A method of treatment for human patients with an acquired hypercoagulable state or acquired protein C deficiency associated with sepsis, purpura fulminans, meningococcal sepsis, bone marrow and other transplantations, severe burns, pregnancy, major surgery, severe trauma, or ARDS, which comprises administering activated protein C providing a highly selective therapeutic agent with a low potential for causing bleeding complications.

This invention relates to medical science particularly the treatment of hypercoagulable states or acquired protein C deficiency with activated protein C.
Protein C is a serine protease and naturally occurring anticoagulant that plays a role in the regulation of hemostasis through its ability to block the generation of thrombin production by inactivating Factors Va and VIIIa in the coagulation cascade. Human protein C is made in vivo primarily in the liver as a single polypeptide of 461 amino acids. This precursor molecule undergoes multiple post-translational modifications including 1) cleavage of a 42 amino acid signal sequence; 2) proteolytic removal from the one chain zymogen of the lysine residue at position 155 and the arginine residue at position 156 to make the 2-chain form of the molecule, (i.e., a light chain of 155 amino acid residues attached through a disulfide bridge to the serine protease-containing heavy chain of 262 amino acid residues); 3) vitamin K-dependent carboxylation of nine glutamic acid residues clustered in the first 42 amino acids of the light chain, resulting in 9 gamma-carboxyglutamic acid residues; and 4) carbohydrate attachment at four sites (one in the light chain and three in the heavy chain). The heavy chain contains the well established serine protease triad of Asp 257, His 211 and Ser 360. Finally, the circulating 2-chain zymogen is activated in vivo by thrombin at a phospholipid surface in the presence of calcium ion. Activation results from removal of a dodecapeptide at the N-terminus of the heavy chain, producing activated protein C (aPC) possessing enzymatic activity.
In conjunction with other proteins, aPC functions as perhaps the most important down-regulator of blood coagulation resulting in protection against thrombosis. In addition to its anti-coagulation functions, aPC has anti-inflammatory effects through its inhibition of cytokine generation (e.g. TNF and IL-1) and also exerts profibrinolytic properties that facilitate clot lysis. Thus, the protein C enzyme system represents a major physiological mechanism of anti-coagulation, anti-inflammation, and fibrinolysis.
Sepsis is defined as a systemic inflammatory response to infection, associated with and mediated by the activation of a number of host defense mechanisms including the cytokine network, leukocytes, and the complement and coagulation/fibrinolysis systems. [Mesters, et al., Blood 88:881-886, 1996]. Disseminated intravascular coagulation [DIC], with widespread deposition of fibrin in the microvasculature of various organs, is an early manifestation of sepsis/septic shock. DIC is an important mediator in the development of the multiple organ failure syndrome and contributes to the poor prognosis of patients with septic shock. [Fourrier, et al., Chest 101:816-823, 1992].
Several encouraging pre-clinical studies using protein C in various animal models of sepsis have been reported. A study in a baboon sepsis model by Taylor, et al., [J. Clin. Invest. 79:918-25, 1987], used plasma-derived human activated protein C. The animals were treated prophylactically (i.e., the aPC was given at the start of the two hour infusion of the LD100 E. coli). Five out of five animals survived 7 days and were considered permanent survivors to the experimental protocol. In control animals receiving an identical infusion of E. coli, five out of five animals died in 24 to 32 hours. The efficacious dose was 7 to 8 mg/kg.
In a lipopolysaccaride (LPS; E. coli) sepsis model in rats [Murakami, et al., Blood 87:642-647, 1996], the pulmonary vascular injury induced by LPS was inhibited by human plasma derived activated protein C at a dose of 100 xcexcg/kg. Furthermore, in a ligation and puncture sepsis model in rabbits, Okamoto, et al., [Gastroenterology 106:A747, 1994], demonstrated that plasma derived human activated protein C was effective in protecting the animals from coagulopathy and organ failure at a dose of 12 xcexcg/kg/hr for nine hours. Due to the species specificity of aPC, results obtained in these animals are not necessarily predictive to the treatment of humans. The efficacious dose level of human activated protein C is extremely variable and unpredictable depending upon the animal model selected. For example, the serum half-life of human activated protein C in humans is 30 to 40 minutes, compared to a half-life of 8 to 10 minutes in baboons and 90 minutes in rabbits.
There have been numerous recent attempts to treat sepsis in humans, for the most part using agents that block inflammatory mediators associated with the pathophysiology of this disease. However, clinical studies with a variety of agents that block inflammatory mediators have been unsuccessful [reviewed in Natanson, et al., Ann. Intern. Med 120:771-783, 1994; Gibaldi, Pharmacotherapy 13:302-308, 1993]. Since many of the mediators involved in inflammation are compensatory responses, and therefore have salutary effects, some investigators have suggested that blocking their action may not be appropriate [e.g., Parrillo, N. Engl. J. Med. 328:1471-1477, 1993].
Recently, blocking DIC has been proposed as a new target for clinical trials in sepsis [e.g., Levi, et al., JAMA 270:975-979, 1993]. However, simply blocking the coagulation defect in sepsis may not be sufficient. As reviewed by Esmon, [Arteriosclerosis and Thromb. 12:135-145, 1992], several antithrombotics have not shown efficacy in the baboon sepsis model, including active site-blocked factor Xa [Taylor, et al., Blood 78:364-368, 1991], hirudin and hirulog [Maraganore, Perspective in Drug Discovery and Design 1:461-478, 1994]. Each of these antithrombotics were able to block the consumptive coagulopathy in the animals but were not able to improve survival. Additionally, investigators in Japan [patent application JP7097335A] have proposed treating coagulopathy associated with hepatic insufficiency, which has the potential of developing DIC-like symptoms, with plasma derived activated protein C.
To date, plasma-derived human protein C zymogen has been used as a successful adjunct to aggressive conventional therapy in the management of twenty-five patients with purpura fulminans in bacterial sepsis of which twenty-two survived (Gerson, et al., Pediatrics 91:418-422, 1993; Smith, et al., Thromb. Haemost, PS1709, p419, 1997; Rintala, et al., Lancet 347:1767, 1996; Rivard, et al., J. Pediatr. 126:646-652, 1995). Gerson, et al., [1993] describe a case study of a treatment of a child with proven gram positive bacteremia and purpura fulminans, who was failing to respond to aggressive conventional treatment. The patient was treated with plasma-derived human protein C zymogen (280 xcexcg/kg bolus+40 xcexcg/kg/hr infusion) resulting in an associated correction of coagulopathy and DIC, and arrest of clinical signs of the development of septic shock-related purpura fulminans. Rintala, et al., [1996] reported the treatment of 2 adults with meningococcal septicemia presented with purpura fulminans. The patients were treated with plasma derived protein C zymogen at 400 xcexcg/kg bolus every six hours for 8-10 days. One died and one survived. Rivard, et al., [1995] reported the treatment of four patients with meningococcemia presented also with purpura fulminans, who all survived following human protein C zymogen therapy. These patients were treated at a dose of 400 xcexcg/kg bolus every six hours. Although the sample size from these studies is small, the mortality associated with meningococcemia presented with purpura fulminans is greater than 50% [Powars, et al., Clin. Infectious Diseases 17:254-261, 1993]. However, because these studies are conducted with human protein C zymogen, they offer little suggestion for establishing the dose and duration of therapy with activated protein C.
In addition to meningococcemia, purpura fulminans and/or DIC have been associated with numerous bacterial, viral, or protozoan infections which include but are not limited to infections caused by Rickettsia (Rocky Mountain Spotted fever, tick bite fever, typhus, etc.) [Graybill, et al., Southern Medical Journal, 66(4):410-413, 1973; Loubser, et al., Annals of Tropical Paediatrics 13:277-280, 1993]; Salmonella (typhoid fever, rat bite fever) [Koul, et al., Acta Haematol, 93:13-19, 1995]; Pneumococci [Carpenter, et al., Scand J Infect Dis, 29:479-483, 1997] Yersina pestis (Bubonic plague) [Butler et al., The Journal of Infectious Disease, 129:578-584, 1974]; Legionella pneumophila (Legionaires Disease); Plasmodium falciparum (cerebral malaria) [Lercari, et al., Journal of Clinical Apheresis, 7:93-96, 1992]; Burkholderia pseudomallei (Melioidosis); Pseudomonas pseudomallei (Melioidosis) [Puthucheary, et al., Transactions of the Royal Society of Tropical Medicine and Hygiene, 86:683-685, 1992]; Streptococci (Odontogenic infections) [Ota, Y., J. Japanese Assoc. Infect. Dis., 68:157-161]; zoster virus [Nguyen, et al., Eur J Pediatr, 153:646-649, 1994]; Bacillus anthracis (Anthrax) [Franz, et al., Journal of the American Medical Assoc., 278(5):399-411, 1997]; Leptospira interrogans (leptospirosis) [Hill, et al., Seminars in Respiratory Infections, 12(1):44-49, 1997]; Staphylococci [Levin, M., Pediatric Nephrology, 8:223-229]; Haemophilus aegyptius (Brazilian purpuric fever); Neisseria (gonococcemia, meningococcemia); and mycobacterium tuberculosis (miliary tuberculosis).
