Evaluation of aldehyde dehydrogenase 1 (ALDH1) as a marker of non-small cell lung cancer (NSCLC) stem cells (SCs) and correlation with prognosis

Abstract

11105 Background: ALDH1 is a cytosolic enzyme responsible for oxidizing intracellular aldehydes, and conversion of retinol to retinoic acid in SCs. ALDH1 has been previously demonstrated to be a marker for SCs in breast cancer (Ginestier C, et al., Cell Stem Cell. 2007 Nov; 1(5):555–67). The Aldefluor assay is based on conversion of a synthetic substrate BAAA, when passively infused into cytosol, to brightly fluorescent BAA by ALDH1. Combined with fluorescence-activated cell sorting (FACS), it has been used successfully in hematopoietic and breast cancer SCs isolation. ALDH1 is also found in NSCLC. We hypothesized that ALDH1 would be a marker for NSCLC SCs and a potential prognostic marker. Methods: NSCLC SCs were isolated from human NSCLC cell lines using the Aldefluor assay and FACS. ALDH1-positive cells were analyzed extensively for SC characteristics. ALDH1 expression in 303 NSCLC biopsy specimens from three independent cohorts of NSCLC patients was then analyzed by immunohistochemisty (IHC) using an ALDH1 antibody (Santa Cruz Biotechnology) and commercially available negative controls. Results: Isolated cancer cells with high ALDH1 activity displayed cancer SCs features, including capacities for proliferation, self-renewal, differentiation; resistance to chemotherapeutic agents including cisplatin, vinorelbine, gemcitabine and docetaxel; expression of the SC surface marker CD133, and high invasiveness. ALDH1-positive cells generated tumors in vivo recapitulating heterogeneity of parental cells. Xenograft tumors derived from ALDH1-positive cells (103 each) were 36±2.9 mm3 on average in size, with some of the cells having lost ALDH1 expression. Xenograft tumors derived from ALDH1-negative cells (105 each) were 4 mm3 on average in size, without ALDH1 reactivation. Statistical analysis of quantitative IHC and clinical data showed ALDH1 expression was correlated with higher stage and grade of NSCLC (p = 0.02). Expression of ALDH1 in stage I NSCLC patients was linked to decreased 5 year cancer-specific survival (62% vs. 96%, p = 0.006) and overall survival (32% vs. 72%, p = 0.009). Conclusions: 1. ALDH1 is a NSCLC SC-associated tumor marker. 2. ALDH1 expression is a negative prognostic marker in early stage NSCLC. No significant financial relationships to disclose.