Evaluation of in vitro efficacy of docetaxel-loaded calcium carbonate aragonite nanoparticles (DTX-CaCO3NP) on 4T1 mouse breast cancer cell line

Cockle shell-derived calcium carbonate nanoparticles have shown promising potentials as slow drug-releasing compounds in cancer chemotherapy. In this study, we evaluated the in vitro efficacy of docetaxel (DTX)-loaded CaCO3NP on 4T1 cell line. This was achieved by evaluating the following: cytotoxicity using MTT assay, fluorescence imaging, apoptosis with Annexin V assay, cell cycle analysis, scanning (SEM) and transmission electron microscopy (TEM), and scratch assay. Based on the results, DTX-CaCO3NP with a DTX concentration of 0.5 μg/mL and above had comparable cytotoxic effects with free DTX at 24 h, while all concentrations had similar cytotoxic effect on 4T1 cells at 48 and 72 h. Fluorescence and apoptosis assay showed a higher (p < 0.05) number of apoptotic cells in both free DTX and DTX-CaCO3NP groups. Cell cycle analysis showed cycle arrest at subG0 and G2/M phases in both treatment groups. SEM showed presence of cellular blebbing, while TEM showed nuclear fragmentation, apoptosis, and vacuolation in the treatment groups. Scratch assay showed lower (p < 0.05) closure in both free DTX and DTX-CaCO3NP groups. The results from this study showed that DTX-CaCO3NP has similar anticancer effects on 4T1 cells as free DTX, and since it has a slow release rate, it is a more preferred substitute for free DTX

Preparation and characterization of cockle shell aragonite nanocomposite porous 3D scaffolds for bone repair

The demands for applicable tissue-engineered scaffolds that can be used to repair load-bearing segmental bone defects (SBDs) is vital and in increasing demand. In this study, seven different combinations of 3 dimensional (3D) novel nanocomposite porous structured scaffolds were fabricated to rebuild SBDs using an extraordinary blend of cockle shells (CaCo3) nanoparticles (CCN), gelatin, dextran and dextrin to structure an ideal bone scaffold with adequate degradation rate using the Freeze Drying Method (FDM) and labeled as 5211, 5400, 6211, 6300, 7101, 7200 and 8100. The micron sized cockle shells powder obtained (75 µm) was made into nanoparticles using mechano-chemical, top-down method of nanoparticles synthesis with the presence of the surfactant BS-12 (dodecyl dimethyl bataine). The phase purity and crystallographic structures, the chemical functionality and the thermal characterization of the scaffolds’ powder were recognized using X-Ray Diffractometer (XRD), Fourier transform infrared (FTIR) spectrophotometer and Differential Scanning Calorimetry (DSC) respectively. Characterizations of the scaffolds were assessed by Scanning Electron Microscopy (SEM), Degradation Manner, Water Absorption Test, Swelling Test, Mechanical Test and Porosity Test. Top-down method produced cockle shell nanoparticles having averagely range 37.8±3–55.2±9 nm in size, which were determined using Transmission Electron Microscope (TEM). A mainly aragonite form of calcium carbonate was identified in both XRD and FTIR for all scaffolds, while the melting (Tm) and transition (Tg) temperatures were identified using DSC with the range of Tm 62.4–75.5 °C and of Tg 230.6–232.5 °C. The newly prepared scaffolds were with the following characteristics: (i) good biocompatibility and biodegradability, (ii) appropriate surface chemistry and (iii) highly porous, with interconnected pore network. Engineering analyses showed that scaffold 5211 possessed 3D interconnected homogenous porous structure with a porosity of about 49%, pore sizes ranging from 8.97 to 337 µm, mechanical strength 20.3 MPa, Young's Modulus 271±63 MPa and enzymatic degradation rate 22.7 within 14 days

Synthesis, characterization, and cytocompatibility of potential cockle shell aragonite nanocrystals for osteoporosis therapy and hormonal delivery

Calcium carbonate is a porous inorganic nanomaterial with huge potential in biomedical applications and controlled drug delivery. This study aimed at evaluating the physicochemical properties and in vitro efficacy and safety of cockle shell aragonite calcium carbonate nanocrystals (ANC) as a potential therapeutic and hormonal delivery vehicle for osteoporosis management. Free and human recombinant parathyroid hormone 1-34 (PTH 1-34)-loaded cockle shell aragonite calcium carbonate nanocrystals (PTH-ANC) were synthesized and evaluated using standard procedures. Transmission electron microscopy and field emission scanning electron microscopy results demonstrated highly homogenized spherical-shaped aragonite nanocrystals of 30±5 nm diameter. PTH-ANC had a zeta potential of −27.6 ± 8.9 mV. The encapsulation efficiency of the formulation was found to be directly proportional to the concentrations of the drug fed. The X-ray diffraction patterns revealed strong crystallizations with no positional change of peaks before and after PTH-ANC synthesis. Fourier transform infrared spectroscopy demonstrated no detectable interactions between micron-sized aragonite and surfactant at molecular level. PTH-ANC formulation was stabilized at pH 7.5, enabling sustained slow release of PTH 1-34 for 168 h (1 week). A 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide cytocompatibility assay in Human Foetal Osteoblast Cell Line hFOB 1.19 showed that ANC can safely support osteoblast proliferation up to 48 h whereas PTH-ANC can safely support the proliferation at 72 h and beyond due to the sustained slow release of PTH 1-34. It was concluded that due to its biogenic nature, ANC is a cytocompatible antiosteoporotic agent. It doubles as a nanocarrier for the enhancement of efficacy and safety of the bone anabolic PTH 1-34. ANC is expected to reduce the cost, dosage, and dose frequency associated with the use of PTH 1-34 management of primary and secondary forms of osteoporosis

