate (DCR) of 81% (95% CI, 72%–87%) in the group of previously treated patients.⁵⁰ In subgroups with BRAF V600E mutation, the ORR was 70%,⁵⁰ the safety profile was handleable. Complementing the previously reported findings, a meta-analysis revealed that there was no statistically significant difference in the ORR between BRAF wild-type mCRC and BRAF mutant mCRC with MSI-H/dMMR (odds ratio [OR] 1.04; 95% CI, 0.48–2.25).⁹ In the later-line therapy of MSI-H/dMMR mCRC with BRAF mutation, ICIs may offer a potential clinical response advantage. A significant response was also shown in the first-line cohort in CheckMate 142 by nivolumab in combination with a low-dose ipilimumab regimen. Patients with BRAF mutations clearly benefited from sustained response with a 24-month PFS rate of 76.5% and ORR (76% per investigator; 82% per blinded indipedent central review).⁵¹ Pembrolizumab monotherapy vs. chemotherapy in treatment-naïve MSI-H/dMMR mCRC patients is the focus of the phase III, multicenter, open-label, international trial Keynote 177.^25,52 While a lack of benefit in mOS between the study arms (not available [NA] vs. 36.7 months, respectively, pembrolizumab monotherapy vs. chemotherapy, hazard ratio [HR] 0.74; 95% CI, 0.53–1.03, P = 0.036) was found due to falling short of the preset statistical boundary, a final analysis reported a significant improvement in mPFS (16.5 months vs. 8.2 months, respectively, pembrolizumab monotherapy vs. chemotherapy, HR 0.59; 95% CI, 0.45–0.79, P = 0.0002). In a subgroup analysis in tumors characterised by BRAF V600E mutation, no OS benefit was seen (HR 0.72; 95% CI, 0.35–1.47) even though data analysis revealed a statistically significant benefit in PFS (HR 0.48; 95% CI, 0.27–0.86). With 60% of patients getting anti-PD-1 or anti-program death ligand 1 (PD-L1) therapy following first-line chemotherapy, the crossover design may be responsible for the lack of appreciable OS benefit. Moreover, the health-related quality of life improved and there were fewer adverse events with pembrolizumab monotherapy.^52,53 In the MSI-H/dMMR mCRC population, the clinical benefit of the BRAF mutant subgroup did not differ significantly from that of the non-BRAF mutation population, according to the scant clinical trial data that are now available, in first-line or later-line treatment. The effectiveness of ICIs with conventional therapies must be compared in more head-to-head trials. The BRAF V600E mutation was, however, recently linked to a deleterious impact on the ICIs response in MSI-H mCRC patients in a real-world retrospective investigation.⁵⁴ Patients with BRAF V600E mutation showed lower ORR (44.4% vs. 74.2% respectively; P = 0.120) but no statistically significant, and the 12- and 24-month PFS rates were much lower than those of BRAF wild-type patients (12-months: 40.0% vs. 73.3%, P < 0.001; 24-months: 26.7% vs. 73.3%, P < 0.001). Still, there were not many BRAF V600E individuals in this subgroup analysis, and the treatment timing was not clear. Large-scale trials in the subgroup with BRAF mutant mCRC are therefore still required to confirm the effectiveness of immunotherapy alone. A fraction of MSI-H/dMMR metastatic patients respond poorly to ICIs, which may be because of acquired or intrinsic resistance. There is still needed to investigate new biomarkers for screening appropriate populations and workable treatment plans. As such, combined therapy approaches have been suggested to overcome immunotherapy resistance. In the first-line treatment of MSI-H/dMMR mCRC with BRAF mutation, the open-label, multicenter SEAMARK study is already underway and compares the efficacy of pembrolizumab alone or with cetuximab (an EGFR inhibitor) and encorafenib (a BRAF inhibitor).

In conclusion, immunotherapy-treated BRAF-mutated MSI-H patients may also have a good prognosis and a long-lasting response, albeit some of them show treatment resistance.

MSS subgroup

While Keynote 016 showed a significant advancement in developing an immunotherapeutic approach for MSI-H/dMMR mCRC, treating MSS/pMMR mCRC with immune checkpoint inhibition alone showed less benefit.^5,26,55,56 Stated otherwise, intrinsic resistance by many mechanisms renders MSS/pMMR mCRC patients resistant to ICIs monotherapy.⁵⁷ Nevertheless, logical combinations based on ICIs still need to be investigated further for MSS/pMMR mCRC by choosing appropriate populations with predictive biomarkers and getting over intrinsic resistance, particularly for patients with BRAF mutation who have a bad prognosis and no efficient treatment plan.

