ES2779402T3 - Treatment methods for patients with heterozygous familial hypercholesterolaemia (heHFH) - Google Patents

Abstract

A proprotein convertase subtilisin / kexin type 9 inhibitor (PCSK9) for use in the treatment of hypercholesterolaemia in a patient with heterozygous familial hypercholesterolaemia (HeFH) who has a serum LDL-C concentration greater than or equal to 70 mg / dL a despite taking a stable daily dose of statin for at least 4 weeks, and a history of documented cardiovascular disease, where the PCSK9 inhibitor is an antibody or antigen-binding fragment thereof that specifically binds to human PCSK9 and comprises an HCVR and an LCVR having the amino acid sequences of SEQ ID NOs: 1 and 6, respectively.

Description

DESCRIPTION

Treatment methods for patients with heterozygous familial hypercholesterolaemia (heHFH)

FIELD OF THE INVENTION

The present invention relates to the field of therapeutic treatments for diseases and disorders that are associated with elevated levels of lipids and lipoproteins. More specifically, the invention relates to the use of PCSK9 inhibitors to treat patients with heterozygous familial hypercholesterolemia who are not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy.

BACKGROUND

Heterozygous familial hypercholesterolemia (HeFH) is a hereditary disorder of lipid metabolism that predisposes affected individuals to cardiovascular (CV) disease. HeFH patients typically have very high levels of low-density lipoprotein cholesterol (LDL-C) - often> 190 mg / dL at diagnosis - which are associated with a high risk of premature CV disease. The results of observational studies have shown that the risk of coronary heart disease (CHD) is reduced in heFH patients receiving statin therapy; however, even with treatment, the risk of CHD is even higher in heFH patients than in the general population. Despite the availability of lipid-lowering therapy (LTL), approximately 80% of patients with heHF do not reach the recommended levels of LDL-C. Given the high CV risk in the heFH population, there is a need to provide patients with more intense cholesterol-lowering therapy.

Current LDL-C-lowering medications include statins, cholesterol absorption inhibitors (eg, ezetimibe [EZE]), fibrates, niacin, and bile acid sequestrants. Statins are the most commonly prescribed, as they have shown a greater ability to lower LDL-C and reduce CHD events. However, many patients at risk for cardiovascular disease (CVD) have poorly controlled low-density lipoprotein cholesterol (LDL-C) despite statin therapy. The anti-PCSK9 antibodies alirocumab and evolocumab have been used in phase 2 clinical trials to treat patients with heFH (Stein et al., N Engl J Med 2012; 366: 1108-18; Kastelein et al., Cardiovasc Drugs Ther (2014) 28: 281-289), but higher risk patient groups with a history of documented cardiovascular disease were not investigated. These patients have been treated with statins supplemented with additional drugs such as ezetimibe (Sniderman et al., JACC Vol. 63, No. 19, 2014: 1935-47).

BRIEF SUMMARY

The present invention provides a proprotein convertase subtilisin / kexin type 9 (PCSK9) inhibitor for use in the treatment of hypercholesterolemia in a patient with heterozygous familial hypercholesterolemia (heFH) having a serum LDL-C concentration greater than or equal to 70 mg / dL despite taking a stable daily dose of statin for at least 4 weeks, and a history of documented cardiovascular disease, where the PCSK9 inhibitor is an antibody or antigen-binding fragment thereof that specifically binds to PCSK9 human and comprises an HCVR and an LCVR having the amino acid sequences of SEQ ID NOs: 1 and 6, respectively.

According to one aspect, the uses of the present invention comprise administering one or more doses of the PCSK9 inhibitor to the patient with heterozygous familial hypercholesterolemia who is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy (i.e., hypercholesterolaemia that is not adequately controlled by statin therapy at the maximum tolerated dose in the absence of a PCSK9 inhibitor, with or without other lipid modifying therapy). According to certain embodiments of the present invention, the PCSK9 inhibitor is administered to the patient with heterozygous familial hypercholesterolemia as an adjunct therapy to the patient's existing statin therapy with or without other lipid-lowering therapy.

According to another aspect, the uses of the present invention comprise selecting a patient with heterozygous familial hypercholesterolemia who is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy (eg, maximum dose statin therapy. tolerated), and administering to the patient one or more doses of a PCSK9 inhibitor in combination with (ie, "in addition to") statin therapy.

A method of treating a patient with heterozygous familial hypercholesterolemia (heFH) that is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy is further disclosed by administering one or more doses of a proprotein convertase inhibitor / subtilisin. kexin type 9 (PCSK9) to the patient, where the patient has inadequate control of hypercholesterolemia despite treatment with statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy in the absence of the PCSK9 inhibitor.

A method of lowering low-density lipoprotein cholesterol (LDL-C) in a patient with heterozygous familial hypercholesterolemia (HeFH) that is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy is further disclosed. administering one or more doses of a proprotein convertase subtilisin / kexin type 9 inhibitor (PCSK9) to the patient, where the patient has inadequate control of hypercholesterolemia despite treatment with statin therapy at the maximum tolerated dose with or without another lipid-lowering therapy in the absence of the PCSK9 inhibitor.

A method of treating hypercholesterolemia in a patient with heterozygous familial hypercholesterolaemia (heFH) is further disclosed that is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy administering one or more doses of a proprotein convertase inhibitor. subtilisin / kexin type 9 (PCSK9) to the patient, where the patient has inadequate control of hypercholesterolemia despite treatment with statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy in the absence of the PCSK9 inhibitor.

A method of improving the serum level of one or more lipid components in a patient with heterozygous familial hypercholesterolaemia (heFH) who is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy by administering one or more is further disclosed. more doses of a proprotein convertase subtilisin / kexin type 9 inhibitor (PCSK9) to the patient, where the patient has inadequate control of the lipid component despite treatment with statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy in absence of PCSK9 inhibitor. In certain aspects, there is a decrease in the serum level of a lipid component selected from the group consisting of LDL-C, Apo B, non-HDL C, total cholesterol, Lp (a), and triglycerides. In certain aspects, there is an increase in the serum level of a lipid component selected from the group consisting of HDL-C and Apo A1.

The diagnosis of heFH can be made by genotyping or by clinical criteria. In some respects, the clinical criteria is either the Simon Broome Register Diagnostic Criteria for Heterozygous Familial Hypercholesterolemia, or the WHO / Dutch Lipid Network criteria with a score> 8.

In certain aspects of the invention, the antibody or antigen-binding fragment thereof that specifically binds to PCSK9 is administered to the patient at a dose of about 75 mg at a frequency of once every two weeks. In some aspects, the dose of approximately 75 mg is maintained if the patient's LDL-C measured after five or more doses is <70 mg / dL. In some aspects, the approximately 75 mg dose is discontinued if the patient's LDL-C measured after five or more doses remains> 70 mg / dL, and the antibody or antigen-binding fragment thereof that specifically binds to PCSK9 is subsequently administered to the patient at a dose of approximately 150 mg at a frequency of once every two weeks. In some aspects, the antibody or antigen-binding fragment thereof that specifically binds to PCSK9 is administered to the patient at a dose of about 150 mg at a frequency of once every two weeks.

In certain aspects of the invention, the PCSK9 inhibitor is administered to the patient in combination with statin therapy at the maximum tolerated dose. In some aspects, statin therapy at the maximum tolerated dose comprises a daily dose of from about 40 mg to about 80 mg of atorvastatin. In some aspects, statin therapy at the maximum tolerated dose comprises a daily dose of about 20 mg to about 40 mg of rosuvastatin. In some aspects, statin therapy at the maximum tolerated dose comprises a daily dose of approximately 80 mg of simvastatin.

In certain aspects of the invention, the PCSK9 inhibitor is administered to the patient in combination with the other lipid-lowering therapy.

In certain aspects of the invention, the method improves at least one parameter associated with hypercholesterolemia selected from the group consisting of: (a) lowering the patient's low-density lipoprotein (LDL-C) cholesterol by at least 40%; (b) reduction of the patient's apolipoprotein B (ApoB) by at least 30%; (c) reduction of the patient's non-high-density lipoprotein (non-HDL C) cholesterol by at least 40%; (d) reduction of the patient's total cholesterol by at least 20%; (e) increase in the patient's high-density lipoprotein cholesterol (HDL-C) by at least 3%; (f) reduction of the patient's triglycerides by at least 5%; (g) reduction of the patient's lipoprotein a (Lp (a)) by at least 20%; and (h) increase in the patient's apolipoprotein A1 by at least 1%.

Other embodiments of the present invention will be apparent from a review of the following detailed description.

BRIEF DESCRIPTION OF THE FIGURES

Figure 1 is a graphical representation of the study design for ODYSSEY FH I (Example 2).

Figure 2 is a graph showing the mean percent change in LDL-C-CM calculated from baseline over time for alirocumab or placebo treatment in the IDT population in the ODYSSEY study. FH I (Example 2). Least squares means (LC) and standard errors (SD) are taken from the MMRM analysis (mixed effects model with repeated measures).

Figure 3 is a graphical representation of the study design for ODYSSEY FH II (Example 3).

Figure 4 is a graph showing the mean percent change of MC (+/- SE) of LDL-C from baseline over time for the IDT population in the ODYSSEY FH II study (Example 3). Least squares means (LC) and standard errors (SE) are taken from the MMRM analysis (mixed effects model with repeated measures).

Figure 5 is a graph showing the mean percent change in MC (+/- SE) of LDL-C from baseline during the efficacy treatment period over time for the mTDI population in the ODYSSEY FH II study ( Example 3).

Figure 6 is a graphical representation of the study design for ODYSSEY HIGH FH (Example 4). The markers in the study design are defined as follows: FU: follow-up; HeFH, heterozygous familial hypercholesterolemia; THL, lipid-lowering therapy; OLE, open label extension.

Figure 7 is a graph showing the mean percent change in MC of LDL-C calculated from baseline with time for treatment with alirocumab or placebo in the IDT population in the ODYSSEY HIGH FH study (Example 4). Least squares means (LC) and standard errors (SE) are taken from the MMRM analysis (mixed effects model with repeated measures).

Figure 8 is a graph showing MC (SE) mean calculated LDL-C values versus time for ODYSSEY FH I and FH II studies. The values indicated in the graph are the change in mean% MC from baseline to Week 24 and Week 52.

Figure 9 is a graph showing mean calculated LDL-C values of MC (SE) versus time for the ODYSSEY FH I and FH II studies. The values indicated below the graph are the numbers of patients analyzed at the various moments in time.

Figure 10 is a graph showing LDL-C levels over time in alirocumab patients according to whether the dose increased to 150 mg Q2W or remained at 75 mg Q2W (TDI analysis).

Figure 11 represents graphs showing subgroup analyzes of LDL-C reductions from baseline to Week 24 (alirocumab vs. placebo) based on demographic characteristics and baseline characteristics (A), statin / THL use. (B), and lipids at baseline (C) (IDT analysis; pooled data from FH I and FH II). Moderate chronic kidney disease (CKD) was defined as an estimated glomerular filtration rate of> 30 and <60 mL / min / 1.73 m2. In FH I, 20/323 and 9/163 patients in the alirocumab and placebo arms had moderate CKD at baseline. The corresponding values in FH II were 2/167 and 1/82. "High intensity" statin dose refers to atorvastatin 40-80 mg or rosuvastatin 20-40 mg.

Figure 12 is a graphical representation of patient disposition in the ODYSSEY HIGH FH study.

Figure 13 is a graph showing the percent change from baseline to Week 24 in LDL-C levels by individual patients in the ODYSSEY HIGH FH study. All patients were on a background statin (at the maximum tolerated level). A subset of patients also received additional lipid-lowering therapy.

Figure 14 represents graphs showing mean calculated MC (SE) LDL-C values versus time for the ODYSSEY HIGH FH study. In part A., the values indicated in the graph are the mean% MC% values (in mg / dL) at Week 24 and Week 52. In part B., the values indicated in the graph are the values mean% MC% (in mg / dL) at Week 24 and Week 78. All patients were on a background statin (at the maximum tolerated level). A subset of patients also received additional lipid-lowering therapy.

