NMR based metabolic profiling of patients with potential celiac disease elucidating early biochemical changes of gluten-sensitivity: A pilot study.

BACKGROUND: The patients with positive celiac disease (CeD) specific serology, but no evidence of intestinal inflammation are defined as potential celiac disease (PCeD) patients. About one-third of PCeD patients develop intestinal inflammation over time. The present study investigated the metabolome of small intestinal biopsies, blood plasma, and urine of patients with PCeD to understand the biochemical changes underlying the CeD. METHODS: The metabolic profiles of small intestinal biopsies, blood plasma, and urine of patients with PCeD (n = 7) were compared with CeD (n = 64) and controls (n = 15) [disease controls (DC) and healthy controls (HC)] using 1H NMR spectroscopy. RESULTS: The intestinal mucosa of PCeD showed lower levels of histidine, glycine, tyrosine, and tryptophan compared to DC. Altered levels of 6 metabolites (glucose, acetate, acetoacetate, ß-hydroxybutyrate, pyruvate, arginine) in blood plasma and two metabolites (succinate and aminohippurate) in urine were observed in PCeD compared to HC. The PLS-DA model built on the concentration of blood plasma showed separate clustering for PCeD and CeD patients. CONCLUSION: Altered metabolic profile of PCeD suggested that gluten intolerance was evident at the metabolic level before the intestinal damage. Altered energy metabolism and lower cytoprotective activity (histidine, glycine, arginine) indicated vulnerability to develop intestinal inflammation in PCeD over time. Our study may provide an insight into early biochemical processes of the progression of PCeD to CeD.

Best practices of handling, processing, and interpretation of small intestinal biopsies for the diagnosis and management of celiac disease: A joint consensus of Indian association of pathologists and microbiologists and Indian society of gastroenterology.

The Indian Association of Pathologists and Microbiologists (IAPM) and Indian Society of Gastroenterology (ISG) decided to make a joint consensus recommendation for handling, processing, and interpretation of SI biopsies for the diagnosis and management of celiac disease (CD) recognizing the inhomogeneous practice of biopsy sampling, orientation, processing, and interpretation. A modified Delphi process was used to develop this consensus document containing a total of 42 statements and recommendations, which were generated by sharing the document draft, incorporating expert's opinion, followed by three cycles of electronic voting as well as a full-day face-to-face virtual ZOOM meeting and review of supporting literature. Of the 42 statements, 7 statements are on small intestinal (SI) biopsy in suspected patients of CD, site and the number of biopsies; 7 on handling, fixative, orientation, processing, and sectioning in pathology laboratories; 2 on histological orientation; 13 statements on histological interpretation and histological grading; 3 on the assessment of follow-up biopsies; 2 statements on gluten-free diet (GFD)-nonresponsive CD; 4 on challenges in the diagnosis of CD; 2 statements each on pathology reporting protocol and training and infrastructure in this area. The goal of this guideline document is to formulate a uniform protocol agreed upon both by the experienced pathologists and gastroenterologists to standardize the practice, improve the yield of small bowel biopsy interpretation, patients' compliance, overall management in CD, and generate unified data for patient care and research in the related field.

Histological analyses of trucut liver biopsies from patients with noncirrhotic portal fibrosis and extra-hepatic portal vein obstruction.

BACKGROUND: Both noncirrhotic portal fibrosis (NCPF) and extrahepatic portal venous obstruction (EHPVO) are important causes of noncirrhotic portal hypertension (PH) in the Asian region. In this study, we analyzed the histopathological changes of liver needle-core biopsies from patients with NCPF and EHPVO. PATIENTS AND METHODS: The patients were diagnosed as per the Asia Pacific Association for the Study of Liver (APASL) criteria. Minimum adequacy criteria for liver core biopsies were defined, and finally, 69 liver biopsies from patients with NCPF and 100 liver biopsies from patients with EHPVO were analyzed. All histological parameters were predefined, and three experienced pathologists analyzed the biopsies after reaching consensus. Institute ethics committee clearance was taken. RESULTS: Although some histological features were overlapping, phlebosclerosis of intra-hepatic branches of the portal vein (PV), periportal aberrant vascular channels, remnant portal tracts, and hepatic fibrosis beyond the portal tracts without the formation of complete hepatic nodules (P < 0.001 for all) were common histological characteristics of NCPF on core-needle liver biopsies; while maintained lobular architecture, nonspecific dilatation of PV branches, absence of intra-hepatic PV phlebosclerosis, aberrant vascular channels, and significant fibrosis were characteristics of EHPVO. CONCLUSIONS: Despite the considerable histological overlap between NCPF and EHPVO, careful histological evaluation, supplemented by clinical features, radiological and biochemical findings can help in making a conclusive diagnosis. Patients with NCPF and EHPVO with clinical jaundice show transaminitis, high serum alkaline phosphatase level, more variceal bleed, and histological evidences of nodular regenerative hyperplasia.

