Synthesis and Characterization of Catechol-Containing Polyacrylamides with Adhesive Properties.

In this study, a row of four analogous dopamine acryl- and methacrylamide derivatives, namely N-(3,4-dihydroxyphenyethyl) acrylamide, N-(3,4-dihydroxyphenyethyl) meth acrylamide, N-phenethyl methacrylamide, N-(4-hydroxyphenethyl) methacrylamide were synthesized and characterized by 1H-NMR and 13C-NMR, followed by further solvent-based radical polymerization with N-hydroxyethyl acrylamide. All copolymers were characterized by 1H-NMR, dynamic differential calorimetry, and gel permeation chromatography. The dependency of the used comonomer ratios to the molecular mass of the corresponding copolymers has been described. The synthesis of the various polymers serves as a feasibility study and provides important data for a future biometric application in the medical field. We synthesized N-(3,4-dihydroxyphenyethyl) acrylamide copolymer up to 80 mol% by free radical polymerization without using any protecting groups. All polymers show identical perfect adhesive properties by a simple scratch test. Further, the monomers were used as a photo reactive glue formulation to test its adherence to a medical titanium surface sample by tensile shear test.

Acrylate functionalized tetraalkylammonium salts with ionic liquid properties.

Acrylate functionalized ionic liquids based on tetraalkylammonium salts with terminal acrylates- and methylacrylates were synthesized. Melting points and ionic conductivity of twenty compounds in six groups were determined. Within one group the effect of three different counterions was investigated and discussed. The groups differ in cationic structure elements because of their functional groups such as acrylate and methacrylate, alkyl residues at the nitrogen and number of quaternary ammonium atoms within the organic cation. The effect of these cationic structure elements has been examined concerning the compiled parameters with a view to qualifying them as components for solid state electrolytes. The newly synthesized ionic liquids were characterized by NMR and FTIR analysis. The exchange of halide ions like bromide as counter ions to weakly coordinating [PF6]⁻, [OTf]⁻ or [TFSI]⁻ reduces the melting points significantly and leads to an ion conductivity of about 10⁻4 S/cm at room temperature. In the case of the dicationic ionic liquid, an ion conductivity of about 10⁻³ S/cm was observed.

Photo-Curing Kinetics of 3D-Printing Photo-Inks Based on Urethane-Acrylates.

In this study, photo-curing kinetics for urethane-acrylate-based photo-inks for 3D printing were evaluated using a photo-differential scanning calorimetry analysis. Initially, the photopolymerization kinetics of di- and monofunctional monomers were separately studied at different temperatures (5-85 °C). Later, the photo-curing kinetics and mechanical properties of photo-inks based on different monomer mixtures (40/60-20/80) were evaluated. The results showed that urethane-dimethacrylate (UrDMA) and urethane-acrylate (UrA) had no light absorption in the region of 280-700 nm, making them a proper crosslinker and a reactive diluent, respectively, for the formulation of 3D-printing photo-inks. The kinetics investigations showed a temperature dependency for the photo-curing of UrDMA, where a higher photopolymerization rate (Rp,max: from 5.25 × 10-2 to 8.42 × 10-2 1/s) and double-bound conversion (DBCtotal: from 63.8% to 92.2%) were observed at elevated temperatures (5-85 °C), while the photo-curing of UrA was independent of the temperature (25-85 °C). Enhancing the UrA content from 60% to 80% in the UrDMA/UrA mixtures initially increased and later decreased the photopolymerization rate and conversion, where the mixtures of 30/70 and 25/75 presented the highest values. Meanwhile, increasing the UrA content led to lower glass transition temperatures (Tg) and mechanical strength for the photo-cured samples, where the mixture of 30/70 presented the highest maximum elongation (εmax: 73%).

Green Chemistry for Biomimetic Materials: Synthesis and Electrospinning of High-Molecular-Weight Polycarbonate-Based Nonisocyanate Polyurethanes.

