Prospects and Challenges of Chitosan-Based Drug Carriers for Anticancer Agents’ Delivery Against Lung Cancer: A Review

Introduction

As a persistent and critical challenge to global public health, lung cancer significantly undermines human health while imposing a substantial economic burden.^1,2 According to statistics from the International Agency for Research on Cancer (IARC), approximately 2.5 million new cases and 1.8 million deaths were attributed to lung cancer worldwide in 2022, making it the leading cause of cancer-related mortality.³ Smoking remains the primary risk factor for lung cancer-related deaths, while genetic susceptibility, occupational exposures (such as asbestos and radon gas), and environmental factors also contribute to its development and progression,^4,5 as depicted in Figure 1. The World Health Organization (WHO) classifies lung cancer into non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC) based on histopathology. NSCLC predominates, accounting for approximately 85% of cases, which include adenocarcinoma, squamous cell carcinoma, and large cell carcinoma.^6,7 Both NSCLC and SCLC are characterized by high proliferation rates, strong early metastatic potential, and poor prognosis.⁸ Furthermore, the high heterogeneity of lung cancer allows it to occur in any part of the bronchial tree, resulting in clinical symptoms and signs that vary depending on the tumor’s origin.⁸

Figure 1 Risk factors and related pathogenesis of lung cancer.

Current treatment modalities for lung cancer include surgery, radiotherapy, chemoradiotherapy, immunotherapy, and targeted therapy. Clinical decision-making is primarily guided by the tumor type and stage, as well as the patients’ physical condition.^9,10 Despite significant advancements in lung cancer therapy over recent decades, persistent challenges--such as severe adverse effects, low bioavailability, and drug resistance--continue to hinder the clinical effectiveness of these interventions.^11–13 For example, chemotherapeutic agents like cisplatin, paclitaxel, and docetaxel (DTX) can cause side effects, including nausea, vomiting, alopecia, and myelosuppression, some of which may be life-threatening. Additionally, targeted and immunotherapeutic drugs may induce adverse reactions such as rash, cough, diarrhea, and visual impairment.¹⁴ Furthermore, long-term treatment can lead to the development of multidrug resistance (MDR), resulting in treatment failure and tumor recurrence.¹⁵ Therefore, the development of innovative drug delivery systems designed to enhance therapeutic efficacy and reduce toxicity has emerged as a critical area of research in lung cancer therapy.

In recent years, nanotechnology-based drug delivery systems have garnered extensive attention in the field of tumor therapy, offering new strategies for lung cancer treatment by improving drug targeting, reducing toxic side effects, and enhancing drug stability and solubility.^15–17 Currently, FDA-approved nanomedicines such as Abraxane® and Pazenirl® are used in the clinical treatment of lung cancer and have shown advantages in improving drug solubility and prolonging blood circulatory time. However, due to the limitations of systemic administration, they still face challenges in terms of targeting and therapeutic index.¹⁷ CS-based nanodelivery systems offer a promising direction to address these shortcomings, leveraging their unique mucoadhesive properties, excellent biocompatibility, and enhanced drug penetration capabilities.¹⁸ Derived from natural sources, CS exhibits properties such as hemostatic, bacteriostatic, anti-cholesterol, and anticancer effects.^19,20 Furthermore, it can be chemically modified to enhance its solubility, drug-loading capacity, bioavailability, and targeting ability.^21,22 Based on its cationic nature, CS can increase adhesion through electrostatic interactions with mucosal surfaces, prolong the contact time between the drug and mucosa, and facilitate cellular internalization.^23,24 Moreover, the positive charge on its surface enables CS to interact with anionic drugs and polyanionic biomolecules (such as siRNA or DNA), thereby facilitating drug encapsulation and controlled release.²⁵ This characteristic has been utilized in various therapeutic fields, particularly in oncology, where it enhances the specificity and efficacy of chemotherapeutic agents while minimizing systemic toxicity.²⁶ Therefore, this review aims to summarize the latest research advancements on CS-based nanodelivery platforms, explore their potential in drug and gene delivery, and delineate the applications of CS nanocomplexes in phototherapeutic strategies for lung cancer.

Chitosan and Its Physicochemical Properties

CS is a linear polysaccharide polymer formed by the linkage of glucose molecules through β-1,4-glycosidic bonds.²⁷ CS is obtained by the deacetylation of chitin, which is widely found in marine crustaceans such as shrimp and crabs.²⁸ Depending on the conditions of deacetylation, CS exhibits a molecular weight (MW) ranging from 10 to 1000 kDa and a degree of deacetylation (DD) between 70% and 95%.²⁹ Due to the protonation of its amino groups, CS dissolves in acidic solutions with a pH below 6.5, while exhibiting poor solubility under neutral and alkaline conditions.³⁰ As the second most abundant natural polymer after cellulose, CS is an ideal candidate material in pharmacology and therapy, owing to its excellent biodegradability, biocompatibility, and non-toxicity.^31,32 Table 1 illustrates the applications of chitosan-based nanocarriers in lung cancer therapy.

