The World Health Organization in 2023 reported that colorectal cancer ranks third as the world’s most common cancer type. Regardless of existing effective technologies for early detection and treatment aimed at lowering mortality rate, over two million new cases are diagnosed yearly, resulting in about one million deaths annually. This places colorectal cancer as the second leading cause of cancer-associated deaths worldwide [1].

Cancer therapy has evolved tremendously over the years from rudimentary surgeries to advanced, personalized, and target-specific therapies. Formerly, cancer was treated through methods like surgery and radiotherapy. These methods were effective for treating localized cancer; they are, however, limited in addressing complications like metastasized cancer, they also often cause harm to the surrounding tissues. By the mid-20th century, advancement in technology and breakthrough in chemotherapy provided more treatment options with better success rates and solutions for the challenges [2]. Today, cancer treatments are increasingly target specific and personalized, these strategies include biomarker testing for cancer, chemotherapy, hormone therapy, photodynamic therapy, immunotherapy (CAR T-cell therapy, cancer vaccines, cytokines, immunomodulators, immune checkpoint inhibitors, monoclonal antibodies), targeted therapy, surgery, stem cell transplant, radiation therapy, hyperthermia [3,4].

Cancer vaccines are a states of the art immunotherapeutic approach developed to prevent and treat cancer. There are two main types of cancer vaccines: preventive and therapeutic cancer vaccines. While preventive cancer vaccine targets infectious agents such as viruses that can lead to cancer, therapeutic vaccines are designed to treat existing cancers. Therapeutic cancer vaccines in general are cancer-associated antigens that are introduced into the body to stimulate immunogenic recognition, attack and destruction of cancer cells by the immune system. In addition to treating fully developed malignancies, cancer vaccines reduce tumor size, block or slow the proliferation of cancer cells, prevent recurrence, and eradicate cancer cells that evade other therapies [5].

Carcinoembryonic antigen (CEA) is an oncofetal antigen that is overexpressed in human adenocarcinomas, especially in gastrointestinal cancer. It is a promising target for gastrointestinal cancer-specific immunotherapy [6,7]. Their antigenic property makes them a good potential target for immunotherapy, this was reported by Larocca and Schlom [8] and Meraviglia-Crivelli et al. [9], who stated that CEA can induce a stronger CD8 T-cell cytotoxic response when animals are immunized with it. In another study, a recombinant vaccinia virus vaccine that encodes complete or internally deleted cDNAs for human CEA induced CEA-specific autoantibodies in seven out of the 32 participants involved in the study. It was reported that the participants previously had no CEA antibodies before vaccination [10].

Insulin-like growth factor-1 receptor (IGF-1R) is another biomarker implicated in the development of several human malignancies [11], including breast, pancreatic, and colorectal cancers [12]. It is reported that IGF-1R is significantly involved in cellular growth, spread, alteration, and apoptotic inhibition [13]. IGF-1R signaling causes tumor cells to be resistant to treatments such as chemotherapy and anti-hormonal therapy.

Many cancer types affecting humans have been found to overexpress IGF-1R, an instance of such observation was reported by Hakam et al. [14], who also wrote in their findings that cases of pancreatic cancer show an overexpression of IGF-1R, this claim was also report by Xu et al. [15], who reported an increased level of IGF-1R in blood plasma for every advanced stage of pancreatic cancer and a total absence of same in the surrounding non-cancerous tissues.

Owing to the role of IGF-1R in cancer cell proliferation and growth, resistance to treatment, its overexpression in specific malignant cells and lack of it in surrounding non-malignant cells, it may be suggested that immunotherapeutic approach involving the epitope IGF-1R could be a better anticancer therapy compared to the traditional methods [13].

There is an increasing amount of data supporting the fact that epitope vaccines can induce immunological response against malignant cells by activating helper T cells (Th), cytotoxic T cells, and B cells. As regards these, IGF-1R has become a prominent molecular target for anticancer therapeutic approaches.

Immunological responses are critically important in fighting cancer, adapting this response in order to treat cancer may be a vital approach in combating the recent rise in cancer morbidity and mortality. To achieve this, antigenic epitopes are introduced to the host immune system through the process referred to as vaccination [16].