Expression patterns and prognostic significances of RRM1 and ERCC1 in pancreatic carcinoma and cholangiocarcinoma.

BACKGROUND: Aggressive pancreatobiliary tumors often require oxaliplatin-based therapies, instead of standard gemcitabine-based therapy and biomarker studies at diagnosis to decide the appropriate therapeutic regimen. The ribonucleotide Reductase catalytic subunit M1 (RRM1) and excision repair cross-complementing gene-1 (ERCC1) are related to DNA synthesis and repair and essential in this regard. However, apart from the therapeutic benefit, their prognostic implication is controversial. METHODS: In this retrospective study, paraffin-embedded tissue from 51 cases of pancreatic cancer and 29 cases of cholangiocarcinoma were evaluated for RRM1 and ERCC1 expression by immunohistochemical technique along with 18 control pancreatic and biliary tissues. The semiquantitatively H score was calculated based on stain distribution and stain intensities. RESULTS: Both RRM1 and ERCC1 expression were high in tumor epithelium than in controls (RRM1: the difference was statistically significant in cholangiocarcinoma (P = 0.008); ERCC1: the difference was statistically significant both in pancreatic and cholangiocarcinoma (P < 0.05)]. However, no correlation was noted between RRM1 and ERCC1-low and high tumors with histological markers of prognosis and overall survival in these patients. CONCLUSIONS: The present study adds further evidence against the controversy that if RRM1 and ERCC1 expression in pancreatic and biliary carcinomas have any prognostic significance apart from their proven therapeutic benefits in these tumors.

Bariatric Surgery in Nonalcoholic Fatty Liver Disease (NAFLD): Impact Assessment Using Paired Liver Biopsy and Fibroscan.

BACKGROUND: Bariatric surgery is associated with a positive impact on the degree of hepatic steatosis and inflammation in nonalcoholic associated fatty liver disease (NAFLD), although its effect on fibrosis is contentious. The role of Fibroscan in the post-bariatric assessment of hepatic steatosis and fibrosis is unclear. OBJECTIVES: This work aims to study the impact of bariatric surgery on the course of NAFLD using both invasive (liver biopsy) and non-invasive tests (biochemical parameters and Fibroscan). METHODS: In this prospective study, the impact of bariatric surgery on the course of NAFLD was assessed using paired liver biopsy (intra-operative and post-bariatric surgery 1-year follow-up). The liver stiffness measurement (LSM) and controlled attenuation parameter (CAP) cutoffs for the assessment of hepatic fibrosis and steatosis, respectively, were calculated in both pre- and post-bariatric settings. RESULTS: Fifty-eight patients (70.7% females, mean age 39.2 years) underwent paired liver biopsy. Post-bariatric surgery 1-year liver biopsy showed significant improvement in all the histopathological parameters of NAFLD. The mean NAFLD Activity Score declined from 2.81 (± 1.08) to 1.31 (± 1.39) post-bariatric surgery. Thirty (51.7%) patients showed improvement in fibrosis, eighteen (31%) no change, and ten (17.2%) had worsening. Worsening of fibrosis was associated with a higher median age of 44.5 versus 38 years (p value = 0.033). The CAP cutoff values for the various stages of hepatic steatosis were higher pre-operatively as compared with those obtained post-bariatric surgery. CONCLUSIONS: Bariatric surgery is associated with significant improvement in histopathological parameters of NAFLD. Fibroscan shows good diagnostic accuracy in detecting advanced stage and grade of NAFLD.

