Kyle P. Kokko

Design, synthesis, and evaluation of the antipsychotic potential of orally bioavailable neurotensin (8–13) analogues containing non-natural arginine and lysine residues

Neurotensin (NT) and its active fragment NT(8-13) elicit behavioral responses typical of clinically used antipsychotic drugs when administered directly to the brain. However, limited peptide stability and oral bioavailability have prevented these compounds from being developed as relevant pharmaceuticals. Recently, our laboratory designed and studied a first-generation NT(8-13) derivative, KK13, that elicited key pharmacokinetic and behavioral responses typical of clinically used antipsychotic drugs when administered to rats parenterally. This compound was the basis for the rational design of a series of second-generation NT(8-13) analogues (KH1-KH30) studied in this paper. Initial screening of these analogues for CNS activity by monitoring hypothermia induction after peripheral administration defined several compounds (KH11, KH24, KH26, and KH28-KH30) that warranted further investigation. Each compound maintained binding affinity for NTR(1), however, only KH24, KH26, and KH28 (as well as KK13) elicited significant hypothermic responses after oral administration. Of these, KH28 demonstrated an oral activity 3-fold greater than any other analogue; hence it was further characterized in a series of rat behavioral assays. KH28 attenuated d-amphetamine induced hyperlocomotion, a hallmark of current clinically effective antipsychotic drugs, after both IP and oral administration. In addition, tolerance to the compound did not develop after repeated daily dosing, as measured by hypothermic induction as well as attenuation of d-amphetamine induced hyperlocomotion. Finally, KH28 did not produce catalepsy, a deleterious side-effect elicited by classical antipsychotic drugs. KH28 is considered to be an ideal compound for further development as a potential novel antipsychotic.

Monitoring neurotensin[8–13] degradation in human and rat serum utilizing matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

A method was developed to quantify neurotensin (NT) fragment [8-13] and a novel NT[8-13] derivative, KK1, in human and rat serum utilizing matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). The method allows for simultaneous quantification of the major NT[8-13] metabolite, NT[9-13] (according to molecular mass), and detection of the major KK1 metabolite, KK1M (according to molecular mass). The degradation rates of NT[8-13] and KK1 were calculated to be 24.1+/-1.0 and 193+/-8min in human serum and 5.90+/-0.22 and 153+/-4min in rat serum, respectively. The method utilizes a novel sample drying technique and spectrum acquisition protocol. In addition, an internal standard dissimilar in structure to the analytes was used. This method may be broadly applicable to the quantification of NT[8-13] and other peptide analogues of varying structure.

Randomized prospective clinical trial comparing reamer irrigator aspirator (RIA) to standard reaming (SR) in both minimally injured and multiply injured patients with closed femoral shaft fractures treated with reamed intramedullary nailing (IMN)

PURPOSE To investigate whether inflammatory markers are improved among patients with traumatic femur fractures who undergo RIA reamed intramedullary nailing (IMN) prior to fixation when compared to patients treated with standard reamed (SR) IMN. METHODS A prospective, randomized, single-blind trial was conducted on patients who had a closed femoral shaft fracture amenable to reamed IMN. Patients were randomized to undergo IMN with standard reaming or IMN with the RIA in a 1:1 ratio. Patients were stratified by Injury Severity Score (ISS) and by presence or absence of chest injury with AIS > 3. Patients had blood samples and bronchioalveolar lavage samples taken at specified time points pre- and postoperatively. Specimens from SR and RIA cohorts were compared for the presence of IL-2, IL-6, IL-8, TNF, and IL-10 in plasma and IL-1b and IL-8 in bronchioalveolar lavage (BAL) samples to determine the relationship between inflammatory markers and intramedullary reaming. RESULTS Nineteen consecutive patients participated in the study with 9 assigned in the RIA group and 10 in SR group. Significant differences existed for ISS between SR and RIA groups (p=0.04). Bronchial lavage data showed no statistical significant differences when RIA and standard reamers were compared and when ISS >16 and <16 were compared, however there were differences for the bronchial IL-8 change when those with chest injury were compared to those without chest injury. Plasma samples showed a trend towards increased IL-6 and IL-10 levels after reaming consistent with the second hit impact. A trend towards higher levels for IL-6 in the SR group was noted at 24 hours post-operatively whereas the IL-10 levels at the post-reaming time point were higher in the RIA group. CONCLUSIONS This prospective study of reamer type indicates that RIA may be protective of systemic inflammation. This is supported by data showing decreased levels of IL-8 in the bronchial washings and increased level of IL-10 in the serum. Reaming and intramedullary fixation may cause an increase in IL-6 levels regardless of reamer type. Further investigations with a larger cohort of patients are desirable.