Conventional synthesis routes for thermoplastic polyurethanes (TPUs) still require the use of isocyanates and tin-based catalysts, which pose considerable safety and environmental hazards. To reduce both the ecological footprint and human health dangers for nonwoven TPU scaffolds, it is key to establish a green synthesis route, which eliminates the use of these toxic compounds and results in biocompatible TPUs with facile processability. In this study, we developed high-molecular-weight nonisocyanate polyurethanes (NIPUs) through transurethanization of 1,6-hexanedicarbamate with polycarbonate diols (PCDLs). Various molecular weights of PCDL were employed to maximize the molecular weight of NIPUs and consequently facilitate their electrospinnability. The synthesized NIPUs were characterized by nuclear magnetic resonance, Fourier-transform infrared spectroscopy, gel permeation chromatography, and differential scanning calorimetry. The highest achieved molecular weight (M w) was 58,600 g/mol. The NIPUs were consecutively electrospun into fibrous scaffolds with fiber diameters in the submicron range, as shown by scanning electron microscopy (SEM). To assess the suitability of electrospun NIPU mats as a possible biomimetic load-bearing pericardial substitute in cardiac tissue engineering, their cytotoxicity was investigated in vitro using primary human fibroblasts and a human epithelial cell line. The bare NIPU mats did not need further biofunctionalization to enhance cell adhesion, as it was not outperformed by collagen-functionalized NIPU mats and hence showed that the NIPU mats possess a great potential for use in biomimetic scaffolds.

Developing non-isocyanate urethane-methacrylate photo-monomers for 3D printing application.

Urethane-methacrylate photo-monomers were prepared via a non-isocyanate route for the 3D printing application. The monomers were synthesized through reacting aliphatic amines, i.e. 1,6-hexanediamine, 1,4-butanediol bis(3-aminopropyl) ether, or n-butylamine, with cyclic carbonates, i.e. ethylene carbonate or propylene carbonate, followed by the methacrylation of the generated hydroxylurethanes. The effects of the chemical structure of monomers on their photo-reactivity and physicomechanical properties of the cured samples were studied. Propylene carbonate generated side methyl groups within the urethane block, which significantly limited the crystallization of the monomers resulting in high photo-reactivity (R p,max = 6.59 × 10-2 s-1) and conversion (DBCtotal = 85%). The ether bonds of 1,4-butanediol bis(3-aminopropyl) ether decreased the intermolecular hydrogen bonding between urethane blocks, which not only improved the photo-reactivity (R p,max = 8.18 × 10-2 s-1) and conversion (DBCtotal = 86%) of the monomer but led to a high crosslinking density (ν c = 5140 mol m-3) and more flexibility for the cured sample. An ink was developed based on the monomers and successfully 3D printed on a digital light processing machine. In the absence of toxic isocyanates and tin compounds, the non-isocyanate route can be employed to develop urethane-methacrylates with desirable photo-reactivity and physicomechanical properties as good candidates to formulate inks for 3D printing of biomedical materials.

Linking proteins with anionic nanoparticles via protamine: ultrasmall protein-coupled probes for magnetic resonance imaging of apoptosis.

Magnetic resonance imaging (MRI) of a target in vivo depends on the surface, size, and particle relaxivity of the target-specific nanoparticles for MRI. Here a new method for decorating very small iron oxide particles (VSOPs) with target-specific ligands is described. The method is based on the electrostatic attraction of the strongly positively charged peptide protamine to the anionic citrate shell of the electrostatically stabilized VSOPs. The protamine coat allows linkage chemistry and chimera technology to functionalize VSOPs or other negative charged surfaces with biologics. Annexin A5 (anxA5)-VSOP utilizing thiol chemistry was generated to couple biologically active anxA5 to VSOPs for in vivo MRI of apoptosis. Annexin A5-VSOP comprises five anxA5 molecules per iron oxide nanoparticle with a high R2 particle relaxivity of 180 000 mM(-1) s(-1) yet small hydrodynamic diameter of only 14.7+/-2.9 nm beneficial for in vivo MRI of extravascular targets.

