Synthesis and physico‐chemical properties of peptides in soil humic substances

Abstract: Soil humic substances (HS) are heterologous, polydispersive, and multi‐functional organometallic macromolecules ubiquitous in soils and sediments. They are key players in the maintenance of the belowground ecosystems and in the bioavailability of both organic and inorganic contaminants. It is widely assumed that the peptidic substructures of HS are readily degraded and therefore do not contribute significantly to interactions with contaminants such as toxic metals. To investigate the turnover of humified peptides, laboratory soil aging experiments were conducted with ¹³C‐glucose or ¹⁵N‐nitrate for 8.5 months. Evidence for random‐coil peptidic structures in the labeled HS was obtained from 2‐D nuclear magnetic resonance (NMR), pyrolysis gas chromatography‐mass spectrometry (pyro‐GC‐MS), and circular dichroism data. Interaction of metals with the peptidic carbonyls of labeled HS was rationalized from the solid‐state NMR data. Detailed ¹³C and ¹⁵N labeling patterns of amino acid residues in the acid hydrolysates of HS acquired from NMR and GC‐MS revealed two pools of peptides, i.e. one extant (unlabeled) and the other, newly humified with little isotopic scrambling (fully labeled). The persistence of pre‐existing peptidic structures indicates their resistance to degradation while the presence of fully labeled peptidic amino acids suggests wholesale incorporation of newly synthesized peptides into HS. These findings are contrary to the general notion that humified peptides are readily degraded.     

Pharmacodynamic differences between species exemplified by the novel anticancer agent CHS 828

When a candidate drug enters clinical trials, decisions regarding dosing are mainly based on animal data. Occasionally, toxicity problems are faced in the clinic because of unexpected species differences in pharmacokinetics or pharmacodynamics between humans and preclinical species. Fludarabine and topotecan are examples of such drugs. In the first clinical trials of the new agent CHS 828, the maximum tolerated dose was reached earlier than expected from animal data. This paper discusses the issue of species differences in the development of anticancer drugs, and preclinical models for detection and quantification of such differences. Pharmacokinetic and hematological toxicity data of CHS 828 from studies in rats and humans are presented. In vitro sensitivity to CHS 828 and some established cytotoxic agents was measured in lymphocytes from humans and rats and in a panel of human and rodent cell‐lines. 10–100 times higher CHS 828 exposure was tolerated by rats than by patients. In both in vitro cell systems, CHS 828 showed higher potency in human cells compared to rodent cells. A species difference was evident also for fludarabine, but not for doxorubicin and cisplatin. CHS 828 pharmacokinetics were similar across species. In conclusion, the lower tolerance of CHS 828 in humans than in rats could be detected in vitro in cultures of peripheral lymphocytes. Preclinical studies of species differences could help the interpretation of in vivo effect studies as well as the choice of starting dose for clinical trials. We suggest peripheral lymphocytes from different species as a potential model system for such studies. Drug Dev. Res. 61:218–226, 2004. © 2004 Wiley‐Liss, Inc.     

Inhibition of β‐amyloid toxicity by short peptides containing N‐methyl amino acids

Abstract: Single N‐methyl amino acid‐containing peptides related to the central hydrophobic region β_16–20 (Lys‐Leu‐Val‐Phe‐Phe) of the β‐amyloid protein are able to reduce the cytotoxicity of natural β_1–42 in PC12 cell cultures. N‐methyl phenylalanine analogs yield statistically significant increments in cell viability (Student's t‐test < 0.01%) and are nontoxic in the same assay. These promising results indicate that these peptide molecules could be a starting point for the development of potential therapeutic compounds for the treatment of Alzheimer's disease.     

Compliance with consensus recommendations for systemic therapy is associated with improved survival of women with node-negative breast cancer.

