Proteins associated with pancreatic cancer survival in patients with resectable pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal disease with a dismal prognosis. However, while most patients die within the first year of diagnosis, very rarely, a few patients can survive for >10 years. Better understanding the molecular characteristics of the pancreatic adenocarcinomas from these very-long-term survivors (VLTS) may provide clues for personalized medicine and improve current pancreatic cancer treatment. To extend our previous investigation, we examined the proteomes of individual pancreas tumor tissues from a group of VLTS patients (survival ≥10 years) and short-term survival patients (STS, survival <14 months). With a given analytical sensitivity, the protein profile of each pancreatic tumor tissue was compared to reveal the proteome alterations that may be associated with pancreatic cancer survival. Pathway analysis of the differential proteins identified suggested that MYC, IGF1R and p53 were the top three upstream regulators for the STS-associated proteins, and VEGFA, APOE and TGFß-1 were the top three upstream regulators for the VLTS-associated proteins. Immunohistochemistry analysis using an independent cohort of 145 PDAC confirmed that the higher abundance of ribosomal protein S8 (RPS8) and prolargin (PRELP) were correlated with STS and VLTS, respectively. Multivariate Cox analysis indicated that 'High-RPS8 and Low-PRELP' was significantly associated with shorter survival time (HR=2.69, 95% CI 1.46-4.92, P=0.001). In addition, galectin-1, a previously identified protein with its abundance aversely associated with pancreatic cancer survival, was further evaluated for its significance in cancer-associated fibroblasts. Knockdown of galectin-1 in pancreatic cancer-associated fibroblasts dramatically reduced cell migration and invasion. The results from our study suggested that PRELP, LGALS1 and RPS8 might be significant prognostic factors, and RPS8 and LGALS1 could be potential therapeutic targets to improve pancreatic cancer survival if further validated.

Quantitative glycoproteomics analysis reveals changes in N-glycosylation level associated with pancreatic ductal adenocarcinoma.

Glycosylation plays an important role in epithelial cancers, including pancreatic ductal adenocarcinoma. However, little is known about the glycoproteome of the human pancreas or its alterations associated with pancreatic tumorigenesis. Using quantitative glycoproteomics approach, we investigated protein N-glycosylation in pancreatic tumor tissue in comparison with normal pancreas and chronic pancreatitis tissue. The study lead to the discovery of a roster of glycoproteins with aberrant N-glycosylation level associated with pancreatic cancer, including mucin-5AC (MUC5AC), carcinoembryonic antigen-related cell adhesion molecule 5 (CEACAM5), insulin-like growth factor binding protein (IGFBP3), and galectin-3-binding protein (LGALS3BP). Pathway analysis of cancer-associated aberrant glycoproteins revealed an emerging phenomenon that increased activity of N-glycosylation was implicated in several pancreatic cancer pathways, including TGF-ß, TNF, NF-kappa-B, and TFEB-related lysosomal changes. In addition, the study provided evidence that specific N-glycosylation sites within certain individual proteins can have significantly altered glycosylation occupancy in pancreatic cancer, reflecting the complexity of the molecular mechanisms underlying cancer-associated glycosylation events.

Chromosomal instability in ulcerative colitis is related to telomere shortening.

Ulcerative colitis, a chronic inflammatory disease of the colon, is associated with a high risk of colorectal carcinoma that is thought to develop through genomic instability. We considered that the rapid cell turnover and oxidative injury observed in ulcerative colitis might accelerate telomere shortening, thereby increasing the potential of chromosomal ends to fuse, resulting in cycles of chromatin bridge breakage and fusion and chromosomal instability associated with tumor cell progression. Here we have used quantitative fluorescence in situ hybridization to compare chromosomal aberrations and telomere shortening in non-dysplastic mucosa taken from individuals affected by ulcerative colitis, either with (UC progressors) or without (UC non-progressors) dysplasia or cancer. Losses, but not gains, of chromosomal arms and centromeres are highly correlated with telomere shortening. Chromosomal losses are greater and telomeres are shorter in biopsy samples from UC progressors than in those from UC non-progressors or control individuals without ulcerative colitis. A mechanistic link between telomere shortening and chromosomal instability is supported by a higher frequency of anaphase bridges--an intermediate in the breakage and fusion of chromatin bridges--in UC progressors than in UC non-progressors or control individuals. Our study shows that telomere length is correlated with chromosomal instability in a precursor of human cancer.

