Perioperative immune responses in cancer patients undergoing digestive surgeries

Aims

We will examine the latest advances in genomic and proteomic laboratory technology. Through an extensive literature review we aim to critically appraise those studies which have utilized these latest technologies and ascertain their potential to identify clinically useful biomarkers.

Methods

An extensive review of the literature was carried out in both online medical journals and through the Royal College of Surgeons in Ireland library.

Results

Laboratory technology has advanced in the fields of genomics and oncoproteomics. Gene expression profiling with DNA microarray technology has allowed us to begin genetic profiling of colorectal cancer tissue. The response to chemotherapy can differ amongst individual tumors. For the first time researchers have begun to isolate and identify the genes responsible. New laboratory techniques allow us to isolate proteins preferentially expressed in colorectal cancer tissue. This could potentially lead to identification of a clinically useful protein biomarker in colorectal cancer screening and treatment.

Conclusion

If a set of discriminating genes could be used for characterization and prediction of chemotherapeutic response, an individualized tailored therapeutic regime could become the standard of care for those undergoing systemic treatment for colorectal cancer. New laboratory techniques of protein identification may eventually allow identification of a clinically useful biomarker that could be used for screening and treatment. At present however, both expression of different gene signatures and isolation of various protein peaks has been limited by study size. Independent multi-centre correlation of results with larger sample sizes is needed to allow translation into clinical practice.     

Characterization of the Human Cervical Mucous Proteome

Introduction

Cervical cancer is among the most common cancers in women worldwide. Discovery of biomarkers for the early detection of cervical cancer would improve current screening practices and reduce the burden of disease.

Objective

In this study, we report characterization of the human cervical mucous proteome as the first step towards protein biomarker discovery.

Methods

The protein composition was characterized using one- and two-dimensional gel electrophoresis, and liquid chromatography coupled with mass spectrometry. We chose to use this combination of traditional biochemical techniques and proteomics to allow a more comprehensive analysis.

Results and Conclusion

A total of 107 unique proteins were identified, with plasma proteins being most abundant. These proteins represented the major functional categories of metabolism, immune response, and cellular transport. Removal of high molecular weight abundant proteins by immunoaffinity purification did not significantly increase the number of protein spots resolved. We also analyzed phosphorylated and glycosylated proteins by fluorescent post-staining procedures. The profiling of cervical mucous proteins and their post-translational modifications can be used to further our understanding of the cervical mucous proteome.     

Differential expression of proteomics models of colorectal cancer, colorectal benign disease and healthy controls

Background

Colorectal cancer (CRC) is often diagnosed at a late stage with concomitant poor prognosis. The hypersensitive analytical technique of proteomics can detect molecular changes before the tumor is palpable. The surface-enhanced laser desorption/ionization-time of flight-mass spectra (SELDI-TOF-MS) is a newly-developed technique of evaluating protein separation in recent years. The protein chips have established the expression of tumor protein in the serum specimens and become the newly discovered markers for tumor diagnosis. The objective of this study was to find new markers of the diagnosis among groups of CRC, colorectal benign diseases (CBD) and healthy controls. The assay of SELDI-TOF-MS with analytical technique of protein-chip bioinformatics was used to detect the expression of protein mass peaks in the sera of patients or controls. One hundred serum samples, including 52 cases of colorectal cancer, 27 cases of colorectal benign disease, and 21 cases of healthy controls, were examined by SELDI-TOF-MS with WCX2 protein-chips.

Results

The diagnostic models (I, II and III) were setup by analyzed the data and sieved markers using Ciphergen - Protein-Chip-Software 5.1. These models were combined with 3 protein mass peaks to discriminate CRC, CBD, and healthy controls. The accuracy, the sensitivity and the particularity of cross verification of these models are all highly over 80%.

Conclusions

The SELDI-TOF-MS is a useful tool to help diagnose colorectal cancer, especially during the early stage. However, identification of the significantly differentiated proteins needs further study.     

