Hyaluronic acid-based drug nanocarriers as a novel drug delivery system for cancer chemotherapy: A systematic review

Graphical abstract

Chemotherapy is the most common treatment strategy for cancer patients. Nevertheless, limited drug delivery to cancer cells, intolerable toxicity, and multiple drug resistance are constant challenges of chemotherapy. Novel targeted drug delivery strategies by using nanoparticles have attracted much attention due to reducing side effects and increasing drug efficacy. Therefore, the most important outcome of this study is to answer the question of whether active targeted HA-based drug nanocarriers have a significant effect on improving drug delivery to cancer cells.

This study aimed to systematically review studies on the use of hyaluronic acid (HA)-based nanocarriers for chemotherapy drugs. The two databases MagIran and SID from Persian databases as well as international databases PubMed, WoS, Scopus, Science Direct, Embase, as well as Google Scholar were searched for human studies and cell lines and/or xenograft mice published without time limit until 2020. Keywords used to search included Nanoparticle, chemotherapy, HA, Hyaluronic acid, traditional medicine, natural medicine, chemotherapeutic drugs, natural compound, cancer treatment, and cancer. The quality of the studies was assessed by the STROBE checklist. Finally, studies consistent with inclusion criteria and with medium- to high-quality were included in the systematic review.

According to the findings of studies, active targeted HA-based drug nanocarriers showed a significant effect on improving drug delivery to cancer cells. Also, the use of lipid nanoparticles with a suitable coating of HA have been introduced as biocompatible drug carriers with high potential for targeted drug delivery to the target tissue without affecting other tissues and reducing side effects. Enhanced drug delivery, increased therapeutic efficacy, increased cytotoxicity and significant inhibition of tumor growth, as well as high potential for targeted chemotherapy are also reported to be benefits of using HA-based nanocarriers for tumors with increased expression of CD44 receptor.

Introduction

Cancer is the second leading cause of death worldwide, after cardiovascular disorders [1]. After decades of impressive efforts to discover the biology of cancer, the most challenging questions still arises in the context of cancer treatment [2]. Despite extensive therapeutic strategies, cancer mortality is rapidly increasing worldwide. Also, the poor efficacy and severe side effects of a variety of these therapeutic strategies in many cases remain an issue [3].

Today, chemotherapy is the most widely used treatment strategy for the vast majority of cancers. There are a variety of chemotherapeutic drugs that are usually cytotoxic and target high-proliferating cells [4]. Cancer cells exhibit uncontrolled proliferation due to bypassing cell cycle checkpoints [5, 6]. Nevertheless, High rates of proliferation are naturally found in hematopoietic cells, cells lining the gastrointestinal tract, and in skin. Therefore, systemic administration of chemotherapeutic drugs that are absorbed by cells based solely on high rates of proliferation is associated with damage to normal, highly proliferating cells [7]. In addition, the use of high doses due to systemic administration, distribution to different tissues, and rapid excretion of the drug may lead to widespread side effects and drug toxicity that can vary in different cases depending on the cancer, drug, dose, and genetic context [8].

Targeted drug delivery is a process that leads to the accumulation of drugs in a specific area of the body. In this process, a specific tissue, type of cell, or even a specific intracellular organelle may be targeted, depending on the drug and the location of the disease [9]. Nanoparticles are commonly used as drug carriers in targeted drug delivery, and their remarkable active targeting efficacy to increase drug accumulation in cancer cells has been reported in various studies [10–13]. Nanocarriers have solved some of the current limitations of systemic drug delivery in the treatment of cancer, including increasing the stability of the drug molecule, continuous drug release, reduced toxicity and side effects, crossing biological barriers to drug delivery, and increased drug access. Hence, this strategy is known as a very effective drug delivery system [14–16].

Hyaluronic acid (HA) is a high molecular weight linear polysaccharide consisting of d-glucuronic acid (D-glucuronic acid) and N-acetyl-d-glucosamine. The repeating sequence is perfectly homogeneous, and is the same in all vertebrate tissues and fluids. The polymer molecular mass is more variable. Most commonly, hyaluronan is synthesized as a high-molecular mass polymer, with an average molecular mass of approximately 1000–8000 kDa [17, 18].