Abnormalities in metabolic pathways in celiac disease investigated by the metabolic profiling of small intestinal mucosa, blood plasma and urine by NMR spectroscopy.

Celiac disease (CeD) is an autoimmune enteropathy caused by gluten intake in genetically predisposed individuals. We investigated the metabolism of CeD by metabolic profiling of intestinal mucosa, blood plasma and urine using NMR spectroscopy and multivariate analysis. The metabolic profile of the small intestinal mucosa was compared between patients with CeD (n = 64) and disease controls (DCs, n = 30). The blood plasma and urinary metabolomes of CeD patients were compared with healthy controls (HCs, n = 39). Twelve metabolites (proline (Pro), arginine (Arg), glycine (Gly), histidine (His), glutamate (Glu), aspartate, tryptophan (Trp), fumarate, formate, succinate (Succ), glycerophosphocholine (GPC) and allantoin (Alln)) of intestinal mucosa differentiated CeD from controls. The metabolome of blood plasma with 18 metabolites (Pro, Arg, Gly, alanine, Glu, glutamine, glucose (Glc), lactate (Lac), acetate (Ace), acetoacetate (AcAc), ß-hydroxybutyrate (ß-OHB), pyruvate (Pyr), Succ, citrate (Cit), choline (Cho), creatine (Cr), phosphocreatine (PCr) and creatinine) and 9 metabolites of urine (Pro, Trp, ß-OHB, Pyr, Succ, N-methylnicotinamide (NMN), aminohippurate (AHA), indoxyl sulfate (IS) and Alln) distinguished CeD from HCs. Our data demonstrated changes in nine metabolic pathways. The altered metabolites were associated with increased oxidative stress (Alln), impaired healing and repair mechanisms (Pro, Arg), compromised anti-inflammatory and cytoprotective processes (Gly, His, NMN), altered energy metabolism (Glc, Lac, ß-OHB, Ace, AcAc, Pyr, Succ, Cit, Cho, Cr and PCr), impaired membrane metabolism (GPC and Cho) and intestinal dysbiosis (AHA and IS). An orthogonal partial least square discriminant analysis model provided clear differentiation between patients with CeD and controls in all three specimens. A classification model built by combining the distinguishing metabolites of blood plasma and urine samples gave an AUC of 0.99 with 97.7% sensitivity, 93.3% specificity and a predictive accuracy of 95.1%, which was higher than for the models built separately using small intestinal mucosa, blood plasma and urine. In conclusion, a panel of metabolic biomarkers in intestinal biopsies, plasma and urine samples has potential to differentiate CeD from controls and may complement traditional tests to improve the diagnosis of CeD.

High potency multistrain probiotic improves liver histology in non-alcoholic fatty liver disease (NAFLD): a randomised, double-blind, proof of concept study.

OBJECTIVE: Pharmacological treatment of non-alcoholic fatty liver disease (NAFLD) is still evolving. Probiotics could be a promising treatment option, but their effectiveness needs to be established. The present study aimed to evaluate the efficacy of a high potency multistrain probiotic in adult patients with NAFLD. METHODS: Thirty-nine liver biopsy-proven patients with NAFLD were randomised in a double-blind fashion to either lifestyle modifications plus an oral multistrain probiotic (675 billion bacteria daily, n=19) or identical placebo (n=20) for 1 year. Lifestyle modifications included regular exercise for all and control of overweight/obesity (with additional dietary restrictions), hypertension and hyperlipidaemia in those with these risk factors. Primary objective of the study was the histological improvement in NAFLD activity score (NAS) and its components and secondary objectives were improvement in alanine transaminase (ALT) and cytokine profile. RESULTS: Thirty (76.9%) out of 39 patients with NAFLD completed the study with 1 year of follow-up. A repeat liver biopsy at 1 year could be done in 10 patients (52.6%) in probiotic group and five patients (25%) in placebo group. In comparison to baseline, hepatocyte ballooning (p=0.036), lobular inflammation (p=0.003) and NAS score (p=0.007) improved significantly at 1 year in the probiotic group. When compared with placebo, the NAS score improved significantly in the probiotic group (p=0.004), along with improvements in hepatocyte ballooning (p=0.05) and hepatic fibrosis (p=0.018). A significant improvement in levels of ALT (p=0.046), leptin (p=0.006), tumour necrosis factor-α (p=0.016) and endotoxins (p=0.017) was observed in probiotic group in comparison to placebo at 1 year. No significant adverse events were reported in the study. CONCLUSION: Patients with NAFLD managed with lifestyle modifications and multistrain probiotic showed significant improvement in liver histology, ALT and cytokines. TRIAL REGISTRATION NUMBER: The clinical trial is registered with CLINICAL TRIAL REGISTRYINDIA (CTRI); http://ctri.nic.in, No. CTRI/2008/091/000074.