18O Labeling for a Quantitative Proteomic Analysis of Glycoproteins in Hepatocellular Carcinoma

Introduction

Quantitative proteomics using tandem mass spectrometry is an attractive approach for identification of potential cancer biomarkers. Fractionation of complex tissue samples into subproteomes prior to mass spectrometric analyses increases the likelihood of identifying cancer-specific proteins that might be present in low abundance. In this regard, glycosylated proteins are an interesting class of proteins that are already established as biomarkers for several cancers.

Materials and Methods

In this study, we carried out proteomic profiling of tumor and adjacent non-cancer liver tissues from hepatocellular carcinoma (HCC) patients. Glycoprotein enrichment from liver samples using lectin affinity chromatography and subsequent ¹⁸O/¹⁶O labeling of peptides allowed us to obtain relative abundance levels of lectin-bound proteins. As a complementary approach, we also examined the relative expression of proteins in HCC without glycoprotein enrichment. Lectin affinity enrichment was found to be advantageous to quantitate several interesting proteins, which were not detected in the whole proteome screening approach. We identified and quantitated over 200 proteins from the lectin-based approach. Interesting among these were fetuin, cysteine-rich protein 1, serpin peptidase inhibitor, leucine-rich alpha-2-glycoprotein 1, melanoma cell adhesion molecule, and heparan sulfate proteoglycan-2. Using lectin affinity followed by PNGase F digestion coupled to ¹⁸O labeling, we identified 34 glycosylation sites with consensus sequence N-X-T/S. Western blotting and immunohistochemical staining were carried out for several proteins to confirm mass spectrometry results.

Conclusion

This study indicates that quantitative proteomic profiling of tumor tissue versus non-cancerous tissue is a promising approach for the identification of potential biomarkers for HCC.     

Quality control and quality assessment of data from surface-enhanced laser desorption/ionization (SELDI) time-of flight (TOF) mass spectrometry (MS)

Background

Proteomic profiling of complex biological mixtures by the ProteinChip technology of surface-enhanced laser desorption/ionization time-of-flight (SELDI-TOF) mass spectrometry (MS) is one of the most promising approaches in toxicological, biological, and clinic research. The reliable identification of protein expression patterns and associated protein biomarkers that differentiate disease from health or that distinguish different stages of a disease depends on developing methods for assessing the quality of SELDI-TOF mass spectra. The use of SELDI data for biomarker identification requires application of rigorous procedures to detect and discard low quality spectra prior to data analysis.

Results

The systematic variability from plates, chips, and spot positions in SELDI experiments was evaluated using biological and technical replicates. Systematic biases on plates, chips, and spots were not found. The reproducibility of SELDI experiments was demonstrated by examining the resulting low coefficient of variances of five peaks presented in all 144 spectra from quality control samples that were loaded randomly on different spots in the chips of six bioprocessor plates. We developed a method to detect and discard low quality spectra prior to proteomic profiling data analysis, which uses a correlation matrix to measure the similarities among SELDI mass spectra obtained from similar biological samples. Application of the correlation matrix to our SELDI data for liver cancer and liver toxicity study and myeloma-associated lytic bone disease study confirmed this approach as an efficient and reliable method for detecting low quality spectra.

Conclusion

This report provides evidence that systematic variability between plates, chips, and spots on which the samples were assayed using SELDI based proteomic procedures did not exist. The reproducibility of experiments in our studies was demonstrated to be acceptable and the profiling data for subsequent data analysis are reliable. Correlation matrix was developed as a quality control tool to detect and discard low quality spectra prior to data analysis. It proved to be a reliable method to measure the similarities among SELDI mass spectra and can be used for quality control to decrease noise in proteomic profiling data prior to data analysis.

     

External jugular vein cutdown approach for chronic indwelling central venous access in cancer patients: A potentially useful alternative

Background

Survival in colorectal cancer may correlate with the degree of systemic inflammatory response to the tumour. Atrial fibrillation may be regarded as an inflammatory complication. We aimed to determine if atrial fibrillation is a prognostic factor in colorectal cancer.

