Reply: 99mTc-labelled Stealth liposomal doxorubicin (Caelyx®) in glioblastomas and metastatic brain tumours

British Journal of Cancer (2002) 86, 660–661. DOI: 10.1038/sj/bjc/6600094 www.bjcancer.co

Aldehyde dehydrogenase 3A1 promotes multi-modality resistance and alters gene expression profile in human breast adenocarcinoma MCF-7 cells

Aldehyde dehydrogenases participate in a variety of cellular homeostatic mechanisms like metabolism, proliferation, differentiation, apoptosis, whereas recently, they have been implicated in normal and cancer cell stemness. We explored roles for ALDH3A1 in conferring resistance to chemotherapeutics/radiation/oxidative stress and whether ectopic overexpression of ALDH3A1 could lead to alterations of gene expression profile associated with cancer stem cell-like phenotype. MCF-7 cells were stably transfected either with an empty vector (mock) or human aldehyde dehydrogenase 3A1 cDNA. The expression of aldehyde dehydrogenase 3A1 in MCF-7 cells was associated with altered cell proliferation rate and enhanced cell resistance against various chemotherapeutic drugs (4-hydroxyperoxycyclophosphamide, doxorubicin, etoposide, and 5-fluorouracil). Aldehyde dehydrogenase 3A1 expression also led to increased tolerance of MCF-7 cells to gamma radiation and hydrogen peroxide-induced stress. Furthermore, aldehyde dehydrogenase 3A1-expressing MCF-7 cells exhibited gene up-regulation of cyclins A, B1, B2, and down-regulation of cyclin D1 as well as transcription factors p21, CXR4, Notch1, SOX2, SOX4, OCT4, and JAG1. When compared to mock cells, no changes were observed in mRNA levels of ABCA2 and ABCB1 protein pumps with only a minor decrease of the ABCG2 pump in the aldehyde dehydrogenase 3A1-expressing cells. Also, the adhesion molecules EpCAM and CD49F were also found to be up-regulated in aldehyde dehydrogenase 3A1expressing cells. Taken together, ALDH3A1 confers a multi-modality resistance phenotype in MCF-7 cells associated with slower growth rate, increased clonogenic capacity, and altered gene expression profile, underlining its significance in cell homeostasis

Incorporating expression data in metabolic modeling: a case study of lactate dehydrogenase

Integrating biological information from different sources to understand cellular processes is an important problem in systems biology. We use data from mRNA expression arrays and chemical kinetics to formulate a metabolic model relevant to K562 erythroleukemia cells. MAP kinase pathway activation alters the expression of metabolic enzymes in K562 cells. Our array data show changes in expression of lactate dehydrogenase (LDH) isoforms after treatment with phorbol 12-myristate 13-acetate (PMA), which activates MAP kinase signaling. We model the change in lactate production which occurs when the MAP kinase pathway is activated, using a non-equilibrium, chemical-kinetic model of homolactic fermentation. In particular, we examine the role of LDH isoforms, which catalyze the conversion of pyruvate to lactate. Changes in the isoform ratio are not the primary determinant of the production of lactate. Rather, the total concentration of LDH controls the lactate concentration.Comment: In press, Journal of Theoretical Biology. 27 pages, 9 figure

Lactate Dehydrogenase-B Is Silenced by Promoter Methylation in a High Frequency of Human Breast Cancers

Objective: Under normoxia, non-malignant cells rely on oxidative phosphorylation for their ATP production, whereas cancer cells rely on Glycolysis; a phenomenon known as the Warburg effect. We aimed to elucidate the mechanisms contributing to the Warburg effect in human breast cancer. Experimental design: Lactate Dehydrogenase (LDH) isoenzymes were profiled using zymography. LDH-B subunit expression was assessed by reverse transcription PCR in cells, and by Immunohistochemistry in breast tissues. LDH-B promoter methylation was assessed by sequencing bisulfite modified DNA. Results: Absent or decreased expression of LDH isoenzymes 1-4, were seen in T-47D and MCF7 cells. Absence of LDH-B mRNA was seen in T-47D cells, and its expression was restored following treatment with the demethylating agent 5'Azacytadine. LDH-B promoter methylation was identified in T-47D and MCF7 cells, and in 25/ 25 cases of breast cancer tissues, but not in 5/ 5 cases of normal breast tissues. Absent immuno-expression of LDH-B protein (<10% cells stained), was seen in 23/ 26 (88%) breast cancer cases, and in 4/8 cases of adjacent ductal carcinoma in situ lesions. Exposure of breast cancer cells to hypoxia (1% O2), for 48 hours resulted in significant increases in lactate levels in both MCF7 (14.0 fold, p = 0.002), and T-47D cells (2.9 fold, p = 0.009), but not in MDA-MB-436 (-0.9 fold, p = 0.229), or MCF10AT (1.2 fold, p = 0.09) cells. Conclusions: Loss of LDH-B expression is an early and frequent event in human breast cancer occurring due to promoter methylation, and is likely to contribute to an enhanced glycolysis of cancer cells under hypoxia

The prognostic value of the hypoxia markers CA IX and GLUT 1 and the cytokines VEGF and IL 6 in head and neck squamous cell carcinoma treated by radiotherapy ± chemotherapy

