The potential impact of Covid-19 on the capacity of routine laboratory tests to detect heparin-induced thrombocytopenia.

In Covid-19, anticoagulation with heparin is often administered to prevent or treat thromboembolic events. Heparin-induced thrombocytopenia (HIT) is a severe complication of heparin treatment, caused by heparin-dependent, platelet activating anti-platelet factor 4 (PF4)/heparin antibodies. Diagnosis of HIT is based on the combination of clinical parameters, allowing to determine the pretest probability, and laboratory testing for anti-PF4/heparin antibodies and confirmatory functional assays, such as the heparin-induced platelet activation (HIPA) test.We report the case of a patient with severe Covid-19 pneumonia requiring ECMO treatment, who developed recurrent clotting of the ECMO filter and a drop in platelet count under heparin treatment. He was therefore suspected to have HIT and the anticoagulation was switched to argatroban. Despite high clinical probability and high titres of anti-PF4/heparin antibodies, the functional HIPA test was negative. Nevertheless, argatroban was continued rather than to reinstate anticoagulation with heparin. Reevaluation 7 days later then demonstrated a strongly positive functional HIPA test and confirmed the diagnosis of HIT. Under anticoagulation with argatroban the patient gradually improved and was finally weaned off the ECMO.In conclusion, this case highlights the critical importance of clinical judgement, exploiting the 4 T score, given that Covid-19 patients may present a different pattern of routine laboratory test results in HIT diagnostics. The possibility of a false negative HIPA test has to be considered, particularly in early phases of presentation. In cases of a discrepancy with high clinical probability of HIT and/or high titre anti-PF4/heparin antibodies despite a negative HIPA test, a reevaluation within 3 to 5 days after the initial test should be considered in order to avoid precipitant reestablishment of unfractionated heparin, with potentially fatal consequences.

Implications of early respiratory support strategies on disease progression in critical COVID-19: a matched subanalysis of the prospective RISC-19-ICU cohort.

BACKGROUND: Uncertainty about the optimal respiratory support strategies in critically ill COVID-19 patients is widespread. While the risks and benefits of noninvasive techniques versus early invasive mechanical ventilation (IMV) are intensely debated, actual evidence is lacking. We sought to assess the risks and benefits of different respiratory support strategies, employed in intensive care units during the first months of the COVID-19 pandemic on intubation and intensive care unit (ICU) mortality rates. METHODS: Subanalysis of a prospective, multinational registry of critically ill COVID-19 patients. Patients were subclassified into standard oxygen therapy ≥10 L/min (SOT), high-flow oxygen therapy (HFNC), noninvasive positive-pressure ventilation (NIV), and early IMV, according to the respiratory support strategy employed at the day of admission to ICU. Propensity score matching was performed to ensure comparability between groups. RESULTS: Initially, 1421 patients were assessed for possible study inclusion. Of these, 351 patients (85 SOT, 87 HFNC, 87 NIV, and 92 IMV) remained eligible for full analysis after propensity score matching. 55% of patients initially receiving noninvasive respiratory support required IMV. The intubation rate was lower in patients initially ventilated with HFNC and NIV compared to those who received SOT (SOT: 64%, HFNC: 52%, NIV: 49%, p = 0.025). Compared to the other respiratory support strategies, NIV was associated with a higher overall ICU mortality (SOT: 18%, HFNC: 20%, NIV: 37%, IMV: 25%, p = 0.016). CONCLUSION: In this cohort of critically ill patients with COVID-19, a trial of HFNC appeared to be the most balanced initial respiratory support strategy, given the reduced intubation rate and comparable ICU mortality rate. Nonetheless, considering the uncertainty and stress associated with the COVID-19 pandemic, SOT and early IMV represented safe initial respiratory support strategies. The presented findings, in agreement with classic ARDS literature, suggest that NIV should be avoided whenever possible due to the elevated ICU mortality risk.