Patients and methods

A prospective colorectal cancer patient database was cross-referenced with the hospital clinical-coding database to identify patients who had underwent colorectal cancer surgery and were in atrial fibrillation pre- or postoperatively.

Results

A total of 175 patients underwent surgery for colorectal cancer over a two-year period. Of these, 13 patients had atrial fibrillation pre- or postoperatively. Atrial fibrillation correlated with worse two-year survival (p = 0.04; log-rank test). However, in a Cox regression analysis, atrial fibrillation was not significantly associated with survival.

Conclusion

The presence or development of atrial fibrillation in patients undergoing surgery for colorectal cancer is associated with worse overall survival, however it was not found to be an independent factor in multivariate analysis.     

Enriched sera protein profiling for detection of non-small cell lung cancer biomarkers

Background

Non Small Cell Lung Cancer (NSCLC) is the major cause of cancer related-death. Many patients receive diagnosis at advanced stage leading to a poor prognosis. At present, no satisfactory screening tests are available in clinical practice and the discovery and validation of new biomarkers is mandatory. Surface Enhanced Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (SELDI-ToF-MS) is a recent high-throughput technique used to detect new tumour markers. In this study we performed SELDI-ToF-MS analysis on serum samples treated with the ProteoMiner? kit, a combinatorial library of hexapeptide ligands coupled to beads, to reduce the wide dynamic range of protein concentration in the sample. Serum from 44 NSCLC patients and 19 healthy controls were analyzed with IMAC30-Cu and H50 ProteinChip Arrays.

Results

Comparing SELDI-ToF-MS protein profiles of NSCLC patients and healthy controls, 28 protein peaks were found significantly different (p < 0.05), and were used as predictors to build decision classification trees. This statistical analysis selected 10 protein peaks in the low-mass range (2-24 kDa) and 6 in the high-mass range (40-80 kDa). The classification models for the low-mass range had a sensitivity and specificity of 70.45% (31/44) and 68.42% (13/19) for IMAC30-Cu, and 72.73% (32/44) and 73.68% (14/19) for H50 ProteinChip Arrays.

Conclusions

These preliminary results suggest that SELDI-ToF-MS protein profiling of serum samples pretreated with ProteoMiner? can improve the discovery of protein peaks differentially expressed between NSCLC patients and healthy subjects, useful to build classification algorithms with high sensitivity and specificity. However, identification of the significantly different protein peaks needs further study in order to provide a better understanding of the biological nature of these potential biomarkers and their role in the underlying disease process.     

Biomarkers of HIV-1 associated dementia: proteomic investigation of sera

Background

Acute lymphoblastic leukemia (ALL) is a common form of cancer in children. Currently, bone marrow biopsy is used for diagnosis. Noninvasive biomarkers for the early diagnosis of pediatric ALL are urgently needed. The aim of this study was to discover potential protein biomarkers for pediatric ALL.

Methods

Ninety-four pediatric ALL patients and 84 controls were randomly divided into a "training" set (45 ALL patients, 34 healthy controls) and a test set (49 ALL patients, 30 healthy controls and 30 pediatric acute myeloid leukemia (AML) patients). Serum proteomic profiles were measured using surface-enhanced laser desorption/ionization-time-of-flight mass spectroscopy (SELDI-TOF-MS). A classification model was established by Biomarker Pattern Software (BPS). Candidate protein biomarkers were purified by HPLC, identified by LC-MS/MS and validated using ProteinChip immunoassays.

Results

A total of 7 protein peaks (9290 m/z, 7769 m/z, 15110 m/z, 7564 m/z, 4469 m/z, 8937 m/z, 8137 m/z) were found with differential expression levels in the sera of pediatric ALL patients and controls using SELDI-TOF-MS and then analyzed by BPS to construct a classification model in the "training" set. The sensitivity and specificity of the model were found to be 91.8%, and 90.0%, respectively, in the test set. Two candidate protein peaks (7769 and 9290 m/z) were found to be down-regulated in ALL patients, where these were identified as platelet factor 4 (PF4) and pro-platelet basic protein precursor (PBP). Two other candidate protein peaks (8137 and 8937 m/z) were found up-regulated in the sera of ALL patients, and these were identified as fragments of the complement component 3a (C3a).