DETAILED DESCRIPTION

It should be understood that the present invention is not limited to the particular methods and experimental conditions described, as such methods and conditions may vary. It should also be understood that the terminology used herein is for the purpose of describing particular aspects only, and is not intended to be limiting, as the scope of the present invention will only be limited by the appended claims.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning that is commonly understood by one of ordinary skill in the art to which this disclosure belongs. As used herein, the term "about", when used in reference to a particular quoted numerical value, means that the value can vary from the quoted value by no more than 1%. By For example, as used herein, the term "about 100" includes 99 and 101 and all values in between (eg, 99.1.99.2, 99.3, 99.4, etc.).

Heterozygous familial hypercholesterolaemia not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy

The present disclosure relates generally to methods and compositions for treating patients with heterozygous familial hypercholesterolemia who are not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy, i.e., hypercholesterolemia not adequately controlled by a therapeutic regimen comprising a maximum daily tolerated dose of a statin. As used herein, the term "not adequately controlled" in reference to hypercholesterolemia means that the concentration of low-density lipoprotein (LDL-C) cholesterol in the patient's serum, total cholesterol concentration, and / or concentration triglycerides is not reduced to a recognized medically acceptable level (taking into account the patient's relative risk of coronary heart disease) after at least 4 weeks on a therapeutic regimen comprising a stable daily dose of a statin. For example, "a patient with hypercholesterolemia that is not adequately controlled by a statin" includes patients with a serum LDL-C concentration greater than approximately 70 mg / dL, 100 mg / dL, 130 mg / dL, 140 mg / dL. , or more (depending on the patient's underlying heart disease risk) after the patient has been on a stable daily statin regimen for at least 4 weeks.

Patients with heterozygous familial hypercholesterolemia who are not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy who are treatable by the methods of the present disclosure may have hypercholesterolemia (for example, an LDL-C concentration in serum greater than or equal to 70 mg / dL in patients with a history of documented cardiovascular disease or a serum LDL-C> 100 mg / dL in patients without a history of documented cardiovascular disease) despite taking a stable daily dose of a statin (with or without other lipid modifying therapy) for at least 4 weeks, 5 weeks, 6 weeks, or more. The hypercholesterolemia of the patient with heterozygous familial hypercholesterolemia may be inadequately controlled by maximum tolerated dose statin therapy (also referred to herein as "a maximum daily tolerated dose therapeutic statin regimen").

As used herein, "statin therapy at the maximum tolerated dose" means a therapeutic regimen comprising the administration of a daily dose of a statin that is the maximum tolerated dose for a particular patient. The maximum tolerated dose means the highest dose of statin that can be administered to a patient without causing unacceptable adverse side effects in the patient. Statin therapy at the maximum tolerated dose includes, but is not limited to, for example, atorvastatin 40-80 mg daily, rosuvastatin 20-40 mg daily, or simvastatin 80 mg (if you are already at this dose for> 1 year). However, patients unable to tolerate the above statin doses could take a lower dose of atorvastatin, rosuvastatin, or simvastatin daily provided there was an acceptable reason for not using the higher dose. Some examples of acceptable reasons for a patient to take a lower dose of statin include: adverse effects with higher doses, older age, low body mass index (BMI), regional practices, local drug label, concurrent medications, and comorbid conditions such as impaired fasting glucose / glucose intolerance.

The present disclosure also includes methods of treating patients with heterozygous familial hypercholesterolemia that are not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy comprising daily administration of other statins such as cerivastatin, pitavastatin, fluvastatin, lovastatin and pravastatin.

Patient selection

The present disclosure includes methods and compositions useful for treating patients with heterozygous familial hypercholesterolemia who are not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy.

The diagnosis of heFH must be made either by genotyping or by clinical criteria. For non-genotyped patients, clinical diagnosis may be based on either the Simon Broome criteria with a criterion for defined HF or the WHO / Dutch Lipid Network criteria with a score> 8 points.

According to the Simon Broome Register Diagnostic Criteria for Heterozygous Familial Hypercholesterolemia, defined familial hypercholesterolemia is defined as: 1) Total C> 6.7 mmol / L (260 mg / dL) or LDL cholesterol greater than 4.0 mmol / L (155 mg / dL) in a child <16 years or total C> 7.5 mmol / L (290 mg / dL) or LDL cholesterol greater than 4.9 mmol / L (190 mg / dL) in an adult . (Levels or pre-treatment or the highest during treatment); more or A) tendinous xanthomas in the patient, or in a 1st degree relative (father, brother, son), or in a 2nd degree relative (grandfather, uncle, aunt); or B) DNA-based evidence of a familial defective LDL or apo B-100 receptor mutation.

According to the Simon Broome Register Diagnostic Criteria for Heterozygous Familial Hypercholesterolemia, possible familial hypercholesterolemia is defined as: 1) Total C> 6.7 mmol / L (260 mg / dL) or higher LDL cholesterol to 4.0 mmol / L (155 mg / dL) in a child <16 years or total C> 7.5 mmol / L (290 mg / dL) or LDL cholesterol greater than 4.9 mmol / L (190 mg / dL) in an adult. (Levels or pre-treatment or the highest in treatment); and at least one of the following: A) family history of MI less than 50 years of age in a 2nd degree relative or less than 60 years of age in a 1st degree relative; and B) a family history of elevated cholesterol> 7.5 mmol / L (290 mg / dL) in an adult relative of the 1st or 2nd degree or> 6.7 mmol / L (260 mg / dL) in a son or brother of less than 16 years of age.

The WHO criteria (Dutch Lipid NetWork clinical criteria) for the diagnosis of heterozygous familial hypercholesterolemia (HeFH) are set out in the examples, as in Table 2.

The patient with heterozygous familial hypercholesterolemia can be selected on the basis of having one or more additional risk factors selected from the group consisting of age (for example, older than 40, 45, 50, 55, 60, 65, 70, 75, or 80 age), race, national origin, gender (male or female), exercise habits (for example, person who exercises regularly, person who does not exercise), other pre-existing medical conditions (for example, type II diabetes, high blood pressure , myocardial infarction, ischemic stroke, etc.), and current medication status (for example, currently taking beta-blockers, niacin, ezetimibe, fibrates, omega-3 fatty acids, bile acid resins, etc.).

Patients with heterozygous familial hypercholesterolemia can be selected based on a combination of one or more of the above selection criteria or therapeutic characteristics.

Administration of a PCSK9 inhibitor as add-on therapy to statin therapy at the maximum tolerated dose

The present disclosure includes methods wherein a patient with heterozygous familial hypercholesterolemia who is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy in the absence of a PCSK9 inhibitor is administered with a PCSK9 inhibitor according to an amount and particular frequency of administration, and wherein the PCSK9 inhibitor is administered as an adjunct to the patient's therapeutic statin regimen. For example, if a patient with heterozygous familial hypercholesterolaemia who is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy comprising, for example, atorvastatin 40-80 mg, the patient with heterozygous familial hypercholesterolemia will be can be administered with a PCSK9 inhibitor at a particular amount and dosage interval while the patient continues his or her stable daily therapeutic statin regimen.

The methods of the present disclosure include complementary therapeutic regimens wherein the PCSK9 inhibitor is administered as adjunct therapy to the same therapeutic statin regimen at the maximum stable daily tolerated dose (i.e., same amount of statin administration) in which the patient he was at risk for heterozygous familial hypercholesterolemia before receiving the PCSK9 inhibitor. The PCSK9 inhibitor can be administered as add-on therapy to a therapeutic statin regimen at the maximum tolerated daily dose comprising a statin in an amount that is higher or lower than the statin dose the patient was on prior to receiving the inhibitor. by PCSK9. For example, after starting a therapeutic regimen comprising a PCSK9 inhibitor administered at a particular frequency and amount of administration, the daily dose of statin administered or prescribed to the patient may (a) remain the same, (b) increase, or (c ) decrease (for example, dose titration up or dose adjustment down) compared to the daily statin dose that the patient at high cardiovascular risk was taking prior to the start of the PCSK9 inhibitor therapy regimen, depending on needs therapies of the patient.

Therapeutic efficacy

The methods of the present disclosure will result in improvement in the serum level of one or more lipid components selected from the group consisting of LDL-C, ApoB, non-HDL-C, total cholesterol, HDL-C, triglycerides, Apo A- 1 and Lp (a). For example, administration of a pharmaceutical composition comprising a PCSK9 inhibitor to a patient with heterozygous familial hypercholesterolaemia who is not adequately controlled by a therapeutic statin regimen at the maximum stable daily tolerated dose (for example, administration of the PCSK9 inhibitor in addition to of statin therapy at the patient's maximum tolerated dose) may result in a mean percentage reduction from baseline in serum low-density lipoprotein cholesterol (LDL-C) of at least about 40%, 41 %, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, or greater; an average percentage reduction from baseline in ApoB of at least approximately 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41% , 42%, 43%, 44%, 45%, or greater; a mean percentage reduction from baseline in non-HDL C of at least about 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, or greater; a mean percentage reduction from baseline in total cholesterol of at least about 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31 %, 32%, 33%, 34%, 35%, or greater; a mean percentage increase from baseline in HDL-C of at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15% or greater; an average reduction in percentage from baseline in triglycerides of at least about 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, or greater; a mean percentage increase from baseline in Apo A-1 of at least approximately 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, or greater ; and / or a mean percentage reduction from baseline in Lp (a) of at least approximately 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29% , 30%, 31%, 32%, 33%, 34%, 35%, or higher.

PCSK9 inhibitors

The methods of the present disclosure comprise administering to a patient with heterozygous familial hypercholesterolemia who is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy a therapeutic composition comprising a PCSK9 inhibitor. As used herein, a "PCSK9 inhibitor" is any agent that binds or interacts with human PCSK9 and inhibits the normal biological function of PCSK9 in vitro or in vivo. Examples of categories of PCSK9 inhibitors include small molecule PCSK9 antagonists, peptide-based PCSK9 antagonists (eg, "peptibody" molecules), and antibodies or antigen-binding fragments of antibodies that specifically bind to human PCSK9. .

The term "proprotein convertase subtilisin / human kexin type 9" or "human PCSK9" or "hPCSK9", as used herein, refers to PCSK9 having the nucleic acid sequence shown in SEQ ID NO: 197 and the amino acid sequence of SEQ ID NO: 198, or a biologically active fragment thereof.

The term "antibody", as used herein, is intended to refer to immunoglobulin molecules comprising four polypeptide chains, two heavy (H) chains, and two light (L) chains interconnected by disulfide bonds, as well as multimers of the themselves (eg IgM). Each heavy chain comprises a heavy chain variable region (abbreviated herein HCVR or V ^h ) and a heavy chain constant region. The heavy chain constant region comprises three domains, C ^h 1, C ^h 2, and C ^h 3. Each light chain comprises a light chain variable region (abbreviated herein LCVR or V ^l ) and a constant region of the light chain. The constant region of the light chain comprises a domain ( ^C1 ). The V ^h and V ^l regions can be further subdivided into regions of hypervariability, termed complementary determining regions (CDRs), interspersed with regions that are more conserved, termed framework regions (FR). Each V ^h and V ^l is composed of three CDRs and four FRs, arranged from the amino end to the carboxy end in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The FRs of the anti-PCSK9 antibody (or antigen-binding portion thereof) may be identical to human germline sequences, or they may be naturally or artificially modified. A consensus amino acid sequence can be defined based on a direct comparative analysis of two or more CDRs.