Indicators of external ventricular drainage-related infections--a retrospective observational study.

BACKGROUND: External ventricular drainage (EVD) is frequently used in different groups of patients in neurocritical care. Despite the frequent use of EVD, no consensus regarding the diagnosis of EVD-related infection currently exists, and diagnosis is commonly based on criteria for the diagnosis of non-EVD-related CNS infections. This study evaluates the diagnostic accuracy of clinical and laboratory parameters for the prediction of EVD-related infection in patients with proven EVD-related infection. METHODS: In two tertiary care centers, data on EVD insertions were matched with a microbiologic database of cultured microorganisms and positive Gram stains of cerebrospinal fluid (CSF) to identify patients with EVD-related infections. Available clinical data and results of blood tests and CSF analysis were retrospectively collected. Predefined potential clinical and laboratory predictors of EVD-related infection were compared between three time points: at the time EVD insertion and 48 h before and at the time of occurrence of EVD-related infection. RESULTS: Thirty-nine patients with EVD-associated infection defined by positive CSF culture or positive CSF Gram stains and concomitant clinical signs of infection were identified. At the time of infection, a significantly higher incidence of abnormal temperature, high respiratory rate, and a slightly but significantly higher incidence of decreased mental state were observed. The assessed blood and CSF parameters did not significantly differ between the different assessment time points. CONCLUSIONS: Our analysis of 39 patients with culture positive EVD-related infection showed that commonly used clinical and laboratory parameters are not reliable infection predictors.

Metabolically programmed iron chelators.

Extensive structure activity relationship (SAR) studies focused on the desferrithiocin [DFT, (S)-4,5-dihydro-2-(3-hydroxy-2-pyridinyl)-4-methyl-4-thiazolecarboxylic acid] pharmacophore have led to three different DFT analogs being evaluated clinically for the treatment of iron overload diseases, for example, thalassemia. The SAR work revealed that the lipophilicity of a ligand, as determined by its partition between octanol and water, logP(app), could have a profound effect on the drug's iron clearing efficiency (ICE), organ distribution, and toxicity profile. While within a given structural family the more lipophilic a chelator the better the ICE, unfortunately, the more lipophilic ligands are often more toxic. Thus, a balance between lipophilicity, ICE, and toxicity must be achieved. In the current study, we introduce the concept of 'metabolically programmed' iron chelators, that is, highly lipophilic, orally absorbable, effective deferration agents which, once absorbed, are quickly converted to their nontoxic, hydrophilic counterparts.

Oxygen kinetics during 6-minute walk tests in patients with cardiovascular and pulmonary disease.

BACKGROUND: The 6-Minute Walk Test (6MWT) is representative of daily-life activities and reflects the functional capacity of patients. The change of oxygen uptake (VO2) in the initial phase of low-intensity exercise (VO2 kinetics) can be used to assess submaximal exercise performance of patients.The objective of the following study was to analyse VO2 kinetics in patients with different pulmonary and cardiovascular diseases. In addition, we investigated the extent to which VO2 kinetics at the onset of the 6MWT were associated with exercise capacity, morbidity and mortality. METHODS: VO2 kinetics of 204 patients and 16 healthy controls were obtained using mobile telemetric cardiopulmonary monitoring during a 6MWT. A new mean response time (MRT) index (wMRT) was developed to quantify VO2 kinetics by correcting MRT for work rate. The differences in wMRT between disease categories as well as the association between wMRT and patients' exercise capacity and outcome - time to hospitalization/death- were tested. RESULTS: The assessment of a robust wMRT was feasible in 86% (244/284) patients. wMRT was increased in patients compared to healthy controls (p <0.001). wMRT was largest in patients with pulmonary arterial hypertension (PAH). There were significant associations between wMRT and exercise capacity in all patients. High wMRT was found to be associated with a high rate of death and re-hospitalization in patients with CHF (p = 0.024). In patients with pulmonary diseases and pulmonary hypertension wMRT was not associated with outcome (p = 0.952). CONCLUSIONS: Submaximal exercise performance of patients is reduced. O2 kinetics at the onset of exercise are associated with exercise capacity in all patients. wMRT was found to be an important prognostic factor in patients with congestive heart failure (CHF), but not with pulmonary diseases.