Conclusion

Platelet factor (PF4), connective tissue activating peptide III (CTAP-III) and two fragments of C3a may be potential protein biomarkers of pediatric ALL and used to distinguish pediatric ALL patients from healthy controls and pediatric AML patients. Further studies with additional populations or using pre-diagnostic sera are needed to confirm the importance of these findings as diagnostic markers of pediatric ALL.     

Nitrous oxide may not increase the risk of cancer recurrence after colorectal surgery: a follow-up of a randomized controlled trial

Background

Even the best cancer surgery is usually associated with minimal residual disease. Whether these remaining malignant cells develop into clinical recurrence is at least partially determined by adequacy of host defense, especially natural killer cell function. Anesthetics impair immune defenses to varying degrees, but nitrous oxide appears to be especially problematic. We therefore tested the hypothesis that colorectal-cancer recurrence risk is augmented by nitrous oxide administration during colorectal surgery.

Methods

We conducted a 4- to 8-year follow-up of 204 patients with colorectal cancer who were randomly assigned to 65% nitrous oxide (n = 97) or nitrogen (n = 107), balanced with isoflurane and remifentanil. The primary outcome was the time to cancer recurrence. Our primary analysis was a multivariable Cox-proportional-hazards regression model that included relevant baseline variables. In addition to treatment group, the model considered patient age, tumor grade, dissemination, adjacent organ invasion, vessel invasion, and the number of nodes involved. The study had 80% power to detect a 56% or greater reduction in recurrence rates (i.e., hazard ratio of 0.44 or less) at the 0.05 significance level.

Results

After adjusting for significant baseline covariables, risk of recurrence did not differ significantly for nitrous oxide and nitrogen, with a hazard ratio estimate (95% CI) of 1.10 (0.66, 1.83), P = 0.72. No two-way interactions with the treatment were statistically significant.

Conclusion

Colorectal-cancer recurrence risks were not greatly different in patients who were randomly assigned to 65% nitrous oxide or nitrogen during surgery. Our results may not support avoiding nitrous oxide use to prevent recurrence of colorectal cancer.

Implications Statement

The risk of colorectal cancer recurrence was similar in patients who were randomly assigned to 65% nitrous oxide or nitrogen during colorectal surgery.

Trial Registration

Current Controlled Clinical Trials NCT00781352 http://www.clinicaltrials.gov     

Colovaginal anastomosis: an unusual complication of stapler use in restorative procedure after Hartmann operation

Background

To support decisions about surgical treatment of elderly patients with cancer, population-based estimates of postoperative mortality (POM) rates are required.

Methods

Electronic records from the Rotterdam Cancer Registry were retrieved for octogenarians and nonagenarians who underwent resection in the period 1987–2000. POM was defined as death within 30 days of resection and both elective and emergency operations were included.

Results

In a series of 5.390 operated patients aged 80 years and older, POM rates were 0.5% for breast cancer, 1.7% for endometrial cancer and 4.2% for renal cancer. For patients with colorectal cancer, POM increased from 8% for the age group 80–84 to 13% for those 85–89 to 20% in nonagenarians. For stomach cancer, the respective figures were 11%, 20% and 44%.

Conclusion

These results show that resections can be performed at acceptable risk in selected elderly patients with cancer.     

A multiplex platform for the identification of ovarian cancer biomarkers

Background

Currently, there are no FDA approved screening tools for detecting early stage ovarian cancer in the general population. Development of a biomarker-based assay for early detection would significantly improve the survival of ovarian cancer patients.

Methods

We used a multiplex approach to identify protein biomarkers for detecting early stage ovarian cancer. This new technology (Proseek^® Multiplex Oncology Plates) can simultaneously measure the expression of 92 proteins in serum based on a proximity extension assay. We analyzed serum samples from 81 women representing healthy, benign pathology, early, and advanced stage serous ovarian cancer patients.