Even though the purpura fulminans, DIC or acquired protein C deficiency conditions in sepsis/septic shock or other infections have been well documented as indicated above, there is little data as to how to treat these patients with activated protein C. Establishing human dose levels using the pre-clinical pharmacology data generated from treatment with activated human protein C in animal models is difficult due to the species specificity properties of the biological actions of protein C.
A variety of transplantation associated thromboembolic complications may occur following bone marrow transplantation (BMT), liver, kidney, or other organ transplantations [Haire, et al., JAMA 274:1289-1295, (1995); Harper, et al., Lancet 924-927 (1988); and Sorensen, et al., J. Inter. Med 226:101-105 (1989); Gordon, et al., Bone Marrow Transplan. 11:61-65, (1993)]. Decreased levels of circulating protein C have been reported after BMT [Bazarbachi, et al., Nouv Rev Fr Hematol 35:135-140 (1993); Gordon, et al., Bone Marrow Trans. 11:61-65 (1993)], renal transplantation [Sorensen, et al., J. Inter. Med 226:101-105 (1989)], and liver transplantation [Harper, et al., Lancet 924-927 (1988)]. This deficiency in protein C contributes to a hypercoagulable state placing patients at risk for thromboembolic complications.
For example, hepatic venocclusive disease (VOD) of the liver is the major dose-limiting complication of pretransplantation regimens for BMT. VOD is presumably the result of small intrahepatic venule obliteration due to intravascular deposition of fibrin. [Faioni, et al., Blood 81:3458-3462 (1993)]. In addition, VOD causes considerable morbidity and mortality following BMT [Collins, et al., Throm. and Haemo. 72:28-33 (1994)]. A decreased level of protein C coincident with the peak incidence of VOD has been reported [Harper, et al., Bone Marrow Trans. 5:39-42 (1990)] and is likely to be a contributing factor to the genesis of this condition
Organ dysfunction after BMT including pulmonary, central nervous system, hepatic or renal, is a complication that occurs in a high percentage of transplant patients [Haire, et al., JAMA 274:1289-1295, (1995)]. A single organ dysfunction in BMT is a strong predictor of multiple organ dysfunction syndrome (MODS) which is the leading cause of death in BMT patients. Disseminated intravascular coagulation (DIC) due to a massive activation of the coagulation system and widespread deposition of fibrin in the microvasculature of various organs is an important mediator in the development of MODS [Fourrier, et al., Chest 101:816-823 (1992)]. Thus, a deficiency in protein C levels in patients who have undergone bone marrow or other organ transplantations leads to a hypercoagulable state that predisposes the patients to venous thromboembolic complications and organ dysfunction. A need currently exists to determine a method of treating humans with a hypercoagulable state associated with organ transplantations utilizing activated protein C.
It has long been recognized that severely burned patients have complications associated with hypercoagulation [Curreri, et al., Ann. Surg. 181:161-163 (1974)]. Burned patients have supranormal in vitro clotting activity and frequently develop DIC which is characterized by the sudden onset of diffuse hemorrhage; the consumption of fibrinogen, platelets, and Factor VIII activity; intravascular hemolysis; secondary fibrinolysis; and biopsy evidence of microthrombi [McManis, et al., J. of Trauma 13:416-422, (1973)]. Recently, it was reported that the levels of protein C were reduced drastically in severely burned patients and that this reduction of the natural anticoagulant may lead to an increase in the risk of DIC [Lo, et al., Burns 20:186-187 (1994)]. In addition, Ueyama, et al., in discussing the pathogenesis of DIC in the early stage of burn injury, concluded that massive thrombin generation and decrease of anticoagulant activity may occur in proportion to the severity of burns [Ueyama, et al., Nippon Geka Gakkai Zasshi 92:907-12 (1991)]. DIC is one of the common complications in patients suffering from severe burn injuries.
Protein C deficiency has been documented in severely burned patients as indicated above, however, there is little data regarding whether protein C replacement therapy would be effective or regarding how to treat these patients with activated protein C.
It is well known that pregnancy causes multiple changes in the coagulation system which may lead to a hypercoagulable state. For example, during pregnancy and post-partum, the risk of venous thrombosis is almost fivefold higher than in the non-pregnant state. In addition, clotting factors increase, natural inhibitors of coagulation decrease, changes occur in the fibrinolytic system, venous stasis increases, as well as increases in vascular injury at delivery from placental separation, cesarean section, or infection [Barbour, et al., Obstet Gynecol 86:621-633, 1995].
Although the risk of a complication due to this hypercoagulable state in women without any risk factors is small, women with a history of thromboembolic events are at an increased risk for recurrence when they become pregnant. In addition, women with underlying hypercoagulable states, including the recent discovery of hereditary resistance to activated protein C, also have a higher recurrence risk [Dahlback, Blood 85:607-614, 1995].
Therefore, it has been suggested that women with a history of venous thromboembolic events who are found to have a deficiency in antithrombin-III, protein C, or protein S, are at an appreciable risk of recurrent thrombosis and should be considered for prophylactic anticoagulant therapy [Conrad, et al., Throm Haemost 63:319-320, 1990].
The conditions of preeclampsia and eclampsia in pregnant women appear to be a state of increased coagulopathy as indicated by an increase in fibrin formation, activation of the fibrinolytic system, platelet activation and a decrease in platelet count [Clin Obstet Gynecol 35:338-350, 1992]. Preeclampsia is thought to be the result of uteroplacental ischemia due to an anomaly of the xe2x80x9cvascular insertionxe2x80x9d of the placenta. Consequences of preeclampsia include hypertension as well as DIC which leads to the release of numerous microthrombi which cause placental, renal, hepatic and cerebral lesions [Rev Fr Gynecol Obstet 86:158-163, 1991]. Furthermore, preeclampsia can lead to a severe and life threatening condition known as the HELLP syndrome which is defined as preeclampsia complicated by thrombocytopenia, hemolysis and disturbed liver function [Rathgeber, et al., Anasth Intensivther Notfallmed 25:206-211, 1990]. Additionally, it has been documented that there is a reduction in protein C levels in pregnant women with severe preeclampsia when compared to normal pregnancies [De Stefano, et al., Thromb Haemost 74:793-794, 1995].
Thus, the risk of venous thromboembolic complications occurring in pregnant women is a major concern, especially in women who have a history of thromboembolic events. Although the possibility of severe complications such as preeclampsia or DIC is relatively low, it has been suggested that it is essential to start therapy of DIC as soon as it has been diagnosed by onset of inhibition of the activated coagulation system [Rathgeber, et al., Anasth Intensivther Notfallmed 25:206-211, 1990]. The complications of preelampsia or DIC is analogous to the situation that occurs in sepsis in that there is a hypercoagulable state and a decrease in the levels of protein C.
Patients recovering from major surgery or accident trauma frequently encounter blood coagulation complications as a result of an induced hypercoagulable state [Watkins, et al., Klin Wochenschr 63:1019-1027, 1985]. Hypercoagulable states are increasingly recognized as causes of venous thromboembolism in surgical patients [Thomas, et al., Am J Surg. 158:491-494, 1989; LeClerc, J. R., Clin Appl Thrombosis/Hemostasis 3(3):153-156, 1997]. Furthermore, this hypercoagulable state can lead to complications with DIC-like symptoms, which is infrequently encountered but, nonetheless, is devastating and often fatal when it occurs. [Collins, et al., Am J Surg. 124:375-380, 1977].
