CN107456573A - Treatment of Bleeding in Hematological Patients and Cancer Patients - Google Patents

Abstract

The present invention provides a method of treating bleeding in a patient prone to bleeding, wherein the method of treatment will reduce the severity of bleeding, the method comprising: the fibrinogen-coated albumin spheres are administered intravenously at a dosage concentration and frequency sufficient to reduce the severity of bleeding in the patient. The invention also provides a product for treating or reducing bleeding, comprising fibrinogen-coated albumin spheres having a diameter in the range of 50 to 500 nanometers and a median diameter of about 140 to 160 nanometers. The products and methods of treatment of the present invention are better able to reduce the severity of bleeding and the amount of bleeding.

Description

Treatment of Bleeding in Hematological Patients and Cancer Patients

technical field

The present invention relates to treatments for controlling or reducing bleeding in blood disease patients and cancer patients. These patients are prone to bleeding due to their disease condition or due to treatment for a hematological or cancer disorder (eg, chemotherapy that produces or increases the likelihood of bleeding). The treatment may be prophylactic or therapeutic, with prophylactic treatment referring to treatment given prior to any active clinical or subclinical bleeding, and therapeutic treatment referring to treatment given when the patient is actively bleeding.

Background technique

There are many types of blood disease patients or cancer patients. Since platelet function and platelet concentration in the body are known to be the main predictors of bleeding, many studies have been conducted to evaluate how low a platelet concentration should be given to a patient for prophylactic platelet transfusion, in the hope that the patient bleeds less frequently or when bleeding Not so serious. One such study is "Bleeding due to thrombocytopenia in acute leukemias and reevaluation of the prophylactic platelet transfusion policy" by Aderka et al. J. Med Sci. 1986 Mar;291(3):147-51).

However, a patient's platelet count is a value obtained at a single point in time when the blood sample is drawn, which does not reveal whether the patient is going through a crisis in which platelets continue to decline severely. Consequently, many studies have revealed that bleeding frequency and clinical severity of bleeding cannot be predicted by patients with platelet counts above or below 20,000 cells/microliter. Furthermore, giving prophylactic platelet transfusions does not prevent such bleeding, so the patient's internal crisis will continue to consume the transfused platelets, resulting in clinical bleeding shortly after the so-called prophylactic transfusion. Such bleeding may be referred to as breakthrough bleeding and will require therapeutic platelet transfusions. Numerous studies have dealt with the so-called magic number of transfused donor platelets, including Wandt, 1998 ("Prophylactic platelet transfusions compared with traditional 20 × 10[9]/L Safety and Cost Effectiveness of a 10×10[9]/L Trigger for 105 Patients with Acute Myeloid Leukemia Prophylactic Platelet Transfusion Compared with the Traditional 20×10[9]/L Trigger: A Prospective Comparative Trial in 105 Patients with Acute Myeloid Leukemia)” (Blood 1998 May, 15:91(10)).

Accordingly, there is a need for products and treatments that reduce the severity and volume of bleeding better than can be done by transfusion of donor platelets.

Contents of the invention

The present invention includes intravenous administration of fibrinogen-coated albumin spheres, which may range in size from 1 micron to 5 microns or from 50 nanometers to 500 nanometers, which administration reduces bleeding in blood disease patients, cancer patients, and other patients, such as by These patients revealed an improvement in the time to bleeding from the surgical incision and in the volume of bleeding. Fibrinogen-coated albumin spheres can be prepared as a "ready-to-use" formulation that is a suspension stable for more than 2 years at room temperature without shaking. Alternatively, the spherical suspension may be lyophilized to a dry powder which would require reconstitution with a liquid compatible with intravenous administration prior to administration to a patient. Patients who may benefit include, but are not limited to, those with acute lymphoblastic leukemia, acute myeloid leukemia, aplastic anemia, myelodysplastic syndrome, idiopathic thrombocytopenic purpura, and those exposed to high doses of radiation . Said dose is at least 4 mg globules per kg of patient body weight.

detailed description

Although specific embodiments of the invention and cases of patients treated will now be described, it should be understood that such embodiments are by way of example only, and merely illustrative of the many possible applications which can only represent the principles of the invention. A small part of the detailed description. Various changes and modifications apparent to those skilled in the art to which the present invention pertains are deemed to be within the spirit, scope and intent of the present invention as further defined in the claims or appended claims.

The manufacture of fibrinogen-coated and non-fibrinogen-coated albumin spheres of medicinal value has been widely disclosed by Dr. Richard C.K. Yen and his works. For example, U.S. Patent No. 5069936 "Manufacturing Protein Microspheres" (Dec. 3, 1991); U.S. Patent No. 9351925 "Submicron Particles for Reducing Blood Transfusion" (May 31, 2016); Large-scale production of ready-to-use suspensions of fibrinogen-coated albumin spheres for the treatment of thrombocytopenic patients" (21 October 2015). Dr. Yen's other patents can be found by searching "Richard C.K. Yen patents" in Google.

However, none of these previous publications describe: how to treat patients based on (a) an effective dose to control or reduce bleeding; (b) the frequency of administering such dose; (c) the beneficial medical outcome obtained by administering an effective dose. These are the subjects of the present invention and disclosure.