Results

Principle component analysis and unsupervised hierarchical clustering separated patients into cancer versus non-cancer subgroups. Data from the Proseek^® plate for CA125 levels exhibited a strong correlation with current clinical assays for CA125 (correlation coefficient of 0.89, 95% CI 0.83, 0.93). CA125 and HE4 were present at very low levels in healthy controls and benign cases, while higher levels were found in early stage cases, with highest levels found in the advanced stage cases. Overall, significant trends were observed for 38 of the 92 proteins (p < 0.001), many of which are novel candidate serum biomarkers for ovarian cancer. The area under the ROC curve (AUC) for CA125 was 0.98 and the AUC for HE4 was 0.85 when comparing early stage ovarian cancer versus healthy controls. In total, 23 proteins had an estimated AUC of 0.7 or greater. Using a naïve Bayes classifier that combined 12 proteins, we improved the sensitivity corresponding to 95% specificity from 93 to 95% when compared to CA125 alone. Although small, a 2% increase would have a significant effect on the number of women correctly identified when screening a large population.

Conclusions

These data demonstrate that the Proseek^® technology can replicate the results established by conventional clinical assays for known biomarkers, identify new candidate biomarkers, and improve the sensitivity and specificity of CA125 alone. Additional studies using a larger cohort of patients will allow for validation of these biomarkers and lead to the development of a screening tool for detecting early stage ovarian cancer in the general population.

     

Constitutive basal and stimulated human small bowel contractility is enhanced in obesity

Background

Between 5 and 10% of the patients undergoing a colonoscopy cannot have a complete procedure mainly due to stenosing neoplastic lesion of rectum or distal colon. Nevertheless the elective surgical treatment concerning the stenosis is to be performed after the pre-operative assessment of the colonic segments upstream the cancer. The aim of this study is to illustrate our experience with the Computed Tomographic Colonography (CTC) for the pre-operative assessment of the entire colon in the patients with stenosing colorectal cancers.

Methods

From January 2005 till March 2009, we observed and treated surgically 43 patients with stenosing colorectal neoplastic lesions. All patients did not tolerate the pre-operative colonoscopy. For this reason they underwent a pre-operative CTC in order to have a complete assessment of the entire colon. All patients underwent a follow-up colonoscopy 3 months after the surgical treatment. The CTC results were compared with both macroscopic examination of the specimen and the follow-up coloscopy.

Results

The pre-operative CTC showed four synchronous lesions in four patients (9.3% of the cases). The macroscopic examination of the specimen revealed three small sessile polyps (3 - 4 mm in diameter) missed in the pre-operative assessment near the stenosing colorectal cancer. The follow-up colonoscopy showed four additional sessile polyps with a diameter between 3 - 11 mm in three patients. Our experience shows that CTC has a sensitivity of 83,7%.

Conclusion

In patients with stenosing colonic lesions, CTC allows to assess the entire colon pre-operatively avoiding the need of an intraoperative colonoscopy. More synchronous lesions are detected and treated at the time of the elective surgery for the stenosing cancer avoiding further surgery later on.     

Reconsidering low-dose aspirin therapy for cardiovascular disease: a study protocol for physician and patient behavioral change

Background

Opinion leaders represent one way to disseminate new knowledge and influence the practice behaviors of physicians. This study explored the stability of opinion leaders over time, whether opinion leaders were polymorphic (i.e., influencing multiple practice areas) or monomorphic (i.e., influencing one practice area), and reach of opinion leaders in their local network.

Methods

We surveyed surgeons and pathologists in Ontario to identify opinion leaders for colorectal cancer in 2003 and 2005 and to identify opinion leaders for breast cancer in 2005. We explored whether opinion leaders for colorectal cancer identified in 2003 were re-identified in 2005. We examined whether opinion leaders were considered polymorphic (nominated in 2005 as opinion leaders for both colorectal and breast cancer) or monomorphic (nominated in 2005 for only one condition). Social-network mapping was used to identify the number of local colleagues identifying opinion leaders.