Combining ICIs and anti VEGF

A step in growth of a lot of solid tumors, is angiogenesis; proangiogenic factors are markedly increased in the BRAF V600E mutant tissue.⁵⁸ Significantly involved in vasculogenesis and angiogenesis, vascular endothelial growth factor (VEGF) causes aberrant and leaky blood vessels in cancer.⁵⁹ Preclinical studies revealed that VEGF might directly mediate immune cells, including regulatory T cells, dendritic cells, tumor-associated macrophages (TAMs), and myeloid-derived suppressor cells (MDSCs), thereby exerting an immunosuppressive impact.^60,61 Hence, by normalising the tumor vasculature and changing the immunological milieu, blocking the VEGF signalling pathway may improve the effectiveness of ICIs.

A precedent for the combination therapy for MSS/pMMR mCRC was established with the REGONIVO trial, a phase Ib/II study, investigate the safety and efficacy of nivolumab in combination with multikinase inhibitor regorafenib (a VEGF receptor [VEGFR]2 inhibitor) in patients with MSS gastrointestinal cancers.⁶² In the mCRC cohort, 25 patients, the mPFS was 7.5 months and the ORR was 36%. Study data on patients with BRAF mutant mCRC are currently scarce, nonetheless. Preliminary findings from the NIVACOR trial, a phase II trial assessing the effectiveness of FOLFOXIRI in combination with bevacizumab and nivolumab in advanced CRC with a mutation in RAS/BRAF, were published.⁶³ The ORR in 52 MSS patients was 78.9%, and the median DOR was 7.59 (95% CI, 6.21–11.43) months. Within the MSS patient subgroup, the mPFS was 9.8 (95% CI, 8.18–15.24) months and the DCR was 96.2%. It is still unknown how the results from advanced CRC with BRAF mutations will turn out. In the meanwhile, a case study revealed that, with a combination therapy of nivolumab and VEGFR inhibitor (bevacizumab, an anti-angiogenic agent), an MSS/pMMR mCRC patient with BRAF V600E mutation had a PFS of more than 17 months.⁶⁴

ICIs with blockade of the MAPK signalling pathway

Growing research indicates that via altering the tumor microenvironment (TME) and anti-tumor immunomodulation, inhibiting the MAPK pathway may work together with immunotherapy.⁶⁵ Clinical improvement is obtained in BRAF mutant melanoma by ICI therapy combined with MAPK pathway inhibition.^66–69 Studies on the simultaneous inhibition of the immune checkpoint and MAPK pathway have also been conducted in mCRCs with BRAF mutations. A phase II trial assessed patients with BRAF V600E mutated mCRC treated with dabrafenib, a BRAF inhibitor, trametinib, a MEK inhibitor, and PDR001, also known as spartalizumab.^70,71 Seventy-three BRAF V600E mutated mCRCs were enrolled; the ORR and DCR were 24.3% and 70.3%, respectively. In 28 MSS mCRC patients who had never treatment with ICIs or BRAF inhibitors, the mPFS was 5.6 months, and the ORR and DCR were 25% (7/28) and 75% (21/28), respectively. Comparing the regimen of dabrafenib plus trametinib to the combination of the PD-1 inhibitor, a good effectiveness was shown.⁷² Compared to pre-treatment biopsies, T cell and other immune cell populations infiltrated tumor biopsies more after therapy.⁷⁰ In MSS, BRAF V600E mutant, unresectable or mCRC patients, the phase I/II trial was conducted for evaluate the dosage, safety, and efficacy of encorafenib (a BRAF inhibitor) combination with cetuximab (an EGFR inhibitor) and nivolumab.⁷³ Results indicated a strong efficacy with ORRs of 96% (95% CI, 78%–100%) and 48% (95% CI, 27%–69%). Of those who respond, the median response time is 7.7 months (95% CI, 4.5–NA). Noteworthy, mPFS and mOS were 7.4 months (95% CI, 7.7–NA) and 15.1 months (95% CI, 5.6–NA), respectively, demonstrating a better efficacy than targeted treatment. Regarding safety profile, patients did well with the combination; 19% (5/26) of them had grade 3 or 4 adverse events. Currently underway, a second randomised phase II trial investigated the benefits of adding nivolumab, in previously treated MSS mCRC patients with BRAF V600E mutation, with encorafenib and cetuximab (NCT05308446). Ultimately, the small clinical trial suggests that patients with BRAF-mutated MSS/pMMR mCRC may benefit from a combination of MAPK inhibition and ICIs. ICI-containing combination therapy has been tested in a number of studies recently in BRAF mutant mCRC (Table 1).