BACKGROUND: Several parameters of the tumor microenvironment, such as hypoxia, inflammation and angiogenesis, play a critical role in tumor aggressiveness and treatment response. A major question remains if these markers can be used to stratify patients to certain treatment protocols. The purpose of this study was to investigate the inter-relationship and the prognostic significance of several biological and clinicopathological parameters in patients with head and neck squamous cell carcinoma (HNSCC) treated by radiotherapy ± chemotherapy. METHODS: We used two subgroups of a retrospective series for which CT-determined tumoral perfusion correlated with local control. In the first subgroup (n = 67), immunohistochemistry for carbonic anhydrase IX (CA IX) and glucose transporter-1 (GLUT-1) was performed on the pretreatment tumor biopsy. In the second subgroup (n = 34), enzyme linked immunosorbent assay (ELISA) was used to determine pretreatment levels of the cytokines vascular endothelial growth factor (VEGF) and interleukin-6 (IL-6) in serum. Correlation was investigated between tumoral perfusion and each of these biological markers, as well as between the markers mutually. The prognostic value of these microenvironmental parameters was also evaluated. RESULTS: For CA IX and GLUT-1, the combined assessment of patients with both markers expressed above the median showed an independent correlation with local control (p = 0.02) and disease-free survival (p = 0.04) with a trend for regional control (p = 0.06). In the second subgroup, IL-6 pretreatment serum level above the median was the only independent predictor of local control (p = 0.009), disease-free survival (p = 0.02) and overall survival (p = 0.005). CONCLUSION: To our knowledge, we are the first to report a link in HNSCC between IL-6 pretreatment serum levels and radioresistance in vivo. This link is supported by the strong prognostic association of pretreatment IL-6 with local control, known to be the most important parameter to judge radiotherapy responses. Furthermore, the combined assessment of CA IX and GLUT-1 correlated independently with prognosis. This is a valuable indication that a combined approach is important in the investigation of prognostic markers

Clinical significance of VEGF-A, -C and -D expression in esophageal malignancies

Vascular endothelial growth factors ( VEGF)- A, - C and - D are members of the proangiogenic VEGF family of glycoproteins. VEGF-A is known to be the most important angiogenic factor under physiological and pathological conditions, while VEGF-C and VEGF-D are implicated in the development and sprouting of lymphatic vessels, so called lymphangiogenesis. Local tumor progression, lymph node metastases and hematogenous tumor spread are important prognostic factors for esophageal carcinoma ( EC), one of the most lethal malignancies throughout the world. We found solid evidence in the literature that VEGF expression contributes to tumor angiogenesis, tumor progression and lymph node metastasis in esophageal squamous cell carcinoma ( SCC), and many authors could show a prognostic value for VEGF-assessment. In adenocarcinoma (AC) of the esophagus angiogenic properties are acquired in early stages, particularly in precancerous lesions like Barrett's dysplasia. However, VEGF expression fails to give prognostic information in AC of the esophagus. VEGF-C and VEGF-D were detected in SCC and dysplastic lesions, but not in normal mucosa of the esophagus. VEGF-C expression might be associated with lymphatic tumor invasion, lymph node metastases and advanced disease in esophageal SCC and AC. Therapeutic interference with VEGF signaling may prove to be a promising way of anti-angiogenic co-treatment in esophageal carcinoma. However, concrete clinical data are still pending

A lactate shuttle system between tumour and stromal cells is associated with poor prognosis in prostate cancer

Background In a malignant tumour, cancer cells are embedded in stromal cells, namely cancer-associated fibroblasts (CAFs). These CAFs are now accepted as important players in cancer dynamics, being involved in tumour growth and progression. Although there are various reports on the interaction between tumour and stromal cells, the clinico-pathological significance of this cross-talk is still largely unknown. In this study, we aimed to characterise the expression of key metabolic proteins involved in glucose transport, pyruvate/lactate shuttle system, glycolytic metabolism and fatty acid oxidation in CAFs and tumour cells in different stages of malignant transformation. We further aimed to contextualise the clinico-pathological significance of these protein expression profiles with reference to known prognostic indicators, including biochemical recurrence in pT stage. Methods Prostate tissues were obtained from 480 patients with a median age of 64 years following radical prostatectomy with no previous hormonal therapy. Tissues were analysed for the expression of several key metabolism-related proteins in glands and surrounding fibroblasts by immunohistochemistry. Reliable markers of prognosis such as pT stage and biochemical recurrence were assessed for each case. Results We observed that prostate cancer cells did not rely mainly on glycolytic metabolism, while there was a high expression of MCT4 and CAIX - in CAFs. This corroborates the hypothesis of the "Reverse Warburg effect" in prostate cancer, in which fibroblasts are under oxidative stress and express CAIX, an established hypoxia marker. We found that alterations in the expression of metabolism-related proteins were already evident in the early stages of malignant transformation, suggesting the continuing alteration of CAFs from an early stage. Additionally, and for the first time, we show that cases showing high MCT4 expression in CAFs with concomitant strong MCT1 expression in prostate cancer (PCa) cells are associated with poor clinical outcome, namely pT3 stage of the tumour. Conclusions In summary, this work demonstrates for the first time the clinico-pathological significance of the lactate shuttle in prostate cancer. It also suggests that other alterations in CAFs may be useful prognostic factors, and further supports the use of MCT1/MCT4 as targets for PCa therapy.NPG received a fellowship from the Portuguese Foundation for Science and Technology (FCT), refs. SFRH/BD/61027/2009. This work was supported by the FCT grant ref. PTDC/SAUMET/113415/2009, under the scope of "Programa Operacional Tematico Factores de Competitividade" (COMPETE) of "Quadro Comunitario de Apoio III" and co-financed by Fundo Comunitario Europeu FEDER. JA was supported by a Boehringer Ingelheim Fonds fellowship