In the present invention, it has been found that intravenous administration of fibrinogen-coated albumin spheres can have a beneficial effect in many patients; the size of the spheres can range from 1 micron to 5 microns or from 50 nanometers to 500 nanometers.

Doses of at least 1.75 mg/kg to at least 14 mg/kg of patient body weight have been found to be safe and produce no detectable side effects, so certainly no adverse effects.

It has been found that intravenous administration of fibrinogen-coated albumin globules at a dose of at least 3.5 mg/kg of patient body weight, either in a ready-to-use formulation or after reconstitution from a lyophilized product, reduces and other patients, as revealed by improvements in the time and volume of bleeding from surgical incisions in these patients.

No detectable difference in medical benefit by administering fibrinogen-coated albumin globules as a "ready-to-use" formulation compared to reconstituted lyophilized powder, except for reconstitution requiring withdrawal from another container fluid, which can then be injected to reconstitute the powder, and the process takes at least 15-30 minutes to complete. Otherwise, the lyophilized powder will contain pellets held together by undissolved sugar in the excipient.

The ready-to-use formulation has been found to be a stable suspension for more than 2 years at room temperature without shaking. In addition, for either formulation, there is no need to cross-match the patient's blood or tissue type: both can be used in all patients, including those who for whatever reason have become heterogeneously immune to platelets or have become resistant to platelets. Those patients receiving platelet transfusions.

Patients who may benefit include, but are not limited to, those with acute lymphoblastic leukemia, acute myeloid leukemia, aplastic anemia, myelodysplastic syndrome, idiopathic thrombocytopenic purpura, and those exposed to high doses of radiation patient. The effective dose for reducing bleeding is at least 3.5 mg of balls per kg of patient body weight.

The benefits of administering fibrinogen-coated albumin microspheres or nanospheres are not limited to the patient, but extend to the group that as a whole must bear the cost of the patient's health. About 20,000 new cases of AML (acute myeloid leukemia) are known in the US, Europe and China. Because these patients required so many platelet transfusions, about 7000 of them became "anti-platelet transfusions." The cost of additional care beyond their chemotherapy is about $100,000/year. See Rubella, "A Mini-review on Platelet Refractoriness," Haematologica: 2005 Feb;90(2):247-53.

The present invention shows data supporting that the present invention can be used in these patients (ALL and AML) in place of or in combination with donor platelets. Annual savings will be approximately $100,000 x $7000 = $700 million. Bleeding itself is known to be a contributing factor in death. Therefore, reducing bleeding will not only save money, but may also improve the survival rate of such patients whose bleeding tends to be increased by their chemotherapy or radiotherapy.

Summary of clinical trials showing benefit of fibrinogen-coated albumin spheres administered by the intravenous route

1.1. Summary of Clinical Trial I

The product name used in the following description is "Fibrinoplate", which is a lyophilized product. However, the same is expected for "Fibrinoplate-S" which is a formulation which is a suspension and which is "ready-to-use" in the sense that the suspension formulation has never been lyophilized and therefore does not require reconstitution before intravenous administration to a patient. result. These designations apply to all clinical trials.

Phase I clinical trial report

Venue: Shanghai Second Medical University RJ Medical Center, Shanghai Second Medical University Statistics Teaching and Research Office

summary

Purpose and experimental design:

In order to evaluate the safety of human fibrinogen albumin microspheres (Fibrinoplate), 24 healthy volunteers were divided into 4 groups. Each volunteer received 1.75 mg/kg, 3.5 mg/kg, 7.0 mg/kg or 14 mg/kg of Fibrinoplate by intravenous infusion. Clinical status including heart rate, respiratory rate, blood pressure, skin condition (presence or absence of ecchymosis) was evaluated prior to the infusion and at 1 hour, 2 hours, 24 hours, ) and recorded EKG, derived CBC, BUN, creatinine, liver function tests, renal function tests, coagulation time, PT, aPTT, fibrinogen concentration, plasma P-selectin and immunoglobulin concentration. During the second week, first month and second month after the infusion, antibodies against fibrinogen were checked and the presence of Fibrinoplate in the serum was checked.

result:

None of the dose groups demonstrated adverse effects that could be related to Fibrinoplate during and after the infusion. Clinical vital signs showed no major changes, and all indicators showed no abnormal changes. Anti-fibrinogen antibodies and Fibrinoplate were not detected at the time points tested.

in conclusion:

Fibrinoplate infused at 4 test doses was safe and well tolerated by volunteers, and Fibrinoplate was not immunogenic.

1.2. Summary of Clinical Trial IIa:

Lyophilized Human Fibrinogen Albumin Microspheres

Phase II clinical trial report

Fibrinoplate (freeze-dried human fibrinogen albumin microspheres), is a platelet substitute for short-term use, using human albumin microspheres with a diameter of less than 3 microns as the carrier of human fibrinogen, which is coated with on the surface of the microsphere.

The Institute of Hematology, Chinese Academy of Medical Sciences conducted preclinical studies that established the safety and efficacy of Fibrinoplate and provided the data necessary to initiate a Phase I clinical study. The RJ Medical Center of Shanghai Second Medical University completed a phase I clinical study, which confirmed the safety of Fibrinoplate.