Hemodynamic assessment of critically ill patients using a miniaturized transesophageal echocardiography probe.

INTRODUCTION: Hemodynamic management in intensive care patients guided by blood pressure and flow measurements often do not sufficiently reveal common hemodynamic problems. Trans-esophageal echocardiography (TEE) allows for direct measurement of cardiac volumes and function. A new miniaturized probe for TEE (mTEE) potentially provides a rapid and simplified approach to monitor cardiac function. The aim of the study was to assess the feasibility of hemodynamic monitoring using mTEE in critically ill patients after a brief operator training period. METHODS: In the context of the introduction of mTEE in a large ICU, 14 ICU staff specialists with no previous TEE experience received six hours of training as mTEE operators. The feasibility of mTEE and the quality of the obtained hemodynamic information were assessed. Three standard views were acquired in hemodynamically unstable patients: 1) for assessment of left ventricular function (LV) fractional area change (FAC) was obtained from a trans-gastric mid-esophageal short axis view, 2) right ventricular (RV) size was obtained from mid-esophageal four chamber view, and 3) superior vena cava collapsibility for detection of hypovolemia was assessed from mid-esophageal ascending aortic short axis view. Off-line blinded assessment by an expert cardiologist was considered as a reference. Inter-rater agreement was assessed using Chi-square tests or correlation analysis as appropriate. RESULTS: In 55 patients, 148 mTEE examinations were performed. Acquisition of loops in sufficient quality was possible in 110 examinations for trans-gastric mid-esophageal short axis, 118 examinations for mid-esophageal four chamber and 125 examinations for mid-esophageal ascending aortic short axis view. Inter-rater agreement (Kappa) between ICU mTEE operators and the reference was 0.62 for estimates of LV function, 0.65 for RV dilatation, 0.76 for hypovolemia and 0.77 for occurrence of pericardial effusion (all P<0.0001). There was a significant correlation between the FAC measured by ICU operators and the reference (r=0.794, P (one-tailed)<0.0001). CONCLUSIONS: Echocardiographic examinations using mTEE after brief bed-side training were feasible and of sufficient quality in a majority of examined ICU patients with good inter-rater reliability between mTEE operators and an expert cardiologist. Further studies are required to assess the impact of hemodynamic monitoring by mTEE on relevant patient outcomes.

The prevalence of cardiac complications and their impact on outcomes in patients with non-traumatic subarachnoid hemorrhage.

Subarachnoid hemorrhage (SAH) is a serious condition, and a myocardial injury or dysfunction could contribute to the outcome. We assessed the prevalence and prognostic impact of cardiac involvement in a cohort with SAH. This is a prospective observational multicenter study. We included 192 patients treated for non-traumatic subarachnoid hemorrhage. We performed ECG recordings, echocardiographic examinations, and blood sampling within 24 h of admission and on days 3 and 7 and at 90 days. The primary endpoint was the evidence of cardiac involvement at 90 days, and the secondary endpoint was to examine the prevalence of a myocardial injury or dysfunction. The median age was 54.5 (interquartile range [IQR] 48.0-64.0) years, 44.3% were male and the median World Federation of Neurological Surgeons (WFNS) score was 2 (IQR 1-4). At day 90, 22/125 patients (17.6%) had left ventricular ejection fractions ≤ 50%, and 2/121 patients (1.7%) had evidence of a diastolic dysfunction as defined by mitral peak E-wave velocity by peak e' velocity (E/e') > 14. There was no prognostic impact from echocardiographic evidence of cardiac complications on neurological outcomes. The overall prevalence of cardiac dysfunction was modest. We found no demographic or SAH-related factors associated with 90 days cardiac dysfunction.