In addition, patients undergoing coronary artery bypass grafting (CABG) [Menges, et al., J Cardiothor Vasc An. 10:482-489, 1996], major spinal surgery [Mayer, et al., Clin Orthop. 245:83-89, 1989], major abdominal surgery [Blamey, et al., Thromb Haemost. 54:622-625, 1985], major orthopedic surgery or arthroplastic surgery of the lower extremities [LeClerc, 1997], or other types of surgery [Thomas, et al., Am J Surg. 158:491-494, 1989], occasionally develop venous thromboembolic complications. Additionally, investigators in Japan have proposed treating microvascular thrombosis associated with spinal cord injury [patent application JP8325161A] with plasma derived protein C at a dose of 1-10 mg/day for an adult, or preferably, 2-6 mg divided by 1-2 times to be administered as a bolus or by intravenous infusion.
It has been suggested that anticoagulant therapy is important as a prophylactic therapy to prevent venous thromboembolic events in major surgery or trauma patients [Thomas, et al., 1989; LeClerc, 1997]. For example, many patients who succumb from pulmonary embolism have no clinical evidence of preceding thromboembolic events and die before the diagnosis is made and the treatment is instituted [LeClerc, 1997]. Existing prophylactic methods e.g., warfarin, low molecular weight heparins, have limitations such as residual proximal thrombosis or the need for frequent dose adjustments.
Adult respiratory distress syndrome [ARDS] is characterized by lung edema, microthrombi, inflammatory cell infiltration, and late fibrosis. Pivotal to these multiple cellular and inflammatory responses is the activation of coagulation resulting in a hypercoagulable state. Common ARDS-associated coagulation disorders include intravascular coagulation and inhibition of fibrinolysis. Fibrin formed by the activation of the coagulation system and inhibition of fibrinolysis presumably contributes to the pathogenesis of acute lung injury. Sepsis, trauma and other critical diseases are important risk factors that lead to ARDS [Hasegawa, et al., Chest 105(1):268-277, 1994].
ARDS is associated with an activation of coagulation and inhibition of fibrinolysis. Considerable clinical evidence exists for the presence of pulmonary vascular microemboli which is analogous to the hypercoagulation that is present in DIC. Therefore, a need currently exists for an effective treatment of this hypercoagulable state associated with ARDS.
For ease of comparison of the dose levels of protein C noted in literature and patent documents, Table I sets forth normalized dose levels of several studies in humans or non-human primates. These data establish dose levels that are higher or lower than the dose levels provided in the present invention. Significantly, the human studies were done utilizing plasma derived protein C zymogen while the non-human primate study utilized recombinant human aPC.
Despite these reports, however, the dosing regime for safe and efficacious therapy in humans suffering from an acquired hypercoagulable state or acquired protein C deficiency associated with sepsis, transplantations, burns, pregnancy, major surgery, trauma, or ARDS, remains unknown. These studies are not predictive of the use of recombinant activated protein C of the present invention in the treatment of hypercoagulable states or acquired protein C deficiency in humans.
The present invention discloses the use of aPC in a clinical trial in severe sepsis patients. In these patients, the r-aPC treated group demonstrated statistical improvement in organ functions, lowering of DIC markers and decrease in mortality as compared to the placebo control group. The doses of aPC used in the severe sepsis patients were 12, 18, 24, and 30 xcexcg/kg/hr in a 48 hour infusion. The doses of 12 and 18 xcexcg/kg/hr were not effective in this study. Surprisingly, the doses of 24 and 30 xcexcg/kg/hr used in this study were efficacious and are considerably and unexpectedly low as compared to published pre-clinical pharmacology data.
In addition, the applicants have found that pre-clinical toxicology studies in non-human primates indicate the safety of aPC for a 96 hour infusion is limited to a top dose of around 50 xcexcg/kg/hr. These data are also unexpected when compared to the prior art. In fact, the dose levels of r-aPC for humans that have been based on previous pre-clinical and clinical studies will be above the toxicological range established in the above toxicological studies.
The present invention provides a method of treating human patients with an acquired hypercoagulable state or acquired protein C deficiency which comprises administering to said patient by continuous infusion for about 24 to about 144 hours a dosage of about 20 xcexcg/kg/hr to about 50 xcexcg/kg/hr of activated protein C.
The invention further provides a method of treating human patients with an acquired hypercoagulable state or acquired protein C deficiency which comprises administering to said patient an effective amount of activated protein C to achieve activated protein C plasma levels in the range of 2 ng/ml to 200 ng/ml.
Thus, the present invention establishes methods utilizing aPC in the treatment of the hypercoagulable state or protein C deficiency associated with sepsis, purpura fulminans, and meningococcemia in human patients.
The present invention establishes methods utilizing aPC to treat the hypercoagulable state or protein C deficiency associated with severe burns.
The present invention establishes methods utilizing aPC to treat the hypercoagulable state or protein C deficiency associated with bone marrow and other organ transplantations.
The present invention establishes methods utilizing aPC to treat the hypercoagulable state or protein C deficiency associated with human patients undergoing or recovering from major surgery or severe trauma.
The present invention establishes methods utilizing aPC to treat the hypercoagulable state or protein C deficiency associated with complications during pregnancy.
The invention further provides a method of treating human patients with an acquired hypercoagulable state or acquired protein C deficiency associated with ARDS.
For purposes of the present invention, as disclosed and claimed herein, the following terms are as defined below.
aPC or activated protein C refers to recombinant activated protein C. aPC includes and is preferably human protein C although aPC may also include other species or derivatives having full protein C proteolytic, amidolytic, esterolytic, and biological (anticoagulant or pro-fibrinolytic) activities. Examples of protein C derivatives are described by Gerlitz, et al., U.S. Pat. No. 5,453,373, and Foster, et al., U.S. Pat. No. 5,516,650, the entire teachings of which are hereby incorporated by reference. Recombinant activated protein C may be produced by activating recombinant human protein C zymogen in vitro or by direct secretion of the activated form of protein C. Protein C may be produced in cells, eukaryotic cells, transgenic animals, or transgenic plants, including, for example, secretion from human kidney 293 cells as a zymogen then purified and activated by techniques known to the skilled artisan.
Treatingxe2x80x94describes the management and care of a patient for the purpose of combating a disease, condition, or disorder and includes the administration of aPC prophylactically to prevent the onset of the symptoms or complications of the disease, condition, or disorder, or administering aPC to eliminate the disease, condition, or disorder.
Continuous infusionxe2x80x94continuing substantially uninterrupted the introduction of a solution into a vein for a specified period of time.
Bolus injectionxe2x80x94the injection of a drug in a defined quantity (called a bolus) over a period of time up to about 120 minutes.
Suitable for administrationxe2x80x94a lyophilized formulation or solution that is appropriate to be given as a therapeutic agent.
Receptaclexe2x80x94a container such as a vial or bottle that is used to receive the designated material, i.e., aPC
Unit dosage formxe2x80x94refers to physically discrete units suitable as unitary dosages for human subjects, each unit containing a predetermined quantity of active material calculated to produce the desired therapeutic effect, in association with a suitable pharmaceutical excipient.
Hypercoagulable statesxe2x80x94excessive coagulability associated with disseminated intravascular coagulation, pre-thrombotic conditions, activation of coagulation, or congenital or acquired deficiency of clotting factors such as aPC.
Zymogenxe2x80x94Protein C zymogen, as used herein, refers to secreted, inactive forms, whether one chain or two chains, of protein C.
Juvenilexe2x80x94a human patient including but not restricted to newborns, infants, and children younger than 18 years of age.
Effective amountxe2x80x94a therapeutically efficacious amount of a pharmaceutical compound.
Purpura fulminansxe2x80x94ecchymotic skin lesions, fever, hypotension associated with bacterial sepsis, viral, bacterial or protozoan infections. Disseminated intravascular coagulation is usually present.
The present invention relates to the treatment or prevention of hypercoagulable states or acquired protein C deficiency associated with sepsis, transplantations, burns, pregnancy, major surgery, trauma, or ARDS, with activated protein C. The aPC can be made by techniques well known in the art utilizing eukaryotic cell lines, transgenic animals, or transgenic plants. Skilled artisans will readily understand that appropriate host eukaryotic cell lines include but are not limited to HEPG-2, LLC-MK2, CHO-K1, 293, or AV12 cells, examples of which are described by Grinnell in U.S. Pat. No. 5,681,932, herein incorporated by reference. Furthermore, examples of transgenic production of recombinant proteins are described by Drohan, et al., in U.S. Pat. No. 5,589,604 and Archibald, et al., U.S. Pat. No. 5,650,503, herein incorporated by reference.
