Hybrid Mesoporous-Microporous Nanocarriers for Overcoming Multidrug Resistance by Sequential Drug Delivery.

We identified drugs or mechanisms targeting ABCB1 (or P-glycoprotein; P-gp)-overexpressing drug-resistant cancer populations, given that these cells play a key role in tumor recurrence. Specifically, we searched for Akt inhibitors that could increase cytotoxicity in P-gp-overexpressing drug-resistant cancer cells. We performed cytotoxicity assays using five cell lines: 1. MCF-7/ADR, 2. KBV20C cancer cells (P-gp overexpression, vincristine [VIC] resistance, and GSK690693-resistance), 3. MCF-7, 4. normal HaCaT cells (non-P-gp-overexpressing, VIC-sensitive, and GSK690693-sensitive), and 5. MDA-MB-231 cancer cells (non-P-gp overexpression, relatively VIC-resistance, and GSK690693-sensitive). Herein, we found that low-dose perifosine markedly and selectively sensitizes both MCF-7/ADR and KBV20C drug-resistant cancer cells exhibiting P-gp overexpression. Compared with other Akt inhibitors (AZD5363, BKM120, and GSK690693), low-dose perifosine specifically sensitized P-gp-overexpressing resistant MCF-7/ADR cancer cells. Conversely, Akt inhibitors (other than perifosine) could enhance sensitization effects in drug-sensitive MCF-7 and HaCaT cells. Considering that perifosine has both an alkyl-phospholipid structure and is an allosteric inhibitor for membrane-localizing Akt-targeting, we examined structurally and functionally similar Akt inhibitors (miltefosine and MK-2206). However, we found that these inhibitors were non-specific, suggesting that the specificity of perifosine in P-gp-overexpressing resistant cancer cells is unrelated to phospholipid localizing membranes or allosteric inhibition. Furthermore, we examined the molecular mechanism of low-dose perifosine in drug-resistant MCF-7/ADR cancer cells. MCF-7/ADR cells exhibited increased apoptosis via G2 arrest and autophagy induction. However, no increase in P-gp-inhibitory activity was observed in drug-resistant MCF-7/ADR cancer cells. Single low-dose perifosine treatment exerted a sensitization effect similar to co-treatment with VIC in P-gp-overexpressing drug-resistant MCF-7/ADR cancer cells, suggesting that single treatment with low-dose perifosine is a more powerful tool against P-gp-overexpressing drug-resistant cancer cells. These findings could contribute to its clinical use as a first-line treatment, explicitly targeting P-gp-overexpressing resistant cancer populations in heterogeneous tumor populations. Therefore, perifosine may be valuable in delaying or reducing cancer recurrence by targeting P-gp-overexpressing drug-resistant cancer cells.

A sequential drug delivery system based on silk fibroin scaffold for effective cartilage repair.

Controlled and sequential drug delivery is a strategy to enhance the therapeutic effectiveness of drugs in a variety of biological processes and disease states. While many different drug delivery systems are developed recently, most cannot generate temporally complex delivery profiles of multiple therapeutics. These temporally complex profiles are critical for applications such as bone regeneration and cancer chemotherapy, where an orchestrated delivery of multiple drugs is required for an optimal outcome. Here, we developed three distinct biomaterial systems that each enable on-demand controlled or sequential drug release. These systems are based on varying external stimulus such as magnetic stimulation, radiofrequency heating, and near infrared (NIR) laser irradiation. The first system is a dual compartment hydrogel composed of an outer gelatin partition and an inner alginate ferrogel. While the outer compartment could be loaded with a recruitment factor to recruit and harbor cells, the inner compartment was capable of retaining and releasing a differentiation factor on-demand. The inner compartment was a biphasic ferrogel and stimulation was conducted using a custom magnetic stimulation set up. It was shown that delayed differentiation factor delivery can enhance mMSCs’ osteo-differentiation outcomes using 2D and 3D cell cultures. The second system is a magnetoliposome (ML) integrated hydrogel system. In this design, different sizes of magnetic iron oxide nanoparticles (IONPs) were used to develop two different MLs: ML-A and ML-B. Cationic and zwitterionic lipids were used to form positively charged liposomes that could electrostatically adsorb the IONPs on their surfaces and form MLs. The ratio of IONP/lipid was optimized to form stable ML-A and ML-B structures. These structures were integrated within 3D alginate hydrogels to enhance stability and provide localized drug delivery. As the different MLs could be stimulated at different frequencies, complex delivery profiles could be generated using these MLs in hydrogels. Controlled and delayed releases of a model drug (FITC-Dextran) from ML-A and ML-B in hydrogels were demonstrated. The third system is a single-walled carbon nanotube (SWCNT) liposome complex (CLC) integrated hydrogel. Here, unique NIR absorbance properties of SWCNTs were used to achieve drug release from liposomal structures. DNA sequences were used to wrap SWCNTs to uniformly disperse them in an aqueous solution and provide negative charge on their surface. These DNA-SWCNTs were then mixed with cationic liposomes to form CLCs. Optimal SWCNT to lipid ratio to form stable CLCs were determined. CLCs were then integrated within hydrogel structures and drug release was controlled using