Risk assessment in the first fifteen minutes: a prospective cohort study of a simple physiological scoring system in the emergency department.

INTRODUCTION: The survival of patients admitted to an emergency department is determined by the severity of acute illness and the quality of care provided. The high number and the wide spectrum of severity of illness of admitted patients make an immediate assessment of all patients unrealistic. The aim of this study is to evaluate a scoring system based on readily available physiological parameters immediately after admission to an emergency department (ED) for the purpose of identification of at-risk patients. METHODS: This prospective observational cohort study includes 4,388 consecutive adult patients admitted via the ED of a 960-bed tertiary referral hospital over a period of six months. Occurrence of each of seven potential vital sign abnormalities (threat to airway, abnormal respiratory rate, oxygen saturation, systolic blood pressure, heart rate, low Glasgow Coma Scale and seizures) was collected and added up to generate the vital sign score (VSS). VSSinitial was defined as the VSS in the first 15 minutes after admission, VSSmax as the maximum VSS throughout the stay in ED. Occurrence of single vital sign abnormalities in the first 15 minutes and VSSinitial and VSSmax were evaluated as potential predictors of hospital mortality. RESULTS: Logistic regression analysis identified all evaluated single vital sign abnormalities except seizures and abnormal respiratory rate to be independent predictors of hospital mortality. Increasing VSSinitial and VSSmax were significantly correlated to hospital mortality (odds ratio (OR) 2.80, 95% confidence interval (CI) 2.50 to 3.14, P < 0.0001 for VSSinitial; OR 2.36, 95% CI 2.15 to 2.60, P < 0.0001 for VSSmax). The predictive power of VSS was highest if collected in the first 15 minutes after ED admission (log rank Chi-square 468.1, P < 0.0001 for VSSinitial;,log rank Chi square 361.5, P < 0.0001 for VSSmax). CONCLUSIONS: Vital sign abnormalities and VSS collected in the first minutes after ED admission can identify patients at risk of an unfavourable outcome.

Desferrithiocin analogue iron chelators: iron clearing efficiency, tissue distribution, and renal toxicity.

The current solution to iron-mediated damage in transfusional iron overload disorders is decorporation of excess unmanaged metal, chelation therapy. The clinical development of the tridentate chelator deferitrin (1, Table 1) was halted due to nephrotoxicity. It was then shown by replacing the 4'-(HO) of 1 with a 3,6,9-trioxadecyloxy group, the nephrotoxicity could be ameliorated. Further structure-activity relationship studies have established that the length and the position of the polyether backbone controlled: (1) the ligand's iron clearing efficiency (ICE), (2) chelator tissue distribution, (3) biliary ferrokinetics, and (4) tissue iron reduction. The current investigation compares the ICE and tissue distribution of a series of (S)-4,5-dihydro-2-[2-hydroxy-4-(polyether)phenyl]-4-methyl-4-thiazolecarboxylic acids (Table 1, 3-5) and the (S)-4,5-dihydro-2-[2-hydroxy-3-(polyether)phenyl]-4-methyl-4-thiazolecarboxylic acids (Table 1, 8-10). The three most effective polyether analogues, in terms of performance ratio (PR), defined as mean ICE(primate)/ICE(rodent), are 3 (PR 1.1), 8, (PR 1.5), and 9, now in human trials, (PR 2.2). At the onset of the clinical trial on 9, no data were available for ligand 3 or 8. This is unfortunate, as 3 has many advantages over 9, e.g., the ICE of 3 in rats is 2.5-fold greater than that of 9 and analogue 3 achieves very high levels in the liver, pancreas, and heart, the organs most affected by iron overload. Finally, the impact of 3 on the urinary excretion of kidney injury molecule-1 (Kim-1), an early diagnostic biomarker for monitoring acute kidney toxicity, has been carried out in rats; no evidence of nephrotoxicity was found. Overall, the results suggest that 3 would be a far superior clinical candidate to 9.