The term "antibody", as used herein, also includes antigen-binding fragments of whole antibody molecules. The terms "antigen-binding portion" of an antibody, "antigen-binding fragment" of an antibody, and the like, as used herein, include any naturally occurring, enzymatically obtainable, synthetic polypeptide or glycoprotein. , or genetically engineered, that specifically binds to an antigen to form a complex. Antigen-binding fragments of an antibody can be derived, for example, from whole antibody molecules using any suitable conventional technique, such as proteolytic digestion or recombinant genetic engineering techniques involving the manipulation and expression of variable and optionally constant domains. DNA-encoding antibodies. Such DNAs are known as and / or readily available from, for example, commercial sources, DNA libraries (including, for example, phage-antibody libraries), or can be synthesized. DNA can be sequenced and manipulated chemically or using molecular biology techniques, for example, to arrange one or more variable and / or constant domains in a suitable configuration, or to introduce codons, create cysteine residues, modify, add or delete amino acids , etc.

Examples of antigen-binding fragments include: (i) Fab fragments; (ii) F (ab ') 2 fragments; (iii) Fd fragments; (iv) Fv fragments; (v) single chain Fv molecules (scFv); (vi) dAb fragments; and (vii) minimal recognition units consisting of amino acid residues that mimic the hypervariable region of an antibody (e.g., an isolated complementarity determining region (CDR) such as a CDR3 peptide), or a FR3-CDR3 peptide. -FR4 limited. Also encompassed within the term "antigen-binding fragment" as used herein are other engineered molecules, such as domain-specific antibodies, single-domain antibodies, domain-deleted antibodies, chimeric antibodies, grafted antibodies. with CDRs, diabodies, tribodies, tetrabodies, minibodies, nanobodies (eg, monovalent nanobodies, bivalent nanobodies, etc.), small modular immunpharmaceuticals (SMIPs), and shark variable IgNAR domains.

An antigen-binding fragment of an antibody will typically comprise at least one variable domain. The variable domain can be of any size or amino acid composition and will generally comprise at least one CDR that is adjacent or in frame with one or more framework region sequences. In antigen-binding fragments that have a V ^h domain associated with a V ^l domain, the V ^h and V ^l domains can be located in any suitable arrangement with respect to each other. For example, the variable region can be dimeric and contain V ^h -V ^h , V ^h -V ^l, or V ^l -V ^l dimers. Alternatively, the antigen-binding fragment of an antibody may contain a monomeric VH or VL domain.

An antigen-binding fragment of an antibody can contain at least one variable domain covalently linked to at least one constant domain. Exemplary variable and constant domain configurations that may be found within an antigen-binding fragment of an antibody of the present disclosure include: (i) V ^h -C ^h 1; (ii) V ^h -C ^h 2; (iii) V ^h -C ^h 3; (iv) V ^h -C ^h 1 -C ^h 2; (v) V ^h -C ^h 1 -C ^h 2 -C ^h 3; (vi) V ^h -C ^h 2 -C ^h 3; (vii)

V ^h -C ^l ; (viii) V ^l -C ^h 1; (ix) V ¹ -C ^h 2; (x) V ^l -C ^h 3; (xi) V ^l -C ^h 1 -C ^h 2; (xii) V ^l -C ^h 1 -C ^h 2 -C ^h 3; (xiii) V ^l -C ^h 2 -C ^h 3; y (xiv any configuration of variable and constant domains, including any of the example configurations listed above, the variable and constant domains can be either directly linked to each other, or they can be linked by a hinge or connector region total or partial. A hinge region may consist of at least 2 (eg, 5, 10, 15, 20, 40, 60 or more) amino acids that result in a flexible or semi-flexible bond between adjacent variable and / or constant domains in a single polypeptide molecule.

Furthermore, an antigen-binding fragment of an antibody may comprise a homodimer or heterodimer (or other multimer) of any of the variable and constant domain configurations listed above in non-covalent association with each other and / or with one or more V ^h domains. or V ^l monomeric (eg by disulfide bond (s)).

As with full length antibody molecules, antigen-binding fragments can be monospecific or multispecific (eg, bispecific). A multispecific antigen-binding fragment of an antibody will normally comprise at least two different variable domains, where each variable domain is capable of specifically binding a separate antigen or a different epitope on the same antigen. Any multispecific antibody format, including the exemplary bispecific antibody formats disclosed herein, can be adapted for use in the context of an antigen-binding fragment of an antibody described herein, using routine techniques. available in the art.

The constant region of an antibody is important in an antibody's ability to bind complement and mediate cell-dependent cytotoxicity. Thus, the isotype of an antibody can be selected based on whether it is desired that the antibody mediates cytotoxicity.

The term "human antibody", as used herein, is intended to include antibodies that have variable and constant regions derived from human germline immunoglobulin sequences. The human antibodies of the disclosure may, however, include amino acid residues not encoded by human germline immunoglobulin sequences (eg, mutations introduced by random or site-specific mutagenesis in vitro or by somatic mutation in vivo), for example For example, in CDRs and in particular CDR3.

However, the term "human antibody", as used herein, is not intended to include antibodies in which CDR sequences derived from the germline of other mammalian species have been grafted onto sequences of the human framework region. like a mouse.

The term "recombinant human antibody", as used herein, is intended to include all human antibodies that are prepared, expressed, created, or isolated by recombinant means, such as antibodies expressed using a recombinant expression vector transfected into a host cell. (described below), antibodies isolated from a recombinant combinatorial library of human antibodies (described below), antibodies isolated from an animal (e.g., a mouse) that is transgenic for human immunoglobulin genes (see, e.g., Taylor et al. (1992) Nucl. Acids Res. 20: 6287-6295) or antibodies prepared, expressed, created, or isolated by any other means that involves splicing of human immunoglobulin gene sequences with other DNA sequences . Such recombinant human antibodies have variable and constant regions derived from human germline immunoglobulin sequences. In certain cases, however, such recombinant human antibodies are subjected to in vitro mutagenesis (or, when a transgenic animal is used for human Ig sequences, to somatic mutagenesis in vivo) and thus the amino acid sequences of the V ^h and Recombinant antibody V ^l are sequences that, although derived from and related to human germline V ^h and V ^l sequences, cannot naturally exist within the human germline antibody repertoire in vivo.

Human antibodies can exist in two forms that are associated with hinge heterogeneity. In one form, an immunoglobulin molecule comprises a stable four-chain construct of approximately

150-160 kDa in which the dimers are held together by an interchain heavy chain disulfide bond. In a second form, the dimers are not linked via interchain disulfide bonds and a molecule of approximately 75-80 kDa is formed composed of a covalently coupled heavy and light chain (half antibody). These forms have been extremely difficult to separate, even after affinity purification.

The frequency of occurrence of the second form in the various isotypes of intact IgG is due to, but is not limited to, structural differences associated with the isotype of the hinge region of the antibody. A single amino acid substitution in the hinge region of human IgG4 can significantly reduce the appearance of the second form (Angal et al. (1993) Molecular Immunology 30: 105) to levels normally observed using

a human IgG1 hinge. The present disclosure encompasses antibodies that have one or more mutations in the hinge, C ^h 2 or C ^h 3 region that may be desirable, for example, in production, to improve the performance of the desired form of antibody.

An "isolated antibody", as used herein, means an antibody that has been identified and separated and / or recovered from at least one component of its natural environment. For example, an antibody that has been separated or removed from at least one component of an organism, or from a tissue or cell in which the antibody exists naturally or occurs naturally, is an "isolated antibody" for the purposes herein. divulgation. An isolated antibody also includes an antibody in situ within a recombinant cell. Isolated antibodies are antibodies that have undergone at least one purification or isolation step. An isolated antibody can be substantially free of other cellular material and / or chemicals.

The term "specifically binds to", or the like, means that an antibody or antigen-binding fragment thereof forms a complex with an antigen that is relatively stable under physiological conditions. Methods of determining whether an antibody specifically binds to an antigen are well known in the art, and include, for example, equilibrium dialysis, surface plasmon resonance, and the like. For example, an antibody that "specifically binds to" PCSK9, as used herein, includes antibodies that bind to PCSK9 or a portion thereof with a K ^{d of} less than about 1000 nM, less than about 500 nM, less than about 300 nM, less than about 200 nM, less than about 100 nM, less than about 90 nM, less than about 80 nM, less than about 70 nM, less than about 60 nM, less than about 50 nM, less than about 40 nM, less than about 30 nM, less than about 20 nM, less than about 10 nM, less than about 5 nM, less than about 4 nM, less than about 3 nM, less than about 2 nM, less than about 1 nM or less than about 0.5 nM, as measured in a surface plasmon resonance assay. An isolated antibody that specifically binds human PCSK9 may, however, have cross-reactivity with other antigens, such as PCSK9 molecules from other (non-human) species.

Anti-PCSK9 antibodies useful for the methods of the present disclosure may comprise one or more amino acid substitutions, insertions and / or deletions in the framework region and / or CDR regions of the variable domains of the heavy and light chain as compared to the ones corresponding germline sequences from which the antibodies were derived. Such mutations can be easily established by comparing the amino acid sequences disclosed herein with germline sequences available from, for example, public antibody sequence databases. The present disclosure includes methods involving the use of antibodies, and antigen-binding fragments thereof, which are derived from any of the amino acid sequences disclosed herein, wherein one or more amino acids within one or more framework regions and / or CDR regions are mutated to the corresponding residue (s) of the germline sequence from which the antibody was derived, or to the corresponding residue (s) of another human germline sequence, or to a conservative amino acid substitution of the corresponding germline residue (s) (such sequence changes are collectively referred to as "germline mutations"). One of ordinary skill in the art, starting from the heavy and light chain variable region sequences disclosed herein, can readily produce numerous antibodies and antigen-binding fragments comprising one or more individual germline mutations or combinations of the same. All framework region and / or CDR residues within the V ^h and / or V ^l domains can be back-mutated to residues found in the original germline sequence from which the antibody was derived. Alternatively, only certain residues are back-mutated to the original germline sequence, for example, only the mutated residues found within the first 8 amino acids of FR1 or within the last 8 amino acids of FR4, or only the mutated residues found within CDR1, CDR2 or CDR3. One or more of the framework region and / or CDR residues are mutated to the corresponding residues of a different germline sequence (i.e., a germline sequence that is different from the germline sequence germ cell from which the antibody was originally derived). In addition, the antibodies of the present disclosure may contain any combination of two or more germline mutations within framework and / or CDR regions, for example, where certain individual residues are mutated to the corresponding residue of a particular sequence of the germline while certain other residues that differ from the original germline sequence remain or are mutated to the corresponding remainder of a different germline sequence. Once obtained, antibodies and antigen-binding fragments containing one or more germline mutations can be readily tested for one or more desired properties such as improved binding specificity, increased binding affinity, enhanced antagonist or agonist biological properties. or enhanced (as the case requires), reduced immunogenicity, etc. The use of antibodies and antigen-binding fragments obtained in this general way is encompassed within the present invention.

The present disclosure also includes methods that involve the use of anti-PCSK9 antibodies comprising variants of any of the HCVR, LCVR and / or CDR amino acid sequences disclosed herein that have one or more conservative substitutions. For example, the present disclosure includes the use of anti-PCSK9 antibodies that have amino acid sequences of HCVR, LCVR and / or CDR with, for example, 10 or minus, 8 or less, 6 or less, 4 or less, etc., conservative amino acid substitutions with respect to any of the HCVR, LCVR and / or CDR amino acid sequences disclosed herein.

The term "surface plasmon resonance", as used herein, refers to an optical phenomenon that allows analysis of interactions in real time by detecting alterations in protein concentrations within a biosensor matrix, for example using the BIAcore ™ system (Biacore Life Sciences, a division of GE Healthcare, Piscataway, NJ).

The term "K ^d ", as used herein, is intended to refer to the equilibrium dissociation constant of a particular antibody-antigen interaction.

The term "epitope" refers to an antigenic determinant that interacts with a specific antigen-binding site in the variable region of an antibody molecule known as a paratope. A single antigen can have more than one epitope. Thus, different antibodies can bind to different areas on an antigen and can have different biological effects. Epitopes can be either conformational or linear. A conformational epitope is produced by spatially juxtaposed amino acids from different segments of the linear polypeptide chain. A linear epitope is one produced by adjacent amino acid residues in a polypeptide chain. In certain circumstances, an epitope can include saccharide moieties, phosphoryl groups, or sulfonyl groups on the antigen.