During the process of malignant tumor treatment, photodynamic therapy (PDT) exerts poor efficacy due to the hypoxic environment of the tumor cells, and long-time chemotherapy reduces the sensitivity of tumor cells to chemotherapy drugs due to the presence of drug-resistant proteins on the cell membranes for drug outward transportation. Therefore, we reported a nano platform based on mesoporous silica coated with polydopamine (MSN@PDA) loading PDT enhancer MnO2, photosensitizer indocyanine green (ICG) and chemotherapeutic drug doxorubicin (DOX) (designated as DMPIM) to achieve a sequential release of different drugs to enhance treatment of malignant tumors. MSN was first synthesized by a template method, then DOX was loaded into the mesoporous channels of MSN, and locked by the PDA coating. Next, ICG was modified by π–π stacking on PDA, and finally, MnO2 layer was accumulated on the surface of DOX@MSN@PDA- ICG@MnO2, achieving orthogonal loading and sequential release of different drugs. DMPIM first generated oxygen (O2) through the reaction between MnO2 and H2O2 after entering tumor cells, alleviating the hypoxic environment of tumors and enhancing the PDT effect of sequentially released ICG. Afterwards, ICG reacted with O2 in tumor tissue to produce reactive oxygen species, promoting lysosomal escape of drugs and inactivation of p-glycoprotein (p-gp) on tumor cell membranes. DOX loaded in the MSN channels exhibited a delay of approximately 8 h after ICG release to exert the enhanced chemotherapy effect. The drug delivery system achieved effective sequential release and multimodal combination therapy, which achieved ideal therapeutic effects on malignant tumors. This work offers a route to a sequential drug release for advancing the treatment of malignant tumors.

The dense extracellular matrix (ECM) of stroma-rich solid tumors acts as a significant barrier to effective chemotherapy by hindering drug penetration. In this study, a supramolecular hydrogel was successfully developed, enabling the codelivery and sequential release of hydrophilic and lipophilic drugs designed to target the ECM. The hydrogel is easy to prepare, has self-healing properties and excellent biocompatibility. Upon administration, the hydrogel first releases pirfenidone to inhibit collagen production, weakening the ECM, followed by the release of paclitaxel, which improves tumor penetration. The effectiveness of this sequential drug delivery system was validated in both oral squamous cell carcinoma and pancreatic cancer models, a classic example of a tumor with abundant ECM. In vitro experiments showed controlled sequential release profiles, whereas in vivo experiments using cell-derived and patient-derived xenograft models revealed that the hydrogel was more effective at tumor suppression compared to traditional methods. Single administration of the hydrogel led to long-term localized drug release, maintaining higher concentrations of chemotherapeutic agents in the tumor tissue and effectively reducing the tumor volume. This study provided a promising strategy to enhance chemotherapy in ECM-dense tumors, offering an efficient and minimally invasive method for localized, sustained-release cancer therapy.