On March 20, 2002, the State Food and Drug Administration (SFDA) of China approved the proposed human Phase II and Phase III clinical studies of Fibrinoplate ( approval number: 2002SL0012 ). RJ Medical Center of Shanghai Second Medical University is one of the most important clinical research centers in China and is responsible for these multi-center clinical trials. The phase II clinical study was completed between June 2002 and December 2003 and was divided into 3 parts: (1) randomized dose evaluation, (2) comparison of results before and after infusion in the same patient and (3) comparison with placebo Comparative randomized double-blind drug evaluation. The results of the randomized dose evaluation study are reported below.

Part 1 of Phase II: Random Dose Evaluation

Venue: Shanghai Second Medical University RJ Medical Center, Peking University People's Medical Center, Shanghai Second Medical University Statistics Department

summary

Objective: To evaluate the efficacy and safety of Fibrinoplate in the treatment of bleeding and bleeding tendency due to thrombocytopenia in patients with acute leukemia (AL)

Patients and Methods:

Acute leukemia (AL) patients with platelet counts equal to or less than 30×10 ⁹ /L and prolonged bleeding time (>8 minutes) were recruited in a random fashion to compare the two doses. Each volunteer received a dose of 3.5 mg/kg or 7.0 mg/kg of reconstituted Fibrinoplate suspension in one intravenous infusion. Bleeding time was the primary clinical endpoint used to determine efficacy. The two doses were evaluated by comparing intragroup and intergroup differences before and after Fibrinoplate infusion using appropriate statistical methods.

result:

A total of 28 patients were recruited. Platelet values and/or bleeding times did not meet the inclusion criteria in 7 patients, therefore, the data obtained from them were not used for statistical analysis and were excluded. Of the 21 patients enrolled for statistical analysis, 12 were in the 3.5 mg dose group and 9 were in the 7.0 mg dose group. In the 3.5 mg dose group, bleeding time was 2315.00±1447.36 seconds before infusion, 1505.00±985.55 seconds at hour 2 (1 hour after infusion), and bleeding time was 25 hours (24 hours after infusion) The time is 1862.50±1127.40 seconds. In the 7.0 mg group, bleeding time was 1963.33±1033.71 seconds before infusion, 1203±509.66 seconds at hour 2 (1 hour after infusion), and 1496.67 seconds at hour 25 (24 hours after infusion) ±659.05 seconds. In the 3.5 mg group, the bleeding time at 2 hours after infusion was 810.00±1466.74 seconds, which was shorter than the bleeding time before infusion (p=0.0283). In the 7 mg group, the bleeding time was shortened by 760.00±761.02 seconds at 2 hours after infusion (p=0.0273). At 2 hours post-infusion, there was no statistically significant difference in reduction in bleeding time between the two dose groups (p=0.9717). At 25 hours post-infusion, the bleeding time was shortened by 452.5±1112.57 seconds (p=0.3633) in the 3.5 mg group and by 466.67±557.45 seconds (p=0.0586) in the 7.0 mg group. At 25 hours post-infusion, there was no statistically significant difference in reduction in bleeding time between the two dose groups (p=0.6437). No adverse events were observed.

in conclusion:

At 3.5 mg/kg and 7.0 mg/kg, Fibrinoplate was effective in shortening the increased bleeding time in all thrombocytopenic AL patients. No statistically significant differences between the two doses were observed and no adverse events were associated with Fibrinoplate infusion.

1.3. Clinical Trial IIb: Summary

Fibrinoplate

Lyophilized Human Fibrinogen Albumin Microspheres

Phase II clinical trial report

Part II: Within-Patient Paired Analysis Before and After Dosing

Venue: Shanghai Second Medical University RJ Medical Center, Peking University People's Medical Center, The First Affiliated Hospital of Zhejiang University School of Medicine, Statistics Department of Shanghai Second Medical University

summary

Objective: To evaluate the efficacy and safety of lyophilized human fibrinogen-coated albumin microspheres (Fibrinoplate) in patients with acute leukemia (AL) who are bleeding or have bleeding tendency due to thrombocytopenia

Patients and Methods:

Acute leukemia (AL) patients with platelet counts ≤30 x 10 ⁹ /L with prolonged bleeding time (as determined by standard bleeding time assays and defined as >8 minutes) were recruited. We used an open-label trial design with a single 7.0 mg/kg dose. The primary clinical endpoint was measured by improvement in bleeding time. We compared bleeding times before and after dosing in each patient (paired test) using appropriate statistical methods.

result:

A total of 35 patients were enrolled; one patient withdrew from the study, leaving a total of 34 patients to complete the study. Bleeding time was significantly shortened at 2 hours post-infusion compared to time 0 (p=0.02482). Bleeding time was not significantly different at 25 hours post infusion compared to time 0 (p=0.05004). No adverse events were observed.

in conclusion:

At 7.0mg/kg, Fibrinoplate can safely shorten the bleeding time associated with thrombocytopenia in patients with acute leukemia.