Chromosomal genotype in breast cancer progression: comparison of primary and secondary manifestations.

The purpose of this study was to compare the chromosomal genotype between breast cancers with and without secondary manifestations and between primary tumors and their secondary manifestations. Eighty six breast cancers, twenty lymph node metastases, ten distant metastases and ten local recurrences were analyzed by comparative genomic hybridization. Tumors with local recurrences showed significant more frequent losses at 2q32 than the tumors without recurrences. Lymph node positive cases showed significant more frequent losses at 9p21 than node negative cases. Lymph node metastases exhibited significant more frequent losses at 7q11, 14q24.3-q31 and 17q22-q24 than their primary tumors. In cases with distant metastases, losses at 5q23 were more frequent than in those without, but not reaching the significance level. The distant metastases showed significant more frequent losses at 5p15, 12q24 and 17q22-q24 than the primary tumors. These results reveal strong evidence that the potential for progression is determined in the primary tumor and that different ways of the development of local recurrences, lymph node and distant metastases exist. After confirmation of the results by interphase FISH on tissue micro arrays, the detection of these specific chromosomal imbalances may contribute to a more individual prediction of prognosis in breast cancer.

Predictive value of Ki67 and p53 in locally advanced rectal cancer: correlation with thymidylate synthase and histopathological tumor regression after neoadjuvant 5-FU-based chemoradiotherapy.

AIM: To investigate the predictive value of Ki67 and p53 and their correlation with thymidylate synthase (TS) gene expression in a rectal cancer patient cohort treated according to a standardized recommended neoadjuvant treatment regimen. METHODS: Formalin fixed, paraffin embedded pre-therapeutical tumor biopsies (n = 22) and post-therapeutical resection specimens (n = 40) from patients with rectal adenocarcinoma (clinical UICC stage II/III) receiving standardized neoadjuvant 5-fluorouracil (5-FU) based chemoradiotherapy were studied for Ki67 and p53 expression by immunohistochemistry and correlated with TS mRNA expression by quantitative TaqMan real-time PCR after laser microdissection. The results were compared with histopathological tumor regression according to a standardized semiquantitative score grading system. RESULTS: Responders (patients with high tumor regression) showed a significantly lower Ki67 expression than non-responders in the pre-therapeutical tumor biopsies (81.2% vs 16.7%; P < 0.05) as well as in the post-therapeutical resection specimens (75.8% vs 14.3%; P < 0.01). High TS mRNA expression was significantly correlated with a high Ki67 index and low TS mRNA expression was significantly correlated with a low Ki67 index in the pre-therapeutical tumor biopsies (corr. coef. = 0.46; P < 0.01) as well as in the post-therapeutical resection specimens (corr. coef. = 0.40; P < 0.05). No significant association was found between p53 and TS mRNA expression or tumor regression. CONCLUSION: Ki67 has, like TS, predictive value in rectal cancer patients after neoadjuvant 5-FU based chemoradiotherapy. The close correlation between Ki67 and TS indicates that TS is involved in active cell cycle processes.

Optimized vascular network by stereolithography for tissue engineered skin.

This paper demonstrates the essential and efficient methods to design, and fabricate optimal vascular network for tissue engineering structures based on their physiological conditions. Comprehensive physiological requirements in both micro and macro scales were considered in developing the optimisation design for complex vascular vessels. The optimised design was then manufactured by stereolithography process using materials that are biocompatible, elastic and surface bio-coatable. The materials are self-developed photocurable resin consist of BPA-ethoxylated-diacrylate, lauryl acrylate and isobornylacrylate with Irgacure® 184, the photoinitiator. The optimised vascular vessel offers many advantages: 1) it provides the maximum nutrient supply; 2) it minimises the recirculation areas and 3) it allows the wall shear stress on the vessel in a healthy range. The stereolithography manufactured vascular vessels were then embedded in the hydrogel seeded with cells. The results of in vitro studies show that the optimised vascular network has the lowest cell death rate compared with a pure hydrogel scaffold and a hydrogel scaffold embedded within a single tube in day seven. Consequently, these design and manufacture routes were shown to be viable for exploring and developing a high range complex and specialised artificial vascular networks.