To be fully active and operable under the present methods, the aPC made by any of these methods must undergo post translational modifications such as the addition of nine gamma-carboxy-glutamates (gamma-carboxylation i.e. Gla content), the addition of one erythro-beta-hydroxy-Asp (beta-hydroxylation), the addition of four Asn-linked oligosaccharides (glycosylation), the removal of the leader sequence (42 amino acid residues) and removal of the dipeptide Lys 156-Arg 157. Without such post-translational modifications, aPC is not fully functional or is non-functional.
The aPC can be formulated according to known methods to prepare pharmaceutically useful compositions. The aPC will be administered parenterally to ensure its delivery into the bloodstream in an effective form by injecting the appropriate dose as continuous infusion for about 24 to about 144 hours. The amount of aPC administered will be from about 20 xcexcg/kg/hr to about 50 xcexcg/kg/hr. More preferably, the amount of aPC administered will be about 22 xcexcg/kg/hr to about 40 xcexcg/kg/hr. Even more preferably the amount of aPC administered will be about 22 xcexcg/kg/hr to about 30 xcexcg/kg/hr. The most preferable amounts of aPC administered will be about 24 xcexcg/kg/hr or about 30 xcexcg/kg/hr.
Alternatively, the aPC will be administered by injecting a portion (⅓ to xc2xd) of the appropriate dose per hour as a bolus injection over a time from about 5 minutes to about 120 minutes, followed by continuous infusion of the appropriate dose for about twenty-three hours to about 144 hours which results in the appropriate dose administered over 24 hours to 144 hours.
Only after carefully controlled clinical studies and exhaustive experimental studies have the applicants discovered that the dose levels of about 20 xcexcg/kg/hr to about 50 xcexcg/kg/hr continually infused for about 24 hours to about 144 hours results in efficacious therapy. The most preferable dose level of aPC to be administered for treating human patients with an acquired hypercoagulable state or acquired protein C deficiency as described herein will be about 24 xcexcg/kg/hr.
Recombinant human protein C (r-hPC) was produced in Human Kidney 293 cells by techniques well known to the skilled artisan such as those set forth in Yan, U.S. Pat. No. 4,981,952, the entire teaching of which is herein incorporated by reference. The gene encoding human protein C is disclosed and claimed in Bang, et al., U.S. Pat. No. 4,775,624, the entire teaching of which is incorporated herein by reference. The plasmid used to express human protein C in 293 cells was plasmid pLPC which is disclosed in Bang, et al., U.S. Pat. No. 4,992,373, the entire teaching of which is incorporated herein by reference. The construction of plasmid pLPC is also described in European Patent Publication No. 0 445 939, and in Grinnell, et al., 1987, Bio/Technology 5:1189-1192, the teachings of which are also incorporated herein by reference. Briefly, the plasmid was transfected into 293 cells, then stable transformants were identified, subcultured and grown in serum-free media. After fermentation, cell-free medium was obtained by microfiltration.
The human protein C was separated from the culture fluid by an adaptation of the techniques of Yan, U.S. Pat. No. 4,981,952, the entire teaching of which is herein incorporated by reference. The clarified medium was made 4 mM in EDTA before it was absorbed to an anion exchange resin (Fast-Flow Q, Pharmacia). After washing with 4 column volumes of 20 mM Tris, 200 mM NaCl, pH 7.4 and 2 column volumes of 20 mM Tris, 150 mM NaCl, pH 7.4, the bound recombinant human protein C zymogen was eluted with 20 mM Tris, 150 mM NaCl, 10 mM CaCl2, pH 7.4. The eluted protein was greater than 95% pure after elution as judged by SDS-polyacrylamide gel electrophoresis.
Further purification of the protein was accomplished by making the protein 3 M in NaCl followed by adsorption to a hydrophobic interaction resin (Toyopearl Phenyl 650 M, TosoHaas) equilibrated in 20 mM Tris, 3 M NaCl, 10 mM CaCl2, pH 7.4. After washing with 2 column volumes of equilibration buffer without CaCl2, the recombinant human protein C was eluted with 20 mM Tris, pH 7.4.
The eluted protein was prepared for activation by removal of residual calcium. The recombinant human protein C was passed over a metal affinity column (Chelex-100, Bio-Rad) to remove calcium and again bound to an anion exchanger (Fast Flow Q, Pharmacia). Both of these columns were arranged in series and equilibrated in 20 mM Tris, 150 mM NaCl, 5 mM EDTA, pH 6.5. Following loading of the protein, the Chelex-100 column was washed with one column volume of the same buffer before disconnecting it from the series. The anion exchange column was washed with 3 column volumes of equilibration buffer before eluting the protein with 0.4 M NaCl, 20 mM Tris-acetate, pH 6.5. Protein concentrations of recombinant human protein C and recombinant activated protein C solutions were measured by UV 280 nm extinction E0.1%=1.85 or 1.95, respectively.
Bovine thrombin was coupled to Activated CH-Sepharose 4B (Pharmacia) in the presence of 50 mM HEPES, pH 7.5 at 4xc2x0 C. The coupling reaction was done on resin already packed into a column using approximately 5000 units thrombin/ml resin. The thrombin solution was circulated through the column for approximately 3 hours before adding MEA to a concentration of 0.6 ml/l of circulating solution. The MEA-containing solution was circulated for an additional 10-12 hours to assure complete blockage of the unreacted amines on the resin. Following blocking, the thrombin-coupled resin was washed with 10 column volumes of 1 M NaCl, 20 mM Tris, pH 6.5 to remove all non-specifically bound protein, and was used in activation reactions after equilibrating in activation buffer.
Purified r-hPC was made 5 mM in EDTA (to chelate any residual calcium) and diluted to a concentration of 2 mg/ml with 20 mM Tris, pH 7.4 or 20 mM Tris-acetate, pH 6.5. This material was passed through a thrombin column equilibrated at 37xc2x0 C. with 50 mM NaCl and either 20 mM Tris pH 7.4 or 20 mM Tris-acetate pH 6.5. The flow rate was adjusted to allow for approximately 20 min. of contact time between the r-hPC and thrombin resin. The effluent was collected and immediately assayed for amidolytic activity. If the material did not have a specific activity (amidolytic) comparable to an established standard of aPC, it was recycled over the thrombin column to activate the r-hPC to completion. This was followed by 1:1 dilution of the material with 20 mM buffer as above, with a pH of anywhere between 7.4 or 6.0 (lower pH being preferable to prevent autodegradation) to keep the aPC at lower concentrations while it awaited the next processing step.
Removal of leached thrombin from the aPC material was accomplished by binding the aPC to an anion exchange resin (Fast Flow Q, Pharmacia) equilibrated in activation buffer (either 20 mM Tris, pH 7.4 or preferably 20 mM Tris-acetate, pH 6.5) with 150 mM NaCl. Thrombin passes through the column and elutes during a 2-6 column volume wash with 20 mM equilibration buffer. Bound aPC is eluted with a step gradient using 0.4 M NaCl in either 5 mM Tris-acetate, pH 6.5 or 20 mM Tris, pH 7.4. Higher volume washes of the column facilitated more complete removal of the dodecapeptide. The material eluted from this column was stored either in a frozen solution (xe2x88x9220xc2x0 C.) or as a lyophilized powder.
The amidolytic activity (AU) of aPC was determined by release of p-nitroanaline from the synthetic substrate H-D-Phe-Pip-Arg-p-nitroanilide (S-2238) purchased from Kabi Vitrum using a Beckman DU-7400 diode array spectrophotometer. One unit of activated protein C was defined as the amount of enzyme required for the release of 1 xcexcmol of p-nitroaniline in 1 min. at 25xc2x0 C., pH 7.4, using an extinction coefficient for p-nitroaniline at 405 nm of 9620 Mxe2x88x921 cmxe2x88x921.