Six-minute walk test enhanced by mobile telemetric cardiopulmonary monitoring.

BACKGROUND: The 6-min walk test (6MWT) is frequently used to assess overall cardiopulmonary fitness and to predict outcome, but it yields little diagnostic information. Portable telemetric devices allow performing the 6MWT with real-time cardiopulmonary monitoring. OBJECTIVES: The study was designed to analyze feasibility, safety and clinical usefulness of a mobile cardiopulmonary monitoring (MOB)-enhanced 6MWT. METHODS: From August 2003 to June 2007, 261 consecutive patients with chronic lung and/or heart disease as well as healthy controls underwent MOB-enhanced 6MWTs. A subgroup of 33 individuals had the test done with and without cardiopulmonary monitoring on independent days. RESULTS: No test-related adverse events occurred throughout the study. Whether the 6MWT was done without or with cardiopulmonary monitoring (n = 33) did not significantly influence the walking distance (WD: 528 ± 183 vs. 525 ± 192 m; nonsignificant). Fifty-nine percent (155/261) of the patients fulfilled the maximal test criteria. Distinct disease-specific exercise response patterns as well as treatable co-pathologies were observed. The validity of response patterns was better in case of a maximal test. CONCLUSION: An MOB-enhanced 6MWT is feasible within daily routine and safe in patients with various diseases. It does not negatively affect WD. MOB is a valuable tool to identify factors limiting exercise in patients irrespective of their underlying disease.

Desferrithiocin analogue uranium decorporation agents.

PURPOSE: Previous systematic structure-activity studies of the desferrithiocin (DFT) platform have allowed the design and synthesis of analogues and derivatives of DFT that retain the exceptional iron-clearing activity of the parent, while eliminating its adverse effects. We hypothesized that a similar approach could be adopted to identify DFT-related analogues that could effectively decorporate uranium. MATERIALS AND METHODS: The decorporation properties of nine DFT-related analogues were determined in a bile duct-cannulated rat model. Diethylenetriaminepentaacetic acid (DTPA) served as a positive control. Selected ligands also underwent multiple and delayed dosing regimens. Uranium excretion in urine and bile or stool was determined by inductively coupled plasma mass spectroscopy (ICP-MS); tissue levels of uranium were also assessed. RESULTS: The two best clinical candidates are (S)-4,5-dihydro-2-[2-hydroxy-4-(3,6,9-trioxadecyloxy)phenyl]-4-methyl-4-thiazolecarboxylic acid [(S)-4'-(HO)-DADFT-PE (9)], with a 57% reduction in kidney uranium levels on oral (p.o.) administration and (S)-4,5-dihydro-2-[2-hydroxy-3-(3,6,9-trioxadecyloxy)phenyl]-4-methyl-4-thiazolecarboxylic acid [(S)-3'-(HO)-DADFT-PE (10)], with a 62% renal reduction on p.o. administration. The majority of the metal excretion promoted by these analogues is in the bile, thus further reducing kidney actinide exposure. CONCLUSIONS: While 9 administered p.o. or subcutaneously (s.c.) immediately post-metal is an effective decorporation agent, withholding the dose (s.c.) until 4 h reduced the activity of the compound. Conversion of 9 to its isopropyl ester may circumvent this issue.

Impact of the 3,6,9-trioxadecyloxy group on desazadesferrithiocin analogue iron clearance and organ distribution.

