Genomic analyses identify molecular subtypes of pancreatic cancer.

Integrated genomic analysis of 456 pancreatic ductal adenocarcinomas identified 32 recurrently mutated genes that aggregate into 10 pathways: KRAS, TGF-ß, WNT, NOTCH, ROBO/SLIT signalling, G1/S transition, SWI-SNF, chromatin modification, DNA repair and RNA processing. Expression analysis defined 4 subtypes: (1) squamous; (2) pancreatic progenitor; (3) immunogenic; and (4) aberrantly differentiated endocrine exocrine (ADEX) that correlate with histopathological characteristics. Squamous tumours are enriched for TP53 and KDM6A mutations, upregulation of the TP63∆N transcriptional network, hypermethylation of pancreatic endodermal cell-fate determining genes and have a poor prognosis. Pancreatic progenitor tumours preferentially express genes involved in early pancreatic development (FOXA2/3, PDX1 and MNX1). ADEX tumours displayed upregulation of genes that regulate networks involved in KRAS activation, exocrine (NR5A2 and RBPJL), and endocrine differentiation (NEUROD1 and NKX2-2). Immunogenic tumours contained upregulated immune networks including pathways involved in acquired immune suppression. These data infer differences in the molecular evolution of pancreatic cancer subtypes and identify opportunities for therapeutic development.

Whole genomes redefine the mutational landscape of pancreatic cancer.

Pancreatic cancer remains one of the most lethal of malignancies and a major health burden. We performed whole-genome sequencing and copy number variation (CNV) analysis of 100 pancreatic ductal adenocarcinomas (PDACs). Chromosomal rearrangements leading to gene disruption were prevalent, affecting genes known to be important in pancreatic cancer (TP53, SMAD4, CDKN2A, ARID1A and ROBO2) and new candidate drivers of pancreatic carcinogenesis (KDM6A and PREX2). Patterns of structural variation (variation in chromosomal structure) classified PDACs into 4 subtypes with potential clinical utility: the subtypes were termed stable, locally rearranged, scattered and unstable. A significant proportion harboured focal amplifications, many of which contained druggable oncogenes (ERBB2, MET, FGFR1, CDK6, PIK3R3 and PIK3CA), but at low individual patient prevalence. Genomic instability co-segregated with inactivation of DNA maintenance genes (BRCA1, BRCA2 or PALB2) and a mutational signature of DNA damage repair deficiency. Of 8 patients who received platinum therapy, 4 of 5 individuals with these measures of defective DNA maintenance responded.

Precision Oncology in Surgery: Patient Selection for Operable Pancreatic Cancer.

OBJECTIVE: We aimed to define preoperative clinical and molecular characteristics that would allow better patient selection for operative resection. BACKGROUND: Although we use molecular selection methods for systemic targeted therapies, these principles are not applied to surgical oncology. Improving patient selection is of vital importance for the operative treatment of pancreatic cancer (pancreatic ductal adenocarcinoma). Although surgery is the only chance of long-term survival, 80% still succumb to the disease and approximately 30% die within 1 year, often sooner than those that have unresected local disease. METHOD: In 3 independent pancreatic ductal adenocarcinoma cohorts (total participants = 1184) the relationship between aberrant expression of prometastatic proteins S100A2 and S100A4 and survival was assessed. A preoperative nomogram based on clinical variables available before surgery and expression of these proteins was constructed and compared to traditional measures, and a postoperative nomogram. RESULTS: High expression of either S100A2 or S100A4 was independent poor prognostic factors in a training cohort of 518 participants. These results were validated in 2 independent patient cohorts (Glasgow, n = 198; Germany, n = 468). Aberrant biomarker expression stratified the cohorts into 3 distinct prognostic groups. A preoperative nomogram incorporating S100A2 and S100A4 expression predicted survival and nomograms derived using postoperative clinicopathological variables. CONCLUSIONS: Of those patients with a poor preoperative nomogram score, approximately 50% of patients died within a year of resection. Nomograms have the potential to improve selection for surgery and neoadjuvant therapy, avoiding surgery in aggressive disease, and justifying more extensive resections in biologically favorable disease.

Hypermutation In Pancreatic Cancer.

Pancreatic cancer is molecularly diverse, with few effective therapies. Increased mutation burden and defective DNA repair are associated with response to immune checkpoint inhibitors in several other cancer types. We interrogated 385 pancreatic cancer genomes to define hypermutation and its causes. Mutational signatures inferring defects in DNA repair were enriched in those with the highest mutation burdens. Mismatch repair deficiency was identified in 1% of tumors harboring different mechanisms of somatic inactivation of MLH1 and MSH2. Defining mutation load in individual pancreatic cancers and the optimal assay for patient selection may inform clinical trial design for immunotherapy in pancreatic cancer.