1.4. Clinical Trial IIc: Summary

Fibrinoplate

Lyophilized Human Fibrinogen Albumin Microspheres

Phase II clinical trial report

Part III: Randomized double-blind drug evaluation compared with placebo

Venue: Shanghai Second Medical University RJ Medical Center, Peking University People's Medical Center, Harbin Hematology Research Center, Tianjin Chinese Academy of Medical Sciences Hematology Research Center, Zhejiang University School of Medicine First Affiliated Hospital, Guangxi Medical University First Affiliated Hospital , Department of Statistics, Shanghai Second Medical University

summary

Objective: To evaluate the efficacy and safety of Fibrinoplate in patients with idiopathic thrombocytopenic purpura (ITP) during an active bleeding episode due to thrombocytopenia or in patients with bleeding tendency due to thrombocytopenia sex

Patients and Methods:

Randomly recruit ITP patients with platelet count ≤30×10 ⁹ /L and prolonged bleeding time (using standard bleeding time determination method, >8 minutes), using double-blind drugs (Fibrinoplate, 7.0mg/kg, 4 vials/dose) Comparative placebo (albumin solution) trial design. The clinical efficacy was evaluated according to the bleeding time. Use appropriate statistical methods to compare the two groups before and after treatment, and to evaluate differences within and between groups.

result:

Of the 101 patients enrolled, 49 received Firinoplate and 52 were in the placebo group. One patient assigned to the placebo group was excluded from the statistical analysis of efficacy as no longer meeting the inclusion criteria (bleeding time <8 seconds).

In the Fibrinoplate group, the bleeding time was 1335.71±714.13 seconds before infusion, 934.29±564.48 seconds at 2 hours after infusion, and 950.10±561.50 seconds at 25 hours after infusion. In the placebo group, the bleeding time was 1425.69±858.66 seconds before infusion, 1420.59±961.05 seconds at 2 hours after infusion, and 1387.25±803.17 seconds at 25 hours after infusion.

Bleeding time was reduced by 401.43±393.3 seconds at 2 hours post-infusion compared to before Fibrinoplate infusion (p=0.00000), while in the placebo group, bleeding time was only reduced by 5.1±314.48 seconds at 2 hours after infusion (p=0.14566). Significant differences that existed between groups were statistically significant (p=0.00000).

At 25 hours after infusion, the bleeding time was reduced by 385.61±607.88 seconds (p=0.00000) in the Fibrinoplate group compared with the bleeding time before infusion, while the bleeding time in the placebo group was only reduced by 38.43±583.29 seconds (p= 0.36350). Differences between groups were also statistically significant (p=0.00000). No adverse effects were observed at any point during the study.

in conclusion:

Fibrinoplate, administered at 7.0 mg/kg, can safely and effectively shorten the bleeding time associated with thrombocytopenia in patients with ITP without any adverse effects.

1.5. Clinical Trial IIIa: Summary

Fibrinoplate

Lyophilized Human Fibrinogen Albumin Microspheres

Phase III clinical trial report

first part

Efficacy study in patients with various difficult-to-treat blood diseases related to thrombocytopenia

Venue: Shanghai Second Medical University RJ Medical Center, Harbin Hematology Research Center, Beijing Z-W China Medical University Research Center, Shanghai China Medical University Affiliated Hospital, Shanghai Second Medical University Statistics Department

summary

Objective: To observe the efficacy of Fibrinoplate in the treatment of patients with acute leukemia (AL), refractory idiopathic thrombocytopenic purpura (ITP) or aplastic anemia (AA) who are actively bleeding or have bleeding tendency due to thrombocytopenia Efficacy and safety in

Patients and methods: Patients were recruited with platelet counts equal to or less than 30 × 10[9]/L. They had prolonged bleeding times (>8 minutes using standard bleeding time assays), and the patients enrolled were AL, ITP, or AA patients. A randomized double-blind drug versus placebo method was used as the primary study method. The drug is Fibrinoplate, 7.0 mg/kg/dose (4 vials/dose); and the placebo is albumin solution or albumin microspheres. Standard bleeding time assays were used, and bleeding time was used as an index to evaluate clinical efficacy. Nonparametric tests were used for statistical analysis of differences within and between groups.

Results: The data showed no statistically significant difference between the two placebo groups, so the two placebo groups were combined into one for the statistical analysis compared to the treatment group. The total number of patients enrolled was 223, of which 112 were in the treatment group and 111 were in the placebo group. Ten cases were excluded and not included in the paired test (PP) analysis. Finally included in the PP analysis were 108 cases in the treatment group and 105 cases in the placebo group. The data from the PP analysis showed that before treatment, the bleeding time of the treatment group was 1431.889±801.9898 seconds, but the bleeding time was shortened to 759.1389±486.9845 seconds at 2 hours after treatment, which was statistically significant (P=0.0000). In the placebo group, the bleeding time before the infusion was 1456.314±871.6902 seconds, and the bleeding time was prolonged to 1509.333±877.0737 seconds at 2 hours after the infusion, which was statistically significant (P=0.0000). At the 25th hour, the bleeding time of the treatment group was shortened to 1121.357±532.0516 seconds, compared with the bleeding time before treatment, the difference was statistically significant (P=0.0000). In the placebo group, the bleeding time was prolonged to 1622.120±849.5374 seconds, which was statistically significantly different from the bleeding time before infusion (P=0.0139). There was a statistically significant difference between the drug group and the placebo group (P=0.0000). No adverse effects that could be related to drug therapy were observed, and 10 patients were discontinued from the study by the investigator due to age or coagulation disorders, resulting in a total of 213 patients completing the study. All 223 patients were included in the statistical analysis.