PURPOSE: The impact of consensus recommendations for systemic therapy on outcome of disease is unclear. We evaluated if compliance with guidelines for systemic adjuvant treatment is associated with improved survival of women with node-negative breast cancer. PATIENTS AND METHODS: The study population included women diagnosed with invasive node-negative breast cancer in Québec, Canada, in 1988 to 1989, 1991 to 1992, and 1993 to 1994. Information was collected by chart review, linkage with administrative databases, and queries to attending physicians. Guidelines from the 1992 St Gallen conference were used as standard of care. Survival was estimated by Kaplan-Meier and Cox proportional hazards analyses. RESULTS: Among 1,541 women, 358 died before December 1999. Median follow-up was 6.8 years. Seven-year event-free and overall survivals were 66% and 81%, respectively. Survival was 88%, 84%, and 74% in women at minimal, moderate, or high risk of recurrence. Virtually all women at minimal risk were treated according to the consensus (98.4% of 370). In comparison, adjusted hazard ratios of death were 1.0 (95% CI, 0.6 to 1.7) and 2.3 (95% CI, 1.3 to 4.0) among women at moderate risk treated according to the consensus or not, respectively. Among women at high risk, adjusted hazard ratios of death were 2.0 (95% CI, 1.4 to 2.8) and 2.7 (95% CI, 1.9 to 3.9), respectively. Both risk category (P <.0005) and compliance with guidelines (P <.0005) were independent significant predictors of survival. CONCLUSION: Treatment according to consensus recommendations is associated with improved survival of women with breast cancer in the community. Promoting the adoption of guidelines for treatment is an effective strategy for disease control.     

Quantified relationship between cellular radiosensitivity, DNA repair defects and chromatin relaxation: a study of 19 human tumour cell lines from different origin.

BACKGROUND AND PURPOSE: There is still confusion in the choice of the molecular assays to predict the radiation response of human cells. The case of tumours appears to be particularly complex, may be because of their instability and heterogeneity. The aim of this study was to investigate quantitatively the relationships between DNA double-strand breaks (DSB) repair, chromatin relaxation and cellular radiosensitivity. Nineteen human tumour cell lines, representing a large spectrum of radiation responses and tissues, were examined. MATERIALS AND METHODS: Intrinsic radiosensitivity was quantified with surviving fraction at 2 Gy (SF2) as an endpoint. Standard and modified pulsed-field gel electrophoresis techniques were employed to assess DSB repair rate and chromatin relaxation. A cell-free assay was chosen to estimate DSB repair activity, independently of chromatin impairment. RESULTS AND CONCLUSIONS: Surviving fraction at 2 Gy (SF2) decreases linearly with the amount of unrepaired DSB and the extent of chromatin relaxation: one additional unrepaired DSB per cell or 1% chromatin decondensation produce a loss of about 1.5% surviving fraction. However, all the cell lines did not obey both correlations, suggesting that DSB repair and chromatin impairments contribute separately to increase the severity of DNA damage involved in cell lethality. Four cell lines groups showing different DSB repair and/or chromatin impairments were defined. Cell lines exhibiting both DSB repair defect and chromatin relaxation are the most radiosensitive.     

Antifolate resistance in a HeLa cell line associated with impaired transport independent of the reduced folate carrier.

Prior studies from this laboratory documented the prevalence of methotrexate (MTX) transport activity with a low pH optimum in human solid tumor cell lines. In HeLa cells, this low pH activity has high affinity for pemetrexed [PMX (Alimta)] and is reduced folate carrier (RFC)-independent because it is not diminished in a RFC-null subline (R5). R5 cells also have residual transport activity, with high specificity for PMX, at neutral pH. In the current study, a R5 subline, R1, was selected under MTX selective pressure at a modest reduction in pH. There was markedly decreased MTX and PMX transport at both pH 5.5 and pH 7.4. When MTX was removed, there was a slow return of transport activity, and when MTX was added back, there was loss of transport at both pH values within 8 weeks. In R1 cells, there was a marked decrease in accumulation of PMX, MTX, and folic acid along with a decrease in growth inhibition by these and other antifolates that require a facilitative process to gain entry into cells. These data demonstrate that (i) RFC-independent transport in HeLa cells at low and neutral pH contributes to antifolate activity (in particular, to PMX activity) and can be diminished by antifolate selective pressure and (ii) the loss of these activities results in marked resistance to PMX, an agent for which there is little or no loss of activity when transport mediated by RFC is abolished. These observations suggest that transport activity in RFC-null HeLa R5 cells at neutral and low pH may reflect the same carrier-mediated process.     

Characterization and multiparameter analysis of visual adverse events in irofulven single-agent phase I and II trials.

