Anorectal Malformations: Finding the Pathway out of the Labyrinth.

Anorectal malformations (ARMs) are a complex group of congenital anomalies involving the distal anus and rectum, as well as the urinary and genital tracts in a significant number of cases. Most ARMs result from abnormal development of the urorectal septum in early fetal life. In most cases, the anus is not perforated and the distal enteric component ends blindly (atresia) or as a fistula into the urinary tract, genital tract, or perineum. ARMs are also present in a great number of syndromes and associations of congenital anomalies. The classification of ARMs is mainly based on the position of the rectal pouch relative to the puborectal sling, the presence or absence of fistulas, and the types and locations of the fistulas. All of this information is crucial in determining the most appropriate surgical approach for each case. Imaging studies play a key role in evaluation and classification of ARMs. In neonates, clinical and radiologic examinations in the first 3 days of life help determine the type of ARM and the need for early colostomy. In older children, preoperative pelvic magnetic resonance imaging is the most efficient diagnostic method for evaluating the size, morphology, and grade of development of the sphincteric musculature.

Prognostic effect of epithelial cell adhesion molecule overexpression in untreated node-negative breast cancer.

PURPOSE: Epithelial cell adhesion molecule (Ep-CAM) recently received increased attention not only as a prognostic factor in breast cancer but also as a potential target for immunotherapy. We examined Ep-CAM expression in 402 consecutive node-negative breast cancer patients with long-term follow-up not treated in the adjuvant setting. EXPERIMENTAL DESIGN: Ep-CAM expression was evaluated by immunostaining. Its prognostic effect was estimated relative to overexpression/amplification of HER-2, histologic grade, tumor size, age, and hormone receptor expression. RESULTS: Ep-CAM status was positive in 106 (26.4%) patients. In multivariate analysis, Ep-CAM status was associated with disease-free survival independent of age, pT stage, histologic grade, estrogen receptor (ER), progesterone receptor (PR), as well as HER2 status (P = 0.028; hazard ratio, 1.60; 95% confidence interval, 1.05-2.44). Recently, so-called triple-negative (HER-2, ER, and PR) breast cancer has received increased attention. We noticed a similar association of Ep-CAM with disease-free survival in the triple-negative group as for the entire cohort. CONCLUSION: In this study of untreated breast cancer patients, Ep-CAM overexpression was associated with poor survival in the entire cohort and in the subgroup of triple-negative breast cancer. This suggests that Ep-CAM may be a well-suited target for specific therapies particularly in HER-2-, ER-, and PR-negative tumors.

Percutaneous radiofrequency ablation of primary intraosseous spinal glomus tumor.

The glomus tumor is a rare, benign, but painful vascular neoplasm arising from the neuromyoarterial glomus. Primary intraosseous glomus tumor is even rarer, with only about 20 cases reported in the literature so far, 5 of which involved the spine. Surgical resection is currently considered the treatment of choice. We herewith present an uncommon case of primary intraosseous spinal glomus tumor involving the right pedicle of the eleventh thoracic vertebra (T11). To our knowledge, this is the first case of primary intraosseous spinal glomus tumor successfully treated by percutaneous CT-guided radiofrequency ablation (RFA).

TCR-induced NF-kappaB activation: a crucial role for Carma1, Bcl10 and MALT1.

T-cell receptor (TCR) engagement induces the maturation of thymocytes and the activation and proliferation of peripheral T cells through signaling pathways that target several transcription factors. The transcription factor nuclear factor-κB (NF-κB) has an essential role in the activation of mature T cells but the signaling pathway leading from TCR stimulation to NF-κB activation is not well defined. Carma1, Bcl10 and MALT1 are recently identified proteins that have an important and previously unexpected role in antigen receptor-induced NF-κB activation and the control of lymphocyte proliferation. We believe that the recent advances in this field could stimulate research for the development of new immunomodulatory drugs and could lead to a better understanding of the molecular mechanisms underlying the formation of lymphomas and potentially of other immune disorders.

A novel interferon-gamma regulated human melanoma-associated antigen, gp33-38, defined by monoclonal antibody Me14-D12. II. Molecular cloning of a genomic probe.