Clonal expansions in ulcerative colitis identify patients with neoplasia.

Chronic inflammation predisposes to a variety of human cancers. Affected tissues slowly accumulate mutations, some of which affect growth regulation and drive successive waves of clonal evolution, whereas a far greater number are functionally neutral and serve only to passively mark expanding clones. Ulcerative colitis (UC) is an inflammatory bowel disease, in which up to 10% of patients eventually develop colon cancer. Here we have mapped mutations in hypermutable intergenic and intronic polyguanine tracts in patients with UC to delineate the extent of clonal expansions associated with carcinogenesis. We genotyped colon biopsies for length altering mutations at 28 different polyguanine markers. In eight patients without neoplasia, we detected only two mutations in a single individual from among 37 total biopsies. In contrast, for 11 UC patients with neoplasia elsewhere in the colon, we identified 63 mutations in 51 nondysplastic biopsies, and every patient possessed at least one mutant clone. A subset of clones were large and extended over many square centimeters of colon. Of these, some occurred as isolated populations in nondysplastic tissue, considerably distant from neoplastic lesions. Other large clones included regions of cancer, suggesting that the tumor arose within a preexisting clonal field. Our results demonstrate that neutral mutations in polyguanine tracts serve as a unique tool for identifying fields of clonal expansions, which may prove clinically useful for distinguishing a subset of UC patients who are at risk for developing cancer.

Investigating neoplastic progression of ulcerative colitis with label-free comparative proteomics.

Patients with extensive ulcerative colitis (UC) have an increased risk of colorectal cancer. Although UC patients generally undergo lifelong colonoscopic surveillance to detect dysplasia or cancer in the colon, detection of cancer in this manner is expensive and invasive. An objective biomarker of dysplasia would vastly improve the clinical management of cancer risk in UC patients. In the current study, accurate mass and time methods with ion intensity-based label-free proteomics are applied to profile individual rectal and colon samples from UC patients with dysplasia or cancer (UC progressors) compared to rectal samples from patients that are dysplasia/cancer free (UC nonprogressors) to identify a set of proteins in the rectum mucosa that differentiate the two groups. In addition to the identification of proteins in UC dysplastic colon tissue, we for the first time identified differentially expressed proteins in nondysplastic rectal tissue from UC progressors. This provides a candidate pool of biomarkers for dysplasia/cancer that could be detected in a random nondysplastic rectal biopsy. Mitochondrial proteins, cytoskeletal proteins, RAS superfamily, proteins relating to apoptosis and metabolism were important protein clusters differentially expressed in the nondysplastic and dysplastic tissues of UC progressors, suggesting their importance in the early stages of UC neoplastic progression. Among the differentially expressed proteins, immunohistochemistry analysis confirmed that TRAP1 displayed increased IHC staining in UC progressors, in both dysplastic and nondysplastic tissue, and CPS1 showed a statistically significant difference in IHC staining between the nonprogressor and progressor groups. Furthermore, rectal CPS1 staining could be used to predict dysplasia or cancer in the colon with 87% sensitivity and 45% specificity, demonstrating the feasibility of using surrogate biomarkers in rectal biopsies to predict dysplasia and/or cancer in the colon.

Protein alterations associated with pancreatic cancer and chronic pancreatitis found in human plasma using global quantitative proteomics profiling.

Pancreatic cancer is a lethal disease that is difficult to diagnose at early stages when curable treatments are effective. Biomarkers that can improve current pancreatic cancer detection would have great value in improving patient management and survival rate. A large scale quantitative proteomics study was performed to search for the plasma protein alterations associated with pancreatic cancer. The enormous complexity of the plasma proteome and the vast dynamic range of protein concentration therein present major challenges for quantitative global profiling of plasma. To address these challenges, multidimensional fractionation at both protein and peptide levels was applied to enhance the depth of proteomics analysis. Employing stringent criteria, more than 1300 proteins total were identified in plasma across 8-orders of magnitude in protein concentration. Differential proteins associated with pancreatic cancer were identified, and their relationship with the proteome of pancreatic tissue and pancreatic juice from our previous studies was discussed. A subgroup of differentially expressed proteins was selected for biomarker testing using an independent cohort of plasma and serum samples from well-diagnosed patients with pancreatic cancer, chronic pancreatitis, and nonpancreatic disease controls. Using ELISA methodology, the performance of each of these protein candidates was benchmarked against CA19-9, the current gold standard for a pancreatic cancer blood test. A composite marker of TIMP1 and ICAM1 demonstrate significantly better performance than CA19-9 in distinguishing pancreatic cancer from the nonpancreatic disease controls and chronic pancreatitis controls. In addition, protein AZGP1 was identified as a biomarker candidate for chronic pancreatitis. The discovery and technical challenges associated with plasma-based quantitative proteomics are discussed and may benefit the development of plasma proteomics technology in general. The protein candidates identified in this study provide a biomarker candidate pool for future investigations.