Differentiating Crohn's disease from intestinal tuberculosis.

Differentiating Crohn's disease (CD) and intestinal tuberculosis (ITB) has remained a dilemma for most of the clinicians in the developing world, which are endemic for ITB, and where the disease burden of inflammatory bowel disease is on the rise. Although, there are certain clinical (diarrhea/hematochezia/perianal disease common in CD; fever/night sweats common in ITB), endoscopic (longitudinal/aphthous ulcers common in CD; transverse ulcers/patulous ileocaecal valve common in ITB), histologic (caseating/confluent/large granuloma common in ITB; microgranuloma common in CD), microbiologic (positive stain/culture for acid fast-bacillus in ITB), radiologic (long segment involvement/comb sign/skip lesions common in CD; necrotic lymph node/contiguous ileocaecal involvement common in ITB), and serologic differences between CD and ITB, the only exclusive features are caseation necrosis on biopsy, positive smear for acid-fast bacillus (AFB) and/or AFB culture, and necrotic lymph node on cross-sectional imaging in ITB. However, these exclusive features are limited by poor sensitivity, and this has led to the development of multiple multi-parametric predictive models. These models are also limited by complex formulae, small sample size and lack of validation across other populations. Several new parameters have come up including the latest Bayesian meta-analysis, enumeration of peripheral blood T-regulatory cells, and updated computed tomography based predictive score. However, therapeutic anti-tubercular therapy (ATT) trial, and subsequent clinical and endoscopic response to ATT is still required in a significant proportion of patients to establish the diagnosis. Therapeutic ATT trial is associated with a delay in the diagnosis of CD, and there is a need for better modalities for improved differentiation and reduction in the need for ATT trial.

Combination of increased visceral fat and long segment involvement: Development and validation of an updated imaging marker for differentiating Crohn's disease from intestinal tuberculosis.

BACKGROUND AND AIM: Computed tomographic (CT) features (long segment, ileocaecal area involvement, and lymph nodes > 1 cm) have demonstrated good specificity but poor sensitivity, while visceral to subcutaneous fat ratio on CT (VF/SC > 0.63) has moderate sensitivity and specificity in differentiating Crohn's disease (CD) and intestinal tuberculosis (ITB). This study aims to develop and validate an updated model incorporating CT features and VF/SC to improve the diagnostic accuracy of imaging in differentiating CD/ITB. METHODS: Computed tomographic features and VF/SC were documented in two cohorts (development [n = 59, follow-up: January 2012 to November 2014] and validation [n = 69, follow-up: December 2014 to December 2015]) of CD/ITB patients diagnosed by standard criteria. Patients with normal CT were excluded. Features significantly different between CD/ITB were incorporated into a model. RESULTS: In both the cohorts, necrotic lymph nodes were exclusive for ITB (23.1% vs 0% and 43.3% vs 0%), while long segment involvement (57.6% vs 7.7%, P < 0.001, and 52.6% vs 16.1%, P < 0.001) and VF/SC ratio > 0.63 (72.7% vs 19.2%, P < 0.001, and 81.6% vs 25.8%, P < 0.001) were significantly more common in CD. A risk score of 2, based upon long segment involvement and VF/SC ratio > 0.63, had an excellent specificity of 100% and 100% and sensitivity of 54% and 50% for CD in development and validation cohorts, respectively. Based upon these features, in 43% patients with the diagnostic dilemma of CD/ITB, a definite diagnosis based only on imaging could be made. CONCLUSION: Necrotic lymph nodes are exclusive for ITB, and the combination of long segment involvement and VF/SC ratio > 0.63 is exclusive for CD, and these features can make a definite diagnosis in 43% patients with a CD/ITB dilemma.