It is naturally present in the extracellular matrix and has attracted much attention as a nanocarrier for drug delivery. The presence of HA on the surface of nanocarriers results in increased efficacy and safety of the drug as well as efficient drug delivery to cancer cells due to the tendency to bind to the CD44 receptor, which is abundantly expressed on the surface of cancer cells [17, 18].

Other molecules are currently being explored to modify NPs surface and to perform active uptake. Exploitation of simple molecules to enhance active uptake also includes the use of carbohydrates. Among these, one of the most studied is hyaluronic acid (HA). This molecule enhances the NPs uptake through its interaction with CD44 protein. recently reported that HA can be easily used to biofunctionalized carbon dots loaded with doxorubicin to enhance drug uptake in CD44 overexpressing cells. Results obtained by competitive assay performed with free HA showed that the HA-modified NPs were internalized through the binding of HA-modified NPs with the CD44 receptor. Furthermore, in vivo experiments showed that the enhanced accumulation into tumor tissue was confirmed [18].

Despite the benefits and attractive biomedical applications, some limitations need to be addressed, including the fact that laboratory synthesis of hyaluronic acid is a difficult and time-consuming process. Also, HA-based active targeted nanoparticles identify only certain types of cancer cells that express specific receptors at the cellular level [18].

Given that the ultimate goal of cancer therapeutic strategies is to completely clear cancer cells or increase the quality of life and lifespan of patients, today there is a special emphasis on developing treatment systems that can identify the majority of cancer cells and introduce cytotoxic drugs into them. The use of nanocarriers based on HA as a drug delivery system has been associated with disagreements about its efficacy in inhibiting tumor growth, improving patient survival, and reducing drug toxicity in various studies. Therefore, the purpose of this study a systematic review of studies using HA-based therapies in cancer treatment. the most important outcome of this study is to answer the question of whether active targeted HA-based drug nanocarriers have a significant effect on improving drug delivery to cancer cells.

Methods

This study was conducted through a systematic search based on Preferred Reporting Items for Systematic Reviews and Meta-Analyzes (PRISMA)(19). The two databases MagIran and SID from Persian databases as well as international databases PubMed, WoS, Scopus, Science Direct, Embase, as well as Google Scholar were searched for human studies and cell lines and/or xenograft mice published without time limit until 2020. Keywords used to search included Nanoparticle, chemotherapy, HA, Hyaluronic acid, traditional medicine, natural medicine, chemotherapeutic drugs, natural compound, cancer treatment, and cancer. A bibliography of identified articles was also searched by hand. To minimize the possibility of bias, all steps of article search, article selection, qualitative evaluation and data extraction were performed independently by two authors. Disagreements between the two authors were resolved by final agreement with the other author.

Inclusion and exclusion criteria

Inclusion criteria included related studies: 1. investigating the effects of hyaluronic acid-based nanoparticles (NPs) on the delivery system of natural drugs in cancer, 2. interventional, 3. with access to their full text, 4. related in vivo and in vitro studies, 5. investigating the receptor CD44, and 6. investigating cancers.

Exclusion criteria included: 1. unrelated studies, 2. without sufficient data, 3. studies in which the drug-nanoparticle platform and the type of study are not mentioned, 4. duplicated studies, 5.uncertainty methods, 6. studies in which the ligand is not hyaluronic acid, 7. studies in which the full text is not available.

Quality assessment and Data extraction

Quality of the papers was evaluated based on selected and relevant items present in the CONSORT checklist that was proper for assessment in this study (items were study design, background and literature review, place and time of study, outcome, inclusion criteria, sample size, and statistical analysis) as mentioned in previous studies. Papers referring to 6 to 7 criteria were considered as high -quality papers, papers that did not mention 2 items and more than 2 items from the 7 items were considered as papers with medium- and low -quality methodology, respectively. Information on all final articles entered into the systematic review process was summarized in a pre-prepared checklist, including article title, first author name, year of publication, study location, study type, ligand and receptor type, drug-NP platform, and compared to non-targeted. 14 articles were included in the final analysis, the information of which is listed in Table 1.