The impact of introducing a 3,6,9-trioxadecyloxyl group at various positions of the desazadesferrithiocin (DADFT) aromatic ring on iron clearance and organ distribution is described. Three DADFT polyethers are evaluated: (S)-4,5-dihydro-2-[2-hydroxy-4-(3,6,9-trioxadecyloxy)phenyl]-4-methyl-4-thiazolecarboxylic acid [(S)-4'-(HO)-DADFT-PE, 3], (S)-4,5-dihydro-2-[2-hydroxy-5-(3,6,9-trioxadecyloxy)phenyl]-4-methyl-4-thiazolecarboxylic acid [(S)-5'-(HO)-DADFT-PE, 6], and (S)-4,5-dihydro-2-[2-hydroxy-3-(3,6,9-trioxadecyloxy)phenyl]-4-methyl-4-thiazolecarboxylic acid [(S)-3'-(HO)-DADFT-PE, 9]. The iron-clearing efficiency (ICE) in rodents and primates is shown to be very sensitive to which positional isomer is evaluated, as is the organ distribution in rodents. The polyethers had uniformly higher ICEs than their corresponding parent ligands in rodents, consistent with in vivo ligand-serum albumin binding studies. Ligand 9 is the most active polyether analogue in rodents and is also very effective in primates, suggesting a higher index of success in humans. In addition, this analogue is also shown to clear more iron in the urine of the primates than many of the other chelators. If this trend were also observed in patients, it would facilitate iron-balance studies in a clinical setting.

Analysis with respect to instrumental variables for the exploration of microarray data structures.

BACKGROUND: Evaluating the importance of the different sources of variations is essential in microarray data experiments. Complex experimental designs generally include various factors structuring the data which should be taken into account. The objective of these experiments is the exploration of some given factors while controlling other factors. RESULTS: We present here a family of methods, the analyses with respect to instrumental variables, which can be easily applied to the particular case of microarray data. An illustrative example of analysis with instrumental variables is given in the case of microarray data investigating the effect of beverage intake on peripheral blood gene expression. This approach is compared to an ANOVA-based gene-by-gene statistical method. CONCLUSION: Instrumental variables analyses provide a simple way to control several sources of variation in a multivariate analysis of microarray data. Due to their flexibility, these methods can be associated with a large range of ordination techniques combined with one or several qualitative and/or quantitative descriptive variables.

The design, synthesis, and evaluation of organ-specific iron chelators.

A series of iron chelators, three (S)-4,5-dihydro-2-(2-hydroxyphenyl)-4-methyl-4-thiazolecarboxylic acid (DADFT) and three (S)-4,5-dihydro-2-(2-hydroxyphenyl)-4-thiazolecarboxylic acid (DADMDFT) analogues are synthesized and assessed for their lipophilicity (log Papp), iron-clearing efficiency (ICE) in rodents and iron-loaded primates (Cebus apella), toxicity in rodents, and organ distribution in rodents. The results lead to a number of generalizations useful in chelator design strategies. In rodents, while log Papp is a good predictor of a chelator's ICE, chelator liver concentration is a better tool. In primates, log Papp is a good predictor of ICE, but only when comparing structurally very similar chelators. There is a profound difference in toxicity between the DADMDFT and DADFT series: DADMDFTs are less toxic. Within the DADFT family of ligands, the more lipophilic ligands are generally more toxic. Lipophilicity can have a profound effect on ligand organ distribution, and ligands can thus be targeted to organs compromised in iron overload disease, for example, the heart.

(S)-4,5-dihydro-2-(2-hydroxy-4-hydroxyphenyl)-4-methyl-4-thiazolecarboxylic acid polyethers: a solution to nephrotoxicity.