Table 1 Applications of Chitosan-Based Nanocarriers in Lung Cancer Therapy

The MW and DD significantly influence the physicochemical properties and biological activity of CS and its derivatives. CS with a high MW can enhance the stability of complexes and prolong the circulation time of nanoparticles in the bloodstream; however, it reduces their intracellular release rate, thereby increasing tumor targeting selectivity. In contrast, low MW chitosan exhibits the opposite effects.^43,44 DD controls the free amino groups in the CS structure, which not only determines its positive charge density and the capacity to affiliate with DNA/siRNA molecules but also facilitates the evasion of nanovehicles from endolysosomal sequestration, thereby improving transfection efficiency.^43,45 CS with a high DD increases the surface charge density of nanoparticles, significantly enhancing cellular uptake and antitumor effects.^43,46 Additionally, CS with lower MW is often associated with a higher degree of deacetylation, exhibiting enhanced tolerance to amino group protonation and solubility, along with increased toxicity towards malignant cells.⁴⁷

The presence of hydroxyl, amino, and other functional groups in CS molecules enables chemical modifications, including acylation, esterification, alkylation, and grafting. These modifications yield chitosan derivatives with desired physical, chemical, and biological properties,⁴⁸ as depicted in Figure 2. Research indicates that CS possesses anticancer activity due to its cationic nature, MW, and DD, while demonstrating minimal toxicity to non-malignant cells.⁴⁹ CS is more readily protonated and soluble in the acidic tumor microenvironment.⁴⁹ Through electrostatic repulsion, CS disperses and undergoes a conformational change from a tightly curled state to an extended linear form, achieving a ζ potential as high as +30 mV.⁵⁰ This property enables CS to effectively interact with negatively charged tumor cell membranes and tumor vascular endothelial cells, which overexpress anionic surface groups such as phospholipids, glycoproteins, and proteoglycans.^51,52 Consequently, CS can modify cell membrane permeability and penetrate intracellularly to exert antitumor effects by inhibiting the formation of matrix metalloproteinase 9 (MMP-9) protein, as demonstrated in in vitro studies of malignant cell lines. In a physiological environment, CS may exhibit reduced affinity for normal cells, indicating its selective targeting of malignant cells.⁵³

Figure 2 Chitosan derivatives and their properties for improved drug delivery.

The viscosity and mucosal adhesion properties of CS play a pivotal role in regulating the entrapment, absorption, and release of drugs. As a cationic polymer, it engages with the negatively charged membranes of cancer cells, thereby prolonging the residence time at the target site and enhancing absorption capacity.^24,54 The positive charge of CS can also prevent therapeutic agents from being degraded by enzymes during lysosomal transport through the proton sponge effect.⁵⁵ Additionally, the positive charge of CS enables it to disrupt the tight junctions between epithelial cells by activating the protein kinase C pathway, which decreases the transepithelial resistance of the epithelial cell membrane⁵² and disrupts connection proteins such as zonula occludens-1 (ZO-1) and occludin, thereby facilitating paracellular and transcellular drug transport.^56,57 As a result, CS-based drug carriers can be strategically designed to release their therapeutic contents within the tumor microenvironment, achieving precise treatment.

Chitosan-Based Targeted Drug Delivery Systems

Traditional drug delivery systems face critical limitations, including non-specific distribution, low bioavailability, severe adverse reactions, and rapid bodily clearance, which collectively hinder their therapeutic efficacy.⁵⁸ To address the aforementioned challenges, CS-based nanocarriers offer a novel approach for targeted cancer therapy. These nanocarriers not only accumulate selectively in tumor tissues via the retention (EPR) effect, but also can be functionalized with targeting molecules (eg, antibodies or peptides) to enhance their specificity and therapeutic potency against cancer cells.⁵⁹ Furthermore, CS nanocarriers can selectively disrupt tumor cell membranes, enhancing the bioavailability of drugs in lung cancer cells while preserving the safety of healthy cells.⁶⁰ Figure 3 illustrates the mechanisms of CS-NPs for delivering chemotherapeutic drugs through stimuli-responsive, passive, and active targeting strategies.

Figure 3 Mechanisms of CS-NPs for delivering chemotherapeutic drugs through stimuli-responsive, passive, and active targeting strategies.

