Optimization of percutaneous biopsy for diagnosis and pretreatment risk assessment of neuroblastoma.

BACKGROUND: Image-guided percutaneous core needle biopsy (PCNB) is increasingly utilized to diagnose solid tumors. The objective of this study is to determine whether PCNB is adequate for modern biologic characterization of neuroblastoma. PROCEDURE: A multi-institutional retrospective study was performed by the Pediatric Surgical Oncology Research Collaborative on children with neuroblastoma at 12 institutions over a 3-year period. Data collected included demographics, clinical details, biopsy technique, complications, and adequacy of biopsies for cytogenetic markers utilized by the Children's Oncology Group for risk stratification. RESULTS: A total of 243 children were identified with a diagnosis of neuroblastoma: 79 (32.5%) tumor excision at diagnosis, 94 (38.7%) open incisional biopsy (IB), and 70 (28.8%) PCNB. Compared to IB, there was no significant difference in ability to accurately obtain a primary diagnosis by PCNB (95.7% vs 98.9%, P = .314) or determine MYCN copy number (92.4% vs 97.8%, P = .111). The yield for loss of heterozygosity and tumor ploidy was lower with PCNB versus IB (56.1% vs 90.9%, P < .05; and 58.0% vs. 88.5%, P < .05). Complications did not differ between groups (2.9 % vs 3.3%, P = 1.000), though the PCNB group had fewer blood transfusions and lower opioid usage. Efficacy of PCNB was improved for loss of heterozygosity when a pediatric pathologist evaluated the fresh specimen for adequacy. CONCLUSIONS: PCNB is a less invasive alternative to open biopsy for primary diagnosis and MYCN oncogene status in patients with neuroblastoma. Our data suggest that PCNB could be optimized for complete genetic analysis by standardized protocols and real-time pathology assessment of specimen quality.

Sex-specific differences in emphysema using a murine antisense oligonucleotide model of α-1 antitrypsin deficiency.

α-1 Antitrypsin (AAT) deficiency is the leading genetic cause of emphysema; however, until recently, no genuine animal models of AAT deficiency existed, hampering the development of new therapies. This shortcoming is now addressed by both AAT-null and antisense oligonucleotide mouse models. The goal of this study was to more fully characterize the antisense oligonucleotide model. Both liver AAT mRNA and serum AAT levels were lower in anti-AAT versus control oligonucleotide-treated mice after 6, 12, and 24 wk. Six and twelve weeks of anti-AAT oligonucleotide therapy induced emphysema that was worse in female than male mice: mean linear intercept 73.4 versus 62.5 µm (P = 0.000003). However, at 24 wk of treatment, control oligonucleotide-treated mice also developed emphysema. After 6 wk of therapy, anti-AAT male and female mice demonstrated a similar reduction serum AAT levels, and there were no sex or treatment-specific alterations in inflammatory, serine protease, or matrix metalloproteinase mRNAs, with the exception of chymotrypsin-like elastase 1 (Cela1), which was 7- and 9-fold higher in anti-AAT versus control male and female lungs, respectively, and 1.6-fold higher in female versus male anti-AAT-treated lungs (P = 0.04). While lung AAT protein levels were reduced in anti-AAT-treated mice, lung AAT mRNA levels were unaffected. These findings are consistent with increased emphysema susceptibility of female patients with AAT-deficiency. The anti-AAT oligonucleotide model of AAT deficiency is useful for compartment-specific, in vivo molecular biology, and sex-specific studies of AAT-deficient emphysema, but it should be used with caution in studies longer than 12-wk duration.

The consequence of undertreatment of patients treated with breast conserving therapy for ductal carcinoma in-situ.

