Alterations in Mitochondrial and Apoptosis‐regulating Gene Expression in Photodynamic Therapy‐resistant Variants of HT29 Colon Carcinoma Cells^¶

Abstract

Photodynamic therapy (PDT) is a novel cancer therapy inducing irreversible photodamage to tumor tissue via photosensitizer‐mediated oxidative cytotoxicity. The cellular and molecular responses associated with PDT are only partially understood. We have reported previously the generation of several photosensitizer‐specific PDT‐resistant cell variants of HT29 human colon adenocarcinoma cells by selecting cells from sequential PDT treatment using different photosensitizers. In this report, we describe the use of messenger RNA (mRNA) differential display to identify genes that were differentially expressed in the parental HT29 cells compared with their resistant variants. In comparison with parental HT29 cells, mRNA expression was increased in the PDT‐resistant cell variants for BNIP3, estrogen receptor‐binding fragmentassociated gene 9, Myh‐1c, cytoplasmic dynein light chain 1, small membrane protein I and differential dependent protein. In contrast, expression in the PDT‐resistant variants was downregulated for NNX3, human HepG2 3′region Mbol complementary DNA, glutamate dehydrogenase, hepatomaderived growth factor and the mitochondrial genes coding for 16S ribosomal RNA (rRNA) and nicotinamide adenine dinucleotide (NADH) dehydrogenase subunit 4. The reduction for mitochondrial 16S rRNA in the PDT‐resistant variants was confirmed by Northern blotting, and the elevated expression of the proapoptotic BNIP3 in the PDT‐resistant variants was confirmed by Northern and Western blotting analysis. We also examined the expression of some additional apoptosis‐regulating genes using Western blotting. We show an increased expression of Bcl‐2 and heat shock protein 27 and a downregulation of Bax in the PDT‐resistant variants. In addition, the mutant p53 levels in the parental HT29 cells were reduced substantially in the PDT‐resistant variants. We suggest that the altered expression in several mitochondria1 and apoptosisregulating genes contributes to PDT resistance.

Resistance of oral cancer cells to 5‐ALA‐mediated photodynamic therapy

“…Despite the marked alteration in the mitochondrial density, we have found no significant expression of antiapoptotic Bcl‐2 protein in PDT‐resistant cells. This result was different from those of other studies about resistance of oral cancer cells to radiotherapy and PDT . Similarly to our result, a previous investigation also demonstrated a secondary effect of Bcl‐2 …”

“…This result was different from those of other studies about resistance of oral cancer cells to radiotherapy 21,22 and PDT. 23 Similarly to our result, a previous investigation also demonstrated a secondary effect of Bcl-2. 16 Survivin is also an antiapoptotic protein that has been related to poor prognosis of head and neck cancer.…”

Cellular and molecular studies of the initial process of the photodynamic therapy in HEp‐2 cells using LED light source and two different photosensitizers

“…Oxidative stress induced by photosensitization elicits numerous biochemical responses. Damage to multiple organelles occurs following PDT (Shen et al, 2007), to varying degrees of intensity dependent on the photosensitizer (Gomer, 1991;Castano et al, 2005). This chemical mediates oxidative stress and induces transcription of a variety of genes including heat shock proteins, heme oxygenase among other proteins Goldberg et al, 2006) affecting and potentially leading to cell death.…”

Current Advances in Photoactive Agents for Cancer Imaging and Therapy

“…Inhibition of efflux pumps has been successful in antibacterial PDT: Treatment with methylene blue PDT is improved when it is combined with a microbial efflux pump inhibitor (158). Additionally, cell lines resistant to mitochondrial PDT show changes to mitochondrial structure and metabolism, though more research is needed to determine the exact molecular mechanisms (159,160). Overall, this is an area where more research is needed in order to interpret potential resistances associated with therapy and improved indicators for cancers where PDT could be an ideal therapeutic option.…”