Previous studies revealed that within a family of ligands the more lipophilic chelators have better iron-clearing efficiency. The larger the log P(app) value of the compound, the better the iron-clearing efficiency. What is also clear from the data is that although the relative effects of log P(app) changes are essentially the same through different families, there are differences in absolute value between families. However, there also exists a second, albeit somewhat more disturbing, relationship. In all sets of ligands, the most lipophilic chelator is always the most toxic. The current study focuses on designing ligands that balance the lipophilicity/toxicity problem while iron-clearing efficiency is maintained. Earlier studies with (S)-4,5-dihydro-2-(2-hydroxy-4-methoxyphenyl)-4-methyl-4-thiazolecarboxylic acid [(S)-4'-(CH(3)O)-DADFT, 6] indicated that this methyl ether was a ligand with excellent iron-clearing efficiency in both rodents and primates; however, it was too toxic. On the basis of this finding, a less lipophilic, more water-soluble ligand than 6 was assembled, (S)-4,5-dihydro-2-[2-hydroxy-4-(3,6,9-trioxadecyloxy)phenyl]-4-methyl-4-thiazolecarboxylic acid [(S)-4'-(HO)-DADFT-PE, 11], a polyether analogue, along with its ethyl and isopropyl esters. The parent polyether and its isopropyl and ethyl esters were all shown to be highly efficient iron chelators in both rodents and primates. A comparison of 11 in rodents with the desferrithiocin analogue (S)-4,5-dihydro-2-(2,4-dihydroxyphenyl)-4-methyl-4-thiazolecarboxylic acid [(S)-4'-(HO)-DADFT, 1] revealed the polyether to be more tolerable, achieving higher concentrations in the liver and significantly lower concentrations in the kidney. The lower renal drug levels are in keeping with the profound difference in the architectural changes seen in the kidney of rodents given 1 versus those treated with 11.

Sarcoidosis is a multisystem disorder with variable prognosis--information for treating physicians.

Sarcoidosis is a chronic granulomatous inflammatory disorder of unknown origin with heterogeneous outcome. In most cases the disease is self-limited, others progress or die from organ involvement, which is often associated with extensive scarring. Although often the primarily affected sites are the lungs and thoracic lymph nodes patients with sarcoidosis must be staged for multiorgan involvement. Modern treatment strategies appraise a critical awareness for the side-effect-ratio of long-term medication. A main interest of research is to identify those patients with unfavourable outcome.

Polyamine-vectored iron chelators: the role of charge.

The utility of polyamines as vectors for the intracellular transport of iron chelators is further described. Consistent with earlier results with polyamine analogues, these studies underscore the importance of charge in the design of polyamine-vectored chelators. Four polyamine conjugates are synthesized, two of terephthalic acid [N(1)-(4-carboxy)benzoylspermine (7) and its methyl ester (6)] and two of (S)-2-(2,4-dihydroxyphenyl)-4,5-dihydro-4-methyl-4-thiazolecarboxylic acid [(S)-4'-(HO)-DADFT] [(S)-4,5-dihydro-2-[2-hydroxy-4-(12-amino-5,9-diazadodecyl-oxy)phenyl]-4-methyl-4-thiazolecarboxylic acid (10) and its ethyl ester (9)]. These four molecules were evaluated in murine leukemia L1210 cells for their impact on cell proliferation (48- and 96-h IC(50) values), their ability to compete with spermidine for the polyamine transport apparatus (K(i)), and their intracellular accumulation. The data revealed that when neutral molecules (cargo fragments) were fixed to the polyamine vector, the conjugates competed well with spermidine for transport and were accumulated intracellularly to millimolar levels. However, this was not the case when the cargo fragments were negatively charged. Metabolic studies of the polyamine-vectored (S)-4'-(HO)-DADFTs in rodents indicated that not only did the expected deaminopropylation step occur, but also a surprisingly high level of oxidative deamination at the terminal primary nitrogens took place. Finally, the iron-clearing efficiency of the (S)-4'-(HO)-DADFT conjugates was determined in a bile-duct-cannulated rodent model. Attaching the ligand to a polyamine vector had a profound effect on increasing the iron-clearing efficiency of this chelator relative to its parent drug.

Partition-variant desferrithiocin analogues: organ targeting and increased iron clearance.

