Unlocking the Potential of Camel Milk-Derived Exosomes as Novel Delivery Systems: Enhanced Bioavailability of ARV-825 PROTAC for Cancer Therapy.

This study investigates the use of camel milk-derived exosomes (CMEs) as carriers for ARV-825, an anticancer agent targeting bromodomain-containing protein 4 (BRD4), in oral chemotherapy. CMEs were isolated and characterized, and ARV-825-loaded CME formulations were prepared and evaluated through various in vitro and in vivo tests. The ARV-825-CME formulation exhibited an entrapment efficiency of 42.75 ± 5.05%, a particle size of 136.8 ± 1.94 nm, and a zeta potential of -32.75 ± 0.70 mV, ensuring stability and sustained drug release. In vitro studies showed a 5.4-fold enhancement in drug release kinetics compared to the free ARV-825 solution. Permeability studies indicated a 3.2-fold increase in apparent permeability, suggesting improved cellular uptake. Cytotoxicity assays demonstrated potent anticancer activity, with IC50 values decreasing by 1.5 to 2-fold in cancer cell lines SF8628 DIPG and H1975R (resistant to Osimertinib). In vivo pharmacokinetic studies in Sprague-Dawley rats revealed superior systemic absorption and bioavailability of ARV-825 from CMEs, with a 2.55-fold increase in plasma concentration and a 5.56-fold increase in AUC. Distribution studies confirmed absorption through the ileum. This research highlights the potential of CMEs as a promising delivery platform for ARV-825, enhancing its therapeutic efficacy and offering a novel approach to cancer treatment.

Cannabidiol and Beta-Caryophyllene Combination Attenuates Diabetic Neuropathy by Inhibiting NLRP3 Inflammasome/NFκB through the AMPK/sirT3/Nrf2 Axis.

BACKGROUND: In this study, we investigated in detail the role of cannabidiol (CBD), beta-caryophyllene (BC), or their combinations in diabetic peripheral neuropathy (DN). The key factors that contribute to DN include mitochondrial dysfunction, inflammation, and oxidative stress. METHODS: Briefly, streptozotocin (STZ) (55 mg/kg) was injected intraperitoneally to induce DN in Sprague-Dawley rats, and we performed procedures involving Randall Sellito calipers, a Von Frey aesthesiometer, a hot plate, and cold plate methods to determine mechanical and thermal hyperalgesia in vivo. The blood flow to the nerves was assessed using a laser Doppler device. Schwann cells were exposed to high glucose (HG) at a dose of 30 mM to induce hyperglycemia and DCFDA, and JC1 and Mitosox staining were performed to determine mitochondrial membrane potential, reactive oxygen species, and mitochondrial superoxides in vitro. The rats were administered BC (30 mg/kg), CBD (15 mg/kg), or combination via i.p. injections, while Schwann cells were treated with 3.65 µM CBD, 75 µM BC, or combination to assess their role in DN amelioration. RESULTS: Our results revealed that exposure to BC and CBD diminished HG-induced hyperglycemia in Schwann cells, in part by reducing mitochondrial membrane potential, reactive oxygen species, and mitochondrial superoxides. Furthermore, the BC and CBD combination treatment in vivo could prevent the deterioration of the mitochondrial quality control system by promoting autophagy and mitochondrial biogenesis while improving blood flow. CBD and BC treatments also reduced pain hypersensitivity to hyperalgesia and allodynia, with increased antioxidant and anti-inflammatory action in diabetic rats. These in vivo effects were attributed to significant upregulation of AMPK, sirT3, Nrf2, PINK1, PARKIN, LC3B, Beclin1, and TFAM functions, while downregulation of NLRP3 inflammasome, NFκB, COX2, and p62 activity was noted using Western blotting. CONCLUSIONS: the present study demonstrated that STZ and HG-induced oxidative and nitrosative stress play a crucial role in the pathogenesis of diabetic neuropathy. We find, for the first time, that a CBD and BC combination ameliorates DN by modulating the mitochondrial quality control system.