PURPOSE: Irofulven (6-hydroxymethylacylfulvene) is a novel agent, derived from illudin S, with potent apoptotic effects in preclinical models. In the Phase I trial evaluating intermittent weekly schedules, visual symptoms were dose limiting. The aim of this analysis was to better characterize the visual adverse events of irofulven and provide treatment guidelines. EXPERIMENTAL DESIGN: Clinical data from 277 patients entered in single-agent Phase I to II clinical trials who received irofulven on days 1 and 15 every 4 weeks; days 1, 8, and 15 every 4 weeks; or days 1 and 8 every 3 weeks were included in this multiparameter analysis. RESULTS: Overall, 74 patients (27%) experienced visual symptoms. The most frequently reported symptoms were flashing lights (12% of patients), blurred vision (9%), and photosensitivity (8%). Grade 3 toxicity was observed in 12 patients (4%). The incidence and severity of visual events were dose dependent, with no grade 3 visual events occurring at doses < or =0.50 mg/kg and grade 1 to 2 events in only 12% and 8% of patients, at doses of < or =0.50 mg/kg and < or =20 mg/m2, respectively. Grade 1 to 2 toxicity was reversible in most patients. Abnormal electroretinogram and abnormal visual fields were noted after irofulven treatment in 24 of 39 patients (62%) and 15 of 26 patients (58%), respectively. All but 1 patient who had electroretinogram assessment received doses >0.50 mg/kg. Clinical examination and visual field assessment were found to be better correlated with symptoms and appear to be more appropriate for surveillance of irofulven retinal symptoms than electroretinograms. CONCLUSIONS: On the basis of retained antitumor activity and reversibility of grade 1 and 2 visual symptoms at lower doses, it appears that an irofulven dose of < or =0.50 mg/kg or < or =20 mg/m2, not to exceed 50 mg in a single dose, given as a 30-minute infusion on days 1 and 8 every 3 weeks or days 1 and 15 every 4 weeks minimizes the frequency and severity of visual symptoms.     

Antibody-based profiling of the phosphoinositide 3-kinase pathway in clinical prostate cancer.

PURPOSE: As kinase inhibitors transition from the laboratory to patients, it is imperative to develop biomarkers that can be used in the clinic. The primary objectives are to identify patients most likely to benefit from molecularly targeted therapies and to document modulation of the drug target. Constitutive activation of the phosphoinositide 3-kinase (PI3K) pathway and its downstream effectors, as a result of PTEN loss or by other mechanisms, occurs in a high proportion of prostate cancers, making it an ideal template for the design of clinical trials involving PI3K pathway inhibitors. Prostate cancers also present unique organ-specific challenges, in that tumors are heterogeneous and diagnostic tissue is extremely limited. EXPERIMENTAL DESIGN: Working within these limitations, we have developed a set of immunohistochemical assays that define activation of the PI3K pathway in clinical samples. Results and CONCLUSIONS: Using both univariate and multivariate analyses, we show that loss of PTEN is highly correlated with the activation of AKT, and this, in turn, is associated with the phosphorylation of S6, one of its main effectors. These three antibodies are potentially able to define a molecular signature of PTEN loss and/or AKT pathway activation in prostate cancer.     

Bipartite parietal bone: A rare cause of plagiocephaly

A case of an infant with an asymmetrical head is presented. On clinical assessment the patient displayed features of deformational plagiocephaly. With the aid of three‐dimensional CT imaging of the skull, a bipartite parietal bone was diagnosed. The prevalence and possible aetiology of a bipartite parietal bone is discussed as well as a brief overview of the common causes of plagiocephaly.     

Incidence and prognostic value of respiratory events in acute leukemia.

Acute respiratory failure and infectious pneumonia are the major causes of death during induction chemotherapy of acute leukemia. However, the causes, incidence and prognostic value of all respiratory events (REs) occurring in this context have never been assessed prospectively. We recruited 65 consecutive patients with newly diagnosed acute leukemia into a 1-year prospective study (December 2000-November 2001) to evaluate the incidence and prognostic value of these events. REs were frequent: 38 were recorded in 30 patients. There was a significant relationship between REs and pre-existing respiratory disease and/or smoking. REs were caused by infection in 34% of cases, by an established cause other than infection in 42% and had an undetermined cause in 24%. Poor early outcome (death within 45 days of starting induction chemotherapy) in patients experiencing an RE was independently associated with a >25/min respiratory rate (P=0.003) and the nonachievement of complete remission (CR) (P<0.0001). Predictors of overall survival in the entire patient population were the absence of CR (P<0.0001), REs (P=0.02) and a > or =2 performance status (P=0.03). In conclusion, REs are frequent during induction chemotherapy of acute leukemia and represent an independent prognostic factor of poor outcome, regardless of their cause.