Altering the lipophilicity (log P(app)) of desferrithiocin analogues can change the organ distribution of the chelators and lead to enhanced iron clearance. For example, alkylation of (S)-2-(2,4-dihydroxyphenyl)-4,5-dihydro-4-methyl-4-thiazolecarboxylic acid [(S)-4'-(HO)-DADFT] and its analogues to more lipophilic compounds, such as (S)-4,5-dihydro-2-(2-hydroxy-4-methoxyphenyl)-4-methyl-4-thiazolecarboxylic acid [(S)-4'-(CH3O)-DADFT], provides ligands that achieved between a 3- and 8-fold increase in chelator concentrations in the heart, liver, and pancreas (the organs most at risk in iron-overload disease) of treated rodents. The 4'-O-methylated compounds are demethylated to their hydroxylated counterparts in rodents; furthermore, this O-demethylation takes place in both rodent and human liver microsomes. The relationship between chelator lipophilicity and iron-clearing efficacy in the iron-overloaded Cebus apella primate is further underscored by a comparison of the iron-clearing efficiency of (S)-2-(2,3-dihydroxyphenyl)-4,5-dihydro-4-methyl-4-thiazolecarboxylic acid [(S)-3'-(HO)-DADFT] and its 3'-(CH3O) counterpart. Finally, these DFT analogues are shown to be both inhibitors of the iron-mediated oxidation of ascorbate as well as effective radical scavengers.

Methoxylation of desazadesferrithiocin analogues: enhanced iron clearing efficiency.

The impact of altering the octanol-water partition properties (log P) of analogues of desazadesferrithiocin, (S)-4,5-dihydro-2-(2-hydroxyphenyl)-4-methyl-4-thiazolecarboxylic acid, on the ligands' iron clearing properties is described. Increasing chelator lipophilicity can both substantially augment iron clearing efficiency in Cebus apella primates as well as alter the mode of iron excretion, favoring fecal over urinary output. The complications of iron overload are often associated with the metal's interaction with hydrogen peroxide, generating hydroxyl radicals (Fenton chemistry) and, ultimately, other related deleterious species. In fact, some iron chelators actually promote this chemistry. All of the compounds synthesized and tested in the current study are shown to be both inhibitors of the iron-mediated oxidation of ascorbate, thus removing the metal from the Fenton cycle, and effective radical scavengers.

Desferrithiocin analogue based hexacoordinate iron(III) chelators.

Traditional thinking has been that hexacoordinate Fe(III) ligands are more effective at preventing iron's interactions with reactive oxygen species, most particularly the Fe(II)-mediated reduction of hydrogen peroxide to the hydroxyl radical (i.e., Fenton chemistry), than are ligands of lower denticity. Thus, a hexacoordinate derivative of the well-characterized tricoordinate ligand (S)-2-(2,4-dihydroxyphenyl)-4,5-dihydro-4-thiazolecarboxylic acid [4'-(HO)-DADMDFT], (S,S)-1,11-bis[5-(4-carboxy-4,5-dihydrothiazol-2-yl)-2,4-dihydroxyphenyl]-4,8-dioxaundecane, was designed with the aid of a molecular modeling program and synthesized. Evaluations both in vitro and in vivo were carried out to determine whether there is any advantage, at the level of prevention of Fenton chemistry, radical trapping, or iron clearance, to constructing a desferrithiocin-based hexacoordinate analogue. The hexacoordinate analogue was more effective at preventing the iron-mediated oxidation of ascorbate at low ligand/metal ratios than was its tricoordinate parent and can function as an excellent radical scavenger. At equivalent iron binding doses in the bile duct cannulated rodent, oral administration of the tricoordinate ligand was 3-fold more effective than was po administration of the hexacoordinate derivative. However, sc administration of the hexacoordinate derivative resulted in an efficiency that was 3 times greater than that of the tricoordinate chelator. Unfortunately, the rodent findings were not substantiated in the primates. The hexacoordinate ligand was only about one-half as efficient as its tricoordinate parent when administered sc. Owing to these results, po dosing was not attempted. Thus, there appears to be no overall advantage to coupling two molecules of 4'-(HO)-DADMDFT to afford a hexacoordinate derivative.