Pancreatic cancer genomes reveal aberrations in axon guidance pathway genes.

Pancreatic cancer is a highly lethal malignancy with few effective therapies. We performed exome sequencing and copy number analysis to define genomic aberrations in a prospectively accrued clinical cohort (n = 142) of early (stage I and II) sporadic pancreatic ductal adenocarcinoma. Detailed analysis of 99 informative tumours identified substantial heterogeneity with 2,016 non-silent mutations and 1,628 copy-number variations. We define 16 significantly mutated genes, reaffirming known mutations (KRAS, TP53, CDKN2A, SMAD4, MLL3, TGFBR2, ARID1A and SF3B1), and uncover novel mutated genes including additional genes involved in chromatin modification (EPC1 and ARID2), DNA damage repair (ATM) and other mechanisms (ZIM2, MAP2K4, NALCN, SLC16A4 and MAGEA6). Integrative analysis with in vitro functional data and animal models provided supportive evidence for potential roles for these genetic aberrations in carcinogenesis. Pathway-based analysis of recurrently mutated genes recapitulated clustering in core signalling pathways in pancreatic ductal adenocarcinoma, and identified new mutated genes in each pathway. We also identified frequent and diverse somatic aberrations in genes described traditionally as embryonic regulators of axon guidance, particularly SLIT/ROBO signalling, which was also evident in murine Sleeping Beauty transposon-mediated somatic mutagenesis models of pancreatic cancer, providing further supportive evidence for the potential involvement of axon guidance genes in pancreatic carcinogenesis.

Callus Distraction Versus Single-Stage Lengthening With Bone Graft for Treatment of Brachymetatarsia: A Systematic Review.

Brachymetatarsia deformity is a cosmetically displeasing anomaly that can become physically symptomatic. The surgical techniques most commonly used to repair the anomaly include single-stage lengthening with a bone graft, callus distraction, or a combination of bone grafting and callus distraction. A systematic review of the published data was performed to compare the outcomes of these 3 surgical procedures. A total of 61 studies reporting the use of callus distraction or single-stage lengthening, or both, for the treatment of brachymetatarsia were included in the present review. The incidence of major postoperative complications after callus distraction, single-stage lengthening, and the combination procedure was 49 (12.62%), 13 (3.72%), and 3 (33.33%), respectively. The number of minor complications with callus distraction, single-stage lengthening, and the combination procedure was 152 (39.18%), 55 (15.76%), and 1 (11.11%); the mean percentage of the original length achieved was 37.36%, 25.98% and 36.00%; and the mean length achieved was 17.5, 13.2, and 14.0 mm, respectively. The healing index (mo/cm) and healing time was 2.31 and 16.04 weeks, 1.90 and 9.35 weeks, and 3.93 and 14.62 weeks for callus distraction, single-stage lengthening, and the combination procedure, respectively. Our findings indicate that the callus distraction technique is associated with greater length gained but results in greater complication rates and requires almost twice the time to heal. Single-stage lengthening with a bone graft was associated with fewer complications and faster healing times than callus distraction but with lesser gains in length. From the information reported in the studies we reviewed, the prevalence of bilateral brachymetatarsia was 44.52%, and the female/male ratio was 13.7:1. Both of these findings seem to contradict the usual data given (72% for bilateral brachymetatarsia and a female/male ratio of 25:1).

Genome-wide DNA methylation patterns in pancreatic ductal adenocarcinoma reveal epigenetic deregulation of SLIT-ROBO, ITGA2 and MET signaling.

The importance of epigenetic modifications such as DNA methylation in tumorigenesis is increasingly being appreciated. To define the genome-wide pattern of DNA methylation in pancreatic ductal adenocarcinomas (PDAC), we captured the methylation profiles of 167 untreated resected PDACs and compared them to a panel of 29 adjacent nontransformed pancreata using high-density arrays. A total of 11,634 CpG sites associated with 3,522 genes were significantly differentially methylated (DM) in PDAC and were capable of segregating PDAC from non-malignant pancreas, regardless of tumor cellularity. As expected, PDAC hypermethylation was most prevalent in the 5' region of genes (including the proximal promoter, 5'UTR and CpG islands). Approximately 33% DM genes showed significant inverse correlation with mRNA expression levels. Pathway analysis revealed an enrichment of aberrantly methylated genes involved in key molecular mechanisms important to PDAC: TGF-ß, WNT, integrin signaling, cell adhesion, stellate cell activation and axon guidance. Given the recent discovery that SLIT-ROBO mutations play a clinically important role in PDAC, the role of epigenetic perturbation of axon guidance was pursued in more detail. Bisulfite amplicon deep sequencing and qRT-PCR expression analyses confirmed recurrent perturbation of axon guidance pathway genes SLIT2, SLIT3, ROBO1, ROBO3, ITGA2 and MET and suggests epigenetic suppression of SLIT-ROBO signaling and up-regulation of MET and ITGA2 expression. Hypomethylation of MET and ITGA2 correlated with high gene expression, which was associated with poor survival. These data suggest that aberrant methylation plays an important role in pancreatic carcinogenesis affecting core signaling pathways with potential implications for the disease pathophysiology and therapy.