Among the cases entered into the PP analysis, there were 58 AA patients, 71 ITP patients and 86 AL patients. By disease grouping, statistical analysis showed that the bleeding time of the treatment group at the 2nd hour and the 25th hour after treatment was shorter than that before treatment, and the difference was statistically significant (P=0.0000). Compared with the placebo group, there is also a statistically significant difference (P=0.0000).

CONCLUSIONS: Fibrinoplate at a dose of 7.0 mg/kg significantly reduced prolonged bleeding time associated with thrombocytopenia in patients with AL, refractory ITP, and AA, with no adverse effects observed.

1.6. Clinical Trial IIIb: Summary

Fibrinoplate, lyophilized human fibrinogen albumin microspheres

Phase III clinical trial report

Part II: Patients with Refractory Thrombocytopenia Associated with Active Bleeding, Efficacy Studies

Venue: Shanghai Second Medical University RJ Medical Center, Beijing Z-W China Medical University Research Center, Jiangsu Zhenjiang (Chaing) First People's Hospital, Harbin Hematology Oncology Research Center, Nantong Medical University Affiliated Hospital, Jiangsu Nanjing Medical University Center, Shandong University Qilu Hospital, Shanghai China Medical University Affiliated Hospital, Soochow University First Affiliated Medical Center, Jiangsu Wu Zee First People's Hospital, Zhejiang University School of Medicine First Affiliated Hospital, People's Liberation Army 307th Medical Center, Shanghai Second Medical Department University Statistics Department

summary

OBJECTIVE: To evaluate patients with acute leukemia (AL) associated with thrombocytopenia with active bleeding, refractory idiopathic thrombocytopenic purpura (ITP), aplastic anemia (AA), myelodysplastic syndrome Efficacy and safety of Fibrinoplate in patients with MDS

Patients and Methods:

All recruited patients had platelet counts equal to or less than 30×10[9]/L. At the time of enrollment, patients were actively bleeding or showed signs of bleeding within 24 hours of enrollment, and they were patients with AL, ITP, AA, or MDS. We use an open-ended evaluation method. The drug is 7.0mg/kg/dose of Fibrinoplate (4 bottles/dose), one dose. We use standard bleeding time assays. We evaluated the change in the number of bleeding sites after treatment compared to before treatment, and we used improvement in bleeding time as an indicator of efficacy. We also used cessation of bleeding as an indicator of efficacy. We tabulated the location and number of bleeding sites before and after treatment.

result

A total of 214 patients were recruited, of whom 15 were excluded. We used 199 cases for PP statistical analysis, some of these patients had been treated or treated with other drugs in the past without success in controlling their bleeding. Compared with the condition before treatment, the number of bleeding sites decreased at the 2nd hour, 5th hour and 25th hour after the treatment, with a statistically significant difference (P=0.0000). The bleeding time before treatment was 1359.749±787.2715 seconds, and it was 864.1608±607.8245 seconds in the second hour after treatment, which was statistically significantly different (P=0.0000). Among the recruited patients, 114 were AL patients, 31 were ITP patients, 25 were AA patients, and 43 were MDS patients. Statistical analysis according to disease subgroups showed that all 4 groups of patients demonstrated a reduction in the number of bleeding sites and a reduction in bleeding time, all statistically significantly different from pre-treatment values.

in conclusion:

Fibrinoplate at a dose of 7.0 mg/kg was effective in the treatment of active bleeding with thrombocytopenia in patients with AA, ITP, MDS and AL. It induces cessation of bleeding and is a reliable drug in refractory patients. No adverse effects were observed.

Comments on blood loss in all clinical trials above:

It is often difficult to measure the volume of blood loss in all of the above patient populations because the bleeding is sudden and most of the blood will be washed away on clothing. In a classic and controlled setting, all blood should be captured with pre-weighed sponges so that the weight (indicating volume) of blood on these absorbent materials can be weighed.

To estimate whether blood loss was reduced by administration of reconstituted lyophilized fibrinogen-coated spheres, we took advantage of the substances used in bleeding tests.

In the conventional method of performing a bleeding test, the patient's arm is first secured with a blood pressure cuff so that the arm is compressed to a standard (mild) pressure. A standard incision is then made in the skin of the forearm using an instrument known as the "bleeding time template" (Ivy method). When blood starts to come out of the surgical incision (a shallow incision about 1mm thick and about 1cm wide), touch the flowing blood with a round piece of filter paper, not the wound itself. After about 15 seconds, the filter paper is rotated so that a new adjacent area of the filter paper is available to absorb the continuous flow of blood. These steps are repeated until a clot forms spontaneously at the wound site without direct pressure on the wound: at that point, the clot is so thick that no blood will flow to the new area of the filter paper. The time it takes for bleeding to stop spontaneously under such standard conditions is called the bleeding time. In healthy human volunteers, normal bleeding times are less than 10 minutes.