Tubercular Intestinal Strictures Show a Poor Response to Anti-Tuberculous Therapy.

BACKGROUND: The literature on resolution of intestinal strictures in patients with intestinal tuberculosis (ITB) after anti-tuberculous therapy (ATT) is sparse and ambivalent. We aimed to assess the frequency of stricture resolution after ATT and its predictors. METHODS: This ambispective cohort study included consecutive ITB patients with strictures who received ATT for ≥6 months and were on regular follow-up between January 2004 and December 2015. Resolution of stricture was assessed at the end of ATT by endoscopy/radiology. RESULTS: Of 286 patients, 128 had strictures, and 106 were finally included (63 males, median age 35 years). The stricture location was distal ileum/ileocecal in 52 (49.1%), colon in 37 (34.9%), ileocolonic in 4 (3.8%), proximal small bowel in 10 (9.4%), and gastroduodenal in 4 (3.8%) patients. Although all patients demonstrated mucosal healing (indicating resolution of active infection), stricture resolution occurred only in 25/106 (23.6%) patients. Symptoms pertaining to stricture (pain abdomen/recurrent SAIO) were present in 104/106 (98%) patients, and after a median of 6 (6-9) months of ATT, these symptoms resolved only in half, 88% (22/25) in patients with stricture resolution and 38% (30/79) in patients with persistent strictures. Colonic strictures had the least resolution (5.4%) followed by proximal small intestinal (20%) and distal ileal/ileocecal (36.5%). Although not statistically significant, stricture resolution was less frequent in patients with multiple strictures, longer strictures (>3 cm), and strictures in which scope was not negotiable prior to ATT. CONCLUSION: Only one-fourth of ITB patients with strictures show resolution of stricture following ATT. The resolution of strictures is dependent on disease location, and majority of them exhibit symptoms pertaining to stricture even after ATT.

Prevalence of celiac disease in Indian patients with irritable bowel syndrome and uninvestigated dyspepsia.

OBJECTIVE: The clinical spectrum of celiac disease (CeD) is wide and its symptoms overlap with those of functional bowel diseases. This study aimed to investigate the relationship among gluten-related disorders, irritable bowel syndrome (IBS) and uninvestigated dyspepsia in Indian patients. METHODS: Patients with IBS and uninvestigated dyspepsia (using Rome III criteria) were tested for immunoglobulin A (IgA) anti-tissue transglutaminase (anti-tTG) antibody and anti-gliadin antibody (AGA). Those with positive anti-tTG antibody were evaluated for the presence of villous abnormalities. Patients who were only IgA AGA-positive were considered to have gluten sensitivity and those with positive anti-tTG antibody and villous atrophy were considered to have CeD. RESULTS: Of 362 patients with IBS, 22 (6.1%) had positive anti-tTG antibody, among whom 3 (0.8%) had CeD and 19 had potential CeD. Of 358 patients with uninvestigated dyspepsia, 18 (5.0%) were anti-tTG antibody-positive and among them 4 (1.1%) had CeD and 14 had potential CeD. AGA was positive in 104 (28.7%) patients with IBS and 68 (19.0%) with uninvestigated dyspepsia, suggesting the presence of gluten sensitivity. CONCLUSION: This study highlights the relationship between IBS or dyspepsia and CeD or gluten sensitivity.

Clinical significance of altered expression of ß-catenin and E-cadherin in oral dysplasia and cancer: potential link with ALCAM expression.

