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Poly(L-histidine)-tagged 5-aminolevulinic acid prodrugs: new photosensitizing precursors of protoporphyrin IX for photodynamic colon cancer therapy.

Johnson RP, Chung CW, Jeong YI, Kang DH, Suh H, Kim I - Int J Nanomedicine (2012)

Bottom Line: ALA-p(L-His)(n) showed high phototoxicity and selectivity in different pH conditions compared with ALA alone.Because the length of the histidine chain increases in the ALA-p(L-His)(n) prodrugs, the PDT effect was found to be more powerful.In particular, high phototoxicity was observed when the cells were treated with ALA-p(L-His)(15), compared with treatment using ALA alone.

View Article: PubMed Central - PubMed

Affiliation: WCU Centre for Synthetic Polymer Bioconjugate Hybrid Materials, Department of Polymer Science and Engineering, Pusan National University, Pusan, Korea.

ABSTRACT

Background: 5-Aminolevulinic acid (ALA) and its derivatives have been widely used in photodynamic therapy. The main drawback associated with ALA-based photodynamic therapy (ALA-PDT) and ALA fluorescence diagnosis results from the hydrophilic nature of ALA and lack of selectivity for tumor versus nontumor cells. The application of certain triggers, such as pH, into conventional sensitizers for controllable (1)O(2) release is a promising strategy for tumor-targeted treatment.

Methods: A series of pH-sensitive ALA-poly(L-histidine) [p(L-His)(n)] prodrugs were synthesized via ring opening polymerization of 1-benzyl-N-carboxy-L-histidine anhydride initiated by the amine hydrochloride group of ALA itself. As an alternative to ALA for PDT, the synthesized prodrugs were used to treat a cultured human colon cancer HCT116 cell line under different pH conditions. The effect of ALA-p(L-His)(n) derivatives was evaluated by monitoring the fluorescence intensity of protoporphyrin IX, and measuring the cell survival rate after suitable light irradiation.

Results: The cytotoxicity and dark toxicity of ALA and synthesized ALA-p(L-His) derivatives in HEK293T and HCT116 cells in the absence of light at pH 7.4 and 6.8 shows that the cell viability was relatively higher than 100%. ALA-p(L-His)(n) showed high phototoxicity and selectivity in different pH conditions compared with ALA alone. Because the length of the histidine chain increases in the ALA-p(L-His)(n) prodrugs, the PDT effect was found to be more powerful. In particular, high phototoxicity was observed when the cells were treated with ALA-p(L-His)(15), compared with treatment using ALA alone.

Conclusion: The newly synthesized ALA-p(L-His)(n) derivatives are an effective alternative to ALA for enhancing protoporphyrin IX production and the selectivity of the phototoxic effect in tumor cells.

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The photodynamic effect of ALA and ALA-p(L-His) prodrugs (1 mM concentration) on HCT116 cells at pH 7.4 and 6.8. 100 μL of serum-free medium containing 1.0 mM prodrugs was added to HCT116 cells following incubation for 4 hours.Notes: The control was treated with serum-free medium in the absence of both ALA and p(L-His)-ALA. After irradiation at a dose of 1.0 J/cm2, the medium was removed and washed with phosphate-buffered saline. Next, a fresh 100 μL of RPMI with 10% phosphate-buffered saline was added and the cells were incubated for a further 24 hours. Cell phototoxicity was determined by MTT assay.Abbreviations: ALA, 5-aminolevulinic acid; p(L-His), poly(L-histidine).
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f5-ijn-7-2497: The photodynamic effect of ALA and ALA-p(L-His) prodrugs (1 mM concentration) on HCT116 cells at pH 7.4 and 6.8. 100 μL of serum-free medium containing 1.0 mM prodrugs was added to HCT116 cells following incubation for 4 hours.Notes: The control was treated with serum-free medium in the absence of both ALA and p(L-His)-ALA. After irradiation at a dose of 1.0 J/cm2, the medium was removed and washed with phosphate-buffered saline. Next, a fresh 100 μL of RPMI with 10% phosphate-buffered saline was added and the cells were incubated for a further 24 hours. Cell phototoxicity was determined by MTT assay.Abbreviations: ALA, 5-aminolevulinic acid; p(L-His), poly(L-histidine).

Mentions: The effect of ALA-p(L-His)-based PDT was evaluated by treating HCT116 cells with a concentration of 1 mM of the prodrug for 4 hours in the dark. After irradiation, the cell survival percentage, compared with control cells, was estimated using the MTT assay. From Figure 5, it is clear that ALA-p(L-His)n has high phototoxicity compared with ALA. Because the length of the histidine chain increases in the ALA-p(L-His)n prodrugs, the PDT effect was found to be more powerful. In particular, high phototoxicity was observed when the cells were treated with ALA-p(L-His)15 compared with treatment using ALA alone. In addition, phototoxicity was higher when the cells were incubated with ALA-p(L-His)10 and ALA-p(L-His)15 at lower pH than at the physiological pH of 7.4. The imidazole ring (pKb about 6.5) of p(L-His) has lone pairs of electrons that endow it with pH-dependent amphoteric properties. Protonation was seen to occur just below physiological pH, indicating that the histidine unit attached to the ALA-p(L-His) n prodrugs can act as a proton receptor and consequently effect internalization into the tumor cells. Both charged and uncharged forms of histidine are present in p(His) at neutral pH, and the concentration ratio of the charged form to the uncharged form is very sensitive to small changes in pH under physiological conditions.40 Moreover, the ionization of p(L-His) switches the nature of the material to becoming more lipophilic, and p(L-His) shows strong endosomolytic properties by its proton sponge effect and/or its interaction with anionic phospholipids comprising the endosomal compartments.22


Poly(L-histidine)-tagged 5-aminolevulinic acid prodrugs: new photosensitizing precursors of protoporphyrin IX for photodynamic colon cancer therapy.