The anti-PCSK9 antibody used in the methods of the present disclosure can be an antibody with pH-dependent binding characteristics. As used herein, the term "pH-dependent binding" means that the antibody or antigen-binding fragment thereof exhibits "reduced binding to PCSK9 at acidic pH compared to neutral pH" (for the purposes of the present disclosure, both expressions can be used interchangeably). For example, antibodies "with pH-dependent binding characteristics" include antibodies and antigen-binding fragments thereof that bind PCSK9 with higher affinity at neutral pH than at acidic pH. The antibodies and antigen-binding fragments of the present disclosure can bind to PCSK9 with an affinity of at least 3, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, or more times higher, at neutral pH than at acidic pH.

According to this aspect of the disclosure, anti-PCSK9 antibodies with pH-dependent binding characteristics can possess one or more amino acid variations relative to the parent anti-PCSK9 antibody. For example, an anti-PCSK9 antibody with pH-dependent binding characteristics may contain one or more histidine substitutions or insertions, for example, in one or more CDRs of a parent anti-PCSK9 antibody. Thus, methods are disclosed which comprise administering an anti-PCSK9 antibody comprising CDR amino acid sequences (eg, heavy and light chain CDRs) that are identical to the CDR amino acid sequences of a parent anti-PCSK9 antibody, except for the substitution of one or more amino acids of one or more CDRs of the parent antibody with a histidine residue. Anti-PCSK9 antibodies with pH-dependent binding can possess, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, or more, histidine substitutions, either within a single CDR of a parental antibody or distributed across multiple (eg, 2, 3, 4, 5 or 6) CDRs of a parental anti-PCSK9 antibody. For example, the present disclosure includes the use of pH-dependent binding anti-PCSK9 antibodies comprising one or more histidine substitutions in HCDR1, one or more histidine substitutions in HCDR2, one or more histidine substitutions in HCDR3, one or more more histidine substitutions in LCDR1, one or more histidine substitutions in LCDR2, and / or one or more histidine substitutions in LCDR3, of a parent anti-PCSK9 antibody.

As used herein, the term "acidic pH" means a pH of 6.0 or less (eg, less than about 6.0, less than about 5.5, less than about 5.0, etc.). ). The term "acidic pH" includes pH values of about 6.0, 5.95, 5.90, 5.85, 5.8, 5.75, 5.7, 5.65, 5.6, 5, 55, 5.5, 5.45, 5.4, 5.35, 5.3, 5.25, 5.2, 5.15, 5.1.5.05, 5.0, or less. As used herein, the term "neutral pH" means a pH of from about 7.0 to about 7.4. The term "neutral pH" includes pH values of about 7.0, 7.05, 7.1,7,15, 7.2, 7.25, 7.3, 7.35, and 7.4.

Preparation of human antibodies

Methods of generating human antibodies in transgenic mice are known in the art. Any such known method can be used in the context of the present disclosure to prepare human antibodies that specifically bind to human PCSK9.

Using VELOCIMMUNE ™ technology (see, for example, US Patent 6,596,541, Regeneron Pharmaceuticals) or any other known method of generating monoclonal antibodies, high affinity chimeric antibodies are initially isolated for PCSK9 having a human variable region and a constant region of mouse. The VELOCIMMUNE® technology involves generation of a transgenic mouse having a genome comprising human heavy and light chain variable regions operably linked to endogenous mouse constant region loci such that the mouse produces an antibody comprising a human variable region and a mouse constant region in response to antigenic stimulation. DNA encoding the antibody heavy and light chain variable regions is isolated and operably linked to DNA encoding the regions human heavy and light chain constants. The DNA is then expressed in a cell capable of expressing the fully human antibody.

Generally, a VELOCIMMUNE ™ mouse is challenged with the antigen of interest, and lymphatic cells (such as B lymphocytes) are recovered from the mice expressing the antibodies. Lymphatic cells can be fused with a myeloma cell line to prepare immortal hybridoma cell lines, and such hybridoma cell lines are screened and selected to identify hybridoma cell lines that produce antibodies specific for the antigen of interest. DNA encoding the heavy chain and light chain variable regions can be isolated and linked to desirable isotypic heavy chain and light chain constant regions. Such an antibody protein can be produced in a cell, such as a CHO cell. Alternatively, DNA encoding antigen-specific chimeric antibodies or light and heavy chain variable domains can be isolated directly from antigen-specific lymphocytes.

Initially, high affinity chimeric antibodies having a human variable region and a mouse constant region are isolated. Antibodies are characterized and selected for desirable characteristics, including affinity, selectivity, epitope, etc., using standard procedures known to those of skill in the art. The mouse constant regions are substituted for a desired human constant region to generate the fully human antibody of the disclosure, eg, non-mutant or modified IgG1 or IgG4. While the selected constant region may vary according to specific use, high affinity antigen binding and target specificity characteristics reside in the variable region.

In general, the antibodies that can be used in the methods of the present disclosure possess high affinities, as described above, when measured by binding to antigen either immobilized on solid phase or in solution phase. Mouse constant regions are substituted for desired human constant regions to generate the fully human antibodies of the disclosure. Although the constant region selected may vary according to specific use, high affinity antigen binding and target specificity characteristics reside in the variable region.

Specific examples of human antibodies or antigen-binding fragments of antibodies that specifically bind to PCSK9 that can be used in the context of the methods of the present disclosure include any antibody or antigen-binding fragment that comprises the three CDRs of the heavy chain (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) that has an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 and 11, or a substantially similar sequence thereof having at least 90%, at least 95%, at least 98%, or at least 99% sequence identity. Alternatively, specific examples of human antibodies or antigen-binding fragments of antibodies that specifically bind to PCSK9 that can be used in the context of the methods of the present disclosure include any antibody or antigen-binding fragment comprising all three CDRs. of the heavy chain (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) having an amino acid sequence selected from the group consisting of SEQ ID NOs 37, 45, 53, 61, 69, 77 , 85, 93, 101, 109, 117, 125, 133, 141, 149, 157, 165, 173, 181 and 189, or a substantially similar sequence thereof that has at least 90%, at least 95%, at less 98% or at least 99% sequence identity. The antibody or antigen-binding fragment may comprise all three light chain CDRs (LCVR1, LCVR2, LCVR3) contained within a light chain variable region (LCVR) having an amino acid sequence selected from the group consisting of SEQ ID NOs 6 and 15, or a substantially similar sequence thereof that has at least 90%, at least 95%, at least 98%, or at least 99% sequence identity. Alternatively, the antibody or antigen-binding fragment may comprise all three light chain CDRs (LCVR1, LCVR2, LCVR3) contained within a light chain variable region (LCVR) having an amino acid sequence selected from the group consisting in SEQ ID NOs 41,49, 57, 65, 73, 81, 89, 97, 105, 113, 121, 129, 137, 145, 153, 161, 169, 177, 185 and 193, or a substantially similar sequence of the same one that has at least 90%, at least 95%, at least 98% or at least 99% sequence identity.

The antibody or antigen-binding fragment thereof may comprise the six CDRs (HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3) of the pairs of heavy and light chain variable region amino acid sequences (HCVR / LCVR ) selected from the group consisting of SEQ ID NOs: 1/6 and 11/15. Alternatively, the antibody or antigen-binding protein may comprise the six CDRs (HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3) of the pairs of heavy and light chain variable region amino acid sequences (HCVR / LCVR ) selected from the group consisting of SEQ ID NOs: 37 / 41,45 / 49, 53/57, 61/65, 69/73, 77 / 81,85 / 89, 93/97, 101/105, 109/113 , 117/121, 125/129, 133/137, 141/145, 149/153, 157/161, 165/169, 173/177, 181/185 and 189/193.

The anti-PCSK9 antibody, or antigen-binding fragment thereof, which can be used in the methods of the present disclosure has amino acid sequences of HCDR1 / HCDR2 / HCDR3 / LCDR1 / LCDR2 / LCDR3 selected from SEQ ID NOs: 2 / 3/4/7/8/10 (mAb316P) and 12/13/14/16/17/18 (mAb300N) (see US Patent Application Publication No. 2010/0166768).

The antibody or antigen-binding fragment thereof may comprise HCVR / LCVR amino acid sequence pairs selected from the group consisting of SEQ ID NOs: 1/6 and 11/15. Alternatively, the antibody or antigen-binding protein may comprise pairs of the HCVR / LCVR amino acid sequence selected from the group consisting of SEQ ID NOs: 37 / 41,45 / 49, 53/57, 61/65, 69 / 73, 77 / 81,85 / 89, 93/97, 101/105, 109/113, 117/121, 125/129, 133/137, 141/145, 149/153, 157/161, 165/169, 173/177, 181/185 and 189/193.

Pharmaceutical compositions and methods of administration

The present disclosure includes methods comprising administering a PCSK9 inhibitor to a patient with heterozygous familial hypercholesterolemia who is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy, wherein the PCSK9 inhibitor is contained within a pharmaceutical composition. The pharmaceutical compositions are formulated with suitable carriers, excipients, and other agents that provide adequate transfer, delivery, tolerance, and the like. A multitude of suitable formulations can be found in the form known to all pharmaceutical chemists: Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, PA. These formulations include, for example, powders, pastes, ointments, jellies, waxes, oils, lipids, lipid-containing vesicles (cationic or anionic) (such as LIPOFECTIN ™), DNA conjugates, anhydrous absorption pastes, oil emulsions in water and water-in-oil, Carbowax emulsions (polyethylene glycols of various molecular weights), semisolid gels and semisolid mixtures containing Carbowax. See also Powell et al. "Compendium of excipients for parenteral formulations" PDA (1998) J Pharm Sci Technol 52: 238-311.

Various delivery systems are known and can be used to deliver the pharmaceutical composition, eg, encapsulation in liposomes, microparticles, microcapsules, recombinant cells capable of expressing the mutant viruses, receptor-mediated endocytosis (see, for example, Wu et al. , 1987, J. Biol. Chem. 262: 4429-4432). Methods of administration include, but are not limited to, intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, and oral routes. The composition can be administered by any convenient route, for example by infusion or bolus injection, by absorption through the epithelial or mucocutaneous linings (eg, oral mucosa, rectal and intestinal mucosa, etc.) and can be administered in conjunction with other biologically active agents.

A pharmaceutical composition of the present disclosure can be administered subcutaneously or intravenously with a standard needle and syringe. Furthermore, with respect to subcutaneous administration, a pen delivery device easily has applications in administering a pharmaceutical composition of the present disclosure. Such a pen delivery device can be reusable or disposable. A reusable pen delivery device generally uses a replaceable cartridge that contains a pharmaceutical composition. Once all of the pharmaceutical composition has been delivered into the cartridge and the cartridge is empty, the empty cartridge can easily be discarded and replaced with a new cartridge containing the pharmaceutical composition. The pen delivery device can then be reused. In a disposable pen delivery device, there is no replaceable cartridge. Instead, the disposable pen delivery device comes pre-filled with the pharmaceutical composition contained in a reservoir within the device. Once the pharmaceutical composition reservoir is emptied, the entire device is discarded.

Numerous reusable pen and autoinjector delivery devices have applications in the subcutaneous administration of a pharmaceutical composition of the present disclosure. Examples include, but are not limited to, AUTOPEN ™ (Owen Mumford, Inc., Woodstock, UK), DISETRONIC ™ pen (Disetronic Medical Systems, Bergdorf, Switzerland), HUMALOG MIX 75/25 ™ pen, HUMALOG ™ pen, HUMALIN pen 70/30 ™ (Eli Lilly and Co., Indianapolis, IN), No Vo PEN ™ I, II and III (Novo Nordisk, Copenhagen, Denmark), NOVOPEN JUNIOR ™ (Novo Nordisk, Copenhagen, Denmark), Bd ™ pen ( Becton Dickinson, Franklin Lakes, NJ), OPTIPEN ™, OPTIPEN PRO ™, OpT iPEN St A r LET ™ and OpTICLIK ™ (Sanofi-aventis, Frankfurt, Germany), to name just a few. Examples of disposable pen delivery devices that have applications in the subcutaneous administration of a pharmaceutical composition of the present disclosure include, but are not limited to, the SOLOSTAR ™ pen (Sanofi-aventis), FLEXPEN ™ (Novo Nordisk), and KWIKPEN. ™ (Eli Lilly), SURECLICK ™ autoinjector (Amgen, Thousand Oaks, c A), PENLET ™ (Haselmeier, Stuttgart, Germany), Ep IPEN (Dey, LP), and HUMIRA ™ pen (Abbott Labs, Abbott Park IL) , to name just a few.