Medication Delivery Utilizing Passive Targeting

Nanocarriers exploit the hyperpermeability of tumor capillaries and the EPR effect for passive targeting to cancerous sites, where they accumulate due to vascular leakage and insufficient lymphatic drainage in the tumor microenvironment.⁶¹ Moreover, the impaired lymphatic function also facilitates the escape of nanocarriers from defective tumors, enabling them to target lung cancer. Passive targeting relies on the physicochemical properties of nanocarriers, such as size, shape, and surface characteristics, and tumor biological features like vascular distribution and leakage.⁶² Nanoparticles with a particle size less than 200 nm effectively evade immune clearance and prolong circulation time in the bloodstream. Moreover, in the acidic tumor microenvironment, CS protonation induces nanoparticle swelling and subsequent drug release into target malignant cells.^25,63 Altwaijry et al prepared CS-modified lipid nanoparticles (ERT-CLNPs) encapsulating Erlotinib (ERT) through probe sonication and the ionic gelation technique. ERT-CLNPs, with a particle size below 200 nm, exhibited low polydispersity, a cationic zeta potential, and high encapsulation efficiency, while demonstrating excellent gastrointestinal and storage stability. Compared with pure ERT, ERT-CLNPs demonstrated significantly enhanced mucoadhesion and intestinal permeability, while exhibiting superior cytotoxicity toward lung cancer cells.⁶⁴ A poly-lactic glycolic acid (PLGA)/CS nanoparticle was synthesized through a single emulsification and evaporation method for targeted delivery of Meclinertant to treat NSCLC. The Meclinertant encapsulated in PLGA/CS NPs exhibited pH-dependent release, with enhanced drug release under acidic conditions, aligning with the characteristics of the tumor microenvironment. Compared to the free drug, PLGA/CS NPs loaded with Meclinertant demonstrated a more significant anti-proliferative effect on A549 cells. Cell uptake and double staining experiments revealed that PLGA NPs coated with CS substantially enhanced cellular uptake capacity and induced apoptosis in A549 cells.⁶⁵ Sun et al encapsulated Bortezomib (BTZ) and Vincristine (VCR) in polyethylene glycol (PEG)/CS nanoparticles (PEG/CSNPs) to enhance drug solubility. The study revealed that compared to free drugs or single-drug-loaded CSNPs, PEG/CSNPs loaded with BTZ and VCR exhibited significantly enhanced cytotoxicity against NCI-H661, NCI-H460, A549, H157, and H1299 lung cancer cells, demonstrating synergistic effects between BTZ and VCR with substantially reduced half-maximal inhibitory concentrations (IC₅₀). FITC/Annexin-V and PI staining assays indicated that PEG/CSNPs induced apoptosis in NCI-H661 cells by upregulating the expression of pro-apoptotic proteins, such as Bax, Cyt-C, and caspase-3, −8, and −9.⁶⁶ Copper-citrate complexes were loaded into CS nanoparticles (CuCC NPs) using the conventional ion gelation method. The nanoparticles enhanced accumulation within tumor tissues via the EPR effect. In response to the acidic and GSH-rich tumor microenvironment, CuCC NPs enabled the controlled release of Cu²⁺ and mediated efficient chemodynamic therapy to effectively kill A549 cells. Additionally, histopathological and hematological analyses confirmed that CuCC NPs induced massive apoptosis in tumor tissues while maintaining an excellent safety profile.⁶⁷ Nijhawan et al employed a Quality-by-Design approach to encapsulate erlotinib hydrochloride (ERT-HCl) within CS/PLGA NPs. Drug release studies revealed that the nanoparticles sustained drug release for up to 72 h, following zero-order release kinetics, which indicated that the release process was governed by Fickian diffusion.⁶⁸ In a separate study, Karami et al employed double nanoemulsion techniques to synthesize a hydrogel nanocarrier, which was formulated with CS, alumina (γ-Al₂O₃), and graphene quantum dots (GQDs) for targeted delivery of quercetin (QC). Cytotoxicity experiments revealed that the interaction between QC and the nanocarrier resulted in a gradual release, effectively inhibiting the growth and proliferation of lung cancer cells, which indicated the potential of the nanocarrier as a novel nanosystem for targeting tumor tissues.⁶⁹ Ray et al created an inhalable nanocomposite (NIC-CS-PCL-NA) comprising niclosamide (NIC) encapsulated within CS-functionalized poly (ε-caprolactone) (PCL) for NSCLC treatment. In vitro experiments revealed that NIC-CS-PCL-NA exhibited dose-dependent cytotoxic effects, which could effectively enhance autophagic flux, inhibit cellular invasiveness, and induce apoptosis in A549 lung cancer cells. In a mouse model of lung cancer, inhaled NIC-CS-PCL-NA was observed to deposit and retain in the lungs, effectively inhibiting tumor growth while exhibiting minimal toxicity to other major organs. These findings position it as a promising treatment for NSCLC.⁷⁰ The incorporation of zinc phthalocyanine-perfluorinated resin (ZnPc-PFR) and compound 1 into CS-fructose (CH-Fru) yielded CH-Fru@ZnPc-PFR@1, which was further complexed with gefitinib to form the nanocomposites (CH-Fru@ZnPc-PFR@1@Gefitinib). This nanocomposite exhibited dose-dependent inhibitory effects against A549, NCI-H1734, and NCI-H1299 NSCLC cells, with the high-dose group (200 μM) demonstrating stronger inhibitory effect than the low-dose group (100 μM). These findings indicated synergistic effects between compound 1 and gefitinib in suppressing the proliferation of NSCLC cells.⁷¹