BACKGROUND: The use of clinical features to allocate adjuvant therapy in the treatment of ductal carcinoma in situ with breast-conserving therapy remains controversial. METHODS: A review of patients with ductal carcinoma in situ treated with breast-conserving therapy was performed. The recurrence rate was examined in relation to patient age, tumor characteristics, Van Nuys Prognostic Index, and the receipt of prescribed adjuvant therapies. RESULTS: Six percent of patients (17 of 294) had developed local recurrences after a median follow-up period of 63 months. Fifty-nine percent of patients (91 of 154) with estrogen receptor-positive tumors did not receive prescribed tamoxifen. Thirty-one percent of patients (45 of 147) with Van Nuys Prognostic Index scores ≥7 did not receive recommended radiation therapy. Receipt of prescribed adjuvant therapy did not result in a decrease in the rate of local recurrence. Patient age was the only factor associated with local recurrence on univariate but not on multivariate analysis (P = .374). CONCLUSIONS: A low rate of local recurrence was achieved despite a large number of patients' not receiving prescribed adjuvant therapies.

The significance of lobular carcinoma in situ and atypical lobular hyperplasia of the breast.

BACKGROUND: The significance of lobular neoplasia (LN), lobular carcinoma in situ (LCIS) and atypical lobular hyperplasia (ALH)) found at core needle biopsy (CNB) of the breast remains uncertain. There is a consistent risk of underestimating malignancy after the diagnosis of LN on CNB. The aim of this study was to determine if patients with a CNB result of LN need surgical excision. METHODS: Patients were identified by searching the institutions pathology database for the terms "lobular carcinoma in situ" and "atypical lobular hyperplasia" over 20 years. Excluded from this study were those with core needle biopsy (CNB) results of ductal carcinoma in situ, atypical ductal hyperplasia, radial scar, or papilloma. Upgrade was defined as final surgical pathology of invasive carcinoma and/or ductal carcinoma in situ that was directly correlated to the site of the initial biopsy containing LN. RESULTS: LN was found at CNB in 285 patients, and 71 % (n = 201) had subsequent surgical excisions. All patients with pleomorphic LCIS (pLCIS) underwent surgical excision. Following patients with pLCIS, patients with the diagnosis of LCIS were most likely to undergo surgical excision (80 %). Final pathology of the surgically excised specimens confirmed LN in 72 % (n = 144). Also, 13 % (n = 26) of the operated patients were upgraded to malignancy, including 8 % of ALH and 19 % of LCIS cases. CONCLUSION: This is the largest series of surgical excisional pathology following LN on CNB ever reported. The likelihood of finding malignancy at surgical excision after CNB showing LN was 13 %. Patients with the diagnosis of LN on CNB should be considered for surgical excision.

The value of mammography within 1 year of conservative surgery for breast cancer.

INTRODUCTION: Guidelines for screening mammography have been established by numerous medical societies. Guidelines have not been established for follow-up mammography for patients who have been treated with breast-conserving surgery. Many radiologists recommend mammography of the treated breast 6 months after completion of treatment. The purpose of this study was to determine the value of interval mammography. METHODS: Patients were identified by searching the breast cancer database for the diagnoses of ductal carcinoma in situ, infiltrating ductal carcinoma and infiltrating lobular carcinoma. Postoperative mammogram dates and results were obtained. Patients with mammography within 8 months of surgery were included in the study. RESULTS: Ductal carcinoma in situ, infiltrating ductal carcinoma, and infiltrating lobular carcinoma were found in 1,000 patients who underwent breast-conserving surgery, and 789 patients had complete mammographic follow-up data available. Postoperative interval mammography was performed in 169 patients (21 %), including 23 patients who had preradiation mammography. Ninety percent of the interval mammograms were BI-RADS 1 to 3 and 10 % were BI-RADS 4 or 5. Two cancers were found on interval mammography (1.2 % of 169) and 4 of 620 (0.6 %) patients who did not have interval mammography were found to have malignancy within 1 year of surgery (1.2 % vs. 0.6 %, P = 0.614). The use of interval mammography was not related to the mammographic findings at diagnosis. Interval mammography did not affect local and distant disease-free survival. CONCLUSIONS: The likelihood of obtaining a significant finding on short interval follow-up mammography after conservative surgery for breast cancer is 1.2 %.