BACKGROUND: Perturbations in cell adhesion molecules are linked to alterations in cadherin-catenin complexes and likely play major roles in invasion and metastasis; their impact on early precancerous stages remains yet unknown. We showed ALCAM overexpression in early oral lesions and its cytoplasmic accumulation in oral squamous cell carcinoma (OSCC) to be a predictor of disease progression and poor prognosis. This study tested the hypothesis that alterations in E-cadherin and ß -catenin expressions are early events in oral tumorigenesis, associated with disease prognosis, and correlate with perturbations in ALCAM expression. METHODS: Expressions of E-cadherin and ß-catenin were analyzed in the same cohort of 105 OSCCs, 76 oral lesions and 30 normal oral tissues by immunohistochemistry and correlated with clinicopathological parameters and prognosis. The effect of siRNA mediated ALCAM knockdown on E-cadherin and ß -catenin was determined using western blot, confocal microscopy and RT-PCR analysis in oral cancer cells. RESULTS: Significant loss of membranous E-cadherin and ß-catenin expression was observed from normal, hyperplasia, dysplasia to OSCCs (p(trend) <0.001); and correlated with cytoplasmic ALCAM accumulation in OSCCs (p  = 0.006). Multivariate analysis revealed ß-catenin membrane loss and ALCAM/ß-catenin(nuclear/cytoplasmic) accumulation to be significant predictors for late clinical stage (p<0.001, OR = 8.7; p = 0.006, OR = 9.9, respectively) and nodal metastasis (p = 0.003, OR = 3.8; p = 0.025, OR = 3.4 respectively). Cox's regression showed E-cadherin membrane loss/ALCAM cytoplasmic expression [p<0.001; HR = 4.8] to be independent adverse prognosticators in OSCCs. siRNA mediated silencing of ALCAM resulted in concurrent increase in E-cadherin and ß-catenin both at the transcript and protein levels. CONCLUSIONS: Losses of E-cadherin and ß-catenin expressions are early events in oral tumorigenesis; their associations with aggressive tumor behavior and disease recurrence underscore their potential as prognostic markers. Correlation of loss of E-cadherin and ß-catenin with cytoplasmic ALCAM accumulation both in vitro and in in vivo suggests that these dynamic changes in cell adhesion system may play pivotal role in oral cancer.

Loss of DLC1 is an independent prognostic factor in patients with oral squamous cell carcinoma.

Deleted in liver cancer (DLC1), a Rho GTPase-activating protein, was observed to be differentially expressed in oral squamous cell carcinoma in comparison with normal tissues using tissue proteomics. In the current study, we investigated the clinical significance of loss of DLC1 expression in different stages of development of oral squamous cell carcinoma to determine its potential as a biomarker for oral dysplasia and prognosis of oral squamous cell carcinoma. Immunohistochemical analysis of DLC1 expression was carried out in oral squamous cell carcinoma patients (n=214), dysplasia (n=51), hyperplastic squamous mucosa (n=45), and histologically normal oral tissues (n=80), and correlated with clinicopathological parameters and disease prognosis over 91 months for oral squamous cell carcinomas. Loss of DLC1 expression was observed in oral squamous cell carcinoma (64%), oral dysplasia (31%), hyperplastic squamous mucosa (22%), and normal mucosa (16%). Significant loss of DLC1 expression was observed in oral squamous cell carcinomas as compared with dysplasia (P<0.001, odds ratio=3.8, 95% CI=2.0-7.3), suggesting it may be an important event involved in cancer progression. Among oral squamous cell carcinomas, the loss of DLC1 expression was significantly associated with poor prognosis (P=0.021, hazards ratio (HR)=1.8, 95% CI=1.1-2.9). Multivariate analysis revealed loss of DLC1 (P=0.023, HR=2.1, 95% CI=1.2-3.9) and histopathological grade (P=0.015, HR=1.7, 95% CI=1.1-2.7) to be independent predictors for disease-free survival in oral squamous cell carcinoma patients in comparison with known prognostic factors, viz. tumor stage, nodal status, and overall stage. Loss of DLC1 expression emerged as an important biomarker for predicting patients diagnosed with oral dysplasia at high risk of transformation upon future validation in longitudinal studies. Loss of DLC1 expression is a poor prognostic marker for oral squamous cell carcinoma patients.

Overexpression of prothymosin alpha predicts poor disease outcome in head and neck cancer.