Medication Delivery Utilizing Active Targeting

Although passive targeting based on the EPR effect lays the groundwork for nanomedicine delivery, its clinical translation is frequently limited by tumor heterogeneity and variations in vascular leakage, which may lead to heterogeneous drug distribution and off-target toxicity.^72,73 To overcome these limitations, researchers can achieve precise drug delivery via ligand-receptor interactions for specific recognition. Nanocarriers functionalized with ligands, such as folic acid (FA), monoclonal antibodies, and aptamers,¹⁰ can selectively recognize overexpressed target receptors on cancer cell surfaces (such as EGFR, folate receptors, sigma receptors) or target specific membrane proteins on immunosuppressive macrophages within the tumor microenvironment, thereby effectively differentiating them from healthy cells.⁷⁴ As an illustration, Feng et al successfully synthesized CS NPs loaded with bacterial vesicles and DOX for targeted drug delivery to lung cancer cells. Through surface modification with FA, the nanoparticles achieved targeted delivery, induced the release of ROS, and activated the STING signaling pathway via mitochondrial DNA damage, thereby promoting dendritic cell (DC) maturation. Consequently, mature DCs recruited CD4+ and CD8+ cytotoxic T cells and enhance the secretion of IFN-β, IFN-γ, and IL-12.⁷⁵ Patel et al successfully synthesized inhalable polymer nanoparticles encapsulating silibinin, functionalized with FA to confer targeting capability against lung cancer cells. Their findings demonstrated that the nanoparticles significantly reduced the viability of A549 cells compared with free silibinin. Histological analyses showed that, by exploiting the overexpression of FA receptors in lung cancer cells, the nanoparticles effectively accumulated in lung tissue, thus significantly inhibiting tumor volume expansion.⁷⁶ Monoclonal antibodies are widely used for targeting cell receptors due to their high specificity in vivo and stability in serum.⁷⁷ Gold nanoparticles conjugated with CS and loaded with lapatinib were utilized for targeted drug delivery against lung cancer cells. The nanoparticles not only demonstrated dose- and time-dependent cytotoxicity towards A549 cells, but also enhanced attachment to the cell surface and internalization into cancer cells due to their positive charge. PCR analysis revealed that the nanoparticles could significantly downregulated the expression levels of LINC01615 and related genes such as IL11 and MMP14, indicating that they effectively destroyed the tumor growth and metastasis pathway.⁷⁸ Imatinib was loaded into CS polyethylene glycol nanoparticles and modified with Ramucirumab antibody to achieve targeted therapy for A549 cells. The results demonstrated enhanced Imatinib release from the nanoparticles in acidic media compared to physiological conditions. Relative to free Imatinib, the nanoparticles exhibited superior pro-apoptotic effects, and induced G1-phase cell cycle arrest in 8.17% of cells.⁷⁹ Najmeh et al designed an aerosol-assisted synthesis method for CS-based nanocarriers (GefNC) loaded with gefitinib. GefNC exhibited pH sensitivity, with enhanced drug release in acidic environments compared to physiological conditions, indicating potential for targeted delivery to cancer cells. MTT assays demonstrated that the IC₅₀ value of GefNC against lung cancer cells was lower than that of free drugs, while the empty nanocarrier did not elicit appreciable cytotoxicity.⁸⁰ Zhang et al fabricated CS iodoacetamide (CsIA)-coated liposome NPs for lung cancer treatment. Trastuzumab (TZ) was conjugated to the nanoparticle surface to target lung cancer cells overexpressing the HER2 receptor, enabling sustained drug release at the tumor sites. Compared with free drug and unconjugated liposome NPs, the TZ-conjugated NPs loaded with SN-38 accumulated extensively within Calu-3 lung adenocarcinoma cells, and significantly potentiated the anticancer effects on Calu-3 cells.⁸¹ Furthermore, peptides or protein ligands have been utilized for the functional modification of CS, enhancing the specific recognition of cancer targets through active targeting strategies.⁸² In another study, a thermally induced aggregation method was employed to load camptothecin (CPT) into a formulation of pH/temperature dual-responsive poly(acrylic acid)-b-poly(N-isopropyl acrylamide) block copolymer, which were then surface-modified with CS and fucoidan (Fu) to form Fu-CPT-NPs for lung cancer treatment. Cell experiments confirmed that Fu-CPT-NPs exhibited excellent biocompatibility and antitumor activity. The NPs effectively delivering CPT to H460 lung cancer cells through the interaction of Fu with P-selectin, enhancing cellular uptake and inducing apoptosis in cancer cells. Animal studies showed that Fu-CPT-NPs inhibited lung tumor growth by promoting tumor cell apoptosis without causing significant tissue damage.⁸³ Kordbacheh et al embedded DOX in a ZIF-8 (Zn, Fe) bimetallic-organic framework (BMOF), whose surface was coated with curcumin-loaded CS-grafted- polycaprolactone (Cs-g-PCL) and subsequently functionalized with transactivator of transcription (TAT) peptide for targeted delivery. Cs-g-PCL regulated the release rate of curcumin through pH-dependent behavior, promoting drug accumulation in tumor tissues, while TAT peptide modification further prolonged drug release duration, establishing a synergistic controlled-release system. In vitro studies indicated that the combined delivery of DOX and Cur significantly reduced the viability of A549 lung cancer cells, with the TAT peptide and DOX-Cur combination exhibiting a synergistic apoptotic induction effect. In vivo studies in tumor-bearing mice showed that the TAT peptide-modified nanosystem significantly inhibited tumor growth compared to the unmodified Cs-g-PCL/Cur/BMOF/DOX, reaching a maximum relative tumor volume inhibition rate of 0.25.⁸⁴ Hu et al designed a copper ferrite nanocomposite, which was functionalized with vitamin E succinic acid (VES)- ethylene glycol CS polymer (GV). The nanocomposites utilized the protonation of GV polymer and polylysine (PL) in the acidic tumor microenvironment to facilitate drug release, along with the active targeting of human serum albumin to specifically induce tumor cell death. The nanocomposites exhibited cytotoxicity against lung cancer cells by elevating intracellular ROS levels, thus triggering cuproptosis and ferroptosis. Moreover, the nanocomposites not only showed superior antitumor efficacy and biocompatibility compared to carboplatin, but also potentially inhibited tumor cell growth by activating the p53 signaling pathway. In vivo studies showed that the nanocomposite effectively suppressed the growth of subcutaneous A549 tumors in nude mice.⁸⁵

Medication Delivery Utilizing Magnetic Targeting

Magnetic targeting is a technique that harnesses external magnetic fields to direct magnetic drug carriers to the target site. For instance, magnetic nanoparticles can direct and concentrate drugs in specific areas of the body under the influence of an external magnetic field.⁸⁶ Controlled by an external magnetic field, composites formed from CS and magnetic nanoparticles are directed to the tumor site, ensuring targeted drug delivery with minimal impact on surrounding healthy tissues.⁸⁷ Yuly et al developed CS-based microencapsulated systems incorporating magnetite nanoparticles and microzeolites for gemcitabine (GEM) delivery in lung cancer therapy. This system exhibited a drug encapsulation efficiency of over 99% and achieved a slow and controlled release of GEM at pH 7.4 and 5.0 for up to 24 hours. Cell viability assays revealed that the cell viability of A549 and H1299 cells treated with the nanoparticles was significantly lower than that of cells treated with pure GEM.⁸⁸ Peptidyl arginine deiminase 4 inhibitor was loaded into magnetic CS NPs to achieve dual-target drug delivery via magnetic targeting and pH-responsive release. In vitro experiments demonstrated that an external magnetic field enhanced the uptake of nanoparticles by cancer cells, inducing tumor cell apoptosis. In vivo studies demonstrated that nanoparticles effectively inhibited the growth and metastasis of lung cancer in Lewis mice by suppressing the expression of citrulline histone (H3cit) under the influence of an external magnetic field.⁸⁹ Additionally, Sheng et al prepared novel core-shell Fe₃O₄ NPs, which were synthesized by functionalizing gold NPs and CS-agarose composites. In vitro experiments showed that nanocomposites inhibited the proliferation of various cancer cell lines, exhibiting IC₅₀ values of 160, 173, 142, and 212 for NCI-H1563, NCI-H1573, NCI-H1975, and NCI-H661 cells, respectively.⁹⁰ A composite capsule was prepared by an ionic gelation method, in which GEM was incorporated into CS-magnetite-zeolite composite matrix, and the synergistic regulation of drug loading and magnetic targeting was realized. The composite capsule exhibited an encapsulation efficiency of over 86% for GEM and showed a sustained release profile over 24 hours. Cell viability assays revealed that the composite capsule exhibited similar cytotoxicity to free GEM but at a 45-fold lower concentration, suggesting its potential to reduce the required dosage and potential side effects.⁹¹

