Majoros et al. X-3 used a PAMAM dendrimer with a folic acid surface group to aid the delivery of the anticancer drug Taxol. Their work used a G5 PAMAM dendrimer which was neutralised by partial acetylation of the surface amino groups. This provided enhanced solubility when in conjugation with fluorescein isothiocyanate and prevented nonspecific targeting interaction during delivery. The remaining amino groups were functionalised by conjugation with fluorescein isothiocyanate (FITC, an imaging agent), folic acid (to target overexpressed folate receptors on cancer cells) and Taxol (anticancer drug). The conjugate was testing at concentrations of 100, 50 and 25nM for 1 hour with two types of KB tumour cells one with up-regulated folate receptors and one down-regulated folate receptors. At concentrations greater than 100nM cytotoxic effects were seen for both KB cell types. Whereas only the KB cells with up-regulated folate receptors were shown to uptake the trifunctional conjugate by fluorescence microscopy. The subsequent rounding and detachment of the KB cells verified the desired cytotoxic effects of the Taxol. In comparison to the down-regulated cells which showed no uptake of dendrimer conjugate and the cells remained viable and attached to the plate. The same effect was observed for both folate starved cells and those grown on a normal medium, therefore the cytotoxic effect was media independent.
At the highest experimental concentration (100nM), folate down-regulated KB cells showed no uptake of the dendrimer conjugate and the number of dead cells was comparable to the untreated, so no toxic effect of the lone dendrimer were observed. In comparison up-regulated KB cells internalised the dendrimer conjugate and the cytotoxic effect of the Taxol was observed. An XTT proliferation assay at 200nM showed unspecific binding and cytotoxic effects in both cell types, whereas at 50nM only folate up-regulated cells were affected. The conjugate without Taxol was completely nontoxic at 200nM.
This work showed that surface functionalisation of PAMAM dendrimers with folic acid was selective towards folate receptor up-regulated tumour cells at low concentrations of 50nM in vitro and that PAMAM dendrimers are nontoxic at concentrations up to 200nM. However, it is unknown as the whether the same effects can be seen in vivo.