Conservative versus Surgical Management of Fifth Metatarsal Diaphyseal Fractures: A Retrospective Review.

BACKGROUND: Historically, distal fifth metatarsal diaphyseal fractures have been treated with conservative management, with only limited research evaluating surgical treatment of these fractures. This study was performed to compare surgical versus conservative treatment of distal fifth metatarsal diaphyseal fractures in athletes and nonathletes. METHODS: A retrospective review of 53 patients with surgical or conservative treatment of isolated fifth metatarsal diaphyseal fractures was performed. Data recorded included age, sex, tobacco use, diagnosis of diabetes mellitus, time to clinical union, time to radiographic union, athletic versus nonathletic status, time to return to full activity, surgical fixation method, and complications. RESULTS: Patients treated surgically had a mean clinical union time of 8.2 weeks, radiographic union time of 13.5 weeks, and return to activity time of 12.9 weeks. Patients treated conservatively had a mean clinical union time of 16.3 weeks, radiographic union time of 25.2 weeks, and return to activity time of 20.7 weeks. Delayed unions and nonunions occurred in 27.0% of patients (10 of 37) treated conservatively and in none in the surgical group. CONCLUSIONS: Surgical treatment significantly decreased time to radiographic union, clinical union, and return to activity by an average of 8 weeks compared with conservative treatment. We suggest that surgical treatment of distal fifth metatarsal fractures is a viable option that may significantly decrease the patient's time to clinical union, radiographic union, and return to activity.

Comparison of Hallux Rigidus Surgical Treatment Outcomes Between Active Duty and Non-Active Duty Populations A Retrospective Review.

BACKGROUND: Our aim in this study was to compare the long-term outcomes of three different surgical procedures for the treatment of hallux rigidus (ie, cheilectomy, decompressive osteotomy, and arthrodesis) between active duty military and non-active duty patients. METHODS: A retrospective review of 80 patients (95 feet) undergoing surgical treatment for hallux rigidus was performed. Telephone survey was used to obtain postoperative outcome measures and subjective satisfaction. Additional data recorded and analyzed included age, sex, status of patient (active duty or non-active duty), grade of hallux rigidus, surgical procedure performed, date of surgery, time to return to full activity, ability to return to full duty, and follow-up time postoperatively. RESULTS: The decompressive osteotomy group had the highest return-to-duty rate, satisfaction rate, and Maryland Foot Scores of all three surgical groups, although these differences were not statistically significant. Active duty and non-active duty patients did not have statistically significant differences in outcomes measures (ie, time to return to full activity, ability to return to full duty, satisfaction, or postoperative Maryland Foot Score) in any of the three surgical groups. CONCLUSIONS: Decompressive osteotomy, cheilectomy, and first metatarsophalangeal joint arthrodesis are all reliable and effective procedures for treatment of hallux rigidus in both active duty military and non-active duty patients. Active duty military personal have a high rate of returning to their prior military activities after surgical treatment of hallux rigidus.

Purification and partial characterisation of recombinant human hepcidin.

Hepcidin is a liver-expressed antimicrobial and iron regulatory peptide. A number of studies have indicated that hepcidin is important for the correct regulation of body iron homeostasis. The aims of this study were to analyse the expression, trafficking and regulation of human hepcidin in an in vitro cell culture system. Human hepcidin was transfected into human embryonic kidney cells. Immunofluorescence and confocal microscopy analysis revealed that recombinant hepcidin localised to the Golgi complex. Recombinant hepcidin is secreted from the cell within 1 h of its synthesis. Recombinant hepcidin was purified from the cell culture medium using ion-exchange and metal-affinity chromatography and was active in antimicrobial assays. Amino-terminal sequence analysis of the secreted peptide revealed that it was the mature 25 amino acid form of hepcidin. Our results show that recombinant myc-His tagged human hepcidin was expressed, processed and secreted correctly and biologically active in antimicrobial assays.