Array-based comparative genomic hybridization in ulcerative colitis neoplasia: single non-dysplastic biopsies distinguish progressors from non-progressors.

Approximately 10% of ulcerative colitis patients develop colorectal neoplasia. At present, identification of this subset is markedly limited and necessitates lifelong colonoscopic surveillance for the entire ulcerative colitis population. Better risk markers are needed to focus surveillance onto the patients who are most likely to benefit. Using array-based comparative genomic hybridization, we analyzed single, non-dysplastic biopsies from three patient groups: ulcerative colitis progressors (n=9) with cancer or high-grade dysplasia at a mean distance of 18 cm from the analyzed site; ulcerative colitis non-progressors (n=8) without dysplasia during long-term surveillance; and non-ulcerative colitis normal controls (n=2). Genomic DNA from fresh colonic epithelium purified from stroma was hybridized to 287 (low-density) and 4342 (higher-density) feature bacterial artificial chromosome arrays. Sample-to-reference fluorescence ratios were calculated for individual chromosomal targets and globally across the genome. The low-density arrays yielded pronounced genomic gains and losses in 3 of 9 (33%) ulcerative colitis progressors but in none of the 10 control patients. Identical DNA samples analyzed on the higher-density arrays, using a combination of global and individual high variance assessments, distinguished all nine progressors from all 10 controls. These data confirm that genomic alterations in ulcerative colitis progressors are widespread, even involving single non-dysplastic biopsies that are far distant from neoplasia. They therefore show promise toward eliminating full colonoscopic surveillance with extensive biopsy sampling in the majority of ulcerative colitis patients.

Genomic biomarkers to improve ulcerative colitis neoplasia surveillance.

No adequate means exist to identify the minority of ulcerative colitis (UC) patients destined to undergo neoplastic progression. Recognition of this subset would advance UC cancer surveillance by focusing the available management options onto the highest risk patients. Three different assays of genomic alterations in nondysplastic UC biopsies show promise for distinguishing patients with neoplasia (UC progressors) from those without (UC nonprogressors), including assays of telomere length, anaphase bridges, and chromosomal fluorescence in situ hybridization. Expanding the number of patients and testing of assays simultaneously in the same biopsy further validated their utility. A panel approach also improved testing outcome. A total of 14 UC progressors was readily separable from 15 UC nonprogressors and 6 normal controls. Chromosomal entropy (ie, the extent of alteration diversity) proved to be the most useful test. By receiver-operating characteristic analysis, mean chromosomal entropy in 28 patients over all four chromosomes yielded 100% sensitivity and 92% specificity for distinguishing progressors from nonprogressors with optimum choice of threshold. Moreover, separation was achieved using only nondysplastic and predominantly rectal (82.8%) biopsies that were remote from neoplasia, suggesting that full colonoscopy with extensive biopsies might be avoided for the majority of UC patients, the nonprogressors. These data further strengthen the concept that genomic biomarkers can distinguish UC progressors from nonprogressors and improve cancer surveillance in UC.

Quantitative proteomics investigation of pancreatic intraepithelial neoplasia.

Patients with pancreatic cancer are usually diagnosed at late stages, when the disease is incurable. Pancreatic intraepithelial neoplasia (PanIN) 3 is believed to be the immediate precursor lesion of pancreatic adenocarcinoma, and would be an ideal stage to diagnose patients, when intervention and cure are possible and patients are curable. In this study, we used quantitative proteomics to identify dysregulated proteins in PanIN 3 lesions. Altogether, over 200 dysregulated proteins were identified in the PanIN 3 tissues, with a minimum of a 1.75-fold change compared with the proteins in normal pancreas. These dysregulated PanIN 3 proteins play roles in cell motility, the inflammatory response, the blood clotting cascade, the cell cycle and its regulation, and protein degradation. Further network analysis of the proteins identified c-MYC as an important regulatory protein in PanIN 3 lesions. Finally, three of the overexpressed proteins, laminin beta-1, galectin-1, and actinin-4 were validated by immunohistochemistry analysis. All three of these proteins were overexpressed in the stroma or ductal epithelial cells of advanced PanIN lesions as well as in pancreatic cancer tissue. Our findings suggest that these three proteins may be useful as biomarkers for advanced PanIN and pancreatic cancer if further validated. The dysregulated proteins identified in this study may assist in the selection of candidates for future development of biomarkers for detecting early and curable pancreatic neoplasia.

