Probing the hydrophobic pocket of farnesyltransferase: aromatic substitution of CAAX peptidomimetics leads to highly potent inhibitors.

Cysteine farnesylation at the carboxylate terminal tetrapeptide CAAX of Ras protein is catalyzed by farnesyltransferase. This lipid modification is necessary for regulatory function of both normal and oncogenic Ras. The high frequency of Ras mutation in human cancers has prompted an intensive study on finding ways of controlling oncogenic Ras function. Inhibition of farnesyltransferase is among the most sought after targets for cancer chemotherapy. We report here the design, synthesis and biological characterization of a series of peptidomimetics as farnesyltransferase inhibitors. These compounds are extremely potent towards farnesyltransferase with IC50 values ranging from subnanomolar to low nanomolar concentrations. They have a high selectivity for farnesyltransferase over the closely related geranylgeranyltransferase-I. Structure-activity relationship studies demonstrated that a properly positioned hydrophobic group significantly enhanced inhibition potency, reflecting an improved complementarity to the large hydrophobic pocket in the CAAX binding site.

Centerfire frangible ammunition: wounding potential and other forensic concerns.

Recently developed frangible ammunition of copper particulate construction in .38 Special, 9 mm, and .223 calibers was evaluated for wounding performance by firing into pigs' heads. The ability to match fired bullets with the corresponding gun was also examined. Results showed that wounds caused by 9-mm and .38 Special frangible bullets were comparable in severity to those caused by regular service ammunition of the same caliber. The recovered 9-mm and .38 Special bullets demonstrated class characteristics but not the individual rifling marks necessary for bullet-to-gun matching. High-velocity .223-caliber rifle bullets fragmented extensively within target tissues, causing severe wounding. Radiologic examination of resulting wounds showed images strikingly similar to the lead "snowstorm" picture caused by high-velocity hunting ammunition.

Ras CAAX peptidomimetic FTI-277 selectively blocks oncogenic Ras signaling by inducing cytoplasmic accumulation of inactive Ras-Raf complexes.

Ras-induced malignant transformation requires Ras farnesylation, a lipid posttranslational modification catalyzed by farnesyltransferase (FTase). Inhibitors of this enzyme have been shown to block Ras-dependent transformation, but the mechanism by which this occurs remains largely unknown. We have designed FTI-276, a peptide mimetic of the COOH-terminal Cys-Val-Ile-Met of K-Ras4B that inhibited potently FTase in vitro (IC50 = 500 pM) and was highly selective for FTase over geranylgeranyltransferase I (GGTase I) (IC50 = 50 nM). FTI-277, the methyl ester derivative of FTI-276, was extremely potent (IC50 = 100 nM) at inhibiting H-Ras, but not the geranylgeranylated Rap1A processing in whole cells. Treatment of H-Ras oncogene-transformed NIH 3T3 cells with FTI-277 blocked recruitment to the plasma membrane and subsequent activation of the serine/threonine kinase c-Raf-1 in cells transformed by farnesylated Ras (H-RasF), but not geranylgeranylated, Ras (H-RasGG). FTI-277 induced accumulation of cytoplasmic non-farnesylated H-Ras that was able to bind Raf and form cytoplasmic Ras/Raf complexes in which Raf kinase was not activated. Furthermore, FTI-277 blocked constitutive activation of mitogen-activated protein kinase (MAPK) in H-RasF, but not H-RasGG, or Raf-transformed cells. FTI-277 also inhibited oncogenic K-Ras4B processing and constitutive activation of MAPK, but the concentrations required were 100-fold higher than those needed for H-Ras inhibition. The results demonstrate that FTI-277 blocks Ras oncogenic signaling by accumulating inactive Ras/Raf complexes in the cytoplasm, hence preventing constitutive activation of the MAPK cascade.

CAAX peptidomimetic FTI-244 decreases platelet-derived growth factor receptor tyrosine phosphorylation levels and inhibits stimulation of phosphatidylinositol 3-kinase but not mitogen-activated protein kinase.