In certain situations, the pharmaceutical composition can be administered in a controlled release system. In one situation, a pump can be used (see Langer, above; Sefton, 1987, CRC Crit. Ref. Biomed. Eng. 14: 201). In another situation, polymeric materials can be used; See, Medical Applications of Controlled Release, Langer and Wise (eds.), 1974, CRC Pres., Boca Raton, Florida. In yet another situation, a controlled release system can be placed in proximity to the target of the composition, thus requiring only a fraction of the systemic dose (see, for example, Goodson, 1984, in Medical Applications of Controlled Release, above , vol. 2, pp. 115-138). Other controlled release systems are discussed in the review by Langer, 1990, Science 249: 1527-1533.

Injectable preparations may include dosage forms for intravenous, subcutaneous, intracutaneous and intramuscular injections, drip infusions, etc. These injectable preparations can be prepared by known methods. For example, injectable preparations can be prepared, for example, by dissolving, suspending or emulsifying the antibody or its salt described above in a sterile aqueous medium or an oily medium conventionally used for injections. As the aqueous medium for injections there are, for example, physiological saline solution, an isotonic solution containing glucose and other auxiliary agents, etc., which can be used in combination with an appropriate solubilizing agent such as an alcohol (for example, ethanol), a polyalcohol (eg, propylene glycol, polyethylene glycol), a nonionic surfactant [eg, polysorbate 80, HCO-50 (hydrogenated castor oil polyoxyethylene (50 mole) adduct)], etc. As the oil medium, for example, sesame oil, soybean oil, etc. are employed, which can be used in combination with a solubilizing agent such as benzyl benzoate, benzyl alcohol, etc. The injection thus prepared is preferably filled into an appropriate ampoule.

Advantageously, the pharmaceutical compositions for oral or parenteral use described above are prepared in pharmaceutical forms in a unit dose suitable to adjust to a dose of the active principles. Such dosage unit dosage forms include, for example, tablets, pills, capsules, injections (ampoules), suppositories, etc.

Dosage

The amount of PCSK9 inhibitor (eg, anti-PCSK9 antibody) administered to a patient with heterozygous familial hypercholesterolemia that is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy according to the methods of the present disclosure it is, in general, a therapeutically effective amount. As used herein, the term "therapeutically effective amount" means a dose of PCSK9 inhibitor that results in a detectable improvement (at least about 5%, 10%, 15%, 20%, 25%, 30% , 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or more from baseline) in one or more parameters selected from the group consisting of LDL-C, ApoB, non-HDL C, total cholesterol, HC-LDL, triglycerides, Apo A-1 and Lp (a).

In the case of an anti-PCSK9 antibody, a therapeutically effective amount can be from about 0.05 mg to about 600 mg, for example, about 0.05 mg, about 0.1 mg, about 1.0 mg, about 1, 5mg, about 2.0mg, about 10mg, about 20mg, about 30mg, about 40mg, about 50mg, about 60mg, about 70mg, about 75mg, about 80mg, about 90mg, about 100mg, about 110mg, about 120mg, about 130mg, about 140mg, about 150mg, about 160mg, about 170mg, about 180mg, about 190mg, about 200mg, about 210mg, about 220mg , about 230 mg about 240 mg, about 250 mg, about 260 mg, about 270 mg about 280 mg, about 290 mg, about 300 mg, about 310 mg, about 320 mg, about 330 mg, about 340 mg, about 350 mg, about 360 mg, about 370 mg, about 380 mg, about 390 mg, about 400 mg, about 410 mg, about 420 mg, about 430mg about 440mg, about 450mg, about 460mg, about 470mg about 480mg, about 490mg, about 500mg, about 510mg about 520mg, about 530mg, about 540mg, about 550mg about 560mg , about 570 mg, about 580 mg, about 590 mg, or about 600 mg, of the anti-PCSK9 antibody.

The amount of anti-PCSK9 antibody contained within individual doses can be expressed in terms of milligrams of antibody per kilogram of the patient's body weight (ie, mg / kg). For example, the anti-PCSK9 antibody can be administered to a patient at a dose of from about 0.0001 to about 10 mg / kg of the patient's body weight.

Combination therapies

As described elsewhere herein, the methods of the present disclosure may comprise administering a PCSK9 inhibitor to patients with heterozygous familial hypercholesterolemia in combination with the patient's previously prescribed stable daily maximum tolerated dose therapeutic statin regimen. Additional therapeutic agents, in addition to a statin, can be administered to the patient in combination with the PCSK9 inhibitor. Examples of such additional therapeutic agents include, for example, (1) an agent that inhibits cholesterol uptake and or re-absorption of bile acids (eg, ezetimibe); (2) an agent that increases lipoprotein catabolism (such as niacin); and / or (3) LXR transcription factor activators that play a role in removing cholesterol such as 22-hydroxycholesterol.

Administration regimes

Multiple doses of a PCSK9 inhibitor (ie, a pharmaceutical composition comprising a PCSK9 inhibitor) may be administered to a subject over a defined time course (eg, in addition to a daily therapeutic statin regimen). This comprises sequentially administering to a patient with heterozygous familial hypercholesterolemia who is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy multiple doses of a PCSK9 inhibitor. As used herein, "administer sequentially" means that each dose of PCSK9 inhibitor is administered to the subject at a different point in time, eg, on different days separated by a predetermined interval (eg, hours, days , weeks or months). The present disclosure includes methods that comprise sequentially administering to the patient with heterozygous familial hypercholesterolemia a single initial dose of a PCSK9 inhibitor, followed by one or more secondary doses of the PCSK9 inhibitor, and optionally followed by one or more tertiary doses of the PCSK9 inhibitor. .

The terms "initial dose", "secondary dose" and "tertiary dose" refer to the sequence of temporal administration of individual doses of a pharmaceutical composition comprising a PCSK9 inhibitor. Thus, the "starting dose" is the dose given at the beginning of the treatment regimen (also called the "starting level dose"); "Secondary doses" are the doses given after the initial dose; and "tertiary doses" are the doses that are administered after the secondary doses. The initial, secondary, and tertiary doses may all contain the same amount of the PCSK9 inhibitor, but in general they can be differentiated from each other in terms of frequency of administration. However, the amount of PCSK9 inhibitor contained in the initial, secondary and / or tertiary doses may vary from each other (eg, adjusted up or down as appropriate) during the course of treatment. Two or more doses (eg, 2, 3, 4, or 5) may be administered at the beginning of the treatment regimen as a "loading dose", followed by subsequent doses that are given on a less frequent basis (eg, "doses of maintenance").

For example, each secondary and / or tertiary dose is administered 1 to 26 (for example, 1, 1 /, 2, 21/2, 3, 31/2, 4, 41/2, 5, 51/2, 6, 6 /, 7, 7 /, 8, 8 /, 9, 9 /, 10, 10 /, 11, 11 /, 12, 12 /, 13, 13 /, 14, 14 /, 15, 15 /, 16, 16 /, 17, 17 /, 18, 18 /, 19, 1 9 /, 20, 20 /, 21, 21 /, 22, 22 /, 23, 23 /, 24, 24 /, 25, 25 /, 26 , 26 /, or more) weeks after the immediately preceding dose. The term "immediately preceding dose", as used herein, means, in a sequence of multiple administrations, the dose of antigen-binding molecule that is administered to a patient prior to administration of the next dose in the sequence without intermediate doses.

This may comprise administering to a patient with heterozygous familial hypercholesterolemia any number of secondary and / or tertiary doses of a PCSK9 inhibitor. For example, only a single secondary dose can be administered to the patient. Thus, two or more (eg, 2, 3, 4, 5, 6, 7, 8, or more) secondary doses can be administered to the patient. Also, only a single tertiary dose can be administered to the patient. Thus, two or more (eg, 2, 3, 4, 5, 6, 7, 8, or more) tertiary doses can be administered to the patient.

In cases involving multiple secondary doses, each secondary dose can be administered at the same frequency as the other secondary doses. For example, each secondary dose can be administered to the patient with heterozygous familial hypercholesterolemia 1 to 2, 4, 6, 8, or more weeks after the immediately preceding dose. Similarly, in cases involving multiple tertiary doses, each tertiary dose can be administered at the same frequency as the other tertiary doses. For example, each tertiary dose can be administered to the patient 1 to 2, 4, 6, 8, or more weeks after the immediately preceding dose. Alternatively, the frequency at which secondary and / or tertiary doses are administered to a patient may vary over the course of the treatment regimen. The frequency of administration can also be adjusted during the course of treatment by a physician depending on the needs of the individual patient following the clinical examination.

The present disclosure includes administration regimens that comprise an upward titration option (also referred to herein as "dose modification"). As used herein, an "up-titration option" means that, after receiving a particular number of doses of a PCSK9 inhibitor, if a patient has not achieved a specified reduction in one or more defined therapeutic parameters , the dose of the PCSK9 inhibitor is increased from here on. For example, in the case of a therapeutic regimen comprising the administration of 75 mg doses of an anti-PCSK9 antibody to a patient with heterozygous familial hypercholesterolemia who is not adequately controlled by statin therapy at the maximum tolerated dose with or without other lipid-lowering therapy at a frequency of once every two weeks, if after 8 weeks (i.e. 5 doses administered in Week 0, Week 2 and Week 4, Week 6 and Week 8), the patient has not reached a concentration of Serum LDL-C less than 70 mg / dL, then the anti-PCSK9 antibody dose is increased to, for example, 150 mg given once every two weeks thereafter (for example, starting at Week 12).

The anti-PCSK9 antibody can be administered to a subject at a dose of about 75 mg every two weeks, for example for at least six doses.

The antibody can be administered to a subject at a dose of approximately 75 mg every two weeks for 12 weeks, and the dose remains at 75 mg every two weeks if, at Week 8, the subject's LDL-C value was lower. at 70 mg / dL.

The antibody can be administered to a subject at a dose of approximately 75 mg every two weeks for 12 weeks, and the dose is adjusted upward to approximately 150 mg every two weeks if, at Week 8, the LDL-C value of the subject was greater than or equal to 70 mg / dL.

The anti-PCSK9 antibody can be administered to a subject at a dose of about 150 mg every two weeks, for example for at least six doses.

Examples

The following examples are proposed in such a way as to provide those of ordinary skill in the art with a full disclosure and description of how to prepare and use the methods and compositions of the disclosure, and are not intended to limit the scope of what is claimed. Efforts have been made to ensure accuracy with respect to the numbers used (eg quantities, temperature, etc.), but some experimental errors and deviations must be taken into account. Unless otherwise indicated, parts are parts by weight, molecular weight is average molecular weight, temperature is in degrees centigrade, and pressure is atmospheric or near atmospheric.

Example 1. Generation of human antibodies against human PCSK9

Human anti-PCSK9 antibodies were generated as described in US Patent No. 8,062,640. The exemplary PCSK9 inhibitor used in the following examples is the human anti-PCSK9 antibody designated "mAb316P", also known as "Alirocumab". mAb316P has the following amino acid sequence characteristics: heavy chain variable region (HCVR) comprising SEQ ID NO: 1; light chain variable domain (LCVR) comprising SEQ ID NO: 6; heavy chain complementarity determining region 1 (HCDR1) comprising SEQ ID NO: 2; HCDR2 comprising SEQ ID NO: 3; HCDR3 comprising SEQ ID NO: 4; light chain complementarity determining region 1 (LCDR1) comprising SEQ ID NO: 7; LCDR2 comprising SEQ ID NO: 8; and LCDR3 comprising SEQ ID NO: 10.