Identifying Abundant Immunotherapy and Other Targets in Solid Tumors: Integrating RNA-seq and Mass Spectrometry Proteomics Data Sets.

RNA-seq and mass-spectrometry proteomics combined with growing data repositories have greatly increased the capacity to identify candidate proteins or protein sequence variants that share properties of ideal therapy targets, which include being abundant in cancer cells, absent or rare in adult organs (especially vital organs), and shared by many patient tumors. RNA-seq and fixed content arrays can identify genes that are overexpressed or misexpressed in cancer. RNA-seq is uniquely suited to identifying cancer-specific sequence variants. We review factors relevant for determining whether products of genes that are abundant or differentially abundant in RNA-seq are concordant or discordant with proteins that are identified as abundant or differentially abundant in mass-spectrometry proteomics assays.

Tumor-Infiltrating Merkel Cell Polyomavirus-Specific T Cells Are Diverse and Associated with Improved Patient Survival.

Tumor-infiltrating CD8+ T cells are associated with improved survival of patients with Merkel cell carcinoma (MCC), an aggressive skin cancer causally linked to Merkel cell polyomavirus (MCPyV). However, CD8+ T-cell infiltration is robust in only 4% to 18% of MCC tumors. We characterized the T-cell receptor (TCR) repertoire restricted to one prominent epitope of MCPyV (KLLEIAPNC, "KLL") and assessed whether TCR diversity, tumor infiltration, or T-cell avidity correlated with clinical outcome. HLA-A*02:01/KLL tetramer+ CD8+ T cells from MCC patient peripheral blood mononuclear cells (PBMC) and tumor-infiltrating lymphocytes (TIL) were isolated via flow cytometry. TCRß (TRB) sequencing was performed on tetramer+ cells from PBMCs or TILs (n = 14) and matched tumors (n = 12). Functional avidity of T-cell clones was determined by IFNγ production. We identified KLL tetramer+ T cells in 14% of PBMC and 21% of TIL from MCC patients. TRB repertoires were strikingly diverse (397 unique TRBs were identified from 12 patients) and mostly private (only one TCRb clonotype shared between two patients). An increased fraction of KLL-specific TIL (>1.9%) was associated with significantly increased MCC-specific survival P = 0.0009). T-cell cloning from four patients identified 42 distinct KLL-specific TCRa/b pairs. T-cell clones from patients with improved MCC-specific outcomes were more avid (P < 0.05) and recognized an HLA-appropriate MCC cell line. T cells specific for a single MCPyV epitope display marked TCR diversity within and between patients. Intratumoral infiltration by MCPyV-specific T cells was associated with significantly improved MCC-specific survival, suggesting that augmenting the number or avidity of virus-specific T cells may have therapeutic benefit. Cancer Immunol Res; 5(2); 137-47. ©2017 AACR.

Palladin mutation causes familial pancreatic cancer and suggests a new cancer mechanism.

BACKGROUND: Pancreatic cancer is a deadly disease. Discovery of the mutated genes that cause the inherited form(s) of the disease may shed light on the mechanism(s) of oncogenesis. Previously we isolated a susceptibility locus for familial pancreatic cancer to chromosome location 4q32-34. In this study, our goal was to discover the identity of the familial pancreatic cancer gene on 4q32 and determine the function of that gene. METHODS AND FINDINGS: A customized microarray of the candidate chromosomal region affecting pancreatic cancer susceptibility revealed the greatest expression change in palladin (PALLD), a gene that encodes a component of the cytoskeleton that controls cell shape and motility. A mutation causing a proline (hydrophobic) to serine (hydrophilic) amino acid change (P239S) in a highly conserved region tracked with all affected family members and was absent in the non-affected members. The mutational change is not a known single nucleotide polymorphism. Palladin RNA, measured by quantitative RT-PCR, was overexpressed in the tissues from precancerous dysplasia and pancreatic adenocarcinoma in both familial and sporadic disease. Transfection of wild-type and P239S mutant palladin gene constructs into HeLa cells revealed a clear phenotypic effect: cells expressing P239S palladin exhibited cytoskeletal changes, abnormal actin bundle assembly, and an increased ability to migrate. CONCLUSIONS: These observations suggest that the presence of an abnormal palladin gene in familial pancreatic cancer and the overexpression of palladin protein in sporadic pancreatic cancer cause cytoskeletal changes in pancreatic cancer and may be responsible for or contribute to the tumor's strong invasive and migratory abilities.