Prognostic value of histologic tumor regression, thymidylate synthase, thymidine phosphorylase, and dihydropyrimidine dehydrogenase in rectal cancer UICC Stage II/III after neoadjuvant chemoradiotherapy.

Histologic tumor regression (TR) in rectal cancer after preoperative chemoradiotherapy (CT/RT) may be useful as a surrogate end point for early treatment efficacy, but little is known about its prognostic value. The aim of this follow-up study was to evaluate whether TR is able to predict prognosis in rectal cancer patients. Furthermore, the prognostic value of thymidylate synthase (TS)-gene, thymidine phosphorylase (TP)-gene, and dihydropyrimidine dehydrogenase (DPD)-gene expression after neoadjuvant CT/RT was determined. Forty patients with rectal cancer cUICC stage II/III, receiving preoperative 5-fluorouracil (5-FU) based CT/RT were studied for therapy-induced TR and categorized as "responders" or "nonresponders" according to their TR-grade. Posttherapeutical TS-gene, TP-gene, and DPD-gene expression on surgical resection specimens was quantified by TaqMan real-time PCR after microdissection. During a median follow-up of 58 months, cancer recurrence occurred in 28%. A significant correlation was seen between disease-free survival and lymph node status (P<0.001). All patients, who developed cancer recurrence had a posttherapeutical positive lymph node status. The majority of patients with cancer recurrence were "responders" (91%) after CT/RT. There was a significant correlation between posttherapeutical TS-gene expression and cancer recurrence within the subgroup of "responders." TS-gene expression was significantly higher in patients with cancer recurrence than in those, who are disease-free up to date (P=0.028). In conclusion, lymph node status remains the most important prognostic marker in rectal cancer patients, whereas posttreatment TR by itself has no prognostic significance. Furthermore, measurement of posttherapeutical TS-gene expression may help to identify patients at higher risk for cancer recurrence.

Blood-Vessel Mimicking Structures by Stereolithographic Fabrication of Small Porous Tubes Using Cytocompatible Polyacrylate Elastomers, Biofunctionalization and Endothelialization.

Blood vessel reconstruction is still an elusive goal for the development of in vitro models as well as artificial vascular grafts. In this study, we used a novel photo-curable cytocompatible polyacrylate material (PA) for freeform generation of synthetic vessels. We applied stereolithography for the fabrication of arbitrary 3D tubular structures with total dimensions in the centimeter range, 300 µm wall thickness, inner diameters of 1 to 2 mm and defined pores with a constant diameter of approximately 100 µm or 200 µm. We established a rinsing protocol to remove remaining cytotoxic substances from the photo-cured PA and applied thio-modified heparin and RGDC-peptides to functionalize the PA surface for enhanced endothelial cell adhesion. A rotating seeding procedure was introduced to ensure homogenous endothelial monolayer formation at the inner luminal tube wall. We showed that endothelial cells stayed viable and adherent and aligned along the medium flow under fluid-flow conditions comparable to native capillaries. The combined technology approach comprising of freeform additive manufacturing (AM), biomimetic design, cytocompatible materials which are applicable to AM, and biofunctionalization of AM constructs has been introduced as BioRap(®) technology by the authors.

Immunohistochemical analysis of thymidylate synthase, thymidine phosphorylase, and dihydropyrimidine dehydrogenase in rectal cancer (cUICC II/III): correlation with histopathologic tumor regression after 5-fluorouracil-based long-term neoadjuvant chemoradiotherapy.

