An “eat me” combinatory nano-formulation for systemic immunotherapy of solid tumors

Abstract

Rational: Tumor immunogenic cell death (ICD), induced by certain chemotherapeutic drugs such as doxorubicin (Dox), is a form of apoptosis potentiating a protective immune response. One of the hallmarks of ICD is the translocation of calreticulin to the cell surface acting as an 'eat me' signal. This manuscript describes the development of a stable nucleic acid-lipid particles (SNALPs) formulation for the simultaneous delivery of ICD inducing drug (Dox) with small interfering RNA (siRNA) knocking down CD47 (siCD47), the dominant 'don't eat me' marker, for synergistic enhancement of ICD. Methods: SNALPs loaded with Dox or siCD47 either mono or combinatory platforms were prepared by ethanol injection method. The proposed systems were characterized for particle size, surface charge, entrapment efficiency and in vitro drug release. The ability of the SNALPs to preserve the siRNA integrity in presence of serum and RNAse were assessed over 48 h. The in vitro cellular uptake and gene silencing of the prepared SNALPs was assessed in CT26 cells. The immunological responses of the SNALPs were defined in vitro in terms of surface calreticulin expression and macrophage-mediated phagocytosis induction. In vivo therapeutic studies were performed in CT26 bearing mice where the therapeutic outcomes were expressed as tumor volume, expression of CD4 and CD8 as well as in vivo silencing. Results: The optimized SNALPs had a particle size 122 ±6 nm and an entrapment efficiency > 65% for both siRNA and Dox with improved serum stability. SNALPs were able to improve siRNA and Dox uptake in CT26 cells with enhanced cytotoxicity. siCD47 SNALPs were able to knockdown CD47 by approximately 70% with no interference from the presence of Dox. The siCD47 and Dox combination SNALPs were able to induce surface calreticulin expression leading to a synergistic effect on macrophage-mediated phagocytosis of treated cells. In a tumor challenge model, 50% of mice receiving siCD47 and Dox containing SNALPs were able to clear the tumor, while the remaining animals showed significantly lower tumor burden as compared to either monotreatment. Conclusion: Therefore, the combination of siCD47 and Dox in a particulate system showed potent anti-tumor activity which merits further investigation in future clinical studies.

Zinc-Loaded Black Phosphorus Multifunctional Nanodelivery System Combined With Photothermal Therapy Have the Potential to Treat Prostate Cancer Patients Infected With COVID-19

“…Although DOX is not the most sensitive chemotherapy agent for prostate cancer, it is the most widely used immunogenic death stimulant ( 17 ). It certainly promotes ICD, a well-defined type of apoptotic process ( 47 ). Released DAMPs can be recognized by pattern recognition receptors (PRRs) and induce antigen-presenting cells(APC) to activate, differentiate, and mature ( 48 ).…”

Role of CD47-SIRPα Checkpoint in Nanomedicine-Based Anti-Cancer Treatment

“…ICD has a number of clearly defined physiological characteristics, including cell surface CRT expression, release of DAMPs such as adenosine triphosphate and heat shock proteins, and release of high mobility group box 1 ( Liu et al, 2016 ). Among these factors, the surface expression of CRT is considered to be the single most important element of ICD ( Abdel-Bar et al, 2021 ). As previously mentioned, the effect of CRT exposure, serving as an “eat me” signal, is considered to be counterbalanced and potentially dampened by CD47 expression ( Chao et al, 2010a ).…”

“…Moreover, the upregulated expression of CD47 on the surface of tumor cells makes it an active targeting site for tumor cells, facilitating nonspecific distributed ICD inducing drugs to target tumor tissues and reduce systemic toxicity ( Tang et al, 2021 ). Therefore, codelivery, which simultaneously removes an inhibitory signal and introduces an activating signal, can produce an enhanced antitumor effect ( Abdel-Bar et al, 2021 ; Lee et al, 2021 ). For example, Abdel-Bar et al reported a stable nucleic acid-lipid particle (SNALP) formulation with the simultaneous delivery of an ICD inducing drug (Dox) with small interfering RNA (siRNA) knocking down CD47 (siCD47) for synergistic enhancement of ICD.…”

“…For example, Abdel-Bar et al reported a stable nucleic acid-lipid particle (SNALP) formulation with the simultaneous delivery of an ICD inducing drug (Dox) with small interfering RNA (siRNA) knocking down CD47 (siCD47) for synergistic enhancement of ICD. In a CT-26-tumor-bearing mouse model, SNALPs synergistically inhibited tumor growth and prolonged the survival ( Abdel-Bar et al, 2021 ).…”

Fibroblast Activation Protein‐α Responsive Peptide Assembling Prodrug Nanoparticles for Remodeling the Immunosuppressive Microenvironment and Boosting Cancer Immunotherapy

“…[18] Tumor cells undergoing ICD trigger to release calreticulin (CRT) and high-mobility group box 1 (HMGB1) to promote DC maturation and activate antigen-specific T cell responses. [19] However, the ICD-induced therapeutic effect is severely abolished by negative immune regulation mechanisms. [20,21] Furthermore, the poor pharmacokinetics of these drugs limit their widespread use in cancer immunotherapy.…”