Characterization of genomic instability in ulcerative colitis neoplasia leads to discovery of putative tumor suppressor regions.

Ulcerative colitis (UC) is an inflammatory disease of the colon that is associated with increased risk of colorectal cancer associated with genomic instability. We have previously demonstrated that genomic instability is present in UC patients with colonic neoplasia, and hypothesized that the chromosomal alterations may be taking place in regions that are susceptible to mutation or that provide a growth advantage to a cell undergoing neoplastic transformation. In this study, we used two polymerase chain reaction (PCR)-based DNA fingerprinting techniques (arbitrarily primed PCR and inter-simple-sequence-repeat PCR) to study the process of genomic instability. The two techniques of DNA fingerprinting cross-validate the instability observed in these studies. We analyzed the molecular basis of 10 commonly altered DNA bands obtained from DNA fingerprints of biopsies from various histologic grades of UC patients with dysplasia or cancer (UC Progressors). We determined that the band changes in the fingerprint truly represent changes in DNA sequence, and that the fingerprinting provides highly reproducible results. Furthermore, our investigation revealed that 40% of alterations involve repetitive sequences. Two frequently deleted sequences in 6q27 and 2q14 were studied further because they were frequently abnormal in the dysplastic and nondysplastic tissue of UC Progressors. The losses from 6q27 and 2q14 were confirmed by loss of heterozygosity and real-time PCR analysis. Both of these regions in chromosomes 6 and 2 are surrounded by highly repetitive and mobile LINE-1 elements, possibly making the region susceptible to mutational change. These regions were affected (lost) in UC Progressors but not in UC patients who were neoplasia free. Loss of heterozygosity at 6q27 has been described in ovarian and other cancers, while the 2q14 region has been implicated in prostate and sporadic colon cancers. Both regions are likely to contain tumor-suppressor genes. In conclusion, the genomic instability in UC Progressors can occur in regions that are susceptible to change and are locations of putative tumor-suppressor genes.

Subacute calorie restriction and rapamycin discordantly alter mouse liver proteome homeostasis and reverse aging effects.

Calorie restriction (CR) and rapamycin (RP) extend lifespan and improve health across model organisms. Both treatments inhibit mammalian target of rapamycin (mTOR) signaling, a conserved longevity pathway and a key regulator of protein homeostasis, yet their effects on proteome homeostasis are relatively unknown. To comprehensively study the effects of aging, CR, and RP on protein homeostasis, we performed the first simultaneous measurement of mRNA translation, protein turnover, and abundance in livers of young (3 month) and old (25 month) mice subjected to 10-week RP or 40% CR. Protein abundance and turnover were measured in vivo using (2) H3 -leucine heavy isotope labeling followed by LC-MS/MS, and translation was assessed by polysome profiling. We observed 35-60% increased protein half-lives after CR and 15% increased half-lives after RP compared to age-matched controls. Surprisingly, the effects of RP and CR on protein turnover and abundance differed greatly between canonical pathways, with opposite effects in mitochondrial (mt) dysfunction and eIF2 signaling pathways. CR most closely recapitulated the young phenotype in the top pathways. Polysome profiles indicated that CR reduced polysome loading while RP increased polysome loading in young and old mice, suggesting distinct mechanisms of reduced protein synthesis. CR and RP both attenuated protein oxidative damage. Our findings collectively suggest that CR and RP extend lifespan in part through the reduction of protein synthetic burden and damage and a concomitant increase in protein quality. However, these results challenge the notion that RP is a faithful CR mimetic and highlight mechanistic differences between the two interventions.

Up-regulation of mitochondrial chaperone TRAP1 in ulcerative colitis associated colorectal cancer.