Example 2: A randomized, double-blind, placebo-controlled, parallel-group study to evaluate the efficacy and safety of alirocumab in patients with heterozygous familial hypercholesterolemia not adequately controlled by their lipid-modifying therapy

Introduction

This study included patients with heterozygous familial hypercholesterolaemia (heFH) with or without a history of documented myocardial infarction (MI) or ischemic stroke.

The aim of the study was to evaluate the efficacy and safety of alirocumab in patients with heFH and who required additional pharmacological management, since their current lipid modifying therapy (TML) failed to achieve the goal of LDL-C treatment.

This study (Figure 1) was conducted to demonstrate in patients with HeFH who are not in their LDL-C target that alirocumab 75 mg Q2W or 75 mg Q2W / 150 mg Q2W as add-on therapy to statin ± other TML causes a statistically significant reduction and clinically significant in LDL-C. This non-target LDL-C population with optimized TML represents the highest risk group with a well-identified unmet medical need that can be treated by adding alirocumab to their LDL-C-lowering therapies.

Objectives of the study

The primary objective of the study was to demonstrate the reduction of LDL-C by alirocumab as add-on therapy to stable maximum tolerated daily statin therapy with or without other TML compared to placebo after 24 weeks of treatment in patients with heHF.

The secondary objectives of the study were: 1) to evaluate the effect of alirocumab 75 mg compared to placebo on LDL-C after 12 weeks of treatment; 2) evaluate the effect of alirocumab on other lipid parameters (ie Apo B, non-HDL C, total C, Lp (a), HDL-C, TG levels and Apo A-1 levels); 3) evaluate the long-term effect of alirocumab on LDL-C; 4) evaluate the safety and tolerability of alirocumab; 5) evaluate the development of anti-alirocumab antibodies; and 6) evaluating the PK of alirocumab.

Study design

It was a randomized, double-blind, placebo-controlled, parallel-group, unbalanced (2: 1, alirocumab: placebo), multicenter, multinational study to evaluate the efficacy and safety of alirocumab in patients with HeHF not adequately controlled with your TML (ie maximum stable tolerated daily statin therapy ± other TML). Not adequately controlled was defined as LDL-C> 70 mg / dL (1.81 mmol / L) at the screening visit (Week -3) in patients with a history of documented cardiovascular disease or LDL-C> 100 mg / dL (2.59 mmol / L) at the screening visit (Week -3) in patients with no history of documented cardiovascular disease. Randomization was stratified by prior history of MI or ischemic stroke [Yes / No], statin therapy (atorvastatin 40 to 80 mg daily or rosuvastatin 20 to 40 mg per day versus simvastatin whatever the daily dose, atorvastatin below 40 mg per day or rosuvastatin below 20 mg per day) and geographical region. After randomization, patients received double-blind study treatment (or alirocumab or placebo) Q2W for a period of 18 months (78 weeks) in addition to stable maximum tolerated daily statin therapy ± other TML. Upward dose adjustment may occur at Week 12 depending on LDL-C levels at Week 8 for patients randomized to alirocumab. After the completion of the 18-month double-blind treatment period, all patients who successfully completed the study had the opportunity to participate in an open-label extension study. Consequently, all patients received alirocumab at entry into the open-label extension study regardless of the study treatment received during the 18-month double-blind treatment period.

The study consisted of 3 periods: screening, double-blind treatment, and follow-up.

The screening period was up to 3 weeks in length that includes an intermediate visit during which the patient (or other designated person such as spouse, relative, etc.) received training to self-inject / inject with placebo for alirocumab. Eligibility assessments were performed to allow for randomization of patients in the study.

The double-blind treatment period (DBTP) was a treatment period in the 18-month randomized double-blind study. The first injection during the double-blind period was made at the site on the day of randomization (Week 0 [D1] - V3). Subsequent injections were made by the patient (self-injection) or another designated person (such as spouse, relative, etc.) at a location preferred by the patient (home ...). Patients randomized to alirocumab received a 75 mg dose of Investigational Pharmaceutical Specialty (IMP) from Randomization (V3) to Week 12 (V6) (i.e. Weeks 0, 2, 4, 6, 8, and 10) . At the Week 12 visit (V6), these patients, in a blinded mode, or: 1) continued with alirocumab 75 mg Q2W from Week 12 onwards until the last injection at Week 76, if LDL-C in Week 8 was <70 mg / dL (1.81 mmol / L); or 2) dose titration up to alirocumab 150 mg Q2W from Week 12 onwards to the last injection at Week 76, if the LDL-C at Week 8 was> 70 mg / dL (1.81 mmol / L ).

The follow-up period (if applicable) was a period of 8 weeks after the end of DBTP for patients who did not consent to participate in the open-label extension study or prematurely discontinued study treatment.

Laboratory measurement of lipid parameters was performed by a central laboratory during the study.

Patients who achieved 2 consecutive calculated LDL-C levels <25 mg / dL (0.65 mmol / L) during the study were monitored and managed.

Statin and other TML (if applicable) should be stable (including dose) for the first 24 weeks of DBTP, except in exceptional circumstances where primary concerns warrant such changes. In Week 24 onwards, the background TML can be modified only under certain conditions as described below.

Patients must be on a stable diet (Therapeutic Lifestyle Changes [TLC] diet of NCEP-ATPIII or equivalent) for the entire duration of the study from selection. Table 1 provides a summary of the TLC diet for high cholesterol.

Table 1

Total fat 25% - 35% of total calories *

Saturated fat * <7% of total calories

Polyunsaturated fat up to 10% of total calories

Monounsaturated fat up to 20% of total calories

Carbohydrates of 50% - 60% of total calories *

Protein ~ 15% of total calories

Cholesterol <200 mg / day (5.172 mmol / day)

Plant sterols 2 g

Soluble fiber such as psyllium 1 0 g - 25 g

* ATP III allows an increase in total fat up to 35 percent of total calories and a reduction in carbohydrate up to 50 percent for people with metabolic syndrome. Any increase in fat intake It must be in the form of either polyunsaturated or monounsaturated fat. Trans fatty acids are another LDL-raising fat that should be kept low.

f Carbohydrate should be derived predominantly from foods rich in complex carbohydrates including grains - especially whole grain fruits and vegetables.

Study duration included a screening period of up to 3 weeks, a 78-week DBTP for efficacy and safety assessment, and an 8-week post-treatment follow-up period after the last DBTP visit for patients who did not give the consent to participate in the open-label extension study or if they prematurely discontinued study treatment. Thus, the maximum study duration per patient was approximately 89 weeks (ie, 20 months) (up to 3 weeks of screening 78 weeks of double-blind treatment 8 weeks of follow-up). End of study per patient was the last planned protocol visit or resolution / stabilization of all SAEs, and AESI, whichever comes first.

Patient selection

The inclusion criteria were: 1) HeFH patients * who were not adequately controlled with a maximum tolerated daily dose of statin ** with or without another TML, at stable doses before the screening visit (Week -3).

* The diagnosis of heFH must be made either by genotyping or by clinical criteria. For non-genotyped patients, clinical diagnosis can be based on either the Simon Broome criteria with a criterion for HF or the WHO / Dutch Lipid NetWork criteria with a score> 8 points.

According to the Simon Broome Register Diagnostic Criteria for Heterozygous Familial Hypercholesterolemia, defined familial hypercholesterolemia is defined as: 1) Total C> 6.7 mmol / L (260 mg / dL) or LDL cholesterol greater than 4.0 mmol / L (155 mg / dL) in a child <16 years or total C> 7.5 mmol / L (290 mg / dL) or LDL cholesterol greater than 4.9 mmol / L (190 mg / dL) in an adult. (Levels or pre-treatment or the highest during treatment); more or A) tendinous xanthomas in the patient, or in a 1st degree relative (father, brother, son), or in a 2nd degree relative (grandfather, uncle, aunt); or B) DNA-based evidence of a familial defective LDL or Apo B receptor mutation.

According to the Simon Broome Register Diagnostic Criteria for Heterozygous Familial Hypercholesterolemia, possible familial hypercholesterolemia is defined as: 1) Total C> 6.7 mmol / L (260 mg / dL) or LDL cholesterol greater than 4.0 mmol / L (155 mg / dL) in a child <16 years or total C> 7.5 mmol / L (290 mg / dL) or LDL cholesterol greater than 4.9 mmol / L (190 mg / dL) in an adult. (Levels or pre-treatment or the highest during treatment); and at least one of the following: A) family history of MI below 50 years of age in a 2nd degree relative or below 60 years of age in a 1st degree relative; and B) family history of elevated cholesterols> 7.5 mmol / L (290 mg / dL) in an adult relative of 1st or 2nd degree or> 6.7 mmol / L (260 mg / dL) in a son or brother due to under 16 years of age.

The WHO criteria (Dutch Lipid NetWork clinical criteria) for diagnosing heterozygous familial hypercholesterolemia (HeFH) are set out in Table 2.

Table 2

Patients who met all the above inclusion criteria were selected by the following exclusion criteria, which are classified and numbered in the following 3 subsections: exclusion criteria related to the study methodology, exclusion criteria related to the active comparator, and / or mandatory background therapies, and exclusion criteria related to alirocumab.