Chitosan Miscellaneous Nanocomplexes in Lung Cancer Therapy

CS plays multiple roles in the development of metal-based, carbon-based, and silica-based hybrid nanoparticles. Due to its cationic nature, the primary amine groups in the CS structure, along with adjacent hydroxyl groups, can reduce metal cations (such as Ag⁺, Au³⁺), thereby promoting the synthesis of metal nanoparticles (MNPs). In addition, the modification of gold NPs, silver NPs, and quantum dots with CS enhances their stability and optical properties while reducing their cytotoxicity.^92,93 Aldosari et al doped gold into modified graphene oxide CS NPs (GO@CSP/Au NPs) for the treatment of lung cancer. MTT assays demonstrated that GO@CSP/Au NPs inhibited A549 cell viability in a dose-dependent manner, with an IC₅₀ value of 35.2 μg/mL, indicating potential antineoplastic efficacy against lung cancer.⁹⁴ Wang et al synthesized silver nanoparticles (AgNPs) mediated by curcumin on CS-modified kaolin. The kaolin minerals promoted the in-situ nucleation of AgNPs on the surface and within the interlayers of the modified kaolin, effectively preventing particle aggregation and achieving a uniform distribution with a particle size range of 20–30 nm. The nanocomposites possessed potent antioxidant activity and exhibited IC₅₀ values of 110 μg/mL, 96 μg/mL and 38 μg/mL against HLC-1, LC-2/ad and PC-14 lung cancer cells, respectively, among which PC-14 showed the strongest anticancer effect.⁹⁵ In another study, the integration of Cu₂O into CS/guar gum (GG) composite hydrogel successfully yielded nanoparticles (CS-GG/Cu2O NPs). This synthesis approach demonstrated CS-GG/Cu2O NPs could inhibit the expression of pivotal proteins (PI3K, AKT, mTOR) in the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling pathway in NCI-H661 lung cancer cells, thereby increasing ROS levels and promoting apoptosis, and inhibiting the cell cycle in lung cancer cells.⁹⁶ Abiraman et al utilized acryloyl CS grafted with [2-(methacryloyloxy)ethyl] trimethyl ammonium chloride acryloyl piperazinium chloride-based polymers as a supporting agent to synthesize spherical zinc oxide nanoparticles (ZnO NPs). The results indicated that, in comparison to uncapped ZnO, the capped ZnO NPs exhibited a more potent inhibitory effect on the A549 cancer, with nanoparticles capped with a hybrid polymer containing an acryloyl ethyl substituent in the piperazine ring exhibiting the maximum inhibitory effect on cancer cells.⁹⁷ Khichar and associates extracted components from Terminalia Arjuna using water and ethanol to functionally modify CS-loaded AgNPs through biological methods. Cytotoxicity assays revealed that the two types of nanoparticles inhibited the viability of A549 lung cancer cells in a dose-and time-dependent manner, with IC₅₀ values of 400 μg/mL and 300 μg/mL after 24 and 72 hours, respectively.⁹⁸ Venkidasamy and coworkers synthesized vanillic acid nanocomposites incorporating silver and CS NPs. Apoptosis assays revealed that the nanocomposites induced cellular apoptosis by upregulating the expression of pro-apoptotic proteins, such as BAX, Casp3, Casp7, and cytochrome C, while concurrently suppressing the gene expression of Bcl-2. Moreover, the nanocomposites exhibited a dose-dependent increase in the expression of the tumor suppressor gene p53, indicating a multi-pathway synergistic mechanism underlying its pro-apoptotic effects.⁹⁹ Thenmozhi et al developed a biosynthesized curcumin-loaded cadmium oxide-silicon dioxide nanocomposites utilizing Pithecellobium dulce leaf extract as a reducing and capping agent. Their research indicated that the nanocomposites exhibited a dose-dependent cytotoxicity effect against various human cancer cell lines, with a particularly pronounced impact on A549 lung cancer cells (IC₅₀ value of 31.10 μg/mL). Furthermore, the nanocomposites demonstrated good antibacterial activity and anti-inflammatory effects.¹⁰⁰

