Colloids and Surfaces B: Biointerfaces2018ResearchDrug Delivery

A novel MUC1 aptamer-modified PLGA-epirubicin-PβAE-antimir-21 nanocomplex platform for targeted co-delivery of anticancer agents in vitro and in vivo

Amirhossein Bahreyni, Mona Alibolandi, Mohammad Ramezani, Atefeh Sarafan Sadeghi, Khalil Abnous, Seyed Mohammad Taghdisi29 September 2026DOI 10.1016/j.colsurfb.2018.12.006

Summary

Conventional chemotherapy for breast and colon cancers suffers from low specificity, rapid drug clearance, and severe toxic side effects on healthy tissues, while chemoresistance and metastasis remain major causes of cancer-related mortality. There is an unmet need for targeted delivery systems that can co-deliver chemotherapeutic agents with oligonucleotide-based therapeutics (e.g., miRNA inhibitors) to enhance efficacy and reduce off-target. Epi Release Profile: - ~80% Epi released at pH 5.5 (tumor tissue/lysosomal pH) after 72 h - ~28% Epi released at pH 7.4 (physiological blood pH) after 72 h - Demonstrates pH-responsive release mediated by PβAE.

Keywords

PLGA Cellular uptake Polymeric Chemotherapy DNA Targeted delivery Tumor microenvironment

Purpose: Conventional chemotherapy for breast and colon cancers suffers from low specificity, rapid drug clearance, and severe toxic side effects on healthy tissues, while chemoresistance and metastasis remain major causes of cancer-related mortality. There is an unmet need for targeted delivery systems that can co-deliver chemotherapeutic agents with oligonucleotide-based therapeutics (e.g., miRNA inhibitors) to enhance efficacy and reduce off-target toxicity. MiR-21 is an oncomiR overexpressed in multiple cancers that promotes tumor growth, migration, invasion, and chemoresistance, making it an attractive target for inhibition alongside conventional chemotherapy.

Hypothesis: If epirubicin (Epi) and antimir-21 are co-delivered via a MUC1 aptamer-modified PLGA-PβAE nanocomplex, then the system will selectively internalize into MUC1-overexpressing cancer cells (MCF-7 and C26), synergistically reduce cell viability and inhibit tumor growth, while sparing MUC1-negative non-target cells (CHO) and reducing systemic toxicity compared to free Epi.

Aims: Primary aim: Design and fabricate a targeted co-delivery system based on two biocompatible polymers — poly(β-amino ester) (PβAE) as a pH-responsive core for antimir-21 condensation and PLGA as a reservoir for epirubicin — with a MUC1 aptamer as the targeting ligand.

Secondary aim 1: Characterize the physicochemical properties of the nanocomplex (particle size, zeta potential, morphology, Epi loading, pH-dependent release profile).

Secondary aim 2: Evaluate in vitro cellular uptake, cytotoxicity, and synergetic effect of Epi + antimir-21 co-delivery in MUC1-positive (MCF-7, C26) vs. MUC1-negative (CHO) cell lines.

Secondary aim 3: Assess in vivo antitumor efficacy in C26 tumor-bearing BALB/c mice compared to free Epi and control groups.

Delivery system:

Component: Polymer Core; Description: Poly(β-amino ester) (PβAE) — pH-responsive, positively charged, biodegradable; condenses antimir-21 via electrostatic interaction

Component: Outer Shell; Description: PLGA (poly(D,L-lactide-co-glycolide)) — biocompatible, biodegradable, FDA-approved; electrostatically deposited onto PβAE-antimir-21 complex; serves as reservoir for Epi

Component: Targeting Ligand; Description: MUC1 aptamer (5TR1 sequence: 5'-NH₂-GAAGTGAAAATGACAGAACACAACA-3') — covalently attached to PLGA surface via NHS-activated groups

Component: Payload 1; Description: Epirubicin (Epi) — anthracycline chemotherapeutic; intercalates DNA, inhibits topoisomerase II, generates ROS

Component: Payload 2; Description: Antimir-21 — oligonucleotide inhibitor of miR-21 (oncomiR); 5'-TCAACATCAGTCTGATAAGCTA-3'