Symptomatic Bipartite Medial Cuneiform: Report of Five Cases and Review of the Literature.

UNLABELLED: Bipartition of the medial cuneiform is a well-described but rarely seen anatomic variant. The majority of literature focuses on anatomic description and incidents based on studies of archeological collections. Symptomatic cases can be overlooked or misdiagnosed initially given the vague complaint of pain either chronic in nature or following an acute injury that could result in a myriad of foot conditions. Treatment ranges from orthotics, immobilization, injection therapy, and surgery. Presented here is a series of 5 cases treated successfully with conservative and surgical measures. LEVELS OF EVIDENCE: Therapeutic, Level IV.

Retained viable plant material in the calcaneus: a case report of a 22-year-old soldier with atypical heel pain.

Foreign bodies can be difficult to diagnose and should be considered in the differential diagnosis of unexplained pain, even in the absence of recalled trauma. We present the case of a 22-year-old male with a painful left heel. The patient did not recall a specific traumatic incident, and there were no clinical signs of trauma or infection. Plain films of the foot were nonrevealing, but magnetic resonance imaging revealed a sinus tract and left calcaneal defect. A biopsy of the calcaneal defect revealed viable woody material embedded and partially integrated with the surrounding bone. Postoperatively the patient's pain completely resolved. This case illustrates the importance of radiopathologic pursuit of an etiology of unexplained foot pain in an otherwise healthy person.

Connective tissue growth factor as a novel therapeutic target in high grade serous ovarian cancer.

Ovarian cancer is the most common cause of death among women with gynecologic cancer. We examined molecular profiles of fibroblasts from normal ovary and high-grade serous ovarian tumors to identify novel therapeutic targets involved in tumor progression. We identified 2,300 genes that are significantly differentially expressed in tumor-associated fibroblasts. Fibroblast expression of one of these genes, connective tissue growth factor (CTGF), was confirmed by immunohistochemistry. CTGF protein expression in ovarian tumor fibroblasts significantly correlated with gene expression levels. CTGF is a secreted component of the tumor microenvironment and is being pursued as a therapeutic target in pancreatic cancer. We examined its effect in in vitro and ex vivo ovarian cancer models, and examined associations between CTGF expression and clinico-pathologic characteristics in patients. CTGF promotes migration and peritoneal adhesion of ovarian cancer cells. These effects are abrogated by FG-3019, a human monoclonal antibody against CTGF, currently under clinical investigation as a therapeutic agent. Immunohistochemical analyses of high-grade serous ovarian tumors reveal that the highest level of tumor stromal CTGF expression was correlated with the poorest prognosis. Our findings identify CTGF as a promoter of peritoneal adhesion, likely to mediate metastasis, and a potential therapeutic target in high-grade serous ovarian cancer. These results warrant further studies into the therapeutic efficacy of FG-3019 in high-grade serous ovarian cancer.

Precision Medicine for Advanced Pancreas Cancer: The Individualized Molecular Pancreatic Cancer Therapy (IMPaCT) Trial.

PURPOSE: Personalized medicine strategies using genomic profiling are particularly pertinent for pancreas cancer. The Individualized Molecular Pancreatic Cancer Therapy (IMPaCT) trial was initially designed to exploit results from genome sequencing of pancreatic cancer under the auspices of the International Cancer Genome Consortium (ICGC) in Australia. Sequencing revealed small subsets of patients with aberrations in their tumor genome that could be targeted with currently available therapies. EXPERIMENTAL DESIGN: The pilot stage of the IMPaCT trial assessed the feasibility of acquiring suitable tumor specimens for molecular analysis and returning high-quality actionable genomic data within a clinically acceptable timeframe. We screened for three molecular targets: HER2 amplification; KRAS wild-type; and mutations in DNA damage repair pathways (BRCA1, BRCA2, PALB2, ATM). RESULTS: Tumor biopsy and archived tumor samples were collected from 93 patients and 76 were screened. To date 22 candidate cases have been identified: 14 KRAS wild-type, 5 cases of HER2 amplification, 2 mutations in BRCA2, and 1 ATM mutation. Median time from consent to the return of validated results was 21.5 days. An inability to obtain a biopsy or insufficient tumor content in the available specimen were common reasons for patient exclusion from molecular analysis while deteriorating performance status prohibited a number of patients from proceeding in the study. CONCLUSIONS: Documenting the feasibility of acquiring and screening biospecimens for actionable molecular targets in real time will aid other groups embarking on similar trials. Key elements include the need to better prescreen patients, screen more patients, and offer more attractive clinical trial options.

Rare open hallux interphalangeal joint dislocations sustained in combatives training: a case series.