We used the same blood-stained filter paper as the assessment of blood loss volume. The method was as follows: 1. Blood was collected from the surgical incision made through the blade of the template by the filter paper method described above. 2. Note and record when bleeding stops (bleeding time). 3. After the blood on the filter paper is completely dry, trace the blood-covered area of the filter paper onto another clean, unsoiled filter paper. 4. Cut the clean filter paper along the trail from the rest of the filter paper to obtain the "blood-free dry filter paper" weight. 5. All blood-stained areas (edges of filter paper covered with blood) were similarly cut out, collected and weighed. 6. The weight of the dried blood lost or shed by the patient is obtained by subtracting the dry weight of the blood-free dry filter paper from the weight of all blood-stained filter paper discs.

Results: For example, the bleeding time of one patient before treatment was 16 minutes. The total weight of the blood-stained sheet was 1081 mg and the total weight of an equivalent area of blood-free dry filter paper was 280 mg. Thus, the weight of dried blood lost by the patient was 801 mg.

The same patient showed an improvement in bleeding time of only 12 minutes after treatment with fibrinogen-coated albumin spheres (still abnormal, but improved). The total weight of the blood-stained sheet was 773 mg and the total weight of an equivalent area of blood-free dry filter paper was 210 mg. Thus, the weight of dried blood lost by the patient was 563 mg.

When data were collected for 100 patients, the data indicated that the volume or weight of blood that the patient lost from the surgical incision was proportional to the patient's bleeding time. Thus, the overall improvement in bleeding time in patients treated with fibrinogen-coated spheres (whether lyophilized or non-lyophilized) demonstrates that the treatment improves bleeding and helps reduce the volume or weight of blood loss , regardless of whether the blood loss is visible to an external observer (ie, clinical or subclinical).

1.7. Patients requiring potentially lethal whole body radiation in preparation for stem cell transplantation

There are potentially many patients with acute radiation syndrome who have been exposed to radiation doses high enough to cause death. Such cases were found in the accident at the industrial nuclear power plant at Chernobyl and in Japan after the detonation of a nuclear bomb. But those cases could resurface as terrorists around the world try to use "dirty bombs" -- ordinary explosives mixed with nuclear material that are primarily used to harm civilians and first responders.

In the field of medicine, there are cancer patients whose cancer cannot be eradicated by conventional chemotherapy. Their only hope is whole-body radiation to kill the latest cancer cells, followed by a transplant of stem cells compatible with the patient's tissue type. Such high doses of radiation would be dangerous, as they could kill patients by themselves; however, in some cases, such high doses of radiation may be the only option to ensure that the cancer does not recur.

It is well known that it is unethical to intentionally irradiate human volunteers in order to know whether any drug or treatment is effective in promoting the survival of such irradiated patients. Therefore, in the United States there is legislation passed to allow the approval of drug treatments or methods based on the "two-animal rule". Demonstrated equivalence of disease entities in both animal models to disease states in human populations in both rodent and non-rodent animal models as well as demonstration of mechanism of efficacy between these two animal models and human species Probably the same time, the US FDA will agree to approve.

Work done in the US was presented at the American Society of Hematology meeting in San Francisco, December 2014 (Sung et al., "Fibrinogen-Coated Nanospheres Prevent Thrombocytopenia-Related Bleeding )”, Blood and Marrow Transplantation Biology, February 2015, Vol. 21, No. 2 Supplement, pp. S111-S113). In one mouse model, intravenous administration of ready-to-use fibrinogen-coated nanospheres (where the spheres had a median diameter of less than 500 nm) increased survival from 5% to 60%.

While the work done in the U.S. has not been reproduced in human populations (e.g., cancer patients or others who require whole body radiation as part of their treatment), we believe that the work done in mice is reproducible in human populations , because the mechanism of bleeding and the function of platelets in hemostasis are highly preserved in all animal species.

In particular, there is evidence beyond Sung's published data that the following observations are true:

(A) Four doses given on days 1, 5, 10, and 15 after the day of radiation will result in higher survival and less bleeding than three doses given on days 1, 5, and 10.

(B) Giving 5 doses on days 1, 5, 10, 15 and 20 after the day of radiation will result in a higher survival rate with less bleeding than giving 4 doses in (A).

(C) A more frequent dosing schedule, for example every 3 days instead of every 5 days (as in A or B) will give better results than dosing at intervals of 4 or more days.

(D) A more frequent dosing regimen, eg, daily or every other day, is better than (C).

Although the data above relate to survival after lethal doses of radiation, there is evidence that more frequent dosing and shorter intervals between dosing will help blood disease and cancer patients whose bleeding is not a direct result of radiation.

Survival is clearly a more challenging situation than "bleed less". The term "bleeding" should also be better defined whenever possible. For example, if someone has a broken toe that is bleeding, the expected requirements for recovery or the need for effective treatment are basically simpler than if someone had their entire leg amputated in an accident and had a shattered femur that bleeds. Therefore, improvements in outcomes observed from changes in the dosing regimen of therapeutic products following lethal doses of whole body radiation should also readily translate into improvements in less challenging situations, such as reduced internal bleeding in non-irradiated patients.

Observation Point:

The following comments are conclusions drawn with regard to the present invention.

CLAIMS 1. The present invention is a method of treating bleeding in a patient with a bleeding tendency, wherein said method of treatment will reduce the severity of the bleeding. The method comprises: whether the patient is obese or lean, intravenously administering fibrinogen-coated albumin spheres at a dose concentration defined as the weight of the spheres administered per kilogram of patient weight (usually in mg meter). The frequency of dosing (ie, one, two, or more doses administered over the course of a patient's treatment) is also important in reducing the severity of bleeding in a patient.