The exclusion criteria related to the study methodology were: 1) patient without a diagnosis of HFhe made either by genotyping or clinical criteria; 2) LDL-C <70 mg / dL (<1.81 mmol / L) at the screening visit (Week -3) and patient with a history of documented cardiovascular disease. Cardiovascular disease was defined as coronary heart disease, ischemic stroke, or peripheral arterial disease; 3) LDL-C <100 mg / dL (<2.59 mmol / L) at the screening visit (Week -3) and patient with no history of documented cardiovascular disease; 4) are not on a stable dose of TML (including statin) for at least 4 weeks and / or fenofibrate for at least 6 weeks, as applicable, prior to the screening visit (Week -3) and from screening to screening. randomization; 5) were currently taking a statin other than simvastatin, atorvastatin, or rosuvastatin; 6) simvastatin, atorvastatin, or rosuvastatin is not taken daily or is not taken in recorded doses; 7) daily doses higher than atorvastatin 80 mg, rosuvastatin 40 mg or simvastatin 40 mg (except for patients with simvastatin 80 mg for more than one year, who are eligible); 8) use of fibrates, other than fenofibrate, within 6 weeks of the screening visit (Week -3) or between screening and randomization visits; 9) use of nutraceutical products or over-the-counter therapies that may affect lipids that have not been in a stable dose / amount for at least 4 weeks prior to the screening visit (Week -3) or between screening visits and of randomization; 10) use of red yeast rice products within 4 weeks of the screening visit (Week -3) or between screening and randomization visits; 11) patient who has received plasmapheresis treatment within 2 months prior to screening visit (Week -3), or plans to receive it during the study; 12) Recent MI (within 3 months before screening visit [Week -3] or between screening and randomization visits), unstable angina leading to hospitalization, percutaneous coronary intervention (PCI), graft surgery of coronary artery bypass (CABG), uncontrolled cardiac arrhythmia, cerebrovascular accident, transient ischemic attack (TIA), carotid revascularization, endovascular procedure or surgical intervention for peripheral vascular disease; 13) plan to undergo scheduled PCI, CABG, carotid or peripheral revascularization during the study; 14) systolic BP> 160 mmHg or diastolic BP> 100 mmHg at screening visit or randomization visit; 15) New York Heart Association (NYHA) history of class III or IV heart failure within the past 12 months; 16) known history of hemorrhagic stroke; 17) age <18 years or legal age of majority at the screening visit (Week -3), whichever is greater; 18) patients not previously informed about a low cholesterol diet before the screening visit (Week -3); 19) newly diagnosed diabetes (within 3 calendar months prior to randomization visit [Week 0]) or poorly controlled (glycated hemoglobin A1c [HbA1c]> 9% at screening visit [Week -3]); 20) Presence of any clinically significant uncontrolled endocrine disease known to influence serum lipids or lipoproteins. Note that patients on thyroid replacement therapy can be enrolled if the dosage has been stable for at least 12 weeks prior to screening and between screening and randomization visits, and the TSH level is within the normal central laboratory range. on the screening visit; 21) history of bariatric surgery within 12 months prior to screening visit (Week -3); 22) unstable weight defined by a variation> 5 kg within 2 months before the screening visit (Week -3); 23) known history of homozygous HF; 24) known history of PCSK9 loss of function (ie, genetic mutation or sequence variation); 25) use of systemic corticosteroids, unless used as replacement therapy for pituitary-adrenal disease with a stable regimen for at least 6 weeks prior to the randomization visit (Week 0). Note that topical, intra-articular, nasal, inhaled, and ophthalmic steroid therapies were not considered 'systemic' and were allowed; 26) use of continuous estrogen or testosterone hormone replacement therapy unless the regimen has been stable in the last 6 weeks prior to the screening visit (Week -3) and with no plans to change the regimen during the study; 27) history of cancer within the last 5 years, except adequately treated basal cell skin cancer, squamous cell skin cancer, or cervical cancer in situ; 28) known history of a positive HIV test; 29) patient who has taken any investigational drug other than alirocumab training placebo kits within 1 month or 5 half-lives, whichever is longer; 30) patient who has previously been treated with at least one dose of alirocumab or any other monoclonal anti-PCSK9 antibody in other clinical trials; 31) patient who withdraws consent during the selection period (patient who does not wish to continue or does not return); 32) conditions / situations such as: a) any clinically significant abnormality identified at the time of selection that, in the discretion of the investigator or any collaborating investigator, would prevent the safe completion of the study or restrict the evaluation of endpoints; for example, major systemic diseases, patients with a short life expectancy; or b) deemed by the investigator or any collaborating investigator as inappropriate for this study for any reason, for example: deemed unable to meet specific protocol requirements, such as scheduled visits; deemed unable to administer or tolerate long-term injections according to the patient or investigator; researcher or any collaborating researcher, pharmacist, study coordinator, other study personnel or relatives of the same directly involved in carrying out the protocol, etc .; presence of any other condition (eg geographic or social), both current and anticipated, that the investigator feels would restrict or limit the patient's participation for the duration of the study; or 33) laboratory data during the screening period (which does not include randomization labs at Week 0): positive test for hepatitis B surface antigen or hepatitis C antibody; positive for beta-hCG in serum or urine pregnancy test (including Week 0) in women of childbearing potential (WOCBP); triglycerides> 400 mg / dL (> 4.52 mmol / L) (1 repeat lab allowed); Estimated glomerular filtration rate (eGFR) <30 mL / min / 1.73 m2 according to the modification of 4 variables of the diet in kidney disease (MDRD) the study equation (calculated by the central laboratory); alanine aminotransferase (ALT) or aspartate aminotransferase (AST)> 3 x upper limit of normal (ULN) range (1 repeat laboratory allowed); CPK> 3 x ULN (1 repeat lab allowed); TSH <lower limit of normal (LLN) or> ULN (1 repeat lab allowed).

Exclusion criteria related to active comparator and / or mandatory background therapies were: 1) all contraindications to background therapies or warnings / precautions for use (where appropriate) as presented in the respective national product labeling.

Exclusion criteria related to alirocumab were: 1) known hypersensitivity to monoclonal antibody or any component of the drug; 2) pregnant or lactating women; or 3) women of childbearing potential not protected by highly effective method (s) of birth control (as defined in the informed consent document and / or in an appendix to the local protocol) and / or who are unwilling or are unable to take a pregnancy test. Note that women of childbearing potential must have a confirmed negative pregnancy test at screening and randomization visits. They must use effective contraception for the entire duration of study treatment, and for 10 weeks after the last IMP intake, and agree to repeat the urine pregnancy test at designated visits. Postmenopausal women must be amenorrheic for at least 12 months.

Coronary heart disease, ischemic stroke, and peripheral arterial disease, as defined in exclusion criteria number 2 related to study methodology, was as follows. Documented history of CHD (includes one or more of the following): acute myocardial infarction (MI); silent myocardial infarction; unstable angina; coronary revascularization procedure (eg, percutaneous coronary intervention [PCI] or coronary artery bypass graft surgery [CABG]); Clinically significant CHD diagnosed by invasive or non-invasive testing (such as coronary angiography, treadmill stress test, stress echocardiography, or nuclear magnetic resonance imaging).

Previous ischemic stroke documented with a focal ischemic neurological deficit that persisted for more than 24 hours, considered not of atherothrombotic origin. CT or MRI should have been done to rule out hemorrhage and nonischemic neurologic disease.

Documented peripheral arterial disease (one of the following criteria must be met): 1) current intermittent claudication (muscle discomfort in the lower extremity produced by exercise that is both reproducible and relieved by rest within 10 minutes) of presumed atherosclerotic origin together with an ankle-brachial index equal to 0 less than 0.90 in any resting leg or 2) history of intermittent claudication (muscle discomfort in the lower limb produced by exercise that is both reproducible and relieved by rest within 10 minutes) in conjunction with endovascular procedure or surgery on one or both legs due to atherosclerotic disease or 3) history of critical limb ischemia along with thrombolysis, endovascular procedure, or surgery on one or both legs due to atherosclerotic disease.

Treatments under study

The sterile alirocumab drug was delivered at a concentration of 75 mg / mL and 150 mg / mL both as 1 mL volume in an autoinjector. The active principle was formulated in histidine, pH 6.0, polysorbate 20 and sucrose.

Sterile placebo for alirocumab was prepared in the same formulation as alirocumab without the addition of protein as 1 mL volume in an autoinjector.

During the double-blind treatment period, alirocumab or placebo was administered subcutaneously Q2W, starting at Week 0 continuing until the last injection (Week 76) 2 weeks before the end of the double-blind treatment period (DBTP). If the injection was scheduled to take place on the same date as the site visit, then the IMP was administered after the blood sampling was completed.

The Investigational Pharmaceutical Specialty (IMP) should ideally have been administered Q2W subcutaneously at approximately the same time of day; however, it was acceptable to have a window period of ± 3 days. Time of day was based on patient preference.

The following drug classes were identified as non-NIMP because the medication was either background therapy or a possible rescue medication: statins (rosuvastatin, atorvastatin, simvastatin); cholesterol absorption inhibitors (ezetimibe); bile acid binding sequestrants (such as cholestyramine, colestipol, colesevelam); nicotinic acid; fenofibrate; and omega-3 fatty acids (> 1000 mg per day).

Patients were randomized to receive either placebo or alirocumab during the double-blind study treatment period using a 1: 2 ratio, with permuted block randomization. Randomization was stratified by previous history of MI or ischemic stroke [Yes / No], statin therapy (atorvastatin 40 to 80 mg daily or rosuvastatin 20 to 40 mg daily versus simvastatin whatever the daily dose, lower atorvastatin to 40 mg per day or rosuvastatin less than 20 mg per day) and geographic region.

Concurrent medication was any treatment received by the patient concurrent with the study (up to the follow-up visit). Medications should be kept to a minimum during the study. However, if these are considered necessary for the well-being of the patient and are unlikely to interfere with IMP, they can be administered at the investigator's discretion, at a stable dose (when possible). In addition to the specific information regarding concurrent medications provided in this section, any other concurrent medication (s) will be permitted. If the patient has an LDL-C> 160 mg / dL (4.14 mmol / L) at the screening visit (Week -3) and is treated with a statin alone, that is, without additional TML, the investigator will need to report the patient's reason for not being on a second TML. For background TML, which includes statins, sites must follow the national product label for safety monitoring and patient management.

Nutraceuticals or OTC therapies that could affect lipids were allowed only if they had been used at a stable dose for at least 4 weeks prior to the screening visit, during the screening period, and were maintained for the first 24 weeks of the double-blind treatment period. After the Week 24 visit, modification to these nutraceuticals or OTC therapies was allowed, but should generally be avoided. Examples of such over-the-counter nutraceuticals or therapies include omega-3 fatty acids at doses <1000 mg, plant stanols such as those found in Benecol, flax seed oil, and psyllium.

Patients must have been on stable maximum tolerated daily recorded statin doses with or without another TML for at least 4 weeks (6 weeks for fenofibrate) prior to the screening visit. During the study, patients should remain on these stable tolerated maximum recorded daily statin doses with or without another TML. From the screening visit (Week -3) to Week 24 of the double-blind treatment period, the background TML should not be changed. No dose adjustment, interruption, or initiation of other statins or other TMLs should take place during this time, except for exceptional circumstances where primary concerns (including, but not limited to, central laboratory-reported triglyceride alert) guarantee such changes, at the discretion of the researcher.

For an LDL-C salvage notification at the Week 24 visit and beyond, that is, an increase in LDL-C> 25% compared to the LDL-C randomization visit on two consecutive occasions, the investigator must have guaranteed that there was no reasonable explanation for insufficient LDL-C control (such as an alternative medical cause such as the use of corticosteroids, etc.) and in particular that: adherence to the diet was appropriate; compliance with background TML was appropriate; and the study treatment was administered as planned. If any of the above could reasonably explain the insufficient control of LDL-C, the investigator must have taken the appropriate action, that is, emphasized the absolute need to comply with the treatment, if a specific interview with a qualified professional of the nutrition and emphasize the absolute need to adhere to the diet, and performed a blinded LDL-C assessment within 1 to 2 months. If none of the reasons listed above were found, or if the appropriate action stopped lowering LDL-C below the alert value, rescue medication may have been introduced.

If the reason for LDL-C could not be found above the threshold value, or if the appropriate action failed to reduce LDL-C below the threshold value, rescue medication may have been introduced. The efficacy of any such changes was to be made based on the lack of rescue threshold from the blinded lipid test at the next routinely scheduled draw. The patients per protocol already received a maximum tolerated dose of statin, so upward adjustment of the statin dose or change was not an option. For additional LDL-C reduction, the investigator might consider adding: a cholesterol absorption inhibitor (ezetimibe), or a bile acid binding sequestrant (the resins cholestyramine and colestipol, or colesevelam, a nonabsorbable polymer) . The following lipid modifying agents might also be considered: fibrate (Note: Caution should be exercised when combining fibrates with other cholesterol-lowering medications such as statins due to the risk of myopathy. When combining a fibrate with a statin, the fenofibrate is fibrate of choice because it does not affect statin glucuronidation. The only fibrate allowed by protocol was fenofibrate); Nicotinic acid (niacin) (Note: Niacin increases blood glucose, but has been shown to be effective in modifying lipid disorders in people with diabetes if glucose control is maintained).

In summary, the background TML should not be changed from screening to the follow-up visit. However, until Week 24, if a confirmed TG alert was reached or if there was overwhelming clinical concern (at the discretion of the investigator), then background TML modification was allowed. In Week 24 and beyond, if a confirmed TG alert was reached, or if a salvage threshold for LDL-C was obtained (and there is no other reasonable explanation), or if there was an overwhelming clinical concern (at the discretion of the investigator) , then a modification of the background TML was allowed.

Women of childbearing must take effective contraception throughout study treatment and for 10 weeks after last IMP injection (eg follow-up visit).

Concurrent medications prohibited from the initial screening visit to the follow-up visit included the following: statins other than simvastatin, atorvastatin, and rosuvastatin; fibrates, other than fenofibrate; and red yeast rice products.

Study endpoints

The primary efficacy endpoint was the percent change in LDL-C calculated from baseline to Week 24, which was defined as: 100 x (LDL-C value calculated at Week 24 - LDL-C value calculated at baseline) / LDL-C value calculated at baseline. The LDL-C value calculated at baseline was the last LDL-C level obtained before the first double-blind IMP injection. The calculated LDL-C at Week 24 was the level of LDL-C obtained within the analysis window of Week 24 and during the period of main efficiency. The primary efficacy period was defined as the time from the first double-blind IMP injection to 21 days after the last double-blind IMP injection or to the upper limit of the Week 24 analysis window, whichever is. First. All calculated LDL-C values (scheduled or unscheduled, fasting or non-fasting) can be used to provide a value for the primary efficacy endpoint if appropriate as defined above.