Table 1.

Summary of information of the final selected articles for the systematic review

References	Compared to non-targeted	Types of study	Drug-NP platform	Receptor	Region	Author , year
[19]	Higher cellular uptake in CD44 overexpressing (SCC7) compared to CD44 negative (NIH3T3); no difference in cellular uptake compared to free drug 30% higher tumor growth inhibition rate compared to free drug	In vitro: murine squamous cell carcinoma cell lines (SCC7) and mouse embryo fibroblast cell lines (NIH3T3) In vivo: cell lines SCC7 xenograft	Doxorubicin- HACE-PEG	CD44 receptor	Korea	Hyun-Jong Cho et al.,2011
[31]	Higher tumor growth inhibition rate; higher survival time	In vivo: Ehrlich ascites tumor-bearing mice	Doxorubicin-PBLG	CD44 receptor	France	Kamal Kumar Upadhyay et al.,2012
[29]	Higher tumor accumulation; higher tumor growth inhibition rate	In vivo: murine melanoma cell lines (B16F10) xenograft	Methotrexate-lipid-based NP	CD44 receptor	USA	Shoshy Mizrahy et al.,2014
[26]	Higher cellular uptake in time-dependent manner; higher cytotoxicity – 1.75-fold for MiaPaCa-2 and twofold for AsPC-1	In vitro: human pancreatic cancer cell lines (MiaPaCa-2, AsPC-1)	3,4-difluorobenzylidene curcumin-styrene maleic acid	CD44 receptor	USA	Kesharwani et al., 2015
[12]	Higher cellular uptake; threefold higher cytotoxicity compared to free drug Higher tumor growth inhibition rate; 3.6-fold and 1.7-fold higher drug accumulation in tumor compared to kidney and liver	In vitro: human colorectal cancer cell lines (HCT-116) In vivo: cell lines HCT-116 xenograft	Topotecan hydrochloride-dendrimer	CD44 receptor	China	Xiaole Qi et al.,2015
[20]	Higher cellular uptake compared to free drug; no difference in cytotoxicity Lower relative tumor volume; higher median survival time	In vitro: doxorubicin-resistant human breast adenocarcinoma cell lines (MCF-7/ADR) In vivo: cell lines MCF-7/ADR xenograft	Doxorubicin- hyaluronic acid-Lys-LA10	CD44 receptor	China	Yinan Zhong et al.,2015
[21]	4.1-Fold higher cellular uptake 2.80-Fold higher tumor accumulation; 31.89% higher tumor growth inhibition rate; higher median survival time	In vitro: human breast adenocarcinoma cell lines (MCF-7) In vivo: murine hepatic carcinoma cell lines (Heps) xenograft	Paclitaxel-micelle	CD44 receptor	China	Shaoping Yin et al.,2015
[22]	Higher cellular uptake; 46.3% higher cytotoxicity compared to free drug Higher in tumor targeting; lower tumor volume	In vitro: human hepatocellular carcinoma cell lines (HepG2) In vivo: cell lines HepG2 xenograft	Doxorubicin- hydroxyl apatite	CD44 receptor	China	Hui Xiong et al.,2016
[23]	10-Fold higher in cellular DOX level; higher cytotoxicity No difference in tumor growth inhibition rate; higher survival time	In vitro: human breast adenocarcinoma cell lines (MCF-7) In vivo: cell lines MCF-7 xenograft	Doxorubicin-PBLG-LA	CD44 receptor	China	Bingfeng Sun et al.,2016
[27]	Higher cellular uptake; eightfold higher cytotoxicity	In vitro: human lung cancer cell lines (A549)	Cisplatin-chitosan	CD44 receptor	USA	Min Sung Suh et al.,2017
[25]	Higher cytotoxicity, 1.35-fold for MDA-MB-231, and 1.1-fold lower cytotoxicity to MCF-7	In vitro: human breast adenocarcinoma cell lines (MCF-7 and MDA-MB-231)	Rapamycin-LbL-LCNP	CD44 receptor	Egypt	May S Freag et al.,2016
[24]	higher tumor growth inhibition rate lower accumulation in the tumor	In vitro: human liver sinusoid endothelial cells In vivo: cells 4T1-bearing mice	PEGylated hyaluronic acid	CD44 receptor	USA	Chao Teng et al.,2019
[30]	higher cytotoxicity	Invivo:4T1 tumor-bearing mice	doxorubicin dehydrochloride	CD44 receptor	China	Jianping Li et al.,2020
[28]	higher accumulate in cancer cells, lower cytotoxicity	In vitro: cells 4T1-bearing mice In vivo: cells 4T1-bearing mice	Doxorubicin hyaluronic acid	CD44 receptor	China	Beibei Lu et al.,2020