The anticoagulant activity of activated protein C was determined by measuring the prolongation of the clotting time in the activated partial thromboplastin time (APTT) clotting assay. A standard curve was prepared in dilution buffer (1 mg/ml radioimmunoassay grade BSA, 20 mM Tris, pH 7.4, 150 mM NaCl, 0.02% NaN3) ranging in protein C concentration from 125-1000 ng/ml, while samples were prepared at several dilutions in this concentration range. To each sample cuvette, 50 xcexcl of cold horse plasma and 50 xcexcl of reconstituted activated partial thromboplastin time reagent (APTT Reagent, Sigma) were added and incubated at 37xc2x0 C. for 5 min. After incubation, 50 xcexcl of the appropriate samples or standards were added to each cuvette. Dilution buffer was used in place of sample or standard to determine basal clotting time. The timer of the fibrometer (CoA Screener Hemostasis Analyzer, American Labor) was started upon the addition of 50 xcexcl 37xc2x0 C. 30 mM CaCl2 to each sample or standard. Activated protein C concentration in samples are calculated from the linear regression equation of the standard curve. Clotting times reported here are the average of a minimum of three replicates, including standard curve samples.
The above descriptions enable one with appropriate skill in the art to prepare aPC and utilize it in the treatment of hypercoagulable states or acquired protein C deficiency associated with but not limited to sepsis, transplantations, burns, pregnancy, major surgery/trauma, and ARDS.

Method of reducing mortality in severe sepsis


The described concepts relate to automated patient disposition. One example can receive a clinician's disposition for a patient. This implementation can perform parameter-based cluster analysis on the patient and a set of patients to identify a sub-set of the patients with which the patient has a high similarity. This example can also cause a graphical user interface to be generated that conveys parameters from the sub-set of the patients and the patient.

The present discussion relates to patient care. For instance, one profoundly fragile but consequential decision in hospitals is related to making decisions around the disposition of a patient. For example, such decisions can relate to when to discharge from the emergency department, versus admit, when to discharge from the hospital, when to transition from one part of the hospital to another, etc. Patient disposition is a complex and subtle decision with profound implications to the patient. One such scenario relates to "Failure to Rescue". Failure to rescue refers to the phenomena where a patient is not transferred to the intensive care unit (ICU) soon enough and instead suffers cardiac or respiratory arrest on a non-ICU floor. Failure to Rescue is estimated to occur across the U.S. at a rate of almost 300,000 cases per year.
In an attempt to remedy this phenomenon, today many emergency medicine rooms (EMRs) manually implement a series of rules to check whether a discharge might be dangerous. These rules may check for critical lab values or abnormal vital signs. The challenge is that there could be hundreds or thousands of rules that need to be written to cover the thousands of known lab tests available today. Yet when all those rules are written, there is also a factorial of interactions between those labs that may also be predictive of danger. Further, new laboratory tests and new knowledge on how to use those tests is entering healthcare at an exponential rate.

Automated clustering for patient disposition


A film contains a first polymer having a plurality of hydrogen bond donating moieties, and a second polymer having a plurality of hydrogen bond accepting moieties. The second polymer is hydrogen bonded to the first polymer.

The present invention relates to polymer films, their assembly, and their controlled destruction.
Polymer films find wide-ranging uses, from non-linear optics to electronic coatings. One way to produce polymer films is through self-assembly of the films, layer-by-layer. This self-assembly relies on electrostatic charge, of alternating sign, in each layer; charges of opposite sign in each layer attract, directing the self-assembly process. The layers may be build up from a solution, and in the assembly process, molecules may become trapped between the layers.
It would be desirable to destroy the polymer films in a controlled fashion, to release the molecules trapped between the layers. Molecules, such as drugs or dyes, could be trapped in the polymer films during the assembly process, and then released during controlled destruction of the polymer films. However, the layers of known self-assembled polymer films contain alternating charge, and therefore any controlled method of destruction of these films would require eliminating or reducing charge on some or all of the layers, in order to overcome the electrostatic attraction that holds the layers together, and to do so is not practical.
In a first aspect, the present invention is a film including a first polymer, and a second polymer hydrogen bonded to the first polymer.
In a second aspect, the present invention is a film including a first polymer having a plurality of hydrogen bond donating moieties, and a second polymer having a plurality of hydrogen bond accepting moieties.
In a third aspect, the present invention is a method of forming a film, including contacting a surface with a first polymer having a plurality of hydrogen bond donating moieties, and contacting the surface with a second polymer having a plurality of hydrogen bond accepting moieties.
The present invention includes a polymer film in which the layers are held together by hydrogen bond formation between the layers. The formation of the hydrogen bonds drives a self-assembly process. The attraction of the layers to each other can be overcome by creating charge on at least some of the layers, the charge creating a repulsive force, or increasing the solubility of the polymers while in contact with a solvent. In the case of creating charge, when the repulsive force between layers exceeds the attractive force from the hydrogen bonds, the polymer film will be destroyed, releasing molecules trapped in the polymer film. Through control of environmental factors, the creation of charge on the layers can be controlled, allowing for controlled destruction of the polymer film, and thereby controlled release of molecules trapped in the film. Unlike films formed in the past, the films are not made of polymers which each contain charged groups of opposite signs; these past films used the electrostatic attraction of the oppositely charged groups to bond the polymers together into the films.
The polymers used to form the films of the present invention include up to three moieties: a hydrogen bond donor, a hydrogen bond acceptor, and optionally, a charge-forming group. Each polymer used must include at least a hydrogen bond donor or a hydrogen bond acceptor; preferably at least one of the polymers used in the film contains a charge-forming group. Preferably, the film is produced using two polymers, with one polymer containing hydrogen bond donors, and the other polymer containing hydrogen bond acceptors, and at least one of the two polymers containing charge-forming groups. In the alternate embodiment of three, or more polymers, each polymer must contain at least hydrogen bond donors or hydrogen bond acceptors; at least one of the polymers used must also contain a charge-forming group.
A hydrogen bond is a relatively weak secondary interaction between a hydrogen atom bound to a more electronegative atom and another atom that is also generally more electronegative than hydrogen and has one or more lone electron pairs, enabling it to act as a base. Hydrogen bonding has been extensively studied (see, for example, xe2x80x9cAdvanced Inorganic Chemistryxe2x80x9d 5th ed., F. Albert Cotton and Geoffrey Wilkinson, pp.90-94 (John Wiley and Sons, 1988)).
Hydrogen bond donors are moieties that contain at least one hydrogen atom that may participate in hydrogen bond formation, and a more electronegative atom bound to the hydrogen atom. Examples of these moieties include, preferably, Oxe2x80x94H and Nxe2x80x94H, and less preferably, Pxe2x80x94H, Sxe2x80x94H. The moiety Cxe2x80x94H may also, less preferably, be a hydrogen bond donor, when the carbon atom is bond to another atom through a triple bond, when the carbon atom is bound through a double bond to O, or when the carbon atom is bound to at least two atoms selected from O, F, Cl and Br.
Hydrogen bond acceptors are moieties that contain an atom more electronegative than hydrogen that also has a lone pair of electrons. Examples of these atoms include preferably N, O and F, and less preferably Cl, Br, I, S and P. Examples of hydrogen bond acceptor moieties include Cxe2x95x90O, Oxe2x80x94H, Nxe2x80x94H, Cxe2x80x94F, Pxe2x95x90O and Cxe2x89xa1N.
Charge forming structures are moieties that can develop charge when exposed to one or more environmental changes. Examples of environmental changes are a change in pH, a change in ionic strength, exposure to an electric field, or exposure to dissolved ions. Examples of moieties that can develop charge under changing pH conditions include acid or base moieties. Examples of moieties that can develop charge under exposure to an electric field include carboxylic acids. Examples of moieties that can develop charge under exposure to dissolved ions include crown ethers (upon exposure to certain alkali metal ions).
Polymers for use in the present invention include polymers containing hydrogen bond donors and/or hydrogen bond acceptors, for example polycarboxylic acids such polyacrylic acid and polymethacrylic acid; polynucleotides such as poly(adenylic acid), poly(uridylic acid), poly(cytidylic acid), poly(uridylic acid) and poly(inosinic acid); polymers of vinyl nucleic acids such as poly(vinyladenine); polyamino acids such as polyglutamic acid and poly(xcex5-N-carbobenzoxy-L-lysine); polyalcohols such as poly(vinyl alcohol); polyethers such as poly(ethylene oxide), poly(1,2-dimethoxyethylene), poly(vinylmethyl ether), and poly(vinylbenzo-18-crown-6); polyketones and polyaldehydes such as poly vinyl butyral and poly(N-vinyl-2-pyrrolidone); polyacrylamides such as polyacrylamide, polymethacrylamide and poly(N-isopropylacrylamide); polyamines such as poly(4-amine)styrene; polyesters such poly(cylohexane-1,4-dimethylene terephthalate) and polyhydroxy methyl acrylate; polyphosphazenes such as poly(bis(methylamino)phosphazene) and poly(bis(methoxyethoxyethoxy)phosphazene; polysaccharides such as carboxymethyl cellulose; and copolymers thereof,
Some examples of preferable pairs of polymer for forming films include those of the following types:
Type 1. Homopolymer of polycarboxylic acid, paired with the specified Polymer B.