Nanosystems composed of CS and quantum dots endow nanoparticles with stability, functionality, cell binding and uptake, while also modulating drug release, increasing tumor drug accumulation, inhibiting degradation, and reducing toxicity associated with inorganic metal ions.^101–103 Tahamtan et al encapsulated QC within CS/starch/GQDs/titanium dioxide porous nanocomposite using a water-in-oil-in-water (W/O/W) multiple emulsion method. The release behavior of QC from the nanocomposites followed the Fickian diffusion mechanism and exhibited responsiveness to environmental stimuli such as pH and temperature. MTT assays showed that the nanocarrier exhibited good biocompatibility; compared to free QC, the drug-loaded nanocomposites significantly enhanced cytotoxicity against the A549 lung cancer cell line.¹⁰⁴ Furthermore, carbon nanomaterials (CNMs) have been widely applied in the biomedical field due to their unique physicochemical properties, including high stability, conductivity, thermal conductivity, and optical properties.¹⁰⁵ The application of CNMs coated with CS has been shown to augment their stability, diminish cytotoxicity, and exhibit pH-responsive release characteristics in tumor microenvironments with pH levels below 6.5, thereby effectively facilitating drug delivery to tumor tissues.^106,107 Fateme et al created a pH-responsive nanocarrier (CS/HNTs/GQDs) composed of CS, halloysite nanotubes (HNT) and GQDs for targeted QC to lung cancer cells. The incorporation of HNTs enhanced QC drug loading capacity and encapsulation efficiency, while the presence of GQDs accelerated QC release kinetics under different pH conditions. MTT assays revealed that CS/HNTs/GQDs@QC exhibited the strongest cytotoxicity against both L929 and A549 cells compared to other groups, with significantly greater effects on A549 cells. Furthermore, CS/HNTs/GQDs@QC induced the highest apoptotic proportion of apoptosis, accounting for 74.4% early apoptosis and 5.15% late apoptosis.¹⁰⁸ AbouAitah et al fabricated multi-walled carbon nanotubes (MWCNTs) loaded with ferulic acid (FUA) and diosgenin (DGN) for lung cancer treatment. Initially, the MWCNTs were functionalized with carboxylic acid or amine functionalities, followed by coating with CS or a CS-stearic acid complex. Cytotoxicity assays showed that, compared with free DNG and FUA, the MWCNTs exhibited a significantly superior inhibitory effect on the viability of A549 cells. The MWCNTs loaded with DGN and FUA exhibited synergistic and additive interactions, but antagonistic effects were observed at elevated concentrations. Consequently, this nanosystem has emerged as a promising drug delivery platform, which has the potential to enhance the therapeutic activity of natural prodrugs.¹⁰⁹

Chitosan-Alginate Nanocarrier in Lung Cancer Therapy

CS and alginate are biopolymers with opposite charges. CS, a cationic polysaccharide, contains amino groups on its molecular chains that can be protonated in an acidic environment, thereby conferring cationic properties to CS. Due to its cationic nature, CS enables electrostatic interactions with negatively charged polymers, providing unique advantages in chemotherapeutic drug delivery. Alginate, an anionic polysaccharide, is composed of α-L-guluronic (G) and β-D-mannuronic (M) acids.¹¹⁰ The carboxyl groups in its structure confer good water solubility and pH responsiveness when the pH exceeds the pKa values of guluronic and mannuronic acids (3.38 and 3.65).¹¹¹ The protonated amino groups of CS electrostatically interact with carboxylate groups of alginate, forming a variety of polyelectrolyte complexes.¹¹² Eldin et al designed a novel core-shell nanocarrier system where aminated mesoporous silica nanoparticles (MSNs) served as the core, encapsulating DTX. An intermediate layer of alginate controlled the release of DTX, while the outermost layer, composed of GMC-loaded PEG-CS, ensured the stealth properties and rapid dissemination of GMC. The core-shell nanocarriers were capable of depositing in the lungs and demonstrated excellent aerosolization behavior. Compared with the individual drugs, the combined use of both drugs within the nanocarriers elicited a stronger cytotoxic effect against A549 cells. Additionally, in vivo biological distribution studies confirmed the accumulation of the nanocarriers in the lungs, and histopathology assessments indicated the biological safety of the core-shell nanocarriers.¹¹³ Gefitinib was loaded into self-assembled CS/alginate nanocarriers, which displayed pH sensitivity, releasing a higher concentration of gefitinib in acidic conditions. MTT assays revealed that the nanocarriers had a significantly lower IC₅₀ value than free gefitinib, with no appreciable cytotoxic effects observed.¹¹⁴ Yaşar et al developed a CS-alginate liposome to encapsulate mallow extract. The anti-proliferative efficacy of the liposome loaded with mallow extract was evaluated using the CCK8 assay in A549 cell. Treatment of A549 cells with these liposomes elicited a dose- and time-dependent cytotoxic response. As the treatment time extended to 48 hours, the amount of extract released from the liposomes gradually increased, resulting in increased cell death.¹¹⁵