An epitope with antigenic properties is a unit from the pathogen or in this case the tumor cell that can induce innate, humoral or cellular immune response [16]. Combining antigenic epitopes to develop a vaccine results into a multiepitope vaccine which is one composed of a series of overlapping chain of amino acids with the potential to elicit cytotoxic T-lymphocytes (CTLs), helper T cells (Th), B cells and also induce efficient immune responses against the targeted tumor.

Multiepitope approach for cancer therapy is gaining recognition because compared to other techniques, it is more convenient, time-saving and cost-effective [17]. Based on the above-mentioned factors, this research was focused on designing a multiepitope vaccine developed by combining CEA and IGF-1R epitope to treat and prevent colorectal cancer.

In this study, we sought to design novel multiepitope vaccine biomolecule that targets colorectal cancer using bioinformatic and immunoformatic approaches to linking CEA and IGF-1R epitopes. The prime rationale in this approach is based on CEA and IGF-1R roles in resistance to targeted therapies, involvement in growth of malignancy and their oncological biomarker status [25,26]. Multiepitope vaccines as a therapeutic tool is becoming progressively accepted because of their efficiency and nontoxic property when paralleled to other treatment modalities [27].

To achieve this, CEA and IGF-1R sequences were selected and retrieved from the NCBI database. The selection was based on their antigenic potential which was predicted on the Scratch database. To develop the vaccine, conserve regions of the retrieved CEA and IGF-1R sequences were screened for and selected to ensure that the vaccine would be active for large variant of the cancer type and would still be active even if mutation should occur [28,29]. Analysis of the conserved sequence for antigenicity revealed that the sequences maintained the antigenic criterion of selection as the original CEA and IGF-1R sequences, hence possessing the potential of an immune stimulant.

To determine their immune-stimulating properties the conserved sequences were screened for CTLs, HTLs, and B cells inducing epitopes as a vaccine should be able of inducing these immune components to initiate humoral and cellular immune response. Epitopes that can initiate the production of these immune components were linked together using suitable linkers, adjuvant was also linked with the epitopes at the N-terminal to enhance the antigenic potential [18]. The complex now formed makes up the in silico primary vaccine structure.

It was also necessary to subject the vaccine’s primary structure to antigenic and allergenic screen as was carried out using the Scratch online tool. Screening result predicted that the vaccine’s primary structure preserved the antigenic potential while remaining nonallergenic. Physicochemical properties prediction also showed that the vaccine properties fell within the accepted threshold [30].

Vaccine structural stability is essential as the vaccine should be able to withstand biological, biochemical and metabolic pressure of the host system, we evaluate the vaccine structural stability by predicting the secondary structure of the vaccine, result obtained showed that the structure fall within accepted threshold, this was on the premise of the alpha helix and beta sheet percentages.

The tertiary structure of a polypeptide plays a major role in its function. We predicted the vaccine tertiary structure by homology modeling. This method predicts the tertiary structure of an unknown polypeptide by comparing it with known structures on the database based on the similarity of their sequences.

In order to validate the predicted tertiary structure, the PDB file was subjected to statistical analysis, a z-score of −4.03, and a Ramachandran plot with 87.5% in the most favored region was obtained. This result is satisfactory as it indicates a peptide that conforms to native protein.

Docking the vaccine peptide with the TLR4 molecule was done to evaluate the vaccine’s ability to bind to TLR4. Binding an antigen with a TLR4 activates inflammatory response and innate immune system. The vaccine polypeptide in this study docked with TLR4 with a binding energy of −317.92 kcal mol⁻¹. The low binding energy indicates a high binding affinity with human TLR4 (Fig. 6).

In conclusion, using an immunoinformatics approach, we identified epitopes that have potential as a multiepitope-based subunit vaccine against cancer. CTL, HTL, and B-cell lymphomas were screened and selected for vaccine design. Finally, the vaccine constructs consist of CTL, HTL, and B-cell epitopes, an adjuvant to elicit strong immune responses and fusion protein linkers. Conservancy analysis of the promiscuous epitopes was used to select epitopes with robust immune response against large variants or mutants of cancer. Molecular docking revealed stronger binding affinity with human TLR4. Our computational analysis suggests that the constructed vaccine is highly immunogenic, safe, and stable in nature.