AIM: To characterize tumor necrosis factor receptor-associated protein 1 (TRAP1) expression in the progression of ulcerative colitis (UC)-associated colorectal cancer. METHODS: Chronic UC is an inflammatory bowel disease that predisposes to colorectal cancer. Immunohistochemical analysis was used to evaluate TRAP1 expression on tissue microarrays containing colonic tissues from 42 UC progressors (patients with cancer or dysplasia) and 38 non-progressors (dysplasia/cancer free patients). Statistical analyses of the TRAP1 immunohistochemistry staining were performed using GraphPad Prism. Differences in the TRAP1 level between non-progressors and progressors were tested for statistical significance using the Mann-Whitney test. Receiver operating characteristic curve method was used to quantify marker performance in distinguishing diseased cases from controls. RESULTS: TRAP1 was up-regulated in the colon tissues from UC progressors, but not in the colon tissues from UC non-progressors. Moreover, up-regulation of TRAP1 preceded the neoplastic changes: it was present in both the dysplastic and non-dysplastic tissues of UC progressors. When TRAP1 staining in rectal tissue was used as a diagnostic marker, it could distinguish progressors from non-progressors with 59% sensitivity and 80% specificity. Our study further showed that the increase of TRAP1 expression positively correlated with the degree of inflammation in the colorectal cancer tissues, which could be related to the increased oxidation present in the colonic mucosa from UC progressors. We then investigated the cellular proteome changes underlying oxidative stress, and found that oxidative stress could induce up-regulation of TRAP1 along with several other negative modulators of apoptosis. CONCLUSION: These results suggest that oxidative stress in long standing UC could lead to the increase of cytoprotective protein TRAP1, which in turn could promote cancer progression by preventing or protecting the oxidative damaged epithelial cells from undergoing apoptosis. TRAP1 could be a potential diagnostic marker for UC associated colorectal cancer.

Altered proteome turnover and remodeling by short-term caloric restriction or rapamycin rejuvenate the aging heart.

Chronic caloric restriction (CR) and rapamycin inhibit the mechanistic target of rapamycin (mTOR) signaling, thereby regulating metabolism and suppressing protein synthesis. Caloric restriction or rapamycin extends murine lifespan and ameliorates many aging-associated disorders; however, the beneficial effects of shorter treatment on cardiac aging are not as well understood. Using a recently developed deuterated-leucine labeling method, we investigated the effect of short-term (10 weeks) CR or rapamycin on the proteomics turnover and remodeling of the aging mouse heart. Functionally, we observed that short-term CR and rapamycin both reversed the pre-existing age-dependent cardiac hypertrophy and diastolic dysfunction. There was no significant change in the cardiac global proteome (823 proteins) turnover with age, with a median half-life 9.1 days in the 5-month-old hearts and 8.8 days in the 27-month-old hearts. However, proteome half-lives of old hearts significantly increased after short-term CR (30%) or rapamycin (12%). This was accompanied by attenuation of age-dependent protein oxidative damage and ubiquitination. Quantitative proteomics and pathway analysis revealed an age-dependent decreased abundance of proteins involved in mitochondrial function, electron transport chain, citric acid cycle, and fatty acid metabolism as well as increased abundance of proteins involved in glycolysis and oxidative stress response. This age-dependent cardiac proteome remodeling was significantly reversed by short-term CR or rapamycin, demonstrating a concordance with the beneficial effect on cardiac physiology. The metabolic shift induced by rapamycin was confirmed by metabolomic analysis.

Global proteomics and pathway analysis of pressure-overload-induced heart failure and its attenuation by mitochondrial-targeted peptides.