Abbreviations: LbL-LCNP, layer-by-layer-liquid crystalline nanoparticle; PBLG, poly(γ-benzyl L-glutamate); PBLG-LA, G-poly(c-benzyl-L-glutamate)-lipoic acid; Lys-LA10, L-lysine methyl ester-lipoic acid;HPAEG, hyper branched poly(2-((2-(acryloyloxy)ethyl)disulfanyl)ethyl 4-cyano-4-(((propylthio)-carbonothioyl)-thio)-pentanoate-co-polyethylene glycol methacrylate; PEG, polyethylene glycol; PBLG, poly(γ-benzyl L-glutamate).

Results

Initially, a total of 213 articles were retrieved. 81 repeated articles, 65 articles inconsistent with inclusion and exclusion criteria, and 53 articles with low quality were excluded. Figure 1 shows the performed process. Finally, 14 articles were selected for this systematic review, including seven in vitro and in vivo studies [12, 19–24], four in vitro studies [25–28], and three in vivo studies [29–31].

Fig. 1.

Flow chart indicating the stages of article selection in this systematic review and meta-analysis (PRISMA 2009)

In comparing the drug toxicity resulting from HA-based drug nanocarriers with free drug use, increased drug toxicity in seven studies [12, 22, 23, 25, 26, 30] and decreased drug toxicity in one study were reported [28]. Also, no difference was observed in one study [20]. Several studies also did not compare drug toxicity resulting from HA-based drug nanocarriers with the free drug [19, 21, 24, 29] (Fig. 1).

In addition, six studies have reported that these HA-based nanocarriers have the potential to inhibit tumor growth [12, 19, 21, 29]. In one study, no difference was observed in the rate of tumor growth inhibition compared to the free drug [23]. Also, the rate of tumor growth inhibition of these nanocarriers was not compared in several studies [20, 22, 25, 26, 30, 32].

Moreover, the use of HA-based drug nanocarriers led to increased cellular uptake of the drug in seven studies [12, 19–23, 26, 32] and increased survival rate in four studies [20, 21, 23] (Table 1).

Discussion

Here, in vivo and in vitro studies on HA-based drug nanocarriers used in cancer chemotherapy were systematically reviewed. The included studies were publications in the period 2011–2020, of which 3 studies were published in 2019–2020 [24, 30, 32]. All included studies were of medium- to high-quality. The present study is the first systematic review focusing on hyaluronic acid-based drug nanocarriers delivery systems for cancer chemotherapy. After systematic review of various studies, it was found that nanoparticles high antitumor properties with great potential for targeted chemotherapy in tumors expressing CD44 receptors [19, 21, 27, 28].

In the field of cancer treatment, there are different therapeutic strategies, the most important of which is chemotherapy using anticancer drugs such as paclitaxel (PTX), doxorubicin (DOX), and cisplatin. Systemic administration of chemotherapy drugs can kill cancer cells by accelerating a specific immune response. Nevertheless, healthy cells in the body are also affected, which can be associated with various side effects [33, 34]. Also, the biostability of these drugs is low and they are easily cleared in the bloodstream with physiological reactions or immune system responses. Therefore, important factors for the use of anticancer drugs in the systemic administration of chemotherapy are that the drugs are delivered to cancer cells with minimal impact on healthy cells and maximum stability in vivo. Nanocarriers have been designed to cover the needs and be able to provide targeted drug delivery systems [35–37].