Results

Response rates for surgeons were 41% (2003) and 40% (2005); response rates for pathologists were 42% (2003) and 37% (2005). Four (25%) of the surgical opinion leaders identified in 2003 for colorectal cancer were re-identified in 2005. No pathology opinion leaders for colorectal cancer were identified in both 2003 and 2005. Only 29% of surgical opinion leaders and 17% of pathology opinion leaders identified in the 2005 survey were considered influential for both colorectal cancer and breast cancer. Social-network mapping revealed that only a limited number of general surgeons (12%) or pathologists (7%) were connected to the social networks of identified opinion leaders.

Conclusions

Opinion leaders identified in this study were not stable over a two-year time period and generally appear to be monomorphic, with clearly demarcated areas of expertise and limited spheres of influence. These findings may limit the practicability of routinely using opinion leaders to influence practice.     

Confounding Effects of Benign Lung Diseases on Non-Small Cell Lung Cancer Serum Biomarker Discovery

Introduction

Lung cancer is the leading cause of cancer-related death worldwide. The discovery of new biomarkers could aid early diagnosis and monitoring of recurrence following tumor resection.

Methods

We have prospectively collected serum from 97 lung cancer patients undergoing surgery with curative intent and compared their serum proteomes with those of 100 noncancer controls (59 disease-free and 41 with a range of nonmalignant lung conditions). We initially analyzed serum from 67 lung cancer patients and 73 noncancer control subjects by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry using immobilized metal affinity capture ProteinChip arrays and subsequently validated our findings with an independent analysis of 30 lung cancer patients and 27 noncancer subjects.

Results

The data from both experiments show many significant differences between the serum proteomes of lung cancer patients and nondiseased control subjects, and a number of these polypeptides have been identified. However, the profiles of patients with benign lung diseases resembled those of lung cancer patients such that very few significant differences were found when these cohorts were compared.

Conclusions

This report provides clear evidence of the need to account for the confounding effects of benign diseases when designing lung cancer serum biomarker discovery projects.     

Quantitative proteomic analysis by iTRAQ<Superscript>®</Superscript> for the identification of candidate biomarkers in ovarian cancer serum

Background

Ovarian cancer is the most lethal gynecologic malignancy, with the majority of cases diagnosed at an advanced stage when treatments are less successful. Novel serum protein markers are needed to detect ovarian cancer in its earliest stage; when detected early, survival rates are over 90%. The identification of new serum biomarkers is hindered by the presence of a small number of highly abundant proteins that comprise approximately 95% of serum total protein. In this study, we used pooled serum depleted of the most highly abundant proteins to reduce the dynamic range of proteins, and thereby enhance the identification of serum biomarkers using the quantitative proteomic method iTRAQ^®.

Results

Medium and low abundance proteins from 6 serum pools of 10 patients each from women with serous ovarian carcinoma, and 6 non-cancer control pools were labeled with isobaric tags using iTRAQ^® to determine the relative abundance of serum proteins identified by MS. A total of 220 unique proteins were identified and fourteen proteins were elevated in ovarian cancer compared to control serum pools, including several novel candidate ovarian cancer biomarkers: extracellular matrix protein-1, leucine-rich alpha-2 glycoprotein-1, lipopolysaccharide binding protein-1, and proteoglycan-4. Western immunoblotting validated the relative increases in serum protein levels for several of the proteins identified.

Conclusions

This study provides the first analysis of immunodepleted serum in combination with iTRAQ^® to measure relative protein expression in ovarian cancer patients for the pursuit of serum biomarkers. Several candidate biomarkers were identified which warrant further development.