Chitosan Nanocomposites in Delivery of Bioactive Compounds for Lung Cancer

Bioactive compounds derived from plants, such as polyphenols, flavonoids, and polysaccharides, possess various biological activities including antibacterial, anti-inflammatory, anti-aging, and anticancer effects.¹¹⁶ They can exhibit antitumor activities by interfering with signaling pathways (eg, PI3K/Akt, NF-κB, TGF β) and regulating cellular metabolism.¹¹⁷ Moreover, their beneficial impacts on cancer treatment have been confirmed in both in vitro and in vivo studies.¹¹⁸ Nevertheless, these phytochemicals possess some physicochemical limitations, such as poor solubility and low bioavailability, as well as pharmacokinetic issues such as short half-life, non-specific biodistribution, and narrow therapeutic window, which restrict their clinical translation.¹¹⁹ CS-based nanocomplexes, with their biocompatibility, modifiability, and sustained-release properties, can effectively encapsulate phytochemicals, enhance their stability and bioavailability, prolong circulation time, improve targeting, and reduce toxicity to normal tissues.¹²⁰ El-Ghannam et al synthesized a beetroot@CS nanocomposite (BR@CS NC) by electrostatic interaction. MTT assays demonstrated that the cytotoxicity of BR@CS NC on A549 cells was dose-dependent, reaching 89% cell mortality at 250 μg/mL, which was significantly higher than that of free BR (5–7%). A549 cells treated with BR@CS NC not only exhibited extensive late-stage apoptosis and numerous necrotic changes but also elevated expression levels of the Caspase 3 (CasP3) and p53 genes.¹²¹ Abulaiti et al fabricated an oleanolic acid-conjugated CS nanocomplex (OAC) and explored its anticancer efficacy against lung cancer. In A549 and NCI-H460 lung cancer cells, OAC significantly inhibited cell viability, migration, and invasion, while inducing apoptosis and autophagy in a dose-dependent manner. Western blot and immunofluorescence analyses revealed that OAC attenuated the phosphorylation of signal transducer and activator of transcription 3 (STAT3) in lung cancer cells, leading to the upregulation of LC3-II protein expression and the formation of GFP-labeled LC3-positive autophagic vacuoles. This indicated that OAC induced autophagy by modulating the STAT3/Bcl-2 pathway, thereby exerting its anticancer effects.¹²² Alali et al developed a spray-dried solid dispersion of apigenin (AGN) based on CS (AC2 SDSD), and evaluated its antiproliferative efficacy against A549 cells. Compared with pure AGN, AC2 SDSD not only exhibited stronger antioxidant activity but also significantly potentiated antiproliferative effects in A549 cells. Its mechanism of action involved the inhibition of the PI3K/Akt signaling pathway, activation of the apoptosis pathway, promotion of cancer cell death, and inhibition of lung tumor growth.¹²³ Gomathi et al created a new drug delivery material by conjugating CS with Spondias pinnata phytocompounds, and evaluated the cytotoxicity of the composites against A549 cells. The results revealed that compared with the leaf extraction of Spondias pinnata, the composites could significantly reduce the cell viability.¹²⁴ CS NPs loaded with star anise extract (SAE) were synthesized by ionic gelation. Compared with free SAE, CSNPs loaded with SAE inhibited NCl-H460 lung cancer cells in a dose-dependent manner, suggesting enhanced cellular uptake of the nanoparticles. In vivo studies demonstrated that CSNPs loaded with SAE exhibited the most potent antitumor activity, as seen by decreased levels of malondialdehyde, tumor protein 53 (p53), tumor necrosis factor-alpha and fibronectin, compared with other treatment groups.¹²⁵

Table 2 Chitosan-Based Nanostructures for Cancer Phototherapy

Chitosan Nanocarriers for Phototherapy and Gene Delivery

Phototherapy includes two modalities: photodynamic therapy (PDT) and photothermal therapy (PTT). Although they differ in mechanisms of action, both ultimately induce tumor cell death to achieve antitumor effects (Table 2 provides an overview of CS-based nanostructures for cancer phototherapy). PDT selectively kills cancer cells through the cytotoxic action of photosensitizers (PS) activated by light.^131–133 Its non-invasive nature, minimal side effects, and targeting advantages have garnered significant attention in clinical applications.^134–136 However, the uneven distribution and poor penetration of PS within tumors^137,138 result in suboptimal efficacy and potential resistance, which hinder the effectiveness of PDT.^139,140 Therefore, researchers have employed the strategy of loading PS onto nanomaterials, which promotes ROS generation, disrupts the oxidative-antioxidative balance, inhibits tumor growth, damages DNA, and ultimately leads to cancer cells death, thereby providing a new approach to enhance the efficacy of PDT.³¹ Guo et al developed a novel probiotic-based PS delivery system (EWC), utilizing Escherichia coli Nissle 1917 (EcN) as the carrier. The system facilitated the electrostatic adsorption of Chlorin e6 (Ce6) to the surface of EcN with the assistance of water-soluble CS. In vitro experiments demonstrated that EWC exhibited low toxicity to normal cells but strong photodynamic cytotoxicity against A549 cells. 3D fluorescence imaging revealed that EWC could penetrate tumor spheroids and effectively inhibit their growth. In mouse models, EWC not only demonstrated good biocompatibility but also enhanced the targeted distribution and accumulation of Ce6 at tumor sites, effectively inhibiting tumor progression upon light stimulation.¹⁴¹ In contrast, PTT utilizes the photothermal conversion capability of near-infrared (NIR) light-responsive nanomaterials to convert light energy into thermal energy, inducing cancer cell death through localized hyperthermia.¹⁴² Its deep tissue penetration capability and minimal damage to normal tissues render it an effective therapeutic strategy.¹⁴³ An et al designed a CS-based hydrogel nanocomposite with NIR light response, which induced apoptosis in lung cancer cells through a triple synergistic mechanism combining photothermal, chemodynamic, and chemotherapeutic effects. The nanocomposite, loaded with carbon nanodots, platinum nanoparticles (PtNPs), and DOX, demonstrated favorable sustained drug release properties. Upon NIR irradiation, carbon nanodots amplified oxidative stress and initiated apoptosis through ROS generation and activation of caspase-3/8/9 pathways. The synergistic effects of PtNPs and DOX involved PtNPs-mediated enhancement of ROS production and DOX-induced amplification of cytotoxic effects, resulting in pronounced cytotoxicity and apoptotic induction in A549 and H1299 cells under NIR stimulation. Consequently, this innovative nanocomposite system offered a promising strategy for targeted lung cancer therapy.¹⁴⁴

