Evidence for distinct mechanisms of uptake and antitumor activity of secretory phospholipase A2 responsive liposome in prostate cancer

Abstract

Secretory phospholipase A2 (sPLA2) cleave phospholipids at sn-2 ester bonds, releasing lysophospholipids and fatty acids, and are over expressed in several pathologies, including inflammation, arthritis, sepsis and breast and prostate cancers. Herein we evaluated the therapeutic activity of liposomes engineered to be responsive to different sPLA2 isoforms compared to clinically used long-circulating (pegylated) sterically stabilized liposomes (SSL) in vitro and in vivo, and assess differences in role of sPLA2 in the mechanism of uptake and delivery of these nanoparticles. Exposing sPLA2 responsive liposomes (SPRL) to sPLA2 increased the release of intraluminal entrapped contents in a time-dependent manner that was inhibited by the sPLA2 inhibitor LY3117272. Treatment of prostate cancer cells with doxorubicin encapsulated in SSL and SPRL resulted in cytotoxicity in LNCaP, DU-145 and PC-3 cells lines comparable to free drug. Interestingly, cytotoxicity was not altered by sPLA2 inhibition. Tracking of drug and liposome delivery using fluorescence microscopy and flow cytometry, we demonstrated that drug uptake was liposome-dependent, as encapsulation of doxorubicin in SPRL resulted in 1.5 to 2-fold greater intracellular drug levels compared to SSL. Liposome uptake was cell-dependent and did not correlate to doxorubicin uptake; however, doxorubicin uptake was generally greatest in PC-3 cells, followed by DU-145 cells and then LNCaP cells. In almost all cases, uptake of one of our formulations, SPRL-E, was greater than SSL. The therapeutic activity of SPRL in vivo was demonstrated using a mouse xenograft model of human prostate cancer, which showed that doxorubicin entrapped within SPRL decreased tumor growth compared to SSL, suggesting that SPRL are more effective at slowing tumor growth than a SSL formulation similar to the FDA approved DOXIL™. Collectively, these data show the therapeutic activity of SPRL compared to SSL, yield insights into the mechanisms of action of these nanoparticles and suggest that SPRL could be useful for treatment of other pathologies that over express sPLA2.

Secretory phospholipase A2 enzymes as pharmacological targets for treatment of disease

“…Recent studies, including those from our own laboratory, have engineered liposomes to target pathologies that overexpress sPLA 2 and enhance the release of drugs at diseased tissue [87-89]. sPLA 2 -targeted nanoparticles are composed of phospholipids that are preferentially cleaved by sPLA 2 , such as those with negatively charged and altered levels of phospholipids containing other zwitterionic head groups and shorter fatty acyl chains [89-91].…”

“…sPLA 2 -targeted nanoparticles are composed of phospholipids that are preferentially cleaved by sPLA 2 , such as those with negatively charged and altered levels of phospholipids containing other zwitterionic head groups and shorter fatty acyl chains [89-91]. This preference is likely a result of the many positively charge amino acids present in sPLA 2 ’s protein sequence (Figure 2B-D), especially at the ICS where these residues are believed to increase the interaction with the negatively charged phospholipid head group.…”

“…There have been many proposed modifications of the phospholipid component of liposomes to improve drug release at sPLA 2 -targeted sites [87, 89]. One such modification was the incorporation of low (> 10%) amount of zwitterionic phosphatidylethanolamine (PE) into sterically stable liposomes (SSL).…”

“…One such modification was the incorporation of low (> 10%) amount of zwitterionic phosphatidylethanolamine (PE) into sterically stable liposomes (SSL). These modified liposomes, termed sPLA 2 responsive liposomes (SPRL), showed enhanced drug release in the presence of Group IIA and III sPLA 2 , and were more effective than SSL (which are similar to clinically used formulations) in limiting hormone refractory prostate cancer tumor growth in mouse models [89]. One interesting finding with SPRL is that their activity or effectiveness was not altered by LY3711727.…”

Overcoming tumor cell chemoresistance using nanoparticles: lysosomes are beneficial for (stearoyl) gemcitabine-incorporated solid lipid nanoparticles

“…[199][200][201] Liposomes that are responsible for sPLA2 (SPRL) were engineered to facilitate liposomal degradation and drug release in tumor tissues. [202][203][204] In a study comparing the uptakes and cytotoxicities of the SPRL encapsulated with DOX and sterically stabilized liposomes encapsulated with DOX, Moch et al suggested that the efficacy of the sPLA2 liposomes are mediated by cell-dependent mechanisms. 203 Recently, Hofmann et al provided evidence supporting the existence of drug delivery into cells without cellular uptake of the nanoparticles through a new "kiss-andrun" mechanism between (polymeric) nanoparticles and the cell membrane.…”