Different types of small proteins or peptides have been used to bind at the nanoparticle surface for targeted delivery of chemotherapy drugs to cancer cells [38]. HA is naturally present in all species and plays a key role in maintaining the molecular structure and function of cell membranes [39]. HA is involved in important biological functions such as cell adhesion, differentiation, migration, and proliferation through specific and non-specific interactions [40]. On the other hand, its highly desirable physical, chemical and biological properties, stability in bloodstream, and the ability to release the drug in target cells with increased therapeutic effect has caused HA to attract much attention as a drug carrier [41].Today, HA and its derivatives have been widely studied as drug carriers for slow and targeted release, dermal absorption, and targeted delivery of drugs. Also, uptake of drug carriers into cancer cells in active targeted drug delivery usually occurs through receptor-mediated endocytosis [42].

CD44 is a major receptor for HA that is expressed on the surface of certain cells. The interaction between HA and CD44 has been investigated in a wide range of cellular functions including cell proliferation, differentiation, migration, angiogenesis and binding of cytokines, chemokines and growth factors to relevant receptors. The interaction between HA and CD44 is also known as a signaling mediator for cell survival and endocytosis [43–45].

There are various studies reporting the ability of HA to target cancer cells, leading to the emergence of new HA-based treatment strategies [46]. Also, there are various reports indicating increased expression of transmembrane receptor CD44 on different cancer cells, especially lung cancers [34]. It has recently been shown that there is a significant relationship between increased CD44 expression and clinical indicators such as tumor grade and tumor cell differentiation in cancer patients [47]. Given the strong tendency of HA to bind to CD44 receptor on cancer cells, much attention was drawn to HA for use as a suitable coating on the surface of drug nanocarriers [48].

The NPs surface modification is a powerful instrument to enhance uptake and biocompatibility, as confirmed by the vast amount of scientific papers focused on this topic. These studies demonstrate that the conjugation of molecules on the NPs surface can effectively enhance biocompatibility both in vivo and in vitro, due to the modification of surface charge and to the inactivation of reactive chemical groups that can affect cellular membrane stability, HA modification ensured stable drug encapsulation in mesoporous carbon nanoparticles in an extracellular environment while increasing colloidal stability, biocompatibility, cell-targeting ability, and controlled cargo release. The cellular uptake experiments of fluorescently labeled mesoporous carbon nanoparticles, with or without HA functionalization, demonstrated that HA-UMCS are able to specifically target cancer cells overexpressing CD44 receptors [47, 48].

through the handling of surface characteristics, the nanoparticles can be transformed in smart platforms, containing therapeutic and imaging agents as well as stealth property, delivering drugs to specific tissues and providing controlled release therapy. This targeted and sustained drug delivery decreases the drug related toxicity and the frequency of treatments [47, 48].

Although many efforts have been made to reduce resistance to chemotherapy drugs in most cancers, the major problem in dealing with ovarian cancer is still drug resistance. Yang et al. (2016) in a study aimed at reducing drug resistance to paclitaxel and improving its efficiency reported that the use of hyaluronate-coated lipid nanoparticles may be a suitable tool to reduce drug resistance and increase the therapeutic efficacy of paclitaxel [49].

Doxorubicin (DOX) is one of the most well-known anthracycline antibiotics used to treat cancer. Despite its widespread clinical use for chemotherapy, it can cause dose-dependent toxicity. Oommen et al. developed the HA-DOX Conjugate through covalent bonding between DOX and HA. According to previous research showing that hyaluronidase activity is considerably reduced in the blood of cancer patients, HA-DOX particles were also stable in the serum of patients. In addition, the in vivo toxicity caused by the initial circulation of the drug in the bloodstream was outstandingly reduced [50]. Therefore, Oommen et al. reported that polymer-drug conjugates could be an effective therapeutic strategy for metastatic tumors [51].