In the above three examples of Polymer B, the motif of proton acceptance in hydrogen bonding, is O . . . HO.
In the above five examples of Polymer B, the motif of proton acceptance, is NCxe2x95x90O . . . HO.
Films formed from these pairs of polymers may be dissolved (the films destroyed) at high pH or in dimethylsulfoxide (DMSO). A film formed from PMA-PVP is stable in tetramethylurea and dimethylformamide (DMF) and dissolves in dimethylacetamide, N-methylpyrrolidone and hexamethylphosphoric triamide. The stability of these films can also be affected by temperature; these films become more stable as the temperature increases in water, but are destabilized as the temperature increases in DMF.
Type 2. Multilayers that include crown ethers as one constituent.
This type of film is destroyed by either high or low pH, depending on the specific monovalent ion present in the environment. For example, there is strong sensitivity to the type of cation present in the environment:
Type 3. Films that include, as one constituent, molecules containing Pxe2x95x90O moieties. The motif of hydrogen bonding is Pxe2x95x90O . . . HO.
For example, the above film dissolves in hexamethylphosphoric triamide.
Type 4. Multilayers that include amino acids as one constituent.
Type 5. Films based on hydrogen bonding between synthetic polynucleotides or vinyl-type polymers containing nucleic acid bases.
These nucleic acid polymers will also form films with naturally occurring RNA (ribonucleic acid), DNA (deoxynucleic acid), as well as synthetic polynucleobdes.
The films may be formed on a surface, such as the surface of a monolithic solid, or particles. The film is formed by dissolving at least one of the polymers into a solvent, to form a solution. The solution is then contacted with the surface, and hydrogen bonding of the polymer with the surface forms a layer of the polymer on the surface. A solution of the second polymer is next contacted with the layer of the first polymer on the surface, forming hydrogen bonds between the polymers, and forming a film. The process of contacting the surface with a solution of the first polymer and then a solution of the second polymer may be repeated until a film of the desired thickness is formed. If more than two polymers are to be used to form the film, a solution of the additional polymers may be contacted with the growing film. If the amount of polymer in the solutions is more than enough to form a layer on the surface, the surface may be rinsed between applications with the different polymer solutions, or removed from the solution. Aerosol deposition is also possible. The environmental conditions under which the film is formed are selected so that there is insufficient charge on the charge-forming groups to prevent film formation, or insufficient charge to cause dissolution of the film. It is also possible to form the film by evaporating the first polymer onto a surface, followed by evaporation of the second polymer onto the surface, and repeating these steps until the desired thickness is achieved; this is possible when the polymers have a molecular weight of at most 5000 g/mol, more preferably at most 2000 g/mol.
Examples of materials, either as monolithic solids, or particles, which may be coated include semiconductors such as silicon or germanium, polymers such as polyethylene and TEFLON, ceramics such as glass or alumina, and minerals such as mica. The films may be coated onto a surface, with an agent contained in the film, which may be released at a later time by destruction of the film. Also, the films may be used to change the properties of the surface (surface roughness, reflectivity, color, hydrophobicity, chemical reactivity, etc.), and optionally, the original surface properties may be restored by destruction of the films.
Any agent can be included in the film, as long as it can be dissolved or dispersed in the solvent in which at least one of the polymers is dissolved, or if the molecule can be evaporated with one of the polymers. Examples of agents include dyes, fragrances, perfumes, surfactants, detergents, chemicals used in environmental remediation, and bioactive agents. The bioactive agents can be any physiologically or pharmacologically active substance or substances optionally in combination with pharmaceutically acceptable carriers and additional ingredients such as antioxidants, stabilizing agents, permeation enhancers, etc. The bioactive agents may be any of the agents that are known to be delivered to the body of a human or an animal (including insects), and plants, and that are preferentially soluble in water. These bioactive agents include drug agents, medicaments, vitamins, nutrients, fertilizers, pesticides, or the like. Included among the types of bioactive agents which meet this description are drug agents, proteins, peptides, genetic material, nutrients, vitamins, food supplements, spices, flavor enhancers, emollients, skin and hair conditioners, sex sterilants, fertility inhibitors and fertility promoters. Examples of drug agents include prochlorperzine edisylate, ferrous sulfate, aminocaproic acid, mecamylamine hydrochloride, procainamide hydrochloride, amphetamine sulfate, methamphetamine hydrochloride, benzamphetamine hydrochloride, isoproterenol sulfate, phenmetrazine hydrochloride, bethanechol chloride, methacholine chloride, pilocarpine hydrochloride, atropine sulfate, scopolamine bromide, isopropamide iodide, tridihexethyl chloride, phenformin hydrochloride, methylphenidate hydrochloride, theophylline cholinate and cephalexin hydrochloride.
Solvents for dissolving the polymers include any liquid in which the polymer has measurable solubility. Solvents include hydrocarbons such as pentane, hexane and toluene; alcohols such as methanol, ethanol, isopropanol and phenol; esters such butyl acetate; aldehydes and ketones such as formaldehyde, acetone and methyl ethyl ketone; dimethyl sulfoxide; carbonates such as propylene carbonate; N-methyl formamide, tetramethylurea, dimethylacetamide, N-methylpyrrolidone, hexamethylphosphoric triamide and dimethyl formamide; supercritical fluids such as carbon dioxide; and preferably water; and mixtures thereof
The films may be destroyed in order to release the agent trapped in the film so that the agent may carry out its own effect (such as in the case of a dye or bioactive agent), or to simply release the polymers of the film into a solution (for example, to change the viscosity of the solution). Furthermore, the films may be used to coat particles, allowing for controlled release of the one or more substances contained in the particles. If one or more of the polymers included in the film contain charge-forming groups, then forming sufficient charge in these groups to overcome the strength of the hydrogen bonding will cause the film to decompose. For example, adjusting the pH of the solution, applying an external electric field, or supplying specific ions, may be used depending on the specific charge-forming groups present. Alterative, heating or the addition of solvents may be sufficient to cause the film to dissolve or decompose, especially in the case where there are no charge-forming groups present in the polymers of the film.

Board of Trustees of University of Illinois


A system and method for use by website operators and advertisers to place interstitial advertisements on websites. The system uses an Internet dispatcher server and a mass storage device containing one or more databases storing advertisement command files pertaining to placements of advertisements that are stored in an advertisement server. The method comprises the steps of selecting ad files stored in a computer ad server, designating the url for the ad file, the url for the website page where the ad will be displayed, the url for the websites to which a user will be directed depending on the user's reaction to the advertisement, storing in a dispatcher server database a data string representing the ad placement commands with a url representing a dispatcher server-based application to execute said ad placement commands, and including said data string on said website for launching said dispatcher server command file.

A. Technical Field
The present invention relates generally to methods and systems for creation and distribution of promotional and informational communications regarding products and services via computer networks. More specifically, the present invention is an Internet-based system and method for distributing of interstitial advertisements, including advertisements comprising rich media files, providing real-time interstitial advertisement viewer activity tracking capability and including a relational database for storing data required for commands that execute selection, dispatching, tracking and display of said interstitial advertisement files previously stored in electronic format in a server. The present invention also relates to use of said system in a multi-level advertising business method that eliminates problems relating to the accurate tracking of sales attributable to advertisements placed on websites.
B. Background
Internet advertising is a multi-billion dollar industry inherently different from television and print advertising. Television advertising impressions are primarily served from networks such as ABC, NBC and CBS and their affiliates. Print publishing is dominated by established national and local periodicals and newspaper publishers. In contrast, the top ten mega portals, such as Yahoo and AOL, serve a minority of all Internet page views. The majority of Internet traffic is served from small special interest and personal websites that operate independently.