Cysteine farnesylation of the Ras carboxyl terminal tetrapeptide CAAX motif (where C = cysteine, A = leucine, isoleucine, or valine, and X = methionine or serine) is required for Ras biological activity. In this report, we describe the effects of inhibitors of farnesyltransferase (FTase), the enzyme responsible for this lipid modification, on platelet-derived growth factor (PDGF) signaling in NIH-3T3 cells. In vitro, the CAAX peptidomimetic FTI-232 exhibits potent inhibition of FTase activity (IC50 = 150 nM) and its carboxyl-methylated counterpart, FTI-244, inhibits Ras processing in vivo. Treatment of NIH-3T3 cells with FTI-244 inhibits PDGF-induced DNA synthesis but not stimulation of mitogen-activated protein kinase (MAPK). However, FTI-244 significantly reduces PDGF-induced tyrosine phosphorylation levels of PDGF receptor (PDGFR) as well as its association with, and activation of, phosphatidylinositol-3-kinase (PI-3-K), a key enzyme in PDGF-induced mitogenesis.

A non-peptide mimetic of Ras-CAAX: selective inhibition of farnesyltransferase and Ras processing.

Cysteine farnesylation of the carboxyl-terminal tetrapeptide CAAX (C = Cys, A = Leu, Ile, or Val, X = Met or Ser) of the oncogene product Ras is required for its malignant transformation activity. As a consequence farnesyltransferase (FTase), the enzyme responsible for this lipid modification, has become one of the most sought-after targets for anticancer drug development. We have recently designed peptide mimics of the COOH-terminal Cys-Val-Ile-Met of KB-Ras where the dipeptide Val-Ile was replaced by aminobenzoic acid derivatives. Although these peptidomimetics are potent inhibitors of FTase in vitro, they retain several undesirable peptide features that hamper their use in vivo. We report here the design, synthesis, and biological activity of the first non-peptide mimetics of CAAX where the tripeptide AAX was replaced by biphenyl derivatives. (R)-4-[N-(3-mercapto-2-aminopropyl)]amino-3'- carboxybiphenyl, where the cysteine is linked to the biphenyl derivative through a secondary amine, contains no amino acids, lacks peptidic features, and has no hydrolyzable bonds. This peptidomimetic is a potent inhibitor of FTase in vitro (IC50 = 50-150 nM) and disrupts Ras processing in whole cells. Furthermore, this non-peptide mimetic of CAAX is highly selective for FTase (666-fold) relative to the closely related geranylgeranyltransferase I. This selectivity is also respected in vivo since the processing of Ras but not the geranylgeranylated Rap1A was disrupted in whole cells. Structure activity relationship studies revealed that FTase recognition and inhibitory potency of CAAX peptidomimetics require free thiol and carboxylate groups separated by a hydrophobic moiety, and that precise positioning of these functional groups must correspond to that of the parent CAAX. The true CAAX peptidomimetic described in this manuscript has several desirable features for further development as a potential anticancer agent. It is not metabolically inactivated by FTase, does not require a pro-drug strategy for inhibition in vivo, and is selective for farnesylation relative to geranylgeranylation.

Patterns of facial resuscitation injury in infancy.

Cardiopulmonary resuscitation (CPR)-related artifacts in pediatric rescue that have the potential for serious complications in surviving patients have been well described in the medical literature. Medically trivial soft-tissue injuries, especially of the face and neck, carry predominantly forensic significance and have received less attention. We describe such injuries in nine of 25 consecutive cases of infants who received CPR, and correlate those injuries with specific rescue maneuvers. Techniques for effective investigation and interpretation of such injuries are suggested.

Blunt trauma of the abdomen in children.

Abdominal trauma in children is usually due to motor vehicle accidents or falls, but child abuse is also a common cause. Injuries to the small intestine are particularly worrisome because symptoms may not appear for hours, days, or even weeks. This can cause significant difficulty for the investigator trying to determine when an injury occurred, or who may have caused it.

Disaster planning for air crashes. A retrospective analysis of Delta Airlines flight 191.

The effectiveness of previously untested disaster plans was demonstrated during the aftermath of the crash of Delta Airlines Flight 191 at D/FW International Airport on Friday, August 2, 1985. These plans, in effect for years and subjected to periodic review, were as yet untried before this first disaster at D/FW International since its opening. This paper outlines the disaster plans, the actual problems encountered, the logistics of a disaster of this kind, and insights gained from the experience.