Formulation of a sustained release docetaxel loaded cockle shell-derived calcium carbonate nanoparticles against breast cancer

Purpose: Here, we explored the formulation of a calcium carbonate nanoparticle delivery system aimed at enhancing docetaxel (DTX) release in breast cancer. Methods: The designed nano- anticancer formulation was characterized thorough X-ray diffraction (XRD), Fourier transformed infrared (FTIR), transmission electron microscopy (TEM) and field emission scanning electron microscopy (FESEM) and Brunauer-Emmett-Teller (BET) methods. The nano- anticancer formulation (DTX- CaCO3NP) was evaluated for drug delivery properties thorough in vitro release study in human body simulated solution at pH 7.4 and intracellular lysosomal pH 4.8. Results: Characterization revealed the successful synthesis of DTX- CaCO3NP, which had a sustained release at pH 7.4. TEM showed uniformly distributed pleomorphic shaped pure aragonite particles. The highest entrapment efficiency (96%) and loading content (11.5%) were obtained at docetaxel to nanoparticles ratio of 1:4. The XRD patterns revealed strong crystallizations in all the nanoparticles formulation, while FTIR showed chemical interactions between the drug and nanoparticles with negligible positional shift in the peaks before and after DTX loading. BET analysis showed similar isotherms before and after DTX loading. The designed DTX- CaCO3NP had lower (p 0.05) effects at 48 h and 72 h. However, the DTX- CaCO3NP released less than 80% of bond DTX at 48 and 72 h but showed comparable effects with free DTX. Conclusions: The results showed that the developed DTX- CaCO3NP released DTX slower at pH 7.4 and had comparable cytotoxicity with free DTX at 48 and 72 h in MCF-7 cells

Characterization, histological evaluation and efficacy of docetaxel-loaded cockle shellderived CaCO3 nanoparticles in vitro and in vivo

Breast cancer is one of the most widely researched cancers worldwide. Currently, therapeutic options aimed at reducing its metastasis have a limitation of toxicity to the patient. Cockle shell-derived calcium carbonate aragonite nanoparticles (CSCaCO3NP) have shown promising potentials as a slow drug releasing compound in cancer chemotherapy. This study explores CSCaCO3NP as a delivery system aimed at enhancing docetaxel (DTX) release in breast cancer. Furthermore, this study also evaluated the in vitro cytotoxicity of DTX-loaded CSCaCO3NP against 4T1 cell line, and in vivo toxicity evaluation of CSCaCO3NP, and the effect of DTX-loaded CSCaCO3NP in 4T1 cancer-bearing Balb/C mice. The nano-anticancer formulation (DTX-CSCaCO3NP) was characterized by various physico-chemical characterizations. The in vitro cytotoxicity evaluation was achieved using different bioassay parameters. The pharmacokinetic study was estimated. The in vivo mice acute and sub-chronic toxicity studies were conducted for 14 days. In the therapeutic study, mice were subcutaneously inoculated with 5 x 105 4T1 cells in the right mammary fat pad. Tumor treatment commenced in two stages, the early and late stages. Mice were grouped into 3 treatment groups of DTX (10 mg/kg), DTXCSCaCO3NP (10 mg/kg) and DTX-CSCaCO3NP (5 mg/kg). The positive and negative control groups were also included. Characterization results revealed that DTX-CSCaCO3NP synthesis was excellent, and had a sustained release at pH 7.4. TEM results showed nanoparticles sizes of 42 nm. The XRD patterns revealed strong crystallizations and pure aragonite particles formulation, while FTIR showed entrapment between the drug and nanoparticles. The DTX-CSCaCO3NP had comparable (p>0.05) cytotoxicity effects as DTX against MCF-7 and 4T1 cells. Fluorescence and apoptosis assay showed higher (p<0.05) number of apoptotic cells in both DTX and DTX-CSCaCO3NP groups. SEM showed the presence of cellular blebbing, while TEM showed nuclear fragmentation, apoptosis and vacuolation. In in vivo toxicity analysis, no significant sign was observed and no mortality was recorded in both study periods. Tumor volume, organ weights, tumour inhibition rates and metastatic cells were significantly (p<0.05) lower in the DTX-CSCaCO3NP (10 mg/kg) group. In conclusion, the results showed that the formulated DTX-CSCaCO3NP released DTX slower at pH 7.4. It was also observed that DTX-CSCaCO3NP has similar anticancer effects on MCF-7 and 4T1 cells as DTX and since it has a slow release, CSCaCO3NP is a promising delivery system for DTX in the 4T1 induced breast cancer model. Hence, DTX-CSCaCO3NP (10 mg/kg) showed high efficacy against breast cancer metastasis

Novel drug delivery systems for loading of natural plant extracts and their biomedical applications

Many types of research have distinctly addressed the efficacy of natural plant metabolites used for human consumption both in cell culture and preclinical animal model systems. However, these in vitro and in vivo effects have not been able to be translated for clinical use because of several factors such as inefficient systemic delivery and bioavailability of promising agents that significantly contribute to this disconnection. Over the past decades, extraordinary advances have been made successfully on the development of novel drug delivery systems for encapsulation of plant active metabolites including organic, inorganic and hybrid nanoparticles. The advanced formulas are confirmed to have extraordinary benefits over conventional and previously used systems in the manner of solubility, bioavailability, toxicity, pharmacological activity, stability, distribution, sustained delivery, and both physical and chemical degradation. The current review highlights the development of novel nanocarrier for plant active compounds, their method of preparation, type of active ingredients, and their biomedical applications