Prior art methods of deriving revenue from Internet advertising fall under two general approaches. The first is a pre-pay or billed cost per impression, or "CPM," or cost per click, or "CPC," model, wherein advertisers pay publishers to place their advertisement on a website according to the number of impressions served, or click actions taken on the advertisement. Usually these transactions are done through a third party agency who receives commissions based on a percentage agency discount offered by the publisher.
For the CPM/CPC model to work, a website with heavy traffic must agree to place an ad that an advertiser will pay for with an established advertising budget. For this method to be effective, the mega portal or network must already be established with significant web traffic to offer advertisers, and an advertiser's budget must be available to offer website publishers to place the advertising.
The second method is a revenue share model. With this model a publisher places an advertisement on his/her website and is compensated with a percentage of the revenue from sales that results from traffic driven to the buy site from the advertisement. The advertiser is not required to pay fees when the ad is placed, but rather, when the add produces a sale.
Current revenue share compensation methods are basically flawed in that their verification methods essentially boil down to trust. The methods for tracking of receipt of and responses to advertisements used in prior systems have addressed in a relatively inefficient manner. Typically, tracking of Internet-based advertising and user responses to same is accomplished by redirecting a user's response. First, a connection is established with a tracking system that records the user's response to the URL of the selected ad or link, and then, the destination IP address URL is provided. If an advertisement placed on a website delivers a customer to a different website for purchase, there is no way for the website in which the advertisement is placed to know for sure what has happened once the user arrives at that buy site. The website in which the advertisement is placed has no way to know for sure and if it is being compensated properly. In addition, the customer may visit the buy site later, making a purchase that would not be linked to the advertisement placed on the other website.
With most Internet advertising revenue models, payments are often tied to a function of the number of web users whom the ad reached. But with web advertisements, accurately ascertaining that number has been difficult and problematic at best, and, given a basic technique employed to do so, relatively error-prone, causing erroneous counts and ad charges. Ads are usually logged as a "user impression" at a web server when the file is served rather than after the browser has completely rendered the advertisement to the user. Serving the ad files does not guarantee that the files will be ultimately and completely rendered by the user's browser. As a result, web server generated "user impression" counts can be grossly over- or under-stated. If a user navigates to a new content page after an advertisement has started playing but before that advertisement completes and, by doing so, prematurely terminated the advertisement, a full impression is nevertheless logged—erroneously—since that advertisement was completely served. Additional errors arise if a proxy server is situated between multiple user computers situated on an intranet or a local area network and a web advertisement server situated on the Internet. In this case, a request from one of the website user computers for the advertisement files will be routed to the proxy server, which, in turn, will direct that request onward to the advertisement web server. The advertisement server serves one complete copy of the advertisement files to the proxy server. The resulting retrieved advertisement files will be locally cached in the proxy server and, from there, provided to the requesting user's computer. Should any of the other user's computers request the same files, the proxy server will provide these files, totally unbeknownst to the advertisement server, from its local cache rather than directing a request from that other user computer back to the advertisement server. Hence, the advertisement server will be totally oblivious to each additional instance in which the proxy server accessed the ad files from its local cache and disseminated the advertisement to any user's computer other than that which first requested the ad. Proxy servers can therefore result in significant under-counting and under-charges to the advertiser. In order for Internet advertisements to effectively generate revenue as an on-going stream of payments to the host of the ads, accurate user accounting is essential to ensure that an advertiser is not over- or under-charged given an extent to which an ad is actually disseminated.
Users of the Internet typically rely on robust applications, such as web browsers and browser technology, to handle and maintain the transfer of information between systems. Existing applications to communicate with a user via the Internet are based on the following two methods:
1) Using an Internet browser (such as Netscape and Internet Explorer, for example) that is installed and run on the user's computer to communicate with a server.
2) Having the user download and install a custom application on the user's computer hard drive to communicate with the server.
In method 1), the Internet browser holds specific objects (such as, for example, the history object of the browser itself, or a custom object supported by the browser) which can be recognized by the desired web server.
In method 2), the custom application installed on the user's computer is used to communicate with the server.
In settings where the intent of the user is to use the browser or application, there is little question given to the issue of engaged system resources or bandwidth usage, as both are fundamental components to the primary activity engaged in by the user. The current practice of launching an additional browser window or dedicated application to accommodate the new media, initiate the connection, or transfer the information requires much more computer resources than are actually needed.
With prior art systems, customer and website computer resources are taxed in requiring the system to launch and maintain an additional application that may well exceed the necessary capabilities required for the desired communication and, in some cases, even the capabilities of the system itself. Additionally, as browsers and networked applications are continuously involved in two-way communication between the website user and website server systems involved in the communications, the bandwidth resources available to the communication effort are monopolized at a level, that although customary, are perhaps more than truly required.
Advertisers seeking to utilize the global reach of the Internet as an effective and efficient medium for disseminating advertisements to consumers have had to deal with various limitations in technology and methods. Internet advertisements are currently in forms ranging from static ads placed on space provided on website pages, links to ads or other websites, as well as banner ads, both static and animated. A banner advertisement is created by embedding specific HTML code for that banner within the HTML coding for a given web page in which the banner is to appear. A website user's browser, as it interprets and sequentially executes the HTML code for a selected page, will compile and execute the embedded code for the banner and display the banner as part of a rendered page and at a specified location thereon.
In implementing a banner, whether static or animated, the HTML coding is downloaded to a the website user's browser. The file may be stored on the same server that stores the HTML file for the web page, or accessed from a remote server. The file may contain a graphic itself, such as in a graphic interchange format or "GIF" file, or a Java applet which, once interpreted and executed by the browser, generates and renders the desired graphic. Such files require time to download and must be downloaded and assembled by the browser on the page prior to that page being fully rendered. Download times for large files, especially when the website user's system has relatively low communications bandwidth, can be long enough to lose the website user's interest. Users have to wait a considerable amount of time before all the page components for multimedia content are fully downloaded to permit that page to be rendered. Such delay during a page transition often results in the user prematurely terminating the download and transitioning to another web page. Advertisers therefore often limit the file size of their banner ads in order to minimize page download and hence latency times. Banners are for this reason often less effective than desired, as evidenced by relatively low user click-through counts generally observed for banner ads.
In an effort to overcome the limitations associated with banners, Internet advertisers have also used interstitial advertisements, which are pre-programmed advertisements that are displayed on websites during the interstitial period between when a website user clicks to access a new web page and when the selected web page is displayed to the user. Such advertisements often include a hotlink to enable the user to "click-through" the advertisement to access the website designated by the website advertiser. The length of an interstitial interval, which is variable, is governed by a variety of factors, including, but not limited to, the number of files required to fully render the new page and the size of each such file, network and server congestion and attendant delays occurring when users activate hotlinks. Prior patents in this field disclose the concept of embedding an advertisement as an information object in a web page file in such a manner that the object will remain hidden and will not be displayed when the file is executed to render the page. Rather than being displayed, the information object is locally cached by the website user's browser during execution of the code for that page. Then, during a transition initiated by website user activation of a hotlink to move from one web page to a next successive web page, referred to as an "interstitial period," the website user's browser accesses the advertisement from its local cache and displays it until such time as that next successive page is downloaded and rendered.
Other efforts have included "polite" downloading, which involves a browser on the website user's computer downloading from a remote advertising system server and ostensibly as a background process, file(s) for a web advertisement only during those intervals when bandwidth utilization of a communication channel connected to the browser is less than a pre-established threshold. Such "polite" downloading is intended to minimally interfere with other communication applications then executing on the website user's computer. The website user's browser displays the downloaded ad(s) to the user only after the user has not interacted with the website for a predefined period of time, such as by neither moving a mouse nor depressing a key on a keyboard during that period. The advertisement server selects those advertisements for downloading to the website user's computer based on a user-ID and preference information of the user and configuration information of the user's computer, which is uploaded from the website user's computer to the advertisement server when a connection is established between the website user's computer and the advertisement server. Though the files associated with an interstitial advertisement can be large, these files are advantageously referenced by the website user's browser during those intervals when the website user's browser is idle and bandwidth utilization of its network connection is relatively low.
By reducing limitations inherent in banners and engendered by download latency, prior forms of interstitial web advertisements, by employing idle time downloading and local caching, provide an improved capability to place advertisements comprising rich media content. However, interstitial advertisements as conventionally implemented continue to have practical deficiencies which limit their use.