Combination drug therapy is commonly used to treat cancer, diabetes, cardiovascular conditions, and infections. However, these therapies face challenges associated with patient compliance and toxicology. Over the past decades, microdevices have emerged as a promising candidate for oral delivery allowing for targeted drug delivery with a tunable drug release. In the present work, engineered and monodisperse dual‐compartment microdevices are developed to achieve a physical separation of two drugs followed by a sequential release in the gastrointestinal tract. As proof‐of‐concept, the compartments are sealed with two pH‐sensitive polymers of different thicknesses to control the sequential release of propranolol and furosemide. In vitro release studies and in vivo absorption studies in rats confirm a sequential drug release from the two compartments. Unlike other proposed approaches, it is highly advantageous that the drugs can be loaded directly as powders, and that their release can be tuned via optimized coatings to achieve the desired release and absorption profiles. Conclusively, this study lays a strong foundation for the future use of microdevices to enable co‐delivery of drugs followed by a sequential release in close proximity in the gastrointestinal tract.

Currently, multidrug combinations are often used clinically to improve the efficacy of oncology chemotherapy, but multidrug combinations often lead to multidrug resistance and decreased performance, resulting in more severe side effects than monotherapy. Therefore, sequential drug release strategies in time and space as well as nanocarriers that respond to the tumor microenvironment have been developed. First, the advantage of the sequential release strategy is that it can load multiple drugs simultaneously to meet their spatiotemporal requirements and stability, thus exerting synergistic effects of two or more drugs. Second, in some cases, sequential drug delivery of different molecular targets can improve the sensitivity of cancer cells to drugs, control the metabolism of cancer cells, and remodel tumor vasculature. Finally, some drug combinations with built-in release control are used for sequential administration. This paper focuses on the use of nanotechnology and built-in control device to construct drug delivery carriers with different stimulation responses, thus achieving the sequential release of drugs. Therefore, the nano-sequential delivery carrier provides a new idea and platform for the therapeutic effect of various drugs and the synergistic effect among the drugs.

Currently, multidrug combinations are often used clinically to improve the efficacy of oncology chemotherapy, but multidrug combinations often lead to multidrug resistance and decreased performance, resulting in more severe side effects than monotherapy. Therefore, sequential drug release strategies in time and space as well as nano-carriers that respond to the tumor microenvironment have been developed. First, the advantage of the sequential release strategy is that they can load multiple drugs simultaneously to meet their spatiotemporal requirements and stability, thus exerting synergistic effects of two or more drugs. Second, in some cases, sequential drug delivery of different molecular targets can improve the sensitivity of cancer cells to drugs. Control the metabolism of cancer cells, and remodel tumor vasculature. Finally, some drug combinations with built-in release control are used for sequential administration. This paper focuses on the use of nanotechnology and built-in control device to construct drug delivery carriers with different stimulation responses, thus achieving the sequential release of drugs. Therefore, the nano-sequential delivery carrier provides a new idea and platform for the therapeutic effect of various drugs and the synergistic effect among drugs.

Time-sequenced drug release, or sequential drug release, represents a pivotal strategy in the synergistic treatment of diseases using nanocomposites. Achieving this requires the rational integration of multiple therapeutic agents within a single nanocomposite, coupled with precise time-controlled release mechanisms. These nanocomposites offer many advantages, including enhanced therapeutic synergy, reduced side effects, attenuated adverse interactions, improved stability and optimized drug utilization. Consequently, research in the field of drug delivery and synergistic therapy has become increasingly important. Currently, sequential drug release research is still in the data collection and basic research stages, and its potential has not yet been fully explored. Although prior studies have explored the sequential drug release strategy in various contexts, a comprehensive review of the underlying mechanisms and their applications in nanocomposites remains scarce. This review categorizes different types of sequential drug release strategies and summarizes diverse nanocomposites, focusing on both physical approaches driven by structural variations and chemical methods based on stimulus-responsive mechanisms. Furthermore, we highlight the major applications of sequential drug release strategies in the treatment of various diseases and detail their therapeutic efficacy. Finally, emerging trends and challenges in advancing sequential drug release strategies based on nanocomposites for disease treatment are also discussed.