In locally advanced rectal cancer, neoadjuvant 5-fluorouracil (5-FU)-based long-term chemoradiotherapy leads to marked tumor reduction and decrease of local recurrence rate. Thymidylate synthase (TS), thymidine phosphorylase (TP), and dihydropyrimidine dehydrogenase (DPD) are known to be important biomarkers to predict tumor response to 5-FU-based therapy. The aim of this study was to examine the correlation between TS, TP, and DPD protein expression and histopathologic tumor regression after neoadjuvant chemoradiotherapy. The results were compared with the recently published mRNA data. Preoperative biopsies (n = 25) and resection specimens (n = 40) from patients with rectal carcinoma (clinical UICC stage II/III) receiving neoadjuvant 5-FU-based chemoradiotherapy were studied for TS, TP, and DPD protein expression by immunohistochemistry using three different scoring systems (intensity, pattern, intensity + pattern). Results were compared with histopathologic tumor regression. A significant correlation between protein expression and tumor response was only seen when both staining intensity and staining pattern were considered. With this method, a significant association was seen between high TS expression in tumor biopsies as well as resection specimens and nonresponse of the tumor to therapy (P = 0.04). Furthermore, low TP expression in the resection specimens was significantly associated with lack of response (P = 0.02). For DPD no significant correlations were found at all. In conclusion, these results suggest that immunohistochemistry like RT-PCR is a suitable method to determine the correlation between TS, TP, and DPD expression and histopathologic tumor regression. However, precise results can only be achieved if staining intensity as well as staining pattern within the tumors are evaluated.

Soft Polymers for Building up Small and Smallest Blood Supplying Systems by Stereolithography.

Synthesis of a homologous series of photo-polymerizable α,w-polytetrahydrofuranether-diacrylate (PTHF-DA) resins is described with characterization by NMR, GPC, DSC, soaking and rheometrical measurements. The curing speeds of the resins are determined under UV light exposure. Young's modulus and tensile strength of fully cured resins show flexible to soft material attributes dependent on the molar mass of the used linear PTHF-diacrylates. Structuring the materials by stereo lithography (SL) and multiphoton polymerization (MPP) leads to tubes and bifurcated tube systems with a diameter smaller than 2 mm aimed at small to smallest supplying systems with capillary dimensions. WST-1 biocompatibility tests ofm polymer extracts show nontoxic characteristics of the adapted polymers after a washing process. Some polymers show shape memory effect (SME).

Fabrication of 2D protein microstructures and 3D polymer-protein hybrid microstructures by two-photon polymerization.

Two-photon polymerization (TPP) offers the possibility of creating artificial cell scaffolds composed of micro- and nanostructures with spatial resolutions of less than 1 µm. For use in tissue engineering, the identification of a TPP-processable polymer that provides biocompatibility, biofunctionality and appropriate mechanical properties is a difficult task. ECM proteins such as collagen or fibronectin, which could mimic native tissues best, often lack the mechanical stability. Hence, by generating polymer-protein hybrid structures, the beneficial properties of proteins can be combined with the advantageous characteristics of polymers, such as sufficient mechanical stability. This study describes three steps toward facilitated application of TPP for biomaterial generation. (1) The efficiency of a low-cost ps-laser source is compared to a fs-laser source by testing several materials. A novel photoinitiator for polymerization with a ps-laser source is synthesized and proved to enable increased fabrication throughput. (2) The fabrication of 3D-microstructures with both systems and the fabrication of polymer-protein hybrid structures are demonstrated. (3) The tissue engineering capabilities of TPP are demonstrated by creating cross-linked gelatin microstructures, which clearly forced porcine chondrocytes to adapt their cell morphology.

Correlation of centrosomal aberrations with cell differentiation and DNA ploidy in prostate cancer.