Unlike banners, interstitial advertisements involve embedding HTML ad code, as a separate non-displayable object, within HTML coding for a web page. Unfortunately, this technique is inflexible and expensive for an advertiser to implement, especially when the advertiser, for whatever reason, seeks to modify ad content. Ad coding is manually inserted into each and every content web page that is to carry advertising. Consequently, insertion of increasingly sophisticated embedded advertising, such as rich media, in existing website content requires a large investment in terms of human resources, time and cost as websites, particularly large sites, increase a number of content pages available for advertising.
"Push" technology has also been used to attempt to improve on methods of placing and processing interstitial advertisements. A "push" application program on the website user's computer establishes a network connection with a "push" web server, typically during off-hours, such as in the late evening or early morning, or at a predefined interval. The "push" web server then downloads, or "pushes" to the push application advertisements and/or other predefined information that are to be played to the user sometime later. The "push" application stores all the "pushed" content files into a local database on a hard disk at the website user's computer and, in response to instructions received from the "push" web server, deletes those previously "pushed" content files which have already been displayed. The "push" application also maintains a user profile which specifies user preferences as to the specific advertising and/or other information the user wants to receive. As such, through each connection, the "push" web server, by selecting content from its database relative to preferences specified in the user profile, attempts to "push" fresh content to the user's computer without duplicating that which was already displayed. Stored "pushed" content is later displayed either on user demand or during those times when the user is not interacting with the system.
While "push" technology reduces download latency by shifting downloads to occur at off-hours this technology also has drawbacks which limit its practical acceptance. In particular, to access "pushed" content, a website user must initially download and install to the website user's computer a separate, platform-specific, software application program, as well as subsequent updates to that program as new push capabilities are released. These application programs can often require significant amounts of computer memory capacity. Since typical Internet users establish relatively slow modem connections to their Internet service providers, these users will find that downloading such relatively large program files, even in compressed form, will consume an inordinate amount of time and is generally impractical while the website user is actively using the user's computer. Downloading and then regularly updating a "push" application will reduce, sometimes considerably, the amount of disk space available to the website user on the user's computer. Furthermore, relatively few website users will undertake the effort of downloading and installing an application program to receive advertisements and other such information.
Other efforts to reduce the latency problems inherent in such internet advertisements have included development of various proprietary media formats. These formats employ increasingly sophisticated data compression, sometimes in conjunction with video and/or audio streaming. Rather than waiting for a media file to fully download prior to its being rendered, streaming permits content in a "streamed" media file to be presented in real-time to the website user as that content arrives at the website user's browser. While this approach clearly provides enhanced richness in content over that obtainable through a conventional banner, it requires a continuous real-time network connection existing to a remote web server. Any network or server congestion which stops the download, even if temporary, can suspend or halt the "streamed" media presentation to the website user prior to its completion. This interruption, if noticeable and sufficiently long, will likely frustrate the website user.
Still other efforts to address the shortcomings of interstitial advertisements in current forms have involved decoupling advertising content from a web content page, downloading advertising files through a browser executing at a website user's computer into browser caches at that computer and in a manner that is transparent to a user situated at the browser and interstitially displaying advertisements through the browser in response to a user click-stream associated with normal user navigation across different web pages. This technique relies on embedding an HTML tag into a referring page. The embedded tag contains two components. One component downloads from a server and then executes an agent implemented as a Java applet at the website user's browser. This agent then downloads advertising files, originating from an advertisement management system residing on a third-party advertisement server, for a given advertisement into the website user's browser disk or RAM cache and subsequently plays those media files through the browser on an interstitial basis and in response to a user click-stream. The other component is a reference, in terms of a web address, of the advertising management system from which the advertising files are to be downloaded. The ad management system selects the given advertisement that is to be downloaded, rather than having that selection or its content being embedded in the web content page. The agent operates independently of the content in any referring web page. Once loaded and started, the agent executes in parallel, with standard browser functionality, continually and transparently requesting and downloading advertisements to the website user's browser cache residing in a user's computer and interstitially playing those advertisements. In response to a user-initiated action which instructs the user's browser to transition to a next successive content web page and which signifies a start of an interstitial interval, the agent, if all the media and player files are then resident on the user's computer hard disk, plays the media files, through the browser and during that interstitial interval, directly from the browser cache. Advertisements are interstitially played typically in the order in which they were downloaded to the website user's browser. Interstitial play from the website user's browser cache advantageously permits previously cached rich advertisement content to be played through the website user's browser without adversely affecting communication link bandwidth then available to the user's browser. Thus, the full available link bandwidth can be used, while an advertisement is being played, to download a next successive content web page.
While this technique provides better capability to place and provide quick display of rich media advertisements, it still has the drawback of requiring storage of advertising files or agents on website user browser disk or RAM cache space. It also delivers ad files only in the order received. In view of the drawbacks associated with various web based advertising techniques known in the art, interstitial web advertising appears to hold the most promise of all these techniques. Yet, the limitations inherent in conventional implementations of interstitial advertising have effectively prevented this form of web advertising from effectively fulfilling its promise.
The present invention improves over prior interstitial advertising systems and methods. The present invention does not embed advertising HTML files within a web page, providing considerable economies to advertisers in saved labor, time and cost in terms of both inserting advertisements into web page files, and later changing any of those advertisements. The present invention functions totally transparently to a website user and which neither inconveniences nor burdens the user. The present invention does not require a website user to download or install on the website user's computer a separate application program specifically to receive web advertising or perform any affirmative act other than normal web browsing to receive such advertising. The present invention does not consume any browser hard disk or RAM space on the user's computer. The present invention also provides proper accounting to an advertiser by accurately and validly ascertaining user impressions of fully rendered advertisements. It also provides the capability to implement a multi-level advertising business method in which accurate tracking of sales attributable to advertisements placed on websites is assured.
The present invention accomplishes this by use of a dispatcher server that allows the system to isolate and manage a database of links to different pages on a website to a separate server (dispatcher server) database. That server basically allows the system users to link or connect pages in whatever order is desired and insert ad pages (interstitials) in between existing pages on a website. To envision this process, imagine pages on a website, but no links in between them—all links going to the dispatcher server that contains a database of page links for the website and knows how to find such pages (links) on the website. Because the website knows where all pages are it can show an ad (which is another page that may be stored somewhere else) and then jump to the next requested page on the website. In order to jump from the interstitial ad page to a specific page on the website, the dispatcher server, using a query string, passes variables (information) on this page location (and other parameters such as tracking parameters) to the ad file. The ad file can be rich media .swf file, but not necessarily—it can also be a regular HTML or ASP file that contains a timeout for viewing the ad content and a tag or the ability to jump to a destination page.
The present invention, in a preferred embodiment, uses file server direct connection technology to process ad viewer activity tracking data. This allows the system to send a tracking string with information (variables) received from the dispatcher server by the ad file directly to a tracking server. File Server Direct Connection, or "FSDC," is a method to establish a direct connection via the Internet between a self-contained file and a custom server-based application for online secured transactions, statistic tracking and server-based data sharing. FSDC uses a .swf file or alternately an HTML file, which can send a query string directly to the specific URL for a desired server without loading variables from the file (or html code) or requests to the history object of the user's Internet browser. The use of the Macromedia Flash file is preferred because of the ability to provide instructional code within the .swf, and because nearly all client computer systems have been enabled with the Flash player as a result of normal Internet browser configuration. It is to be understood that any type of file that is capable of operation without the initiation of an additional application could be used in place of Flash and the .swf file; however, presently, such is the preferred method of operation. As directed by its internal coding, the .swf ad file establishes a connection, via TCP/IP, with the specified server which is the tracking server, as discussed later. FSDC is described and claimed in the applicant's pending U.S. non-provisional patent application entitled "Method and System For File Server Direct Connection," application Ser. No. 10/316,431 filed on Dec. 11, 2002 and published on Jun. 26, 2003 (Publication No. 20030120727), the specification and figures of which are incorporated herein by reference. The coding of the .swf file establishes a connection to the tracking server so that a tabulation may be made, appropriately noting the response to the ad. As the communication to the tracking server is only made once the ad file has been opened, this is preferred over the prior art methodologies, which initiate the communication for tabulation prior to completing the link to the desired ad or site. In many cases, such tabulation is premature, as the user may close the window before the earlier desired material is delivered.

Internet-based system and method for distributing interstitial advertisements