In a study by Zhong et al., DOX-loaded HA-based nanoparticles showed markedly lower side effects compared to the free drug. In their study, HA cross-linked nanoparticles offered great potential in cancer treatment due to the excellent biological compatibility, CD44-targetability, and effective reversal of drug resistance [20]. Liu et al. (2016) coated hyaluronic acid on the surface of nanostructured lipid carriers (NLCs) and delivered doxorubicin and Baikaline drugs specifically and co-transmission to breast tumors. The results of their study indicated that the drug co-transmission delivery system based on active targeting led to a synergistic anticancer effect of the two drugs [52]. In another study, Zhang et al. reported HA-coated lipid nanoparticles as high-potential carriers for increased biocompatibility and identified this targeted lipid system as a suitable candidate for drug delivery of natural compounds [53].

In an in vitro and in vivo study of Beibei Lu et al. doxorubicin showed high antitumor activity and targeted affinity for CD44 receptors as well as high durability in bloodstream with fewer side effects. The pro-drug micelles of doxorubicin produced in the study of Beibei Lu et al. showed a high potential for targeted treatment compared to the free drug [32] and their results were consistent with various previous studies [12, 21, 24, 29]. Nevertheless, although survival was increased in a study by Zhong et al., no difference was observed in inhibiting tumor growth [20].

Moreover, the molecular weight of HA directly affects the efficacy of active targeted drug nanocarriers. HA with two molecular weights of 9.5 kDa and 35 kDa was used to conjugate polymeric micelles to deliver paclitaxel to breast adenocarcinoma cells that showed high expression of CD44. Cellular uptake of the drug in conjugation with 9.5 kDa HA was significantly higher than that of HA with a molecular weight of 35 kDa, which in turn was significantly higher than that of the free drug [21]. Also, enhancing cell uptake of anticancer drugs through the receptor-mediated endocytosis by targeting ligand-conjugated nanoparticles improves drug efficacy, which has been associated with increased tumor growth inhibition, cytotoxicity, and survival for HA-conjugates [12, 19–24, 29, 32]. In the study of Zhong et al., No difference in cytotoxicity was observed [20].

Overall, the main challenge in cancer chemotherapy is the side effects of the drugs that can pose serious risks to patient health. Although targeted drug delivery with nanocarriers conjugated to small polymers has been able to solve some of these challenges, their use for long-term treatment can have side effects. Therefore, there are still challenges and limitations to the use of nanoparticles in medicine. Future research hopes to develop the prospect of using nanocarrier strategies to deliver drugs by reducing their synthesis costs and increasing their pharmacokinetic properties in addition to overcoming the limitations of their use. One of the limitations of the present systematic review is the exclusion of some studies that were presented as thesis and the results were not reported in the form of articles. Also, the lack of access to the full text of some studies was another limitation of the study. Furthermore, meta-analysis was not possible due to the diversity in the analysis of different studies and the qualitative expression of the results.

Conclusion

Nanotechnology-based drug delivery systems offer new perspectives on cancer treatment. The development of biomaterials and their targeting for cancer cells using nanotechnology has raised considerable hopes for targeted drug delivery and overcoming the limitations of conventional chemotherapy. Polymeric nanoparticles have been introduced as preferred products for nanocarriers of anticancer drugs due to their easy manufacturing process, biocompatibility and biodegradability. Although the loading of hydrophilic compounds still faces challenges, given the diversity in the fabrication and structure of nanocarriers, it is hoped that it will be possible to encapsulate a variety of molecules. Finally, there are various reports on the importance of using lipid nanoparticles with a suitable coating of hyaluronic acid for targeted delivery of the drug to the target tissue without side effects on other tissues, enhancing drug delivery and increasing therapeutic efficacy.

Abbreviations

SID

Scientific Information Database

MESH

Medical Subject Headings

WoS

Web of Science

PRISMA

Preferred Reporting Items for Systematic Reviews and Meta-Analysis

HA

Hyaluronic acid

Author’s contributions

SHR and NS and KM contributed to the design. MM and EV and FA and MJ prepared the manuscript. SD and SHR assisted in designing the study, and helped in the, interpretation of the study. All authors have read and approved the content of the manuscript.

Funding

Not applicable.

Data availability

Datasets are available through the corresponding author upon reasonable request.

Declaration

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no conflict of interest.