Glioma is the most common cerebral malignancy with high morbidity and mortality. Despite the current treatment measures such as surgery, radiation, and chemotherapy, the prognosis and mortality of patients has not improved significantly (1–3). The annual death rate in China is as high as 30,000. Currently, the adverse reactions to glioma drugs are more prominent, and drug resistance is readily developed (1, 4, 5). To overcome the limits of existing therapies, there is a pressing need for a treatment strategy that can selectively target cancer tissues and avoid non-target tissues.

In addition, the blood-brain barrier (BBB) is a formidable obstacle for the transport of most administered therapeutics to the brain (6, 7), and most anti-tumor drugs have difficulty passing the BBB and the blood-brain tumor barrier (BBTB), it is a major hurdle in the development of targeted drugs for glioma (8–10). Therefore, choosing a carrier that can pass through the BBB is very important for glioma treatment.

Filamentous bacteriophages (Ff phage) are nano-scale viruses that infect bacteria and are not harmful to humans (11–13). Ff phage fd, M13, and f1 are stable under harsh conditions and can be manufactured with uniform specifications and low cost (14–17). As well, Ff phage has genetic flexibility. In 1985, Smith et al. reported phage display technology to display a variety of proteins, antibodies, and peptides on the phage coat proteins. Subsequently, phage display libraries were injected intravenously into laboratory animals to screen the targeting peptides (18). Moreover, Ff phage could enter the central nervous system (CNS) without visible toxic effects (19, 20), it can pass through the BBB as a drug carrier, when administered intranasally or through convection-enhanced delivery (CED), and has great research potential for the treatment of brain diseases (21–24).

Furthermore, the phage display library is used quickly and directly to screen peptides targeting tumor and anti-tumor antibodies. To date, there are numerous studies by the phage library screening tumor targeting peptides for target therapy and immunotherapy (25–27), and it has established the method for screening glioma targeting peptides across the BBB, guiding the immunotherapy in patients. These phages and peptides targeting glioma cells could avoid or reduce the toxic effects of anti-cancer drugs (28–31). At the same time, phages, carrying targeted peptides and antibodies, stimulate the immune response and play an immunotherapy role (28, 32–35).

In summary, this review will clarify the strategy for applying Ff phage nanoparticles to glioma treatment. It can be used to direct clinical treatment of tumors and provide new ideas for personalized disease therapy.

Ff phage is a biological nanomaterial with a length of about 1 um and a diameter of about 7 nm (14, 36). It could specifically infect bacteria and is present in the human body and harmless to humans. Ff phage is made of single-stranded circular DNA and coat proteins. The main coat protein pVIII is located on the phage side and minor coat proteins (pIII, pV I, pVII, and pIX) are located at both tips ( Figure 1 ).

Phage display is to insert the DNA sequence of the exogenous peptide into the phage coat protein gene and express the peptide on the surface of the phage along with the expression of the coat protein. The phage displaying peptide still has protein assembly and infection activity (37–39). Based on phage display technology, phage libraries were built and used to select targeting phages, such as tumor-targeted peptides, which improved research efficiency and reduced costs. In addition, these targeted peptides developed functions of cell-targeting, tumor-homing, and cell-penetrating (40–42). Phage libraries were usually screened by using molecules, cells, and tissues in vitro or in animals and human patients (38, 43, 44).

Traditional chemotherapeutics have poor accuracy on tumor cells and are prone to adverse reactions. Therefore, the targeted therapy is particularly important for tumor therapy (45–47). Peptides specifically binding to tumor tissues, as carriers to direct drugs to tumor tissues, significantly improved the accuracy of drug targeting (48–51). Although monoclonal antibodies as vectors were successfully applied to anti-tumor, the high molecular weight of antibodies might reduce efficiency (52–55), while the phage peptide library has the benefits of screening for small molecular weight peptides, which can compensate for antibody deficiencies that are widely used in the diagnosis and treatment of glioma.

The screened peptides could combine with markers for imaging. Wang et al. developed an HO-8910 ovarian cancer cell targeting peptide (NPMIRRQ) from the phage library, which demonstrated the ability to selectively bind ovarian cancer cells using immunofluorescence and immunohistochemical assays (56).

The screened peptides could also couple with chemotherapeutic drugs or some gene, and then be used in the tumor-targeted treatment or gene treatment; Du et al. obtained the A54 peptide (AGKGTPSLETTP) by in vivo phage display for hepatocarcinoma and conjugated it with doxorubicin for in vivo targeted therapy. The study showed the A54-doxorubicin reduced the tumor size and prolongated the long-term survival rate (57).

Furthermore, some specific binding peptides that inhibit tumor growth, invasion, and metastasis, could be used to treat tumors directly. Zhou et al. isolated the peptide, SWQIGGN, from a Ph.D.-C7C phage library with the ovarian cancer cell HO-8910 (58). They found that the peptide controlled cancer cell migration, viability, adhesion capacity, invasion, and tumor growth in vivo.

Currently, the screening of tumor-targeted peptides is widely used in targeting the treatment of tumors, such as lung cancer, stomach cancer, liver cancer, colon cancer, and prostate cancer.

Poor permeability of the cellular plasma membrane to a drug or gene is the main barrier for targeted delivery, while the nature of vectors affects the efficiency of drug delivery in tumors and tumor-affected tissues. Therefore, construction and selection of drug vectors is one of the most important steps of tumor therapy. Ff phage could deliver genes and peptides to mammalian cells, and the structure of Ff phage results in more efficient cellular attachment and ensuing membrane penetration. It has been successfully used in treatment under the U.S. Food and Drug Administration (FDA) process (www.fda.gov), and methods for isolating, storing, and producing phages are now becoming more available and better developed under the ATCC (www.atcc.org) and PHE (www.gov.uk/government/organisations/public-health-england) collections.