2. The present invention is also a method of treating bleeding in patients who are hematologic patients, cancer patients, and radiation patients suffering from one or a combination of the following conditions: idiopathic thrombocytopenic purpura, acute lymphoblastic leukemia , acute myeloid leukemia, aplastic anemia, myelodysplastic syndrome, and whole-body radiation at doses sufficient to be fatal. Together with acute myeloid leukemia, acute lymphoblastic leukemia is called a group: acute leukemia.

3. Considering the possibility that different races may respond differently to drug treatments and treatment modalities, the present invention selected primarily from Asian patients. Thus, the present invention is a method of specifically treating hemorrhage in Asians, although effectiveness in other races is not excluded.

4. The present invention is a method of treating bleeding, wherein said bleeding is not due to a surgical procedure, such as in an operating room, but is clinical bleeding, such as epistaxis, that can be observed by said patient or a trained observer . More serious internal bleeding such as bleeding in the head will need to be seen from changes in neurologic conditions or MRI images.

5. The present invention is a method of treating bleeding, wherein said bleeding is due to a surgical incision, whether the incision is made during surgery or by the blade of a bleeding time measuring device. In other words, the hemorrhage is a wound anywhere on the body produced by a surgical or sharp instrument.

6. The present invention is a method of treating hemorrhage, which is subclinical hemorrhage, including undetectable by an unaided outside observer but caused by changes in laboratory values (for example, a decrease in platelet count and a decrease in red blood cell count or A drop in hemoglobin concentration) detectable internal bleeding.

7. The present invention is a method for treating bleeding, wherein the dosage concentration is at least 3.5 mg balls per kilogram of patient body weight, and the dosage concentration can fully reach the safe dosage concentration for patients. High levels may be 4 times the effective dose, for example 4 x 8 mg/kg (eg, use 8 mg spheres/kg in irradiated animals).

8. The present invention is a method for treating bleeding, wherein the administration frequency (or repetition frequency) is at least once at any time before the patient starts bleeding or at any time during the patient's bleeding.

9. The present invention is a method of treating bleeding, wherein the frequency of administration is more than once, and the drug treatment is repeated daily or less frequently until the bleeding stops.

10. The present invention is a method of treating bleeding, wherein the improvement in the severity of the bleeding is measured by the number of days the patient bleeds while receiving medical care, or by the number of bleeding sites, including the number and size of ecchymosis and petechiae ) to measure the change.

11. The present invention is a method of treating bleeding wherein the severity of the bleeding is measured by an improvement in the patient's bleeding time.

12. The present invention is also a method of treating bleeding in a patient with a bleeding tendency which will reduce the volume of blood loss or the weight of blood shed from the patient.

13. The present invention is also a method of reducing blood loss volume due to hemorrhage in patients with hematological disorders, cancer patients, and radiation-exposed patients suffering from one or a combination of the following conditions: including idiopathic thrombocytopenic Purpura, Acute Lymphoblastic Leukemia, Acute Myeloid Leukemia, Aplastic Anemia, Myelodysplastic Syndrome, and Whole Body Radiation at Doses Sufficient to Lethal. Acute lymphoblastic leukemia is called together with acute myeloid leukemia as a group: acute leukemia.

14. Since it is possible that different races may respond differently to drug treatment and treatment forms, and the subject of the present invention is mainly selected from Asian patients, the present invention is a method to specifically reduce the blood loss volume or blood loss during bleeding in Asians. The weight approach, though, does not preclude validity in other races.

15. The present invention is a method of reducing the volume of bleeding, wherein said bleeding is not due to a surgical incision, such as in an operating room, but is observable by said patient or a trained observer Clinical bleeding, such as epistaxis. More serious internal bleeding such as bleeding in the head will need to be seen from changes in neurologic conditions or MRI images.

16. The present invention is a method of reducing the volume of blood lost during bleeding, wherein said bleeding is due to a surgical incision, whether the incision is made during the surgical procedure or by the blade of a bleeding time measuring device/instrument resulting incision. In other words, the hemorrhage is a wound anywhere on the body produced by a surgical or sharp instrument.

17. The present invention is a method of reducing bleeding that is subclinical, including changes in laboratory values (e.g., a decrease in platelet count and a decrease in red blood cell count or A drop in hemoglobin concentration) detectable internal bleeding.

18. The present invention is a method of reducing the volume of blood loss due to hemorrhage, wherein the effective dosage concentration is at least 3.5 mg spheres per kilogram of patient body weight, and the effective dosage concentration can fully reach the safe dosage concentration for the patient. High levels may be 4 times the effective dose, eg 4 x 8 mg spheres/kg (eg 8 mg spheres/kg in irradiated animals).

19. The present invention is a method of reducing bleeding, wherein the frequency of administration (or repetition frequency) is at least 1 at any time before the patient begins to bleed or at any time while the patient is bleeding.

20. The present invention is a method of reducing the amount of blood lost due to hemorrhage, wherein the frequency of administration is more than once and the drug treatment is repeated daily or less frequently until the bleeding ceases.