With the emergence of chemoresistance, gene delivery technology has witnessed substantial advancements in exploring new avenues for cancer treatment and enhancing drug sensitivity.^145,146 However, naked gene delivery faces challenges such as easy degradation in the bloodstream and poor cellular internalization, resulting in low transfection efficiency.¹⁴⁷ To overcome these challenges, researchers have proposed strategies utilizing nanoscale structures to achieve precise gene targeting.^148,149 Furthermore, CSNPs have emerged as ideal vectors in the field of gene delivery because of their excellent biocompatibility, biodegradability, and the ability to protect nucleic acids. The cationic nature of CS enables it to interact electrostatically with negatively charged nucleic acids, forming stable complexes that protect genetic material from enzymatic degradation and facilitate cellular uptake of nucleic acids.¹⁵⁰ In cancer treatment, small interfering RNA (siRNA) downregulates the expression of oncogenes by binding to complementary mRNA, inhibiting cancer cell proliferation, inducing cell death, and preventing lymphatic metastasis.¹⁴² Zhang et al created a novel gene vector, vitamin E succinate-CS-histidine (VES-CTS-His, VCH), which was combined with STAT3-shRNA recombinant plasmid by electrostatic interactions to form stable particles. The nanoparticles exhibited typical nanoscale dimensions, allowing them to accumulate in tumor tissues via the EPR effect. They could enter tumor cells through endocytosis. VCH exhibited good pH responsiveness, allowing it to dissociate in the acidic microenvironment of tumor tissues and release the recombinant plasmid. By leveraging the RNA interference effect, the expression of STAT3 protein was downregulated, which inhibited the PI3K/mTOR signaling pathway. This action significantly enhanced the effect of inducing apoptosis and inhibiting growth in NSCLC cells, thereby achieving the goal of tumor treatment.¹⁵¹ Furthermore, several studies have combined chemotherapeutic drugs with siRNA in nanoscale drug delivery systems, which can synergistically enhance the anticancer effects and improve treatment efficiency. DOX and Bcl-2 siRNA were incorporated into a pH-sensitive nanofibrous hybrid formulation consisting of liposome, CS, poly (ethylene oxide), and PCL (CS/PEO/PCL) for treating lung cancer. Cellular experiments indicated that the liposome-incorporated CS/PEO/PCL core-shell nanofibers loaded with DOX and Bcl-2 achieved sustained release of DOX and siRNA, and mediated the maximum down-regulation of Bcl-2 siRNA, which promoted apoptosis and reduced the viability of A549 lung cancer cells. In vivo experiments showed that mice treated with CS/PEO/PCL nanofibers incorporated with DOX-Bcl-2 liposomes could inhibit tumor growth to the greatest extent without changing their body weight.¹⁵² Sun et al synthesized a novel multifunctional micelle (VCPH), which is composed of VES, CS, histidine and polyethylene glycol monomethyl ether. DOX was incorporated into the self-assembled VCPH micelles, and the recombinant pGPU6/GFP/Neo STAT3-shRNA (pDNA) was loaded by electrostatic action, thereby forming dual-loaded nanoparticles (DOX/VCPH/pDNA). The nanoparticles exhibited pH-responsive drug release behavior, with a higher release rate in acidic environments compared to neutral conditions. The nanoparticles upregulated PTEN protein expression, which inhibited the PI3K/AKT pathway and induced apoptosis of adjacent tumor cells. They also downregulated CD31 protein expression, inhibited angiogenesis, and decreased tumor cell viability. In addition, the nanoparticles significantly downregulated the expression level of STAT3 protein, thus enhancing the efficacy of chemotherapy.¹⁵³ The construction and development of this dual-load collaborative delivery system provides important enlightenment for the combined treatment of tumors and offers an effective approach for the precise treatment of NSCLC.

Conclusion and Future Perspective

Lung cancer remains a formidable challenge in oncology, necessitating innovative and targeted treatment approaches. Traditional treatments are costly and are associated with issues such as non-specificity, low bioavailability, and rapid clearance. CS-based nanodelivery systems have emerged as promising platforms for lung cancer therapy due to their unique biocompatibility, cationic properties, and functional versatility. Extensive research has been conducted on CS nanocarriers to explore their distinctive properties and multifunctionality, particularly in targeted cancer therapy and drug delivery. These nanocarriers efficiently load chemotherapeutic drugs, increasing the intracellular accumulation of drugs, enhancing cytotoxic effects, reducing chemoresistance, and minimizing side effects. They also deliver biomacromolecules such as nucleic acids, proteins, and peptides, prolonging their circulation time, enhancing intracellular accumulation, and protecting them from degradation. Notably, despite the delivery of natural active substances being largely overlooked, studies have demonstrated the potential of CS nanocarriers to inhibit lung cancer. The combination of phototherapy and chemotherapy mediated by CS nanocarriers can synergistically contribute to lung cancer treatment. Furthermore, CS nanocarriers can be surface-modified to achieve precise release of active drugs in the TEM, reducing toxicity to normal tissues.

However, CS-based nanoformulations face several limitations. The drug-loading capacity of CS-based nanoformulations depends on the physicochemical properties of the drug and the preparation techniques. Additionally, due to the aggregation characteristics of nanocarriers, the stability of the formulation in physiological environments is reduced, leading to issues such as drug crystalline transformation, heterogeneity in particle size distribution, and burst release effects. Although CS-based nanoformulations have been proven effective in animals, their clinical translation faces multiple challenges. On one hand, the current safety profile of CS nanocarriers remains insufficient, and there is a lack of adequate clinical data to confirm their safety and efficacy in humans. On the other hand, species-dependent immune responses and the differences between mouse models and human pathophysiology collectively hinder their progression toward clinical application. Additionally, the industrialization process encounters bottlenecks such as the low water solubility of CS, limited drug-loading capacity, and difficulties in quality control for large-scale production. Future research could focus on improving the solubility and drug-loading capacity of CS through physical and chemical modifications, and constructing multi-component tumor-homing nanosystems. This approach would enhance the scalability of production, tumor-targeting ability, and therapeutic efficacy of CS-based nanoformulations, thereby providing a solid foundation for their clinical translation and industrial application.