Component: Nanocomplex Size; Description: 210.4 ± 10.14 nm (MUC1 aptamer-modified); 204.7 ± 3.7 nm (unmodified)

Component: Zeta Potential; Description: +9.7 ± 0.2 mV (MUC1 aptamer-modified); +2.29 ± 0.34 mV (unmodified); PβAE-antimir-21 conjugate: +19.8 ± 3.13 mV

Component: Epi Loading Efficiency; Description: ~1%

Component: Key Design Feature; Description: Neutral charge of PLGA-Epi-PβAE-antimir-21 nanocomplex (without MUC1 aptamer) prevents non-specific internalization; pH-sensitive PβAE triggers drug release at tumor tissue pH (5.5)

Approach: In Vitro Model: - Cell lines: MCF-7 (human breast carcinoma, MUC1-positive), C26 (murine colon carcinoma, MUC1-positive), CHO (Chinese hamster ovary, MUC1-negative) - Treatment groups: Epi alone, MUC1 aptamer-modified nanocomplex, nanocomplex (without aptamer), MUC1 aptamer-modified nanocomplex without Epi, MUC1 aptamer-modified nanocomplex without antimir-21, MUC1 aptamer-modified nanocomplex without Epi and antimir-21 - Doses: Epi at IC₅₀ concentrations — 3.5 μM (MCF-7), 2.0 μM (C26), 2.5 μM (CHO) - Incubation: 3 h treatment, then 72 h in fresh medium for MTT; 4 h for uptake studies

In Vivo Model: - Animals: Female BALB/c mice (8 weeks, 25–30 g), n = 5 per group - Tumor model: C26 cells (5 × 10⁵) injected into right flank; treatment initiated at ~50 mm³ tumor volume - Groups (6): (1) PBS control, (2) MUC1 aptamer-modified nanocomplex (1.2 mg/kg Epi), (3) Epi alone (1.2 mg/kg), (4) nanocomplex without antimir-21, (5) nanocomplex without Epi, (6) nanocomplex (unmodified) - Administration: Intravenous tail vein injection (single dose) - Monitoring: Tumor volume measured every 4 days for 20 days

Key methods:

Technique: ¹H NMR & GPC; Purpose: Confirm PβAE synthesis; determine molecular weight (Mₙ = 1516, Mw = 1820, PDI = 1.2)

Technique: Agarose gel electrophoresis (2.5%); Purpose: Assess antimir-21 condensation by PβAE; confirm MUC1 aptamer conjugation on nanocomplex surface

Technique: Dynamic light scattering (DLS); Purpose: Measure particle size and zeta potential

Technique: Scanning electron microscopy (SEM); Purpose: Determine morphology and homogeneity (spherical, ~200 nm)

Technique: UV-Vis absorbance at 480 nm; Purpose: Quantify Epi loading and release

Technique: MTT assay; Purpose: Evaluate in vitro cytotoxicity (cell viability %)

Technique: Flow cytometry (BD Accuri C6); Purpose: Quantify cellular uptake (FL2 log intensity for Epi fluorescence)

Technique: Fluorescence microscopy; Purpose: Visualize intracellular distribution of nanocomplex

Technique: In vivo tumor volume measurement; Purpose: Assess antitumor efficacy (vernier scale, every 4 days)

Key results: Epi Release Profile: - ~80% Epi released at pH 5.5 (tumor tissue/lysosomal pH) after 72 h - ~28% Epi released at pH 7.4 (physiological blood pH) after 72 h - Demonstrates pH-responsive release mediated by PβAE protonation and increased Epi solubility at acidic pH

Cellular Uptake (Flow Cytometry — FL2 log intensity): - MCF-7: Untreated = 298 ± 15; Epi = 1238 ± 120; MUC1 aptamer-modified nanocomplex = 2023 ± 160 (p < 0.05 vs. Epi) - C26: Untreated = 1395 ± 110; Epi = 6280 ± 230; MUC1 aptamer-modified nanocomplex = 17300 ± 530 (p < 0.05 vs. Epi) - CHO: Untreated = 1259 ± 46; Epi = 5980 ± 660; MUC1 aptamer-modified nanocomplex = 2740 ± 70 (significantly lower than Epi, p < 0.05)