Table 1 . Clinical Trials Exploring Combination Therapies Based on ICIs in BRAF Mutant Metastatic Colorectal Cancer.

NCT number	Title	Phase	Status	Intervention	ICIs related arms	Condition
NCT05019534	A Phase I study on tolerance and safety of vemurafenib film-coated tablets, cetuximab solution for infusion and camrelizumab protocol (VCC) in the after line therapy of BRAF V600E mutation/MSS metastatic colorectal cancer	Phase I	Recruting	Drug: vemurafenib; cetuximab; camrelizumab	Single arm: vemurafenib, cetuximab combined with camrelizumab	Microsatellite stable, BRAF V600E-mutated, metastatic colorectal cancer
NCT05308446	Randomized Phase II trial of encorafenib and cetuximab with or without nivolumab (NSC #748,726) for patients with previously treated, microsatellite stable, BRAF V600E metastatic and/or unresectable colorectal cancer	Phase II	Recruting	Drug: cetuximab; encorafenib; nivolumab	Experimental (arm I): encorafenib, cetuximab, nivolumab; Active comparator (arm II): encorafenib, cetuximab	Microsatellite stable, BRAF V600E metastatic and/or unresectable colorectal cancer
NCT04044430	Phase I/II trial of encorafenib, binimetinib, and nivolumab in microsatellite stable BRAF V600E metastatic colorectal cancer	Phase I/II	Active, not recruting	Drug: binimetinib; encorafenib; nivolumab	Single arm: encorafenib, binimetinib, nivolumab	Microsatellite stable, BRAF V600E metastatic colorectal cancer
NCT04653480	Surufatinib and toripalimab combined with chemotherapy for second-line treatment of advanced RAS/BRAF mutant and microsatellite stable colorectal cancer	Phase II	Recruiting	Drug: surufatinib; toripalimab; chemotherapy	Single arm: surufatinib, toripalimab, and chemotherapy	Microsatellite stable, RAS/BRAF mutant advanced colorectal cancer
NCT04294160	A Phase Ib, multicenter, open-label dose escalation and expansion platform study of select drug combinations in adult patients with advanced or metastatic BRAF V600 colorectal cancer	Phase Ib	Recruiting	Drug: dabrafenib; LTT462; spartalizumab; tislelizumab	Backbone arm 1: dabrafenib + LTT462; triplet arm 4: dabrafenib + LTT462 + spartalizumab; triplet arm 6: dabrafenib + LTT462 + tislelizumab	Advanced or metastatic BRAF V600 colorectal cancer

NCT, National Clinical Trial; ICIs, immune checkpoint inhibitors; MSS, microsatellite instability..

Cells of the immune system

A dynamic system of cellular including immune cells, fibroblasts, stromal cells and extracellular matrix is known as TME, a crucial component that influences the effectiveness of immunotherapy.^74,75 The immune microenvironment of tumor (TIME) is composed of immune cells (CD8+T cells, CD4+T cells), MDSCs, anti-inflammatory macrophages, natural killer (NK) cells and associated non-cellular components and is essential to the development of tumors and their evasion of immune therapy.⁷⁶ Improved ICI efficacy in BRAF mutant mCRC patients depends on understanding the properties of TIME and managing the role of immunosuppressive variables in the TME.

Tumor-infiltrating lymphocytes (TILs)

TILs include CD8+ T cells, CD4+ helper T cells, regulatory T cells, B cells, and are essential for host immune response to tumor cells.^77,78 It is proposed that the density of TILs may perform as a marker to some degree for predicting prognosis in a range of malignancies and response to ICI therapy.^77,79 Unfortunately, there are currently no data on the predictive utility of TIL density in evaluating the effectiveness of immunotherapy in BRAF mutant CRC; yet a higher density of TILs was shown to be linked with a positive prognosis independent of BRAF mutation status.^80,81 We addressed the limited information and contradicting findings about the density of TILs in BRAF mutant CRC. BRAF mutation status was not significantly connected with the density of CD8+ intratumor cell-infiltrating lymphocytes in a study included 24 patients with BRAF mutant CRC (P = 0.090).⁸² In another work, CD45RO+ T cells were greater in 114 BRAF mutant CRCs (P = 0.0006), but the density of CD8+ and CD3+, as well as FOXP3+ T cells, was not significantly correlated with BRAF mutation.⁸³ TIL density in the periphery of tumor infiltration has been linked, in the context of anti-PD1 therapy, to an anti-PD1 response more closely than in those with central infiltration. Infiltrative margins of BRAF-mutated advanced colorectal tumors had a significantly higher density of FOXP3+ T cells (P < 0.001), but the centre of these tumors had lower densities of CD4+ and FOXP3+ T cells (P = 0.011 and P < 0.001, respectively).⁸⁴