Johnson RP, Chung CW, Jeong YI, Kang DH, Suh H, Kim I - Int J Nanomedicine (2012)

The photodynamic effect of ALA and ALA-p(L-His) prodrugs (1 mM concentration) on HCT116 cells at pH 7.4 and 6.8. 100 μL of serum-free medium containing 1.0 mM prodrugs was added to HCT116 cells following incubation for 4 hours.Notes: The control was treated with serum-free medium in the absence of both ALA and p(L-His)-ALA. After irradiation at a dose of 1.0 J/cm2, the medium was removed and washed with phosphate-buffered saline. Next, a fresh 100 μL of RPMI with 10% phosphate-buffered saline was added and the cells were incubated for a further 24 hours. Cell phototoxicity was determined by MTT assay.Abbreviations: ALA, 5-aminolevulinic acid; p(L-His), poly(L-histidine).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3367496&req=5

f5-ijn-7-2497: The photodynamic effect of ALA and ALA-p(L-His) prodrugs (1 mM concentration) on HCT116 cells at pH 7.4 and 6.8. 100 μL of serum-free medium containing 1.0 mM prodrugs was added to HCT116 cells following incubation for 4 hours.Notes: The control was treated with serum-free medium in the absence of both ALA and p(L-His)-ALA. After irradiation at a dose of 1.0 J/cm2, the medium was removed and washed with phosphate-buffered saline. Next, a fresh 100 μL of RPMI with 10% phosphate-buffered saline was added and the cells were incubated for a further 24 hours. Cell phototoxicity was determined by MTT assay.Abbreviations: ALA, 5-aminolevulinic acid; p(L-His), poly(L-histidine).
Mentions: The effect of ALA-p(L-His)-based PDT was evaluated by treating HCT116 cells with a concentration of 1 mM of the prodrug for 4 hours in the dark. After irradiation, the cell survival percentage, compared with control cells, was estimated using the MTT assay. From Figure 5, it is clear that ALA-p(L-His)n has high phototoxicity compared with ALA. Because the length of the histidine chain increases in the ALA-p(L-His)n prodrugs, the PDT effect was found to be more powerful. In particular, high phototoxicity was observed when the cells were treated with ALA-p(L-His)15 compared with treatment using ALA alone. In addition, phototoxicity was higher when the cells were incubated with ALA-p(L-His)10 and ALA-p(L-His)15 at lower pH than at the physiological pH of 7.4. The imidazole ring (pKb about 6.5) of p(L-His) has lone pairs of electrons that endow it with pH-dependent amphoteric properties. Protonation was seen to occur just below physiological pH, indicating that the histidine unit attached to the ALA-p(L-His) n prodrugs can act as a proton receptor and consequently effect internalization into the tumor cells. Both charged and uncharged forms of histidine are present in p(His) at neutral pH, and the concentration ratio of the charged form to the uncharged form is very sensitive to small changes in pH under physiological conditions.40 Moreover, the ionization of p(L-His) switches the nature of the material to becoming more lipophilic, and p(L-His) shows strong endosomolytic properties by its proton sponge effect and/or its interaction with anionic phospholipids comprising the endosomal compartments.22

Bottom Line: ALA-p(L-His)(n) showed high phototoxicity and selectivity in different pH conditions compared with ALA alone.Because the length of the histidine chain increases in the ALA-p(L-His)(n) prodrugs, the PDT effect was found to be more powerful.In particular, high phototoxicity was observed when the cells were treated with ALA-p(L-His)(15), compared with treatment using ALA alone.

View Article: PubMed Central - PubMed

Affiliation: WCU Centre for Synthetic Polymer Bioconjugate Hybrid Materials, Department of Polymer Science and Engineering, Pusan National University, Pusan, Korea.

ABSTRACT

Background: 5-Aminolevulinic acid (ALA) and its derivatives have been widely used in photodynamic therapy. The main drawback associated with ALA-based photodynamic therapy (ALA-PDT) and ALA fluorescence diagnosis results from the hydrophilic nature of ALA and lack of selectivity for tumor versus nontumor cells. The application of certain triggers, such as pH, into conventional sensitizers for controllable (1)O(2) release is a promising strategy for tumor-targeted treatment.

Methods: A series of pH-sensitive ALA-poly(L-histidine) [p(L-His)(n)] prodrugs were synthesized via ring opening polymerization of 1-benzyl-N-carboxy-L-histidine anhydride initiated by the amine hydrochloride group of ALA itself. As an alternative to ALA for PDT, the synthesized prodrugs were used to treat a cultured human colon cancer HCT116 cell line under different pH conditions. The effect of ALA-p(L-His)(n) derivatives was evaluated by monitoring the fluorescence intensity of protoporphyrin IX, and measuring the cell survival rate after suitable light irradiation.

Results: The cytotoxicity and dark toxicity of ALA and synthesized ALA-p(L-His) derivatives in HEK293T and HCT116 cells in the absence of light at pH 7.4 and 6.8 shows that the cell viability was relatively higher than 100%. ALA-p(L-His)(n) showed high phototoxicity and selectivity in different pH conditions compared with ALA alone. Because the length of the histidine chain increases in the ALA-p(L-His)(n) prodrugs, the PDT effect was found to be more powerful. In particular, high phototoxicity was observed when the cells were treated with ALA-p(L-His)(15), compared with treatment using ALA alone.

Conclusion: The newly synthesized ALA-p(L-His)(n) derivatives are an effective alternative to ALA for enhancing protoporphyrin IX production and the selectivity of the phototoxic effect in tumor cells.

Show MeSH
Related in: MedlinePlus