Chemically and physically stable multidrug-loaded layer-by-layer (LbL) films are promising candidates for sequential and on-demand drug release at concentrations suitable for various applications. The synergistic effect of the sequential release of drugs may enhance their therapeutic efficacy in treating skin cancer and other complex medical conditions. In this study, we prepared LbL films by alternating the deposition of cationic linear polyethylenimine, camptothecin (CPT)-loaded gold nanorods (GNRs), anionic poly(styrenesulfonate), and doxorubicin (DOX) based on electrostatic interactions. The film exhibited loading of CPT and DOX, which could be tuned according to the requirements of the application by changing the parameters of the LbL process. Herein, CPT was encapsulated in GNRs and showed good stability and absorption in the near-infrared (NIR) range (650-900 nm). The prepared LbL film showed a pH-dependent DOX release. Subsequently, the functionalized GNRs showed excellent photothermal properties, which assisted the on-demand release of CPT upon NIR irradiation with further release of DOX. Our results suggest that the LbL approach for sequential drug release can be an effective drug delivery platform owing to its cytocompatibility, anticancer effects, and stimuli-responsive properties.

A pH-responsive polycarbonate nanoplatform enables sequential drug release for enhanced apoptotic cascade synergy in non-small cell lung cancer therapy.

Janus acoustically responsive scaffolds for sequential drug release with phase-programmed steady and pulsatile kinetics.

Multidrug resistance (MDR) facilitates tumor recurrence and metastasis, which has become a main cause of chemotherapy failure in clinical. However, the current therapeutic effects against MDR remain unsatisfactory, mainly hampered by the rigid structure of drug-resistant cell membranes and the uncontrolled drug release. In this study, based on a sequential drug release strategy, we engineered a core-shell nanoparticle (DOX-M@CaP@ATV@HA) depleting cholesterol for reverse tumor MDR. DOX-M@CaP@ATV@HA could accurately target tumor cells due to the active targetability of hyaluronic acid (HA) toward CD44 receptors. The calcium phosphate (CaP) shell was cleaved in the lysosomal acidic environment so that the cholesterol-lowering drug atorvastatin (ATV) was rapidly released to diminish cholesterol and P-glycoprotein (P-gp) level on the membrane, thereby boosting tumor cell drug uptake. Next, doxorubicin (DOX) was gradually released from the hydrophobic core of the mPEG-DSPE micelle, inflicting irreversible DNA damage and triggering apoptosis. The nanosystem was proven both in vitro and in vivo to reverse MDR effectively and exhibited a remarkable therapeutic efficacy on drug-resistant tumors with high biosafety. In conclusion, DOX-M@CaP@ATV@HA effectively reverses MDR via cholesterol depletion, which provides an innovative strategy for tumor MDR treatment.

Multifunctional all-in-one drug delivery systems for tumor targeting and sequential release of three different anti-tumor drugs

The maintenance of robust ratiometric loading of dual therapeutic agents and fine-tuning release kinetics for consistent in vitro and in vivo optimization of combination effects is vital for discovering new anticancer drug combinations and remains challenging. Smart nanomedicine strategies have been investigated for this purpose, but most of the reported strategies focus either on ratiometric delivery or on unimodal sequential release of the two different agents, which hampers effective optimization of combination effects. Herein we report a sequential drug release system based on nanoformulated mutual prodrugs constructed by the formation of ketal linkages with different acid sensitivities, thus enabling the acid-triggered release of two anticancer drugs, paclitaxel and gemcitabine, in various sequences. We found that in several cell lines, the sequence of drug release substantially affected the combination effects; specifically, in A549 cells, time-staggered release profiles showed enhanced synergistic effects relative to those of a simultaneous release profile. Moreover, in vivo assessment of the antitumor efficacy of the nanoformulations in A549 xenograft models indicated that the best therapeutic effects were obtained with time-staggered release profiles, which was consistent with the in vitro results. Our strategy for precisely controlled sequential drug release can be expected to facilitate the screening of optimal drug combinations and maximize combination effects both in vitro and in vivo.