OBJECTIVE: To analyze the centrosomal abnormalities in correlation with DNA ploidy and clinicopathologic data in prostate cancer. STUDY DESIGN: Formalin-fixed, paraffin-embedded material from 63 prostate cancers (PCa) and 10 normal control cases were studied. Centrosomal features (number, area and shape) were assessed by immunohistochemistry with a gamma-tubulin monoclonal antibody. For each case centrosomal features were assessed in 100 cells, and the mean and median value was calculated. Statistical analysis was done by Student's t test, Mann-Whitney U test and multivariate analysis. The colocalization of gamma-tubulin and pericentrin at the centrosome was proven by double immunofluorescence staining. The DNA ploidy status was analyzed on Feulgen-stained, disintegrated paraffin sections using the OPTIMAS-based work station (Media Cybernetics, Silver Spring, Maryland, U.S.A.). RESULTS: PCa cells showed centrosomal aberrations when compared to normal tissue. Poorly differentiated PCa showed more centrosomal abnormalities than well differentiated PCa (p < 0.05). Twenty-seven percent PCa were DNA nondiploid and 73% PCa were DNA diploid, respectively, just as all control specimens. DNA nondiploid status correlates with centrosomal abnormalities (p < 0.05). pT4 tumors showed significantly more centrosomes than pT2 and pT3 tumors (p < 0.05). CONCLUSION: Changes in centrosome features indicate disturbed centrosome function and are significantly correlated with loss of differentiation in PCa. This is the first image analysis study of centrosome features in PCa, confirming that centrosome defects are involved in the acquisition of chromosomal aberrations in PCa.

Comparative genomic hybridization-based oncogenetic tree model for genetic classification of breast cancer.

OBJECTIVE: To describe a genetic progression pathway in breast cancer by a maximum likelihood-based tree model representing the dependencies between chromosomal imbalances. STUDY DESIGN: One hundred six cases were studied by comparative genomic hybridization, followed by maximum likelihood estimation of an oncogenetic tree model. RESULTS: The tree model identified 3 clusters with correlated chromosomal imbalances. The first cluster included losses at 4q, 5q, 6q, 9p, 13q and a gain at 17q; the second cluster included gains at 1q, 8q, 16p and 20q; the third cluster included losses at 8p, 11q, 16q and 18q. The imbalances nearest the root of the tree were the loss at 13q (cluster 1), the gain at 1q (cluster 2) and the loss at 18q (cluster 3), reflecting an early change in breast cancer evolution. Cox regression analysis revealed the tumor stage and the grade as relevant for overall survival (p = 0.001) and the tumor stage, the grade and the loss at 16q as relevant for disease-free survival (p = 0.001). CONCLUSION: Methods like oncogenetic tree analysis provide insights into the genetic progression of breast cancer and may extract relevant markers detected by screening methods like comparative genomic hybridization for further studies.

Prognostic significance of immunohistochemical RhoA expression on survival in pancreatic ductal adenocarcinoma: a high-throughput analysis.

Among all human carcinomas, pancreatic cancer has one of the worst survival rates. Most patients will die of this cancer shortly after diagnosis, and currently, surgery is the only potential cure. Ductal adenocarcinoma is the most common histologic type. The search for prognostic parameters has progressed from mere physical or histomorphological tumor properties to molecular parameters. These, in turn, might point toward new therapeutic strategies. The K-ras oncogene is known to play a role in early stages of ductal adenocarcinoma carcinogenesis, and ras homologues are differentially expressed in cancerous versus normal ductal cells. RhoA belongs to a family of ras homologues comprising RhoA, RhoB, and RhoC. It is a guanosine triphosphatase associated with the cytoskeleton that seems to be involved in epithelial mesenchymal transition, a process of dedifferentiation. Immunohistologic RhoA expression was studied in a tissue microarray of 94 pancreatic ductal adenocarcinomas and correlated with clinicopathologic parameters and follow-up. RhoA protein expression, measured as labeling intensity or evaluated as percentage of reactive tumor cells, correlated with overall survival. A multivariate analysis demonstrated that RhoA protein expression is independent from other known prognostic parameters such as tumor size or grade. Moreover, a score combining RhoA expression with tumor size and grade resulted in a highly significant increase in the prognostic value for the overall survival of patients with pancreatic ductal adenocarcinoma.