A comprehensively assessment of immunological environment in CRC BRAF mutated, was conducted by Cen et al.⁸⁵ The expression of CD8+ T cells was considerably higher in BRAF mutant colon cancer patients than in wild-type ones (P < 0.001) in this study included 43 BRAF mutant colon cancer patients.⁸⁵ The fraction of CD8+ T cells was considerably higher in BRAF-mutated colon cancer tissues than in BRAF wild-type colon cancer tissues (P < 0.01) according to data from The Cancer Genome Atlas (TCGA) datasets, which included 59 BRAF-mutated and 337 BRAF wild-type colon cancer patients.⁸⁵ By contrast, there were no appreciable variations in the percentage of CD8+ T cells between groups according to the Gene Expression Omnibus (GEO) datasets (51 BRAF-mutated, 441 BRAF wild-type).⁸⁵ Furthermore, exhibiting this paradox are CD4+ T cells. The researchers further found that colon tumors harbouring a BRAF mutation had reduced tumor purity (P = 0.0003), a greater stromal score (P = 0.02), an immune score (P < 0.0001), and an Estimation of STromal and Immune cells in MAlignant Tumor tissues using Expression data (ESTIMATE) score (P = 0.0001). The TILs level may not be sufficient to completely characterise the complicated mechanism that is TIME of BRAF mutant CRC. Thus, this problem must be considered together with other elements like CMS classification and MSI status. Digiacomo et al⁸⁶ assessed the TILs in 22 MSI and 37 MSS BRAF-mutated mCRC and found variations in TILs between distinct subtypes of the disease. Results indicated that in MSI compared to MSS tumors, CD8+ and CD3+ lymphocytes, both intratumoral lymphatic invasion (ILI) and peritumoral lymphatic invasion (PLI), were more prevalent (CD8, P = 0.0001 and P < 0.0001; CD3, P = 0.003 and P = 0.0003; ILI and PLI, respectively). A greater CD3+ and CD8+ lymphocyte infiltration was found in class 2 BRAF-mutated mCRC cases than in class 3 cases (P = 0.033) by Schirripa et al.³³

Tumor-associated macrophages

TAMs are essential elements of TME that, by direct and indirect interactions with other immune cells, contribute to tumor immunosuppression, development, invasion, metastasis, angiogenesis, and drug tolerance.⁸⁷ While M2 macrophages are believed to influence immunosuppression and the course of cancer, M1 macrophages are pro-inflammatory and anti-tumor.⁸⁸ First identified as M2-like macrophages, TAMs were poor indicators for various cancer prognoses.⁸⁸ On BRAF mutant CRC, however, not many studies have been reported. One study found that whereas CD68+ M1 macrophages showed no difference (mean ± standard deviation, 18.43 ± 13.53 vs. 20.96 ± 15.34, respectively, P = 0.664), CD163+ M2 macrophages were significantly increased in BRAF V600E mutant CRC tumor compared to a wild-type tumor (mean ± 5.93 vs. 3.67 ± 3.02, respectively, P = 0.040).⁸⁹ M1 macrophages (P < 0.001, in both datasets) were higher in BRAF mutated patients than that in wild-type patients; no significant differences of M2 macrophages between BRAF mutant patients and BRAF wild-type patients (P > 0.05, in both datasets) were found in a second study combining 110 BRAF-mutated and 798 wild-type colon cancer patients from TCGA datasets (59 BRAF-mutated, 337 BRAF wild-type) and GEO datasets (51 BRAF-mutated, 441 BRAF wild-type).⁸⁵ High tumor infiltration CD68+ macrophages had no predictive value for BRAF mutant mCRC in another study of a subgroup of patients with unresectable mCRC based on BRAF status (data not shown).⁸⁸ In BRAF wild type mCRC with higher tumor infiltration CD68+ macrophages, however, a better mOS (high tumor infiltration CD68+ macrophages vs. low tumor infiltration CD68+ macrophages patients, 26 vs. 15 months, P = 0.002) was observed.⁸⁸ Explanation of the features of TAMs in this subgroup of CRC for prospective immunotherapy and prognosis still needs more research.