Laparoscopic distal pancreatectomy.

BACKGROUND: Laparoscopic management of distal pancreatic malignancies has been slow to gain a foothold in all but high-volume tertiary referral centers. The aim of this study was to assess the safety and outcomes of laparoscopic distal pancreatectomy (LDP) performed in a low-volume community hospital by a diverse group of surgeons, none of whom have a specialized laparoscopic background. METHODS: We conducted a retrospective review of all patients who underwent open distal pancreatectomies (ODPs) and LDPs between August 2001 and June 2008. Data included type of surgery, open versus laparoscopy, demographics, operative time, blood loss, length of hospital stay, histopathologic diagnosis, postoperative complications, American Society of Anesthesiologists score, and mortality. RESULTS: Twenty-seven patients with pancreatic masses underwent distal pancreatic resection during the study period. Fifty-nine percent (n = 16) underwent LDP, and 41% (n = 11) underwent ODP. Mean patient age was 66 y (range, 40 to 86) for the LDP group and 62 (range, 40 to 84) for the ODP group. Mean operative time was 231 min (range, 195 to 305) for LDP and 240 (range, 150 to 210) for the ODP technique. Mean length of stay for LDP and ODP was 8 (range, 3 to 22) and 12 d (range, 5 to 2), respectively. Morbidity was 25% (n = 4) in the LDP group and 36% (n = 4) in the ODP group. None of the differences between the LDP and ODP groups were statistically significant. No mortalities occurred in either group. CONCLUSION: This study supports the idea that LDP can be safely and effectively performed by any surgeon comfortable with basic laparoscopy and may not require specialized training or a specialized center. Further data are required to make more definitive conclusions.

In vivo structure-activity relationship study of dorsomorphin analogues identifies selective VEGF and BMP inhibitors.

The therapeutic potential of small molecule signaling inhibitors is often limited by off-target effects. Recently, in a screen for compounds that perturb the zebrafish embryonic dorsoventral axis, we identified dorsomorphin, the first selective inhibitor of bone morphogenetic protein (BMP) signaling. Here we show that dorsomorphin has significant "off-target" effects against the VEGF (vascular endothelial growth factor) type-2 receptor (Flk1/KDR) and disrupts zebrafish angiogenesis. Since both BMP and VEGF signals are known to be involved in vascular development, we sought to determine whether dorsomorphin's antiangiogenic effects are due to its impact on the BMP or VEGF signals through the development of analogues that target BMP but not VEGF signaling and vice versa. In a structure-activity relationship (SAR) study of dorsomorphin analogues based primarily on their effects on live zebrafish embryos, we identified highly selective and potent BMP inhibitors as well as selective VEGF inhibitors. One of the BMP inhibitors, DMH1, which exclusively targets the BMP but not the VEGF pathway, dorsalized the embryonic axis without disrupting the angiogenic process, demonstrating that BMP signaling was not involved in the angiogenic process. This is one of the first full-scale SAR studies performed in vertebrates and demonstrates the potential of zebrafish as an attractive complementary platform for drug development that incorporates an assessment of in vivo bioactivity and selectivity in the context of a living organism.

Design and synthesis of isoform-selective phospholipase D (PLD) inhibitors. Part II. Identification of the 1,3,8-triazaspiro[4,5]decan-4-one privileged structure that engenders PLD2 selectivity.

This Letter describes the synthesis and structure-activity relationships (SAR) of isoform-selective PLD inhibitors. By virtue of the installation of a 1,3,8-triazaspiro[4,5]decan-4-one privileged structure, PLD inhibitors with nanomolar potency and an unprecedented 40-fold selectivity for PLD2 over PLD1 were developed. Interestingly, SAR for this diverged from our earlier efforts, and dual PLD1/2 inhibitors were also discovered within this series.

Design and synthesis of isoform-selective phospholipase D (PLD) inhibitors. Part I: Impact of alternative halogenated privileged structures for PLD1 specificity.