Charge propagation in "ion channel sensors" based on protein-modified electrodes and redox marker ions.

The mechanism of charge propagation in "ion channel sensors" (ICSs) consisting of gold electrodes modified with a layer of charged proteins and highly charged redox-active marker ions in solution was investigated by electrochemical techniques, QCM and AFM. The study is based on seven proteins (concanavalin A, cytochrome c, glucose oxidase, lysozyme, thyroglobulin, catalase, aldolase, and EF1-ATPase) in combination with seven electroactive marker ions ([Fe(CN)6]3-, [Fe(CN)6]4-, [Ru(NH3)6]3+, mono-, di-, and trimeric viologens), as well as a series of suppressor and enhancer ions leading to the following general statements: (i) electrostatic binding of charged marker ions to the domains of the protein is a prerequisite for an electrochemical current and (ii) charge propagation through the layer consists of electron hopping along surface-confined marker ions into the pores between adsorbed proteins. It is further shown that (iii) marker ions and suppressor ions with identical charge compete for oppositely charged sites on the protein domain, (iv) electrostatically bound multilayers of marker or enhancer ions with alternating charge form on a charged protein domain, and (v) self-exchange and exergonic ET catalysis between adsorbed marker ions and marker ions in solution take place. In addition to fundamental insight into the mechanism of charge propagation, valuable information for the design, optimization, and tailoring of new biosensors based on the ICS concept is demonstrated by the current findings.

Thymidylate synthase, thymidine phosphorylase, dihydropyrimidine dehydrogenase expression, and histological tumour regression after 5-FU-based neo-adjuvant chemoradiotherapy in rectal cancer.

Pre-operative 5-fluorouracil (5-FU)-based chemoradiotherapy in locally advanced rectal cancer (UICC-II/III) may significantly reduce local tumour mass. Response to pre-operative treatment, however, varies significantly. Thymidylate synthase (TS), thymidine phosphorylase (TP), and dihydropyrimidine dehydrogenase (DPD) are thought to be important predictors for the efficiency of 5-FU-based treatment. The aim of this study was to determine the correlation between TS-, TP-, and DPD-gene expression and the response to 5-FU-based long-term pre-operative chemoradiotherapy assessed by histopathological tumour regression. Additionally, the predictive value of intra-tumoural TS-, TP-, and DPD-gene expression in pre-operative rectal tumour biopsies was assessed by correlation with the histopathological regression grade. Formalin-fixed, paraffin wax-embedded pre-operative biopsies (n = 14) and surgical resection specimens (n = 40) from patients with rectal carcinoma (clinical UICC stage II/III) receiving neo-adjuvant 5-FU-based chemoradiotherapy were studied for TS-, TP-, and DPD-gene expression by quantitative TaqMan real-time PCR after laser microdissection. Results were compared with standardized histopathological tumour regression analysis. There was a significant association between low TS-gene expression in pre-operative tumour biopsies and tumour response (p = 0.02). TS- and TP-gene expression was significantly lower in resection specimens of responders than of non-responders (p = 0.02) when microdissection was used. Statistical significance was even higher when TS and TP were combined (p = 0.0001). For the DPD gene, no significance was found at all. In conclusion, this study shows that TS gene expression in a pretreatment biopsy predicts the response of local rectal cancer to neo-adjuvant 5-FU-based chemoradiotherapy in a high percentage. Moreover, intra-tumoural TS- and TP-gene expression in surgical rectal specimens after neo-adjuvant chemoradiotherapy correlates significantly with histopathological tumour regression when microdissection is applied.