Ff phage might be an ideal carrier for drug therapy and immunotherapy. First, an exogenous gene could be inserted into the Ff phage genome. Meanwhile, the peptide displayed on the Ff phage presents its natural conformation and the phage has a strong resistance to physical and chemical factors (59–61). Second, the phage displayed exogenous peptides or chemical modifications, which could combine with inorganic nanomaterials/drugs, to form phage-nanocomplexes and drug-loaded phages. It is well utilized in photodynamic cancer therapy (62–64). Some researchers used the Fd phage to display a cancer-targeting peptide on pVIII major coat protein, and then conjugated photosensitizer at the N-terminal end of the targeting peptides, and demonstrated that the complex of phage-photosensitizers was able to selectively target and kill SKBR3 tumor cells in vitro (65). Third, the displayed Ff phage triggers every arm of the immune response. Berardinis et al. engineered fd to target mouse dendritic cells (DCs), and activated the innate and adaptive responses without the need of exogenous adjuvants (66). The study has also shown that phage could induce the IL-2 and IFN-γ cytokines, which were useful in tumor immunotherapy (67). Fourth, drug conjugated phage increases the half-life in the blood steam (68), while the toxicity and side effects of hazardous drugs are reduced in combination with the Ff phage.

In a word, Ff phage, as a carrier of therapeutic reagents, has more advantages in targeted therapy, with high specificity, high sensitivity, and reproducibility.

Gliomas are aggressive brain tumors and challenging therapeutic cancers that have high mortality (69). The 5-year survival rate of glioma is very low (70), and the prognosis of glioblastoma patients is poor with a median survival of less than 1 year. Recently, cancer research in the U.K. showed that 40% of brain tumor patients survive their cancer for 1 year and more than 10% survive their cancer for 5 years or more.

At present, the clinical therapies for gliomas are surgical therapy, radiation therapy, chemotherapy, gene therapy, and other comprehensives (1, 71–74). However, it is easy to relapse after these treatments, and the patients’ survival rates are not significantly improved. Immunotherapy is useful for treating tumors, the mAbs bevacizumab, rituximab, and trastuzumab were already widely used against tumors outside the brain (75–79). But the current immunotherapy for medical glioma is costly and inefficient.

BBB is another significant barrier to the delivery of targeted treatments for brain tumors. Indeed, more than 98% of low-molecular-weight candidate drugs and almost 100% of large therapeutic candidate drugs cannot cross the BBB (80). There is an urgent need for a carrier that carries drugs across the BBB. Phage display technology can be used for the construction of peptide libraries to screen for glioma tumor-targeting peptides and peptides across the BBB. Surface functionalization with these peptides is a sophisticated way to develop drug delivery platforms that cross the BBB and target glioma.

The rod-shaped nanoparticles have higher avidity and selectivity for endothelial cells and increase the specificity and vascular targeting for brain endothelium (96). Increasing the length-to-diameter ratio of Ff phage results in more effective cellular attachment and ensuing membrane penetration. The phage maintains the biological activity of the peptide displayed on the phage vector, these properties make Ff phage suitable for use as a vector in the treatment of central nervous diseases (97), and research proved that phages carrying antibodies effectively label Aβ plaques is an efficient and nontoxic delivery vector to the brain and is useful for the treatment of Alzheimer’s disease in vivo (68).

Gliomas are the most common primary brain tumors. Effective treatment of glioma is hampered by the presence of both BBB and BBTB. In this review, we presented an Ff phage approach to enhance the permeability of drugs through BBB and BBTB.

Although Ff phages have the problem of further optimization and improvement in separation and purification, they also have a number of advantages. Ff phage has a greater level of safety, it is not reproduce naturally in mammalian hosts, and it expresses a wide range of peptides on coat proteins using genetic engineering techniques to attach targeting peptides and antibodies.

Phage display is a high throughput screening strategy to construct peptide libraries that are used to screen glioma targeting peptides. These peptides might cross the BBB/BBTB and target tumors. It can also be used as a drug or drug carrier after being modified. Furthermore, Ff phage is an ideal transport carrier to CNS across the BBB, it has the anti-tumor ability and could be genetically modified to display glioma homing motifs and conjugated to cytotoxic drugs. Moreover, Ff phage displaying targeting peptide has stronger tumor penetrating ability, a higher load of drug delivery ability, and lower toxicity.

Developing carrier-based Ff phage as a drug delivery system can solve the problem of going through the BBB and BBTB. In short, Ff phage display technology is a powerful method of developing highly effective target drug delivery carriers. In addition, it opens the door to the development of personalized therapy agents in the future.

YW: Conceptualization; JS: Methodology and analysis; JC: Data Curation and analysis; CZ: Resources; XL: Visualization; WY: Writing; RC: Supervision; TG: Writing-Reviewing and Editing.

This study was supported by the grant from the National Key R&D Program of China (Grant #2018YFC1311600), National Natural Science Foundation of China (81901880), Natural Science of Science and Technology Division Jilin (20200201505JC and 20190103099JH), Education deparment research project of Jilin (JJKH20201016KJ)and program of Jilin finance deparment (2019SRCJ003). Jilin provincial health project (2020SC2T091).

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.