This Letter describes the synthesis and structure-activity-relationships (SAR) of isoform-selective PLD inhibitors. By virtue of the installation of alternative halogenated piperidinyl benzimidazolone privileged structures, in combination with a key (S)-methyl group, novel PLD inhibitors with low nM potency and unprecedented levels of PLD1 isoform selectivity (approximately 1700-fold) over PLD2 were developed.

Total synthesis of Ciliatamides A-C: stereochemical revision and the natural product-guided synthesis of unnatural analogs.

The first total synthesis of Ciliatamides A-C was completed, leading to a revision of the reported stereochemistry from (S,S) to the (R,R) enantiomers. Due to the expedited route, a library of over 50 unnatural ciliatamide analogs was also prepared.

Allosteric modulation of kinases and GPCRs: design principles and structural diversity.

Allosteric binding sites, as opposed to traditional orthosteric binding sites, offer unparalleled opportunities for drug discovery by providing high levels of selectivity, mimicking physiological conditions, affording fewer side effects because of desensitization/downregulation, and engendering ligands with chemotypes divergent from orthosteric ligands. For kinases, allosteric mechanisms described to date include alteration of protein kinase conformation blocking productive ATP binding which appear 'ATP competitive' or blocking kinase activation by conformational changes that are 'ATP non-competitive'. For GPCRs, allosteric mechanisms impart multiple modes of target modulation (positive allosteric modulation (PAM), negative allosteric modulation (NAM), neutral cooperativity, partial antagonism (PA), allosteric agonism and allosteric antagonism). Here, we review recent developments in the design principles and structural diversity of allosteric ligands for kinases and GPCRs.

The design of inhibitors for medicinally relevant metalloproteins.

A number of metalloproteins are important medicinal targets for conditions ranging from pathogenic infections to cancer. Many but not all of these metalloproteins contain a zinc(II) ion in the protein active site. Small-molecule inhibitors of these metalloproteins are designed to bind directly to the active site metal ions. In this review several metalloproteins of interest are discussed, including matrix metalloproteinases (MMPs), histone deacetylases (HDACs), anthrax lethal factor (LF), and others. Different strategies that have been employed to design effective inhibitors against these proteins are described, with an effort to highlight the strengths and drawbacks of each approach. An emphasis is placed on examining the bioinorganic chemistry of these metal active sites and how a better understanding of the coordination chemistry in these systems may lead to improved inhibitors. It is hoped that this review will help inspire medicinal, biological, and inorganic chemists to tackle this important problem by considering all aspects of metalloprotein inhibitor design.

A new role for old ligands: discerning chelators for zinc metalloproteinases.

In an effort to identify promising non-hydroxamate inhibitors of matrix metalloproteinases (MMPs) several new zinc-binding groups (ZBGs) based on pyridine-derived or aza-macrocycle chelators have been examined. Fluorescence-based enzyme assays have been used to determine the IC50 values for these ZBGs against MMP-1, MMP-3, and anthrax lethal factor (LF). Many of these ligands were found to be remarkably potent, with IC50 values as much as 185-fold lower than that found for acetohydroxamic acid. These ligands are proposed to be more selective "warheads" for the inhibition of metalloenzymes that contain Zn2+ versus other metal ions at their active site.

Heterocyclic zinc-binding groups for use in next-generation matrix metalloproteinase inhibitors: potency, toxicity, and reactivity.

In an effort to improve the zinc-chelating portion of matrix metalloproteinase (MMP) inhibitors, we have developed a family of heterocyclic zinc-binding groups (ZBGs) as alternatives to the widely used hydroxamic acid moiety. Elaborating on findings from an earlier report, we performed in vitro inhibition assays with recombinant MMP-1, MMP-2, and in a cell culture assay using neonatal rat cardiac fibroblast cells. In both recombinant and cell culture assays, the new ZBGs were found to be effective inhibitors, typically 10-100-fold more potent than acetohydroxamic acid. The toxicity of these chelators was examined by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium salt cytotoxicity assays, which demonstrate that most of these compounds are nontoxic at concentrations of almost 100 microM. To address the possible interaction of sulfur-containing ZBGs with biological reductants, the reactivity of these chelators with 5,5'-dithiobis(2-nitrobenzoic acid) was examined. Finally, thione ZBGs were shown to be effective inhibitors of cell invasion through an extracellular matrix membrane. The data presented herein suggest these heterocyclic ZBGs are potent, nontoxic, and biocompatible compounds that show promise for incorporation into a new family of MMP inhibitors.