BACKGROUND: We investigated the protective effects of mitochondrial-targeted antioxidant and protective peptides, Szeto-Schiller (SS) 31 and SS20, on cardiac function, proteomic remodeling, and signaling pathways. METHODS AND RESULTS: We applied an improved label-free shotgun proteomics approach to evaluate the global proteomics changes in transverse aortic constriction (TAC)-induced heart failure and the associated signaling pathway changes using ingenuity pathway analysis. We found that 538 proteins significantly changed after TAC, which mapped to 53 pathways. The top pathways were in the categories of actin cytoskeleton, mitochondrial function, intermediate metabolism, glycolysis/gluconeogenesis, and citrate cycle. Concomitant treatment with SS31 ameliorated the congestive heart failure phenotypes and mitochondrial damage induced by TAC, in parallel with global attenuation of mitochondrial proteome changes, with an average of 84% protection of mitochondrial and 69% of nonmitochondrial protein changes. This included significant amelioration of all the ingenuity pathway analysis noted above. SS20 had only modest effects on heart failure and this tracked with only partial attenuation of global proteomics changes; furthermore, actin cytoskeleton pathways were significantly protected in SS20, whereas mitochondrial and metabolic pathways essentially were not. CONCLUSIONS: This study elucidates the signaling pathways significantly changed in pressure-overload-induced heart failure. The global attenuation of TAC-induced proteomic alterations by the mitochondrial-targeted peptide SS31 suggests that perturbed mitochondrial function may be an upstream signal to many of the pathway alterations in TAC and supports the potential clinical application of mitochondrial-targeted peptide drugs for the treatment heart failure.

Clonal expansions and short telomeres are associated with neoplasia in early-onset, but not late-onset, ulcerative colitis.

BACKGROUND: Patients with ulcerative colitis (UC) are at risk of developing colorectal cancer. We have previously reported that cancer progression is associated with the presence of clonal expansions and shorter telomeres in nondysplastic mucosa. We sought to validate these findings in an independent case-control study. METHODS: This study included 33 patients with UC: 14 progressors (patients with high-grade dysplasia or cancer) and 19 nonprogressors. For each patient, a mean of 5 nondysplastic biopsies from proximal, mid, and distal colon were assessed for clonal expansions, as determined by clonal length altering mutations in polyguanine tracts, and telomere length, as measured by quantitative PCR. Both parameters were compared with individual clinicopathological characteristics. RESULTS: Clonal expansions and shorter telomeres were more frequent in nondysplastic biopsies from UC progressors than nonprogressors, but only for patients with early-onset of UC (diagnosis at younger than 50 years of age). Late-onset progressor patients had very few or no clonal expansions and longer telomeres. A few nonprogressors exhibited clonal expansions, which were associated with older age and shorter telomeres. In progressors, clonal expansions were associated with proximity to dysplasia. The mean percentage of clonally expanded mutations distinguished early-onset progressors from nonprogressors with 100% sensitivity and 80% specificity. CONCLUSIONS: Early-onset progressors develop cancer in a field of clonally expanded epithelium with shorter telomeres. The detection of these clones in a few random nondysplastic colon biopsies is a promising cancer biomarker in early-onset UC. Curiously, patients with late-onset UC seem to develop cancer without the involvement of such fields.

Pan-colonic field defects are detected by CGH in the colons of UC patients with dysplasia/cancer.

BAC arrays were used to evaluate genomic instability along the colon of patients with ulcerative colitis (UC). Genomic instability increases with disease progression and biopsies more proximal to dysplasia showed increased instability. Pan-colonic field copy number gain or loss involving small (<1Mb) regions were detected in most patients and were particularly apparent in the UC progressor patients who had dysplasia or cancer. Chromosomal copy gains or losses affecting large regions were mainly restricted to dysplastic biopsies. Areas of significant chromosomal losses were detected in the UC progressors on chromosomes 2q36, 3q25, 3p21, 4q34, 4p16.2, 15q22, and 16p13 (p-value⩽0.04). These results extend our understanding of the dynamic nature of pan-colonic genomic instability in this disease.

Pilot study of blood biomarker candidates for detection of pancreatic cancer.

OBJECTIVES: Biomarkers that detect pancreatic cancer at earlier stages could improve the outcome of this deadly disease. METHODS: We investigated a dozen biomarker candidates for their potential as pancreatic cancer blood biomarkers using enzyme-linked immunosorbent assays. RESULTS: Among them, the macrophage migration inhibitory factor and osteopontin blood tests were nearly perfect in distinguishing pancreatic cancer cases from healthy controls (100% and 95% sensitivity, respectively, at 100% specificity). Five biomarker candidates were then tested on an expanded set of diseased controls, which included sera from patients with pancreatitis. The sensitivity dropped significantly for all 5 candidate markers. CONCLUSIONS: Our results suggest that biomarker candidates could fail in various steps of biomarker development. Earlier knowledge of candidate biomarker flaws could lead to strategies to overcome the flaw or alternatively lead to earlier termination of biomarkers that are prone to failure in the later phases of validation testing.