Synergy-delivery Of 17-AAG And SARD279 Using Lipid Nanoparticles To Inhibit Proliferation Of Prostate Cancer Cells

Prostate cancer is one of the most common malignant tumors in the male genitourinary system,which is a serious threat to the life and health of men.Postoperative adjuvant hormone therapy is still a first-line treatment strategy for patients with prostate cancer,in which hormone therapy drugs are mainly divided into androgen deprivation drugs and androgen receptor(AR)antagonists.With the progress of treatment,most prostate cancers will gradually become castration-resistant prostate cancer(CRPC),making androgen deprivation drugs ineffective.AR antagonists can also lead to drug resistance due to long-term use,resulting in treatment failure.Therefore,there is an urgent need to develop new therapeutic strategies for prostate cancer.Heat shock protein(HSPs)plays an important role in the progression of cancer cells,For example,Hsp90 promotes the growth and proliferation of cancer cells by regulating a variety of signal pathways.In recent years,a class of small molecule anticancer drugs by inhibiting the activity of Hsp90 has attracted wide attention.Tanespimycin(17-AAG)is an effective Hsp90 inhibitor,which can effectively bind to Hsp90 and inhibit its chaperone function,thus inhibiting the growth of cancer cells.It has entered clinical research and is a promising anti-tumor drug.However,the molecular chaperone that maintains cancer cell growth is a complex network,and when the function of Hsp90 is suppressed,cancer cells can make up for the damage of Hsp90by expressing other kinds of molecular chaperones.It has been found that 17-AAG can significantly induce the expression of Hsp70 in cancer cells,thus reducing the effect of17-AAG.Protein targeted degradation(PTD)is a rapidly developing technology in recent years.Small molecular drugs developed by PTD have shown outstanding efficacy in tumor therapy.Among them,the hydrophobic tag SARD279 can target AR binding and induce ubiquitination of AR through recruitment of Hsp70,which is degraded through the proteasome pathway,ultimately effectively inhibiting the proliferation of AR resistant antagonist prostate cancer cells.However,the expression level of Hsp70 in cancer cells is low,while SARD279 can play a role only at high concentrations,which seriously limits its use in vivo.Based on the above analysis,the authors believe that the combination of 17-AAG and SARD279 may be an effective strategy for the treatment of prostate cancer:on the one hand,17-AAG induces apoptosis of prostate cancer cells by inhibiting the activity of Hsp90;on the other hand,in order to resist the inhibitory effect of 17-AAG on Hsp90,overexpressed Hsp70 can make SARD279 degrade AR at lower doses,which can effectively solve the problem of drug dose.In theory,this strategy of"drinking poison to quench thirst"can effectively solve the disadvantages of a single use of 17-AAG or SARD279.However,17-AAG and SARD279 are both hydrophobic drugs with relatively large molecular weight,short half-life of circulation in vivo and limited tumor targeting ability.In recent years,liposome-based drug delivery carriers have been widely used in cancer treatment because of their targeting and good biocompatibility,which can improve drug solubility and reduce drug permeability.In summary,by using Michael addition reaction,we combined the hydrophilic head group containing amine with the hydrophobic tail group,and synthesized 18 kinds of bioreducible liposomes by changing the structure of the fat head group and tail group.Heat shock protein inhibitors(17-AAG)and androgen receptor targeted degradants(SARD279)and were introduced into liposomes by hydrophobic self-assembly to construct nano-drug delivery system,inhibiting the proliferation of prostate cancer cells and thus hinder the progression of CRPC.The structure of the target compound was determined by ~1HNMR.The results of dynamic light scattering(DLS)and scanning electron microscope(SEM)showed that the prepared nanoparticles were spherical with a diameter of about 100-200 nm.The drug loading of nanoparticles and the responsive drug release under the condition of glutathione reduction were detected by ultra-micro ultraviolet spectrophotometer.The best carrier for drug delivery was screened by Western blot and the feasibility of the combination of the two drugs was verified.The results showed that the introduction of 17-AAG promoted the overexpression of Hsp70in LNCAP cells and led to the degradation of more AR by SARD279 at low concentration.The ability of nanoparticles to degrade AR protein was detected by immunofluorescence confocal method.The toxicity of nanoparticles in vitro was evaluated by MTT method and the results showed that the toxicity to normal cells was low and that to prostate cancer cells was high.At the same time,the synergistic effect of 17-AAG and SARD279 on prostate cancer cell line LNCAP was evaluated by MTT method.The results showed that 17-AAG and SARD279 had a synergistic inhibitory effect on the growth of LNCAP cells.The combination of 17-AAG and SARD279inhibited cell growth and cell number more effectively than 17-AAG or SARD279alone.The combination of 17-AAG and SARD279 may be a promising strategy to overcome the drug resistance of CRPC in LNCAP.