     

Identification of Early Intestinal Neoplasia Protein Biomarkers Using Laser Capture Microdissection and MALDI MS

Obtaining protein profiles from a homogeneous cell population in tissues can significantly improve our capability in protein biomarker discovery. In this study, unique protein profiles from the top and bottom sections of mouse crypts and Apc^Min+/− adenomas were obtained using laser capture microdissection (LCM) combined with MALDI MS. Statistically significant protein peaks with differential expression were selected, and a set of novel protein biomarkers were identified. Immunohistochemistry was performed to confirm the differentially expressed protein biomarkers found by LCM combined with MALDI MS. To validate the relevance of the findings in human colorectal cancer (CRC), S100A8 was further confirmed in human CRC using immunohistochemistry. In addition, S100A8 was found to have an increased expression at different human CRC stages (Duke''s A–D) compared with controls at both protein (n = 168 cases) and RNA (n = 215 cases) levels. Overall LCM combined with MALDI MS is a promising method to identify intestinal protein biomarkers from minute amounts of tissue. The novel protein biomarkers identified from the top and bottom crypts will increase our knowledge of the specific protein changes taking place during cell migration from the crypt bottom to top. In addition, the identified cancer protein biomarkers will aid in the exploration of colorectal tumorigenesis mechanisms as well as in the advancement of molecularly based diagnosis of colorectal cancer.Obtaining protein profiles from a pure cell population can significantly improve our capability in protein biomarker discovery. Laser capture microdissection (LCM)¹ is an important tool for acquisition of specific cells of interest from heterogeneous tissue sections (1,2). Obtaining protein profiles from laser-microdissected cells has been explored using multidimensional liquid chromatography-mass spectrometry or gel electrophoresis (3–5). However, usually ∼30,000 cells are required for these approaches (6–8). Direct analysis of laser-microdissected cells using MALDI MS is a sensitive, accurate, and high throughput technique to obtain protein profiles from limited numbers of cells (∼100 cells) (9–11). Combining LCM and MALDI MS is a promising tool for discovery of protein biomarkers from minute tissue structures.Individuals with advanced colorectal cancer (CRC) continue to have a poor prognosis despite recent improvements in treatment. As a result, early diagnosis and improved understanding of the pathogenesis of CRC are vital for improved clinical outcomes. Identification of early CRC protein markers has the potential to result in earlier diagnosis, more accurate prognosis, and improved treatment for individuals with CRC. Colorectal neoplasia is believed to arise from the colonic crypt (12,13). The colon is a self-renewing epithelium that consists of an actively dividing, relatively undifferentiated crypt base and a non-dividing, differentiated surface compartment (14–16). Small numbers of stem cells reside in the base of the crypt where daughter cells differentiate into absorptive cells, goblet cells, tuft cells, and endocrine cells. Absorptive cells and goblet cells, in particular, migrate to the luminal surface where they are eventually shed into the colonic lumen. Because the top and bottom crypts contain cells at different stages of differentiation, the protein profiles of these cells are likely to be different.Adenomatous polyps originate from crypts and represent important precursor lesions in the adenoma-carcinoma sequence (12). The Apc^Min+/− mouse model, and other similar models based on truncating mutations in Apc, have been widely used to study intestinal neoplasia. This model contains a germ line mutation in the adenomatous polyposis coli (Apc) gene (17), leading to numerous adenomas in the intestine (18). In humans, loss of APC is often an early event in the cascade of genetic mutations that lead to colorectal neoplasia (19). Approximately 80% of all sporadic human CRC have inactivating mutations in the APC gene (20). Similarly human familial adenomatous polyposis patients have germ line mutations in the APC gene, and loss of heterozygosity leads to the development of hundreds of adenomatous polyps in the colon and rectum. Thus, the Apc^Min+/− mouse is a promising mouse model for the discovery of potential early CRC markers using a proteomics approach.As a discovery study, we aimed to identify a set of proteomic profiles that can differentiate murine top and bottom crypts and Apc^Min+/− adenomas using LCM combined with MALDI MS. Specific protein biomarkers were identified using tandem mass spectrometry. Finally the relevance of the identified protein biomarkers was examined in human CRC at both protein and mRNA levels.