Hallux interphalangeal joint dislocations are a very rare occurrence, and open medial dislocations of the hallux interphalangeal joint, to our knowledge, have not been previously reported in the literature. We report two open medial dislocations, one with fracture, that were sustained within a year of each other at the same military installation. Both patients presented were active duty soldiers that were involved in barefoot combatives and caught their hallux in the fold of the mats while simultaneously experiencing a "twisting force" applied to their foot by their combatives partner. Each soldier required surgical intervention and healed uneventfully, able to return to full activities in an average of 10 weeks with no residual pain. Two injuries of this rarity occurring with the same mechanism of injury within a year at the same military base raise concerns about the surface and shoegear being used for combatives training. The initial analysis of these two separate but similar cases points to the fact that injuries to the foot and toes may be reduced by using seamless mats and/or wearing closed-toed shoes (wrestling style) during combatives training.

Histomolecular phenotypes and outcome in adenocarcinoma of the ampulla of vater.

PURPOSE: Individuals with adenocarcinoma of the ampulla of Vater demonstrate a broad range of outcomes, presumably because these cancers may arise from any one of the three epithelia that converge at that location. This variability poses challenges for clinical decision making and the development of novel therapeutic strategies. PATIENTS AND METHODS: We assessed the potential clinical utility of histomolecular phenotypes defined using a combination of histopathology and protein expression (CDX2 and MUC1) in 208 patients from three independent cohorts who underwent surgical resection for adenocarcinoma of the ampulla of Vater. RESULTS: Histologic subtype and CDX2 and MUC1 expression were significant prognostic variables. Patients with a histomolecular pancreaticobiliary phenotype (CDX2 negative, MUC1 positive) segregated into a poor prognostic group in the training (hazard ratio [HR], 3.34; 95% CI, 1.69 to 6.62; P < .001) and both validation cohorts (HR, 5.65; 95% CI, 2.77 to 11.5; P < .001 and HR, 2.78; 95% CI, 1.25 to 7.17; P = .0119) compared with histomolecular nonpancreaticobiliary carcinomas. Further stratification by lymph node (LN) status defined three clinically relevant subgroups: one, patients with histomolecular nonpancreaticobiliary (intestinal) carcinoma without LN metastases who had an excellent prognosis; two, those with histomolecular pancreaticobiliary carcinoma with LN metastases who had a poor outcome; and three, the remainder of patients (nonpancreaticobiliary, LN positive or pancreaticobiliary, LN negative) who had an intermediate outcome. CONCLUSION: Histopathologic and molecular criteria combine to define clinically relevant histomolecular phenotypes of adenocarcinoma of the ampulla of Vater and potentially represent distinct diseases with significant implications for current therapeutic strategies, the ability to interpret past clinical trials, and future trial design.

Dehydroepiandrosterone sulfate and estrone sulfate reduce GABA-recurrent inhibition in the hippocampus via muscarinic acetylcholine receptors.

Several recent studies have established a role for estrogens in ameliorating specific neurodegenerative disorders, mainly those associated with the cholinergic neurons of the basal forebrain and their targets in the cortex and hippocampus. We have previously demonstrated that endogenous and exogenous application of the neurosteroid dehydroepiandrosterone sulfate (DHEAS) markedly reduces GABA-mediated recurrent inhibition and synchronizes hippocampal unit activity to theta rhythm (Steffensen (1995) Hippocampus 5:320-328). In this study, we evaluated the role of muscarinic receptors in mediating the effects of DHEAS and estrone sulfate (ES), the principal circulating estrogen in humans, on short-latency-evoked potential responses, paired-pulse inhibition (PPI), paired-pulse facilitation, and GABA interneuron activity in the dentate gyrus and CA1 subfields of the rat hippocampus. In situ microelectrophoretic application of the muscarinic M2 subtype cholinergic receptor agonist cis-dioxolane, DHEAS, and ES markedly reduced PPI in the dentate and CA1 that was blocked by the M2 receptor antagonist gallamine. Similar to DHEAS, microelectrophoretic administration of ES increased population spike amplitudes, without increasing excitatory transmission, but this effect was not blocked by gallamine. Microelectrophoretic application of cis-dioxolane and ES markedly increased the firing rate of dentate hilar interneurons and CA1 oriens/alveus interneurons and enhanced their synchrony to hippocampal theta rhythm. These findings suggest that select GABA-modulating neurosteroids and neuroactive estrogen sulfates alter septohippocampal cholinergic modulation of hippocampal GABAergic interneurons mediating recurrent, but not feedforward, inhibition of hippocampal principal cell activity.