The human Me14-D12 antigen is a cell surface glycoprotein regulated by interferon-gamma (IFN-gamma) on tumor cell lines of neuroectodermal origin. It consists of two non-convalently linked subunits with apparent mol. wt sizes of 33,000 and 38,000. Here we describe the molecular cloning of a genomic probe for the Me14-D12 gene using the gene transfer approach. Mouse Ltk- cells were stably cotransfected with human genomic DNA and the Herpes Simplex virus thymidine kinase (TK) gene. Primary and secondary transfectants expressing the Me14-D12 antigen were isolated after selection in HAT medium by repeated sorting on a fluorescence activated cell sorter (FACS). A recombinant phage harboring a 14.3 kb insert of human DNA was isolated from a genomic library made from a positive secondary transfectant cell line. A specific probe derived from the phage DNA insert allowed the identification of two mRNAs of 3.5 kb and 2.2 kb in primary and secondary L cell transfectants, as well as in human melanoma cell lines expressing the Me14-D12 antigen. The regulation of Me14-D12 antigen by INF-gamma was retained in the L cell transfectants and could be detected both at the level of protein and mRNA expression.

Decreased binding of peptides-MHC class I (pMHC) multimeric complexes to CD8 affects their binding avidity for the TCR but does not significantly impact on pMHC/TCR dissociation rate.

The CD8 coreceptor plays a crucial role in both T cell development in the thymus and in the activation of mature T cells in response to Ag-specific stimulation. In this study we used soluble peptides-MHC class I (pMHC) multimeric complexes bearing mutations in the CD8 binding site that impair their binding to the MHC, together with altered peptide ligands, to assess the impact of CD8 on pMHC binding to the TCR. Our data support a model in which CD8 promotes the binding of TCR to pMHC. However, once the pMHC/TCR complex is formed, the TCR dominates the pMHC/TCR dissociation rates. As a consequence of these molecular interactions, under physiologic conditions CD8 plays a key role in complex formation, resulting in the enhancement of CD8 T cell functions whose specificity, however, is determined by the TCR.

The proteolytic activity of the paracaspase MALT1 is key in T cell activation

The paracaspase MALT1 is pivotal in antigen receptor-mediated lymphocyte activation and lymphomagenesis. MALT1 contains a caspase-like domain, but it is unknown whether this domain is proteolytically active. Here we report that MALT1 had arginine-directed proteolytic activity that was activated after T cell stimulation, and we identify the signaling protein Bcl-10 as a MALT1 substrate. Processing of Bcl-10 after Arg228 was required for T cell receptor-induced cell adhesion to fibronectin. In contrast, MALT1 activity but not Bcl-10 cleavage was essential for optimal activation of transcription factor NF-kappaB and production of interleukin 2. Thus, the proteolytic activity of MALT1 is central to T cell activation, which suggests a possible target for the development of immunomodulatory or anticancer drugs

The effect of sodium tetrathionate stabilization on the distribution of three nuclear matrix proteins in human K562 erythroleukemia cells.

Using both conventional fluorescence and confocal laser scanning microscopy we have investigated whether or not stabilization of isolated human erythroleukemic nuclei with sodium tetrathionate can maintain in the nuclear matrix the same spatial distribution of three polypeptides (M(r) 160 kDa and 125 kDa, previously shown to be components of the internal nuclear matrix plus the 180-kDa nucleolar isoform of DNA topoisomerase II) as seen in permeabilized cells. The incubation of isolated nuclei in the presence of 2 mM sodium tetrathionate was performed at 0 degrees C or 37 degrees C. The matrix fraction retained 20-40% of nuclear protein, depending on the temperature at which the chemical stabilization was executed. Western blot analysis revealed that the proteins studied were completely retained in the high-salt resistant matrix. Indirect immunofluorescence experiments showed that the distribution of the three antigens in the final matrix closely resembled that detected in permeabilized cells, particularly when the stabilization was performed at 37 degrees C. This conclusion was also strengthened by analysis of cells, isolated nuclei and the nuclear matrix by means of confocal laser scanning microscopy. We conclude that sodium tetrathionate stabilization of isolated nuclei does not alter the spatial distribution of some nuclear matrix proteins.

A nonapeptide encoded by human gene MAGE-1 is recognized on HLA-A1 by cytolytic T lymphocytes directed against tumor antigen MZ2-E.

We have reported the identification of human gene MAGE-1, which directs the expression of an antigen recognized on a melanoma by autologous cytolytic T lymphocytes (CTL). We show here that CTL directed against this antigen, which was named MZ2-E, recognize a nonapeptide encoded by the third exon of gene MAGE-1. The CTL also recognize this peptide when it is presented by mouse cells transfected with an HLA-A1 gene, confirming the association of antigen MZ2-E with the HLA-A1 molecule. Other members of the MAGE gene family do not code for the same peptide, suggesting that only MAGE-1 produces the antigen recognized by the anti-MZ2-E CTL. Our results open the possibility of immunizing HLA-A1 patients whose tumor expresses MAGE-1 either with the antigenic peptide or with autologous antigen-presenting cells pulsed with the peptide.

In vivo localization of a bispecific antibody which targets human T lymphocytes to lyse human colon cancer cells.