Synthesis, structure and spectroscopy of new thiopyrone and hydroxypyridinethione transition-metal complexes.

The coordination chemistry of several O,S mixed donor ligands, namely thiopyrone and hydroxypyridinethione chelators, with a variety of middle and late first-row transition-metal ions is described. Complexes of 3-hydroxy-2-methyl-4-thiopyrone (thiomaltol) with cobalt(II), copper(II) and zinc(II); 3-hydroxy-1,2-dimethyl-4(1H)-pyridinethione (3,4-HOPTO) with iron(III), nickel(II), copper(II) and zinc(II); and 3-hydroxy-1-methyl-2(1H)-pyridinethione (3,2-HOPTO) with iron(III), nickel(II), copper(II) and zinc(II) have been synthesized and characterized. The structures, absorbance spectroscopy, cyclic voltammetry and superconducting quantum interferometer device (SQUID) measurements of selected metal complexes, as well as ligand protonation constants, are reported. Most of the metal complexes show coordination geometries indicative of a strong trans influence by the O,S chelators. The data presented herein provide the most detailed study of the transition-metal coordination chemistry of both thiopyrone and hydroxypyridinethione O,S donor ligands to date, and provide the basis for the investigation of these ligands in realm of biological inorganic chemistry.

Addressing lead toxicity: complexation of lead(II) with thiopyrone and hydroxypyridinethione O,S mixed chelators.

The lead(II) ion is regarded as a serious environmental contaminant. A considerable need exists to develop selective ligands for remediation of this metal ion. Herein, the coordination chemistry of lead(II) is investigated with three O,S donor ligands: thiomaltol, 3-hydroxy-1-methyl-2(1H)-pyridinethione (3,2-HOPTO), and 3-hydroxy-1,2-dimethyl-4(1H)-pyridinethione (3,4-HOPTO). The X-ray structures of [Pb(thiomaltolato)(2)] and [Pb(3,4-HOPTO)(2)] have been solved, revealing the expected 4-coordinate geometries. Electronic spectra have been obtained for the lead(II) complexes with all three ligands. Preliminary solution studies show that the thiomaltol ligand binds lead(II) preferentially over magnesium(II) and calcium(II); however, [Pb(thiomaltolato)(2)] is not stable in the presence of 1 equiv of EDTA. Tetradentate ligands derived from these O,S chelators are expected to generate higher affinity ligands for lead(II) sequestration.

New beginnings for matrix metalloproteinase inhibitors: identification of high-affinity zinc-binding groups.

In an effort to identify promising non-hydroxamate inhibitors of matrix metalloproteinases (MMPs), several new zinc-binding groups (ZBGs) based on pyrone, pyrothione, hydroxypyridinone, and hydroxypyridinethione chelators have been examined. Structural studies with tris(pyrazolyl)borate model complexes show that these ligands bind to the MMP active site zinc(II) ion in a bidentate fashion, similar to that found with hydroxamate-based inhibitors. Fluorescence- and colorimetric-based enzyme assays have been used to determine the IC50 values for these ZBGs against MMP-3; mixed O,S-donor ligands were found to be remarkably potent, with IC50 values as much as 700-fold lower than that found for acetohydroxamic acid. Inhibitory activity was found to parallel metal binding affinity as determined in titrations with model complexes. These results demonstrate that MPIs based on new ZBGs are feasible and may indeed improve the overall performance of inhibitors designed against these important medicinal targets.