BACKGROUND: In our recent study, tissue proteomic analysis of oral pre-malignant lesions (OPLs) and normal oral mucosa led to the identification of a panel of biomarkers, including prothymosin alpha (PTMA), to distinguish OPLs from histologically normal oral tissues. This study aimed to determine the clinical significance of PTMA overexpression in oral squamous cell hyperplasia, dysplasia and head and neck squamous cell carcinoma (HNSCC). METHODOLOGY: Immunohistochemistry of PTMA protein was performed in HNSCCs (n = 100), squamous cell hyperplasia (n = 116), dysplasia (n = 50) and histologically normal oral tissues (n = 100). Statistical analysis was carried out to determine the association of PTMA overexpression with clinicopathological parameters and disease prognosis over 7 years for HNSCC patients. RESULTS: Our immunohistochemical analysis demonstrated significant overexpression of nuclear PTMA in squamous cell hyperplasia (63.8%), dysplasia (50%) and HNSCC (61%) in comparison with oral normal mucosa (p(trend)<0.001). Chi-square analysis showed significant association of nuclear PTMA with advanced tumor stages (III+IV). Kaplan Meier survival analysis indicated reduced disease free survival (DFS) in HNSCC patients (p<0.001; median survival 11 months). Notably, Cox-multivariate analysis revealed nuclear PTMA as an independent predictor of poor prognosis of HNSCC patients (p<0.001, Hazard's ratio, HR = 5.2, 95% CI = 2.3-11.8) in comparison with the histological grade, T-stage, nodal status and tumor stage. CONCLUSIONS: Nuclear PTMA may serve as prognostic marker in HNSCC to determine the subset of patients that are likely to show recurrence of the disease.

Nuclear S100A7 is associated with poor prognosis in head and neck cancer.

BACKGROUND: Tissue proteomic analysis of head and neck squamous cell carcinoma (HNSCC) and normal oral mucosa using iTRAQ (isobaric tag for relative and absolute quantitation) labeling and liquid chromatography-mass spectrometry, led to the identification of a panel of biomarkers including S100A7. In the multi-step process of head and neck tumorigenesis, the presence of dysplastic areas in the epithelium is proposed to be associated with a likely progression to cancer; however there are no established biomarkers to predict their potential of malignant transformation. This study aimed to determine the clinical significance of S100A7 overexpression in HNSCC. METHODOLOGY: Immunohistochemical analysis of S100A7 expression in HNSCC (100 cases), oral lesions (166 cases) and 100 histologically normal tissues was carried out and correlated with clinicopathological parameters and disease prognosis over 7 years for HNSCC patients. Overexpression of S100A7 protein was significant in oral lesions (squamous cell hyperplasia/dysplasia) and sustained in HNSCC in comparison with oral normal mucosa (p(trend)<0.001). Significant increase in nuclear S100A7 was observed in HNSCC as compared to dysplastic lesions (p = 0.005) and associated with well differentiated squamous cell carcinoma (p = 0.031). Notably, nuclear accumulation of S100A7 also emerged as an independent predictor of reduced disease free survival (p = 0.006, Hazard ratio (HR = 7.6), 95% CI = 1.3-5.1) in multivariate analysis underscoring its relevance as a poor prognosticator of HNSCC patients. CONCLUSIONS: Our study demonstrated nuclear accumulation of S100A7 may serve as predictor of poor prognosis in HNSCC patients. Further, increased nuclear accumulation of S100A7 in HNSCC as compared to dysplastic lesions warrants a large-scale longitudinal study of patients with dysplasia to evaluate its potential as a determinant of increased risk of transformation of oral premalignant lesions.

A phase II study of gemcitabine and oxaliplatin (Oxigem) in unresectable gall bladder cancer.