A bispecific MAb was derived from the fusion of a hybridoma producing MAb CD3 with a hybridoma producing MAb L-DI (which is directed against a 41-kDa glycoprotein expressed in most gastro-intestinal and pancreatic carcinomas). Bispecific antibody molecules were isolated from parental antibody molecules by the use of hydroxylapatite-HPLC and shown to target human cytolytic T lymphocytes, irrespective of their original specificity, to specifically lyse human colon carcinoma cells. Localization studies in vivo using nude mice bearing human colon carcinoma xenografts showed significant accumulation of the HPLC-purified 125I-labelled bispecific antibodies into the tumor compared to 131I-labelled control CD3 antibody.

Vemurafenib in patients with BRAF(V600) mutation-positive melanoma with symptomatic brain metastases: final results of an open-label pilot study.

BACKGROUND & AIM: Brain metastases are frequent in patients with metastatic melanoma, indicating poor prognosis. We investigated the BRAF kinase inhibitor vemurafenib in patients with advanced melanoma with symptomatic brain metastases. METHODS: This open-label trial assessed vemurafenib (960mg twice a day) in patients with BRAF(V600) mutation-positive metastatic melanoma with non-resectable, previously treated brain metastases. The primary end-point was safety. Secondary end-points included best overall response rate, and progression-free and overall survival. RESULTS: Twenty-four patients received vemurafenib for a median treatment duration of 3.8 (0.1-11.3) months. The majority of discontinuations were due to disease progression (n=22). Twenty-three of 24 patients reported at least one adverse event (AE). Grade 3 AEs were reported in four (17%; 95% confidence interval [CI], 4.7-37.4%) patients and included cutaneous squamous cell carcinoma in four patients. Median progression-free survival was 3.9 (95% CI, 3.0-5.5) months, and median survival was 5.3 (95% CI, 3.9-6.6) months. An overall partial response (PR) at both intracranial and extracranial sites was achieved in 10 of 24 (42%; 95% CI, 22.1-63.4) evaluable patients, with stable disease in nine (38%; 95% CI, 18.8-59.4) patients. Of 19 patients with measurable intracranial disease, seven (37%) achieved >30% intracranial tumour regression, and three (16%; 95% CI, 3.4-39.6%) achieved a confirmed PR. Other signs of improvement included reduced need for corticosteroids and enhanced performance status. CONCLUSIONS: Vemurafenib can be safely used in patients with advanced symptomatic melanoma that has metastasised to the brain and can result in meaningful tumour regression.

Colon-specific deletion of epithelial sodium channel causes sodium loss and aldosterone resistance.

Aldosterone promotes electrogenic sodium reabsorption through the amiloride-sensitive epithelial sodium channel (ENaC). Here, we investigated the importance of ENaC and its positive regulator channel-activating protease 1 (CAP1/Prss8) in colon. Mice lacking the αENaC subunit in colonic superficial cells (Scnn1a(KO)) were viable, without fetal or perinatal lethality. Control mice fed a regular or low-salt diet had a significantly higher amiloride-sensitive rectal potential difference (∆PDamil) than control mice fed a high-salt diet. In Scnn1a(KO) mice, however, this salt restriction-induced increase in ∆PDamil did not occur, and the circadian rhythm of ∆PDamil was blunted. Plasma and urinary sodium and potassium did not change with regular or high-salt diets or potassium loading in control or Scnn1a(KO) mice. However, Scnn1a(KO) mice fed a low-salt diet lost significant amounts of sodium in their feces and exhibited high plasma aldosterone and increased urinary sodium retention. Mice lacking the CAP1/Prss8 in colonic superficial cells (Prss8(KO)) were viable, without fetal or perinatal lethality. Compared with controls, Prss8(KO) mice fed regular or low-salt diets exhibited significantly reduced ∆PDamil in the afternoon, but the circadian rhythm was maintained. Prss8(KO) mice fed a low-salt diet also exhibited sodium loss through feces and higher plasma aldosterone levels. Thus, we identified CAP1/Prss8 as an in vivo regulator of ENaC in colon. We conclude that, under salt restriction, activation of the renin-angiotensin-aldosterone system in the kidney compensated for the absence of ENaC in colonic surface epithelium, leading to colon-specific pseudohypoaldosteronism type 1 with mineralocorticoid resistance without evidence of impaired potassium balance.

A tumor-specific cellular environment at the brain invasion border of adamantinomatous craniopharyngiomas.