In vivo behavioral effects of stable, receptor-selective neurotensin[8-13] analogues that cross the blood-brain barrier

A set of neurotensin[8-13] (NT[8-13]) analogues (KK1-19) has been evaluated in various pre-clinical assays relevant for further development of these compounds as potential antipsychotics. Initial screening of these compounds for induction of hypothermia following systemic (I.V.) injection in rats, an indirect method commonly utilized to measure the central nervous system (CNS) activity of NT[8-13] analogues, identified three peptides, KK1, KK13 and KK14, capable of crossing the blood-brain barrier (BBB). KK1 features 2(S)-azido-7-aminoheptanoic acid (AAHA) in the Arg(8) position and represents the first monosubstituted NT[8-13] analogue that crosses the BBB. KK13 and KK14 both feature AAHA in the Arg(8) position and tert-Leu in the Ile(12) position while KK14 includes a Trp substituted for Tyr(11). When I.P. administered, only the latter two analogues induced a significant hypothermic response. KK13 (1mg/kg) inhibited amphetamine-induced hyperlocomotion after I.P. injection; this assay is highly predictive for potential antipsychotics. Chronic dosing (5mg/kg) of this compound over 5 consecutive days failed to induce hypothermic tolerance while the same dose failed to induce measurable catalepsy. KK13 is thus the first NT[8-13] analogue described to date that demonstrates inhibition of amphetamine-induced hyperlocomotion without inducing catalepsy while maintaining day-to-day hypothermic potency.

Selectivity enhancement induced by substitution of non-natural analogues of arginine and lysine in arginine-based thrombin inhibitors.

Seven non-natural analogues of arginine and lysine have been substituted in an established arginine-based thrombin inhibitor. Four of the new compounds exhibited significant thrombin inhibition (K(i)s 0.53-3.95 microM) and were subsequently tested for selectivity against trypsin. The two best compounds gave selectivity ratios of 962 and 525 (trypsin/thrombin), improving upon the parent compound.

Asymmetric Synthesis of ω‐Bromo‐2(S)‐Methyl Acids as Precursors for Novel Arginine, Lysine, and Mercapto Residues

Abstract A series of ω‐bromo‐2(S)‐methyl acids has been synthesized as precursors of novel arginine (Arg), lysine (Lys), and mercapto analogues. These intermediates contain α‐methyl groups and are designed to mask the N‐terminal amine when incorporated in pharmaceutically relevant peptides.

Personal economic impact of performing elective Saturday hand surgery

Background: The economic impact of performing elective hand surgery on Saturdays has yet to be studied. The purpose of this study was to evaluate patient preferences and factors for Saturday hand surgery and to analyze the personal economic and societal costs regarding missed days of work for elective hand surgery. Methods: An anonymous quality improvement survey was distributed to 125 consecutive patients who were planning to undergo elective outpatient hand surgery. Demographics included age, gender, zip code, education, occupation, income level, and interest in Saturday hand surgery. IBM-SPSS Statistics 20 for Windows (SPSS, Chicago, IL) was used for data analysis. Results: Seventy-eight (62.4%) patients responded they would want elective hand surgery performed on a Saturday. Of those who reported income (n=66), the average daily salary was estimated to be

Biomechanical and range of motion analysis of two proximally fixed locking plate systems for fixation of proximal humeral fractures

269.50. If these patients had been given the opportunity to have Saturday hand surgery, a total of

In Vitro Analysis of Stable, Receptor-Selective Neurotensin[8−13] Analogues

17,787 in lost income or paid leave could have been saved. We did not identify any significant factors that correlated with a patient’s decision to undergo elective Saturday hand surgery. Conclusions: Over half (62.4%) of our respondents would request Saturday elective hand surgery if offered at our institution. If 62.4% of patients requiring hand surgery at our institution were to elect for Saturday surgery, we estimated a savings of over

METHODS AND COMPOSITIONS FOR BONE HEALING BY PERIOSTIN

100,000 in lost wages or paid leave annually. This study shows that there is a patient population that would be interested in elective Saturday surgery, and providing availability could help to take care of this market.