Combination therapy and noninvasive imaging with a dual therapeutic vector expressing MDR1 short hairpin RNA and a sodium iodide symporter.
We investigated the feasibility of using combination gene therapy and noninvasive nuclear imaging after expression of the human sodium iodide symporter (hNIS) and inhibition of the multidrug resistance (MDR1) gene in colon cancer cells.


METHODS
HCT-15 cells were stably transfected with a dual expression vector, in which the hNIS gene, driven by a constitutive cytomegalovirus promoter, has been coupled to an MDR1 short hairpin RNA (shRNA) cassette. Cell lines stably expressing the hNIS gene and MDR1 shRNA (designated MN-61 and MN-62) were produced, and the expression of the NIS gene and MDR1 shRNA was examined by Western blotting, reverse transcription-polymerase chain reaction, and immunostaining. The functional activities of MDR1 shRNA were determined by paclitaxel uptake and sensitivity to doxorubicin. Functional NIS expression was confirmed by the uptake and efflux of (125)I and the cytotoxicity of (131)I. The effect of the combination of (131)I and doxorubicin was determined by an in vitro clonogenic assay. In vivo NIS expression was examined by small-animal PET with (124)I.


RESULTS
The shMDR-NIS-expressing cells showed a significant decrease in the expression of MDR1 messenger RNA and its translated product, P-glycoprotein. The inhibition of P-glycoprotein expression by shRNA enhanced the intracellular accumulation of paclitaxel, the cellular retention of which is mediated by P-glycoprotein, thereby increasing sensitivity to the anticancer drug. The shMDR-NIS-expressing cells showed a significant increase of (125)I uptake, which was completely inhibited by KClO(4). Although the iodide efflux rate was rapid, the cell survival rate was markedly reduced by (131)I treatment. Interestingly, the combination of doxorubicin and a radioiodide ((131)I) displayed synergistic cytotoxicity that correlated with MDR1 inhibition and NIS expression in shMDR-NIS-expressing cells. Furthermore, in mice with shMDR-NIS-expressing tumor xenografts, small-animal PET with (124)I clearly visualized shMDR1-NIS-expressing tumors.


CONCLUSIONS
We developed a dual expression vector with the NIS gene and MDR1 shRNA. This study represents a promising first step in investigations of the potential use of a combination of the NIS gene and MDR1 shRNA as a new therapeutic strategy allowing RNA interference-based gene therapy, NIS-based radioiodine therapy, and in vivo monitoring based on NIS imaging.
Park
Seung-Yoon
SY
Department of Biochemistry, School of Medicine, Dongguk University, Kyungju, Republic of Korea.
Kwak
Wonjung
W
Thapa
Narendra
N
Tapha
Narendra
N
Jung
Mi-Yeon
MY
Nam
Ju-Ock
JO
So
In-Seop
IS
Kim
So-Youn
SY
Yoo
Jeongsoo
J
Lee
Jaetae
J
Kim
In-San
IS
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics*; D000818:Animals; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000235:genetics / Q000628:therapy*; D003131:Combined Modality Therapy; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics*; D051379:Mice; D012313:RNA / Q000235:genetics* / Q000627:therapeutic use*; D027981:Symporters / Q000235:genetics*; D014162:Transfection / Q000379:methods; D016896:Treatment Outcome; D000099209:Sodium-Iodide Symporters
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D012313:RNA; D027981:Symporters; D000099209:Sodium-Iodide Symporters