Craniopharyngiomas (CP) are benign epithelial tumors of the sellar region and can be clinicopathologically distinguished into adamantinomatous (adaCP) and papillary (papCP) variants. Both subtypes are classified according to the World Health Organization grade I, but their irregular digitate brain infiltration makes any complete surgical resection difficult to obtain. Herein, we characterized the cellular interface between the tumor and the surrounding brain tissue in 48 CP (41 adaCP and seven papCP) compared to non-neuroepithelial tumors, i.e., 12 cavernous hemangiomas, 10 meningiomas, and 14 metastases using antibodies directed against glial fibrillary acid protein (GFAP), vimentin, nestin, microtubule-associated protein 2 (MAP2) splice variants, and tenascin-C. We identified a specific cell population characterized by the coexpression of nestin, MAP2, and GFAP within the invasion niche of the adamantinomatous subtype. This was especially prominent along the finger-like protrusions. A similar population of presumably astroglial precursors was not visible in other lesions under study, which characterize them as distinct histopathological feature of adaCP. Furthermore, the outer tumor cell layer of adaCP showed a distinct expression of MAP2, a novel finding helpful in the differential diagnosis of epithelial tumors in the sellar region. Our data support the hypothesis that adaCP, unlike other non-neuroepithelial tumors of the central nervous system, create a tumor-specific cellular environment at the tumor-brain junction. Whether this facilitates the characteristic infiltrative growth pattern or is the consequence of an activated Wnt signaling pathway, detectable in 90% of these tumors, will need further consideration.

Demonstration of an interferon gamma-dependent tumor surveillance system in immunocompetent mice.

This study demonstrates that endogenously produced interferon gamma (IFN-gamma) forms the basis of a tumor surveillance system that controls development of both chemically induced and spontaneously arising tumors in mice. Compared with wild-type mice, mice lacking sensitivity to either IFN-gamma (i.e., IFN-gamma receptor-deficient mice) or all IFN family members (i.e., Stat1-deficient mice) developed tumors more rapidly and with greater frequency when challenged with different doses of the chemical carcinogen methylcholanthrene. In addition, IFN-gamma-insensitive mice developed tumors more rapidly than wild-type mice when bred onto a background deficient in the p53 tumor-suppressor gene. IFN-gamma-insensitive p53(-/-) mice also developed a broader spectrum of tumors compared with mice lacking p53 alone. Using tumor cells derived from methylcholanthrene-treated IFN-gamma-insensitive mice, we found IFN-gamma's actions to be mediated at least partly through its direct effects on the tumor cell leading to enhanced tumor cell immunogenicity. The importance and generality of this system is evidenced by the finding that certain types of human tumors become selectively unresponsive to IFN-gamma. Thus, IFN-gamma forms the basis of an extrinsic tumor-suppressor mechanism in immunocompetent hosts.

Mediastinal lymph node clearance after docetaxel-cisplatin neoadjuvant chemotherapy is prognostic of survival in patients with stage IIIA pN2 non-small-cell lung cancer: a multicenter phase II trial.

PURPOSE: A multicenter, phase II trial investigated the efficacy and toxicity of neoadjuvant docetaxel-cisplatin in locally advanced non-small-cell lung cancer (NSCLC) and examined prognostic factors for patients not benefiting from surgery. PATIENTS AND METHODS: Ninety patients with previously untreated, potentially operable stage IIIA (mediastinoscopically pN2) NSCLC received three cycles of docetaxel 85 mg/m2 day 1 plus cisplatin 40 mg/m2 days 1 and 2, with subsequent surgical resection. RESULTS: Administered dose-intensities were docetaxel 85 mg/m2/3 weeks (range, 53 to 96) and cisplatin 95 mg/m2/3 weeks (range, 0 to 104). The 265 cycles were well tolerated, and the overall response rate was 66% (95% confidence interval [CI], 55% to 75%). Seventy-five patients underwent tumor resection with positive resection margin and involvement of the uppermost mediastinal lymph node in 16% and 35% of patients, respectively (perioperative mortality, 3%; morbidity, 17%). Pathologic complete response occurred in 19% of patients with tumor resection. In patients with tumor resection, downstaging to N0-1 at surgery was prognostic and significantly prolonged event-free survival (EFS) and overall survival (OS; P =.0001). At median follow-up of 32 months, the median EFS and OS were 14.8 months (range, 2.4 to 53.4) and 33 months (range, 2.4 to 53.4), respectively. Local relapse occurred in 27% of patients with tumor resection, with distant metastases in 37%. Multivariate analyses identified mediastinal clearance (hazard ratio, 0.22; P =.0003) and complete resection (hazard ratio, 0.26; P =.0006) as strongly prognostic for increased survival. CONCLUSION: Neoadjuvant docetaxel-cisplatin is effective and tolerable in stage IIIA pN2 NSCLC. Resection is recommended only for patients with mediastinal downstaging after chemotherapy.