PURPOSE: There is a need for effective chemotherapy protocols for gall bladder cancer (GBC). Gemcitabine has antitumor activity in pancreatic cancer. Oxaliplatin is effective in GI cancers. Based on evidence of synergy between these two, we designed this study to evaluate efficacy of this combination in unresectable GBC. DESIGN: Unresectable GBC was enrolled for single center phase II study. Drugs gemcitabine 900 mg/m2 and oxaliplatin 80 mg/m2 IV infusion (Oxigem) on days 1 and 8 every 3 weeks for a maximum of six cycles or unacceptable toxicity which ever was earlier. MATERIALS AND METHODS: Fifty patients were enrolled and analysis was restricted to 48 who were treated. Median age was 50 years and 31 patients were females. RESULTS: CR 3 (6.2%), PR 7 (15%), SD 17 (35.4%), and PD 18. One had complete pathological response. Median OS and PFS were 7.5 and 3 months, respectively. OS in responders was 10.5 versus 4 months in non-responders (p<0.0000). Eleven patients (23%) survived for a year or more. There was no toxic death and grade III/IV toxicity seen in 10 (22%) patients: diarrhea 3, vomiting 2, neutropenia and thrombocytopenia 5 patients. CONCLUSION: This combination of Oxigem effective in unresectable GBC. It may even induce complete pathological response. One-year survival was 20%. There is a need for controlled trial to assess efficacy of this combination.

Heterogeneous ribonucleoprotein K is a marker of oral leukoplakia and correlates with poor prognosis of squamous cell carcinoma.

Oral leukoplakia is a heterogeneous lesion with risk of cancer development; there are no biomarkers to predict its potential of malignant transformation. Tissue proteomic analysis of oral leukoplakia using iTRAQ labeling liquid chromatography-mass spectrometry showed overexpression of heterogeneous ribonucleoprotein K (hnRNP K), a transformation-related RNA-binding protein, in leukoplakia in comparison with normal tissue. Herein, we investigated the clinical significance of hnRNP K in identification of oral leukoplakic lesions in early stages and as a prognostic marker in head-and-neck/oral squamous cell carcinomas (HNOSCCs). Immunohistochemical analysis of hnRNP K was performed in 100 HNOSCCs, 199 leukoplakias and 55 nonmalignant tissues and correlated with clinicopathologic parameters and disease prognosis over 6 years for HNOSCCs. hnRNP K nuclear expression increased from normal tissues to leukoplakia, and frank malignancy (p < 0.001). Cytoplasmic hnRNP K increased significantly from leukoplakia to HNOSCCs (p < 0.001) and was associated with poor prognosis of HNOSCCs (p = 0.011) by Kaplan-Meier analysis. The most important finding of our follow-up study is that cytoplasmic hnRNP K is an independent predictor of disease recurrence in HNOSCC patients. In conclusion, nuclear hnRNP K may serve as a potential marker for early diagnosis, whereas its cytoplasmic accumulation can help to identify a subgroup of HNOSCC patients with poor prognosis, suggesting its putative utility in clinical management of HNOSCC.

Cytoplasmic accumulation of activated leukocyte cell adhesion molecule is a predictor of disease progression and reduced survival in oral cancer patients.

Activated leukocyte cell adhesion molecule (ALCAM) has been proposed to function as a cell surface sensor for cell density, controlling the transition between local cell proliferation and tissue invasion in cancer progression. Herein, we determined ALCAM expression in 107 oral squamous cell carcinomas (OSCCs), 78 oral lesions (58 hyperplasias and 20 dysplasias) and 30 histologically normal oral tissues using immunohistochemistry and correlated with clinicopathological parameters. Significant increase in ALCAM immunopositivity was observed from normal oral mucosa, hyperplasia, dysplasia to OSCCs (p(trend) < 0.001). Increased ALCAM expression was observed in cytoplasm of epithelial cells as early as in hyperplasia (p = 0.001, OR = 3.8). Sixty-five of 107 (61%) OSCCs showed significant overexpression of ALCAM protein in cytoplasm/membrane of tumor cells (p = 0.043; OR = 3.3) in comparison with the normal oral tissues. Among OSCCs, cytoplasmic ALCAM was associated with advanced tumor size, tumor stage and tobacco consumption. Importantly, cytoplasmic ALCAM was an independent predictor of poor prognosis of OSCCs in multivariate analysis (p = 0.012, OR = 6.2). In an attempt to understand the molecular basis of cytoplasmic localization of ALCAM, 14-3-3 zeta and 14-3-3 sigma were identified as its novel binding partners in oral cancer cells. In conclusion, increased expression of ALCAM is an early event in oral tumorigenesis; its cytoplasmic accumulation in tumor cells is a predictor of poor prognosis of OSCCs, underscoring its potential as a candidate prognostic marker for oral cancer.