Expression of PD-L1

Transmembrane protein PD-L1, sometimes referred as CD274 or B7 homolog 1, is produced on cancer cells and, by binding to PD-1 in T cells, suppresses the immune system. Now well acknowledged, immune checkpoint blockade (ICB) against PD-L1 or PD-1 at used to treat non-small cell lung cancer, melanoma, esophageal cancer, breast cancer, renal cancer, and gastric cancer.^90,91 One of predictive indicators of response to ICIs is PD-L1 expression.⁹² Anti-PD-1/PD-L1 therapy is apparently ineffective for a significant number of CRC patients.⁹³ To the best of our knowledge, MSI-H is a biomarker to predict how well anti-PD-1/PD-L1 therapy will work in CRC.⁵⁵ Remarkably, MSI colon cancer had higher PD-L1 expression levels than the MSS cohort.^94,95 It is therefore now a crucial clinical topic if PD-L1 expression could function as another useful biomarker to predict the effectiveness of ICB therapy in CRC. Specifically, 9%–15% of CRC patients express PD-L1.⁹⁶

Poorly differentiated and solid/medullary histology, MSI-H or dMMR, and BRAF mutations signatures of the serrated neoplasia pathway of colorectal adenocarcinomas, are linked to activation of PD-L1 in CRC cells.^95,97,98 Cen et al⁸⁵ discovered, in tissue samples from 43 patients with colon cancer, that BRAF mutant colon cancer had significantly increased expression of PD-L1, PD-1, CTLA4, lymphocyte-activation gene 3, and TIM3, which is similar with results from 396 colon cancers from TCGA datasets. Furthermore, the connection between BRAF mutation and PD-L1 expression has been investigated recently.^99,100 BRAF V600E produced a significant level of PD-L1 expression in MSS CRC cell lines,¹⁰¹ raising the possibility that PD-L1 expression is not limited to MSI-H BRAF-mutated CRC. Although no data are available for the subset of BRAF mutations, there is no clear correlation between PD-L1 expression and the effectiveness of anti-PD-1/PD-L1 immunotherapy in CRC.^49,50 Other immune-independent functions of PD-L1 expression in BRAF mutant CRC were found. Feng et al¹⁰² shown that in colon tumors, oncogenic BRAF V600E can transcriptionally upregulate intrinsic PD-L1 expression, which by increasing BIM and BIK proteins, improves chemotherapy-induced apoptosis. Feng et al¹⁰¹ demonstrated how c-JUN and YAP promote PD-L1 expression in colon tumors with BRAF V600E.

Tertiary lymphoid structures (TLS)

Mostly developing in infectious diseases, inflammatory disorders, autoimmune syndromes, and tumors are TLS.¹⁰³ As a part of the TME, TLS becomes implicated in tumor growth and metastasis.¹⁰³ Moreover, TLS may function as a prognostic biomarker in human malignancies and as a predictive biomarker for ICIs.^103,104 A higher degree of TLS density was associated with colorectal tumors with MSI-H and/or BRAF mutations (median: 0.61 vs. 0.45, rank-sum P = 0.03), according to Posch et al¹⁰⁵ TLS mostly exhibited in the peripheral region of CRC tumor tissues (97%).

Patients with BRAF mutant CRC progresses quickly, has a bad prognosis, and responds poorly to conventional treatment. While the BEACON study shown that mCRC patients with BRAF V600E mutations can benefit clinically from a BRAF inhibitor plus an EGFR inhibitor ± MEK inhibitor, the proven ORR is only 20%.¹⁶ Elez et al¹⁰⁶ discovered that improved ORR and PFS were achieved by BRAF mutant mCRC with MSS and ring finger protein 43 mutation. New treatment possibilities must be investigated, for BRAF mutant mCRC patients who are not benefited by targeted combination therapy. Durable responses have been obtained by immunological checkpoint inhibitor therapy in certain MSI-H/dMMR CRC patients. Subgroup analysis of clinical trials supports the advantage for MSI-H patients with BRAF-mutated mCRC; more research in this population is required to confirm. Not all MSI-H/dMMR patients, nevertheless, react effectively to immunological checkpoint inhibitor therapy. The investigation of new predictive biomarkers for screening immunotherapy-sensitive populations other than MSI-H should continue since BRAF mutant CRC is a diverse tumor subtype with varying efficacy for ICB therapy. Patients with MSS/pMMR CRC are usually known to not react to ICIs. Combination therapy with ICIs may be another approach to enhance prognosis for mCRC patients with MSS/pMMR and MSI-H/dMMR who do not react to ICIs. Blocking the MAPK pathway works in concert with immunotherapy. Promising results have been seen in mCRC patients with MSS/pMMR BRAF mutations with immunotherapy and MAPK pathway inhibition. To be sure of it, nevertheless, extensive prospective phase III clinical trials are still required. Furthermore, more combination treatments, like ICIs with radiation, chemotherapy, and tumor vaccines, are still needed to boost the host immune system and get over the immunotherapy insensitivity of particular patients.

None.

No potential conflict of interest relevant to this article was reported.