Enhanced oral bioavailability and in vitro evaluation of cannabidiol camel milk-derived exosome formulation in resistant MDA-MB-231 and MDA-MB-468 breast cancer cells.

The potential of camel milk-derived exosomes (CMDE) to enhance the bioavailability of Cannabidiol (CBD) was investigated. CBD-CMDE formulation was prepared using an established procedure and its particle size was 138.4 ± 4.37 nm, and CBD entrapment efficiency of 56.56 ± 4.26 %. In-vitro release studies showed release of 78.27 ± 5.37 % and 46.42 ± 4.75 % CBD from CMDE and control CBD formulation respectively in pH 6.8 at 24 hr. The apparent permeability (Papp) of CBD-CMDE was found to be enhanced by 3.95-fold with Papp of 22.9*10-6 ± 0.34 cm/sec as compared to control CBD formulation with Papp of 5.8*10-6 ± 0.65 cm/sec in MDCK cells. CBD-CMDE was found to be more potent than CBD in 2D cytotoxicity assay with IC50 values of 3.6 ± 0.54 µM, 3.88 ± 0.54 µM and 7.53 ± 0.59 µM, 7.53 ± 0.59 µM against Doxorubicin (DOX) resistant MDA-MB-231 and Rapamycin (RM) resistant MDA-MB-468 breast cancer cells respectively. Moreover, 3D spheroids assay results demonstrated CBD-CMDE with IC50 values of 14 ± 0.85 µM, 15 ± 0.07 µM as compared to CBD alone with IC50 values of 25 ± 0.93 µM, 34.7 ± 0.08 µM in MDA-MB-231 DOX RT cells and MDA-MB-468 RM RT cells respectively. In-vivo PK studies showed enhanced bioavailability of CBD from CBD-exosomes with AUC(0-24h) of 1350.56 ± 187.50 h.ng/mL as compared to CBD control formulation with AUC(0-24h) of 351.95 ± 39.10 h.ng/mL with a single oral dose of 12 mg/kg. The data indicate that CMDE significantly improved the oral bioavailability of CBD. Overall, CMDE can be used to enhance the oral absorption of poorly bioavailable APIs.

Combined Role of Interleukin-15 Stimulated Natural Killer Cell-Derived Extracellular Vesicles and Carboplatin in Osimertinib-Resistant H1975 Lung Cancer Cells with EGFR Mutations.

In this study, we evaluated IL-15 stimulated natural killer cell-derived EVs (NK-EVs) as therapeutic agents in vitro and in vivo in Osimertinib-resistant lung cancer (H1975R) with EGFR mutations (L858R) in combination with carboplatin (CBP). NK-EVs were isolated by ultracentrifugation and characterized by nanoparticle tracking analysis, and atomic force microscopy imaging revealed vesicles with a spherical form and sizes meeting the criteria of exosomal EVs. Further, Western blot studies demonstrated the presence of regular EV markers along with specific NK markers (perforin and granzyme). EVs were also characterized by proteomic analysis, which demonstrated that EVs had proteins for natural killer cell-mediated cytotoxicity (Granzyme B) and T cell activation (perforin and plastin-2). Gene oncology analysis showed that these differentially expressed proteins are involved in programmed cell death and positive regulation of cell death. Further, isolated NK-EVs were cytotoxic to H1975R cells in vitro in 2D and 3D cell cultures. CBP's IC50 was reduced by approximately in 2D and 3D cell cultures when combined with NK-EVs. The EVs were then combined with CBP and administered by i.p. route to H1975R tumor xenografts, and a significant reduction in tumor volume in vivo was observed. Our findings show for the first time that NK-EVs target the PD-L1/PD-1 immunological checkpoint to induce apoptosis and anti-inflammatory response by downregulation of SOD2, PARP, BCL2, SET, NF-κB, and TGF-ß. The ability to isolate functional NK-EVs on a large scale and use them with platinum-based drugs may lead to new clinical applications. The results of the present study suggest the possibility of the combination of NK-cell-derived EVs and CBP as a viable immunochemotherapeutic strategy for resistant cancers.