In Vitro Cytotoxicity (MTT — Cell Viability %):

Treatment: MUC1 aptamer-modified nanocomplex; MCF-7: 20.67 ± 0.6%; C26: 9.51 ± 0.14%; CHO: 82.77 ± 0.85%

Treatment: Nanocomplex (no aptamer); MCF-7: 83.58 ± 3.3%; C26: 88.63 ± 5.29%; CHO: 93.61 ± 1.59%

Treatment: MUC1 aptamer-modified nanocomplex without Epi; MCF-7: 60.56 ± 0.67%; C26: 58.74 ± 0.84%; CHO: 85.68 ± 2.14%

Treatment: Epi alone; MCF-7: 55.44 ± 0.2%; C26: 54.86 ± 4.92%; CHO: 45.77 ± 0.4%

Treatment: MUC1 aptamer-modified nanocomplex without antimir-21; MCF-7: 42.12 ± 4.98%; C26: 38.18 ± 3.48%; CHO: 94.47 ± 0.67%

Treatment: MUC1 aptamer-modified nanocomplex without Epi and antimir-21; MCF-7: 86.6 ± 2.16%; C26: 92.16 ± 2.65%; CHO: 95.68 ± 3.44%

In Vivo Antitumor Efficacy (Tumor Volume at Day 20, mm³): - PBS control: 4807.73 ± 74.89 mm³ - Nanocomplex (no aptamer): 3845.99 ± 180.57 mm³ - Epi alone: 2619.11 ± 156.54 mm³ - MUC1 aptamer-modified nanocomplex without antimir-21: 1537.99 ± 76.13 mm³ - MUC1 aptamer-modified nanocomplex without Epi: 1869.4 ± 37.58 mm³ - MUC1 aptamer-modified nanocomplex (full system): 739.28 ± 71.47 mm³ (p < 0.05 vs. all other groups)

Interpretation: The authors claim that the MUC1 aptamer-modified PLGA-Epi-PβAE-antimir-21 nanocomplex represents a novel, biocompatible, and biodegradable targeted co-delivery platform that significantly enhances antitumor efficacy through: (1) selective MUC1 receptor-mediated endocytosis into cancer cells, (2) pH-responsive Epi release in the acidic tumor microenvironment, and (3) synergistic inhibition of tumor growth by combining chemotherapeutic (Epi) and oligonucleotide (antimir-21) agents. The system reduced tumor growth in vivo far more effectively than free Epi and other control treatments, while minimizing toxicity to MUC1-negative non-target cells. The neutral surface charge of the unmodified nanocomplex prevented non-specific internalization, highlighting the essential role of the MUC1 aptamer for targeting.

Limitations: Not explicitly stated by the authors. Obvious limitations include: - Only one in vivo tumor model (C26 in BALB/c mice) was tested; no human xenograft or orthotopic model validation. - Short follow-up period (20 days) with no survival analysis or long-term toxicity assessment. - Relatively low Epi loading efficiency (~1%), which may limit translational scalability. - No detailed biodistribution or pharmacokinetic data reported. - No comparison with standard-of-care combination therapies or clinically approved nanoformulations (e.g., Doxil, Abraxane). - The synergetic mechanism of Epi + antimir-21 (e.g., specific downstream target validation such as PTEN or PDCD4) was not investigated at the molecular level. - Only a single dose of treatment was administered in vivo; no dose-response or multi-dose regimen evaluation.

Biochemical Pharmacology*, Volume 173, 2020, Article 113648 DOI/

7-Ketocholesterol and 7β-Hydroxycholesterol — In Vitro and Animal Models, Biological Activities, and Toxicity-Preventing Molecules

Nature Reviews Drug Discovery

Engineering Precision Nanoparticles for Drug Delivery

Materials* (MDPI; inferred from format)

Generation of Well-Defined Micro/Nanoparticles via Advanced Manufacturing Techniques for Therapeutic Delivery

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