21. The invention is a method of reducing bleeding, wherein the improvement in bleeding volume is measured by: (a) a smaller decrease in hemoglobin concentration in vivo compared to a control group not receiving treatment; or (b) compensatory breakthrough or (c) the number of days a patient is bleeding while receiving medical care; or (d) a change in the number of bleeding sites, including the number and size of ecchymosis and petechiae.

22. The present invention is a method of reducing bleeding, wherein the severity of bleeding is measured by improvement in bleeding time in a patient or blood loss during bleeding time assessment.

23. The present invention is also a product for the treatment or reduction of bleeding comprising fibrinogen-coated albumin spheres with a diameter of 1-5 microns and a median diameter of about 2 microns.

24. The present invention is also a product for treating or reducing bleeding comprising fibrinogen-coated albumin globules having a diameter of 50-500 nanometers and a median diameter of about 140-160 nanometers.

25. The present invention is also a product for the treatment or reduction of bleeding, wherein said product is a ready-to-use suspension in which said spheres have never been lyophilized.

26. The present invention is also a product for the treatment or reduction of bleeding wherein said product is a lyophilized product which will require reconstitution with a human plasma compatible fluid prior to administration to a patient in need of hemostasis.

Claims (24)

1. A method of treating bleeding in a patient prone to bleeding, wherein said method of treatment will reduce the severity of the bleeding, said method comprising: administering Frequent intravenous administration of fibrinogen-coated albumin spheres. 2. The method of treating bleeding according to claim 1, wherein said patients include blood disease patients, cancer patients, and radiation-treated patients suffering from a plurality of the following conditions: idiopathic thrombocytopenic purpura, acute lymphocytic Leukemia, acute myeloid leukemia, aplastic anemia, myelodysplastic syndrome, and whole-body radiation at doses sufficient to be fatal. 3. The method of treating bleeding according to claim 2, wherein said patient is of Asian ethnicity. 4. The method of treating bleeding according to claim 1, wherein said bleeding is not surgical bleeding, but clinical bleeding that can be observed by said patient or a trained observer. 5. The method of treating bleeding according to claim 1, wherein said bleeding is surgical bleeding, including a wound anywhere on the body caused by a surgical instrument. 6. The method for treating bleeding according to claim 1, wherein said bleeding is subclinical bleeding, including internal bleeding that cannot be detected by an unaided external observer but can be detected by changes in laboratory values. 7. The method of treating bleeding according to claim 1, wherein said dosage concentration is at least 3.5 mg spheres per kilogram of patient body weight. 8. The method for treating bleeding according to claim 1, wherein the administration frequency is at least once at any time before the patient starts bleeding or at any time during the bleeding of the patient. 9. The method of treating bleeding according to claim 1, wherein the frequency of administration is more than once a day and repeated every day until bleeding stops. 10. The method of treating bleeding according to claim 1 , wherein the severity of the bleeding is measured by the number of days the patient bleeds while receiving medical care, or by the number of bleeding sites, said bleeding sites The number includes the number and size of ecchymosis and petechiae. 11. The method of treating bleeding according to claim 1, wherein the severity of said bleeding is measured by improvement in bleeding time of said patient. 12. A method of treating bleeding in a patient with a bleeding tendency, wherein said method of treatment will reduce the volume of blood lost. 13. The method of treating bleeding according to claim 12, wherein said patients include blood disease patients, cancer patients, and radiation-treated patients suffering from a plurality of the following conditions: idiopathic thrombocytopenic purpura, acute lymphocytic Leukemia, acute myeloid leukemia, aplastic anemia, myelodysplastic syndrome, and whole-body radiation at doses sufficient to be fatal. 14. The method of treating bleeding according to claim 12, wherein said patient is of Asian ethnicity. 15. The method of treating bleeding according to claim 12, wherein said bleeding is not a surgical bleeding, but a clinical bleeding that can be observed by the patient or a trained observer. 16. The method of treating bleeding according to claim 12, wherein said bleeding is surgical bleeding, including a wound anywhere on the body caused by a surgical instrument. 17. The method of treating bleeding according to claim 12, wherein said bleeding is subclinical bleeding, including internal bleeding not detectable by an unaided external observer but detectable by changes in assay values. 18. The method of treating bleeding according to claim 12, wherein said dosage concentration is at least 3.5 mg spheres per kilogram of patient body weight. 19. The method of treating bleeding according to claim 12, wherein the dosing frequency is at least once at any time before the patient starts bleeding or any time during the patient is bleeding. 20. The method of treating bleeding according to claim 12, wherein the frequency of administration is more than once a day and repeated every day until bleeding stops. 21. A product for treating or reducing bleeding comprising fibrinogen-coated albumin spheres, said spheres having a size ranging from 1 to 5 microns and a median diameter of about 2 microns. 22. A product for treating or reducing bleeding comprising fibrinogen-coated albumin spheres, said spheres having a diameter in the range of 50-500 nanometers with a median diameter of about 140-160 nanometers. 23. A product for the treatment or reduction of bleeding, wherein said product is a ready-to-use suspension in which said spheres have never been lyophilized. 24. A product for treating or reducing bleeding, wherein said product is a lyophilized product that will require reconstitution with a fluid compatible with human plasma prior to administration to a patient in need of hemostasis.