The secondary key efficacy endpoints were: 1) percent change in LDL-C calculated from baseline to Week 12: similar definition and rules as for the primary efficacy endpoint, except that C- LDL calculated at Week 12 was the level of LDL-C obtained within the Week 12 analysis window and during the 12 week efficacy period. The 12-week efficacy period was defined as the time from the first double-blind IMP injection to contact with the IVRS re-supply at Visit 6 or up to 21 days after the last double-blind IMP injection, whichever comes first. The blood sample collected on the day of contact with the IVRS re-supply of Visit 6 was considered as before the assessment; 2) the percentage change in Apo B from baseline to Week 24, using the same definition and rules as for the primary endpoint; 3) the percent change in non-HDL C from baseline through Week 24, using the same definition and rules as for the primary endpoint; 4) the percent change in total C from baseline to Week 24, using the same definition and rules as for the primary endpoint; 5) the percentage change in Apo B from baseline to Week 12, using the same definition and rules as for the percent change in LDL-C calculated from baseline to Week 12; 6) percent change in non-HDL-C from baseline through Week 12, using the same definition and rules as for percent change in LDL-C calculated from baseline through Week 12; 7) the percent change in total C from baseline through Week 12, using the same definition and rules as for the percent change in LDL-C calculated from baseline through Week 12; 8) the percent change in LDL-C calculated from baseline to Week 52, using definitions and rules that were similar to those used for the primary endpoint substituting Week 24 for Week 52. Note that the period of 52-week efficacy was defined as the time from the first double-blind IMP injection to 21 days after the last double-blind IMP injection, or up to the upper limit of the Week 52 analysis window, whichever First; 9) the proportion of patients who reach the goal of LDL-C at Week 24, that is, LDL-C <70 mg / dL (1.81 mmol / L) in case of previous CVD or <100 mg / dL ( 2.59 mmol / L) for patients without prior CVD, defined as: (number of patients whose calculated LDL-C value at Week 24 reaches the LDL-C target / number of patients with modified intention-to-treat (mTDI ) population) * 100, using the definition and rules used for the primary endpoint; 10) the proportion of patients achieving LDL-C <70 mg / dL (1.81 mmol / L) at Week 24; 11) the percent change in Lp (a) from baseline to Week 24, using the same definition and rules as for the primary endpoint; 12) the percent change in HDL-C from baseline to Week 24, using the same definition and rules as for the primary endpoint; 13) percent change in HDL-C from baseline through Week 12, using the same definition and rules as for percent change in LDL-C calculated from baseline through Week 12; 14) the percent change in Lp (a) from baseline to Week 12, using the same definition and rules as for the percent change in LDL-C calculated from baseline to Week 12; 15) percent change in fasting TG from baseline to Week 24, using the same definition and rules as for the primary endpoint; 16) percent change in fasting TG from baseline to Week 12, using the same definition and rules as for percent change in LDL-C calculated from baseline to Week 12; 17) the percentage change in Apo A-1 from baseline to Week 24, using the same definition and rules as for the primary endpoint; and 18) the percentage change in Apo A-1 from baseline to Week 12, using the same definition and rules as for the percent change in LDL-C calculated from baseline to Week 12.

Other secondary efficacy endpoints were: 1) percent change in LDL-C calculated from baseline to Week 78, using definitions and rules that were similar to those used for the primary endpoint substituting Week 24 by Week 78. The 78 week efficacy period was defined as the time from the first double-blind injection of IMP to 21 days after the last injection of double-blind IMP, or to the upper limit of the analysis window. from Week 78, whichever comes first; 2) the proportion of patients who reach the LDL-C goal at Weeks 12, 52, and 78, that is, LDL-C <70 mg / dL (1.81 mmol / L) in case of previous CVD or <100 mg / dL (2.59 mmol / L) for patients without previous CVD; 3) the proportion of patients achieving LDL-C <100 mg / dL (2.59 mmol / L) at Week 24; 4) the proportion of patients achieving LDL-C <100 mg / dL (2.59 mmol / L) at Week 12; 5) the proportion of patients achieving LDL-C <70 mg / dL (1.81 mmol / L) at Week 12; 6) the absolute change in calculated LDL-C (mg / dL and mmol / L) from baseline through Weeks 12, 24, 52, and 78; 7) percentage change in Apo B, non-HDL C, total C, Lp (a), HDL-C, fasting TG, and Apo A-1 from baseline to Weeks 52 and 78; 8) the change in the Apo B / Apo A-1 ratio from baseline to Weeks 12, 24, 52 and 78; 9) the proportion of patients with Apo B <80 mg / dL (0.8 g / L) at Weeks 12, 24, 52 and 78; 10) the proportion of patients with C no h Dl <100 mg / dL at Weeks 12, 24, 52 and 78; and 11) the proportion of patients with calculated LDL-C <70 mg / dL (1.81 mmol / L) and / or> 50% reduction in calculated LDL-C (if calculated LDL-C> 70 mg / dL [ 1.81 mmol / L]) at Weeks 12, 24, 52 and 78.

Other endpoints were: evaluations of anti-alirocumab antibodies, highly sensitive C-reactive protein, glycated hemoglobin A1c, EQ-5D questionnaire, pharmacogenetics and pharmacokinetics. Anti-alirocumab antibodies included antibody status (positive / negative) and antibody titers. Serum samples were collected periodically throughout the study for the determination of anti-alirocumab antibodies. The first sample scheduled at the randomization visit was obtained prior to IMP injection (predose). Additional sample (s) were taken from patients who had a titer of 240 or higher for antialirocumab antibody at the follow-up visit 6 to 12 months after the last dose, and approximately every 3 to 3 months thereafter. 6 months until titer falls below 240 again. Percentage change in high sensitivity C-reactive protein (hs-CRP) was measured at baseline and Weeks 24, 52, and 78. EQ-5D is a measure standardized health status developed by the EuroQol Group to provide a simple generic measure of health for clinical and economic assessment. The EQ-5D as a health-related quality of life measure defines health in terms of 5 dimensions: mobility, self-care, usual activities, pain / discomfort, anxiety / depression. Each dimension can adopt one of three responses (3 ordinal levels of severity): "no problem" (1); "some problems" (2); "serious problems" (3); General health status is defined as a five-digit number. The health states defined by the 5-dimensional classification can be converted into corresponding index scores that quantify health status, where 0 represents 'death' and 1 represents 'perfect health'.

Study procedures

For all visits after Day 1 / Week 0 (randomization visit), a time period of a certain number of days was allowed. The window period for visits at Weeks 12 and 24 was ± 3 days, at Weeks 52 and 78 it was ± 5 days, and for all other site visits it was ± 7 days during the double-blind treatment period. , and follow-up period. A window period of 3 days was allowed for the randomization visit (Day 1 / Week 0) and ± 7 days for the injection training visit during the selection period (Week -1). For all visits after Day 1 / randomization visit, if a visit date is changed, then the next visit must take place according to the original schedule.

Security

Occurrence of treatment emergent adverse events (TEAEs) reported by the patient or observed by the investigator, serious adverse events (SAEs), TEAEs leading to treatment discontinuation, AEs of special interest (local injection site reactions, allergic events, selected neurological events and cardiovascular events confirmed by validation result), manifestation of PCSA (potentially clinically significant abnormalities) in laboratory parameters, exploratory analyzes for patients with 2 consecutive calculated LDL-C <25 mg / dL (<0.65 mmol / L) and for changes in glycemic control, including diabetes.

Statistical methods

Determination of sample size:

A total sample size of 45 patients (30 on alirocumab and 15 on placebo) had 95% power to detect a difference in mean percent change in LDL-C of 30% with a two-sided significance level of 0.05 and assuming a common standard deviation of 25%, and all of these 45 patients having an assessable primary endpoint. However, to meet legislative requirements throughout the program, the sample size to assess the safety of alirocumab was increased. To have at least 225 patients with alirocumab followed for 12 months in this study, and assuming a dropout rate of 10% compared to the first 3-month period and a dropout rate of 20% compared to the remaining 9-month period, the final total sample size increased to 471 with a 2: 1 randomization ratio (alirocumab 314: placebo 157).

Exact time of analysis:

The first-stage analyzes include efficacy endpoints through Week 52 (final efficacy analysis) and interim safety analysis, which was performed on all safety data up to the common study deadline (Weekly visit 52 of the last patient). Analysis of lipid data beyond Week 52 was descriptive. These results are presented in this document.

The second (final) stage analysis will be performed at the end of the study and will consist of the final analysis of the efficacy endpoints up to Week 78 and the final safety analyzes.

Analysis populations:

The primary efficacy analysis population was the intention-to-treat (IDT) population, defined as all randomized patients who had an assessable primary endpoint, that is, those with an LDL-C value calculated at the available baseline level. , and at least one calculated LDL-C value available within one of the analysis windows up to Week 24 (including all calculated LDL-C values on treatment and no treatment).

The secondary efficacy analysis population was the modified intention-to-treat (mTDI) population, defined as all randomized patients who took at least one dose or part of a dose of the double-blind investigational pharmaceutical specialty (IMP) and who had a calculated LDL-C value available at baseline and at least one within one of the analysis windows through Week 24 during the efficacy treatment period. The efficacy treatment period was defined as the time from the first double-blind injection of IMP to 21 days after the last double-blind injection.

The safety population included all randomized patients who received at least one dose or part of a dose of the double-blind IMP.

Efficacy analysis:

Primary efficacy endpoint analyzes were performed using an IDT approach (based on the IDT population defined above), including all lipid data, regardless of whether the patient was continuing therapy or not. This corresponds to IDT estimates, defined for primary and secondary key endpoints. In addition, analyzes were also performed using an during treatment approach (based on the mTDI population defined above), including lipid data collected during the efficacy treatment period. This corresponds to estimates during treatment of key secondary endpoints.

The IDT approach analyzed all patients, regardless of their compliance with treatment; assessed the benefit of the treatment strategy and reflected the effect as far as possible in a patient population. The during treatment approach looked at the effect of treatment, restricted to the period during which patients actually received the treatment. It evaluated the benefit that a treatment would achieve in patients who comply with the treatment until the moment of time considered.

Efficacy analyzes were performed by treatment as randomized.

All measurements, scheduled or unscheduled, fasting or non-fasting, were assigned to analysis windows to provide an evaluation for time points from Week 4 to Week 78.

Regarding the primary efficacy analysis (TDI approach), the percentage change in LDL-C calculated from baseline to Week 24 was analyzed using a mixed effects model with repeated measures (MMRM) approach. All available post-baseline data from the Week 4 through Week 52 analysis windows were used and the MMRM accounted for missing data. The model included the fixed categorical effects of treatment group (placebo vs alirocumab), randomization strata (according to IVRS), moment of time (Week 4 to Week 52), interaction of treatment by moment of time and interaction of strata by moment time, as well as the continuous fixed covariates of baseline LDL-C value and baseline value interaction by time. This model provided estimates of least squares means (LS means) adjusted to baseline at Week 24 for both treatment groups with their corresponding standard errors and 95% confidence intervals. To compare the alirocumab group with the placebo group, a contrast instruction was used to test for differences in these estimates at the 5% alpha level.

A hierarchical procedure was defined to test secondary key endpoints while controlling for multiplicity (using the previous order of secondary key endpoints). The first key secondary endpoint was the percent change in LDL-C calculated from baseline to Week 24 using an on-treatment approach.

Continuous secondary variables anticipated to have a normal distribution (ie, lipids other than TGs and Lp (a)) were analyzed using the same MMRM model as for the primary endpoint. Continuous endpoints that were anticipated to have a non-normal distribution (i.e. TGs and Lp (a)) were analyzed using the multiple imputation approach for manipulating missing values followed by the robust endpoint regression model. of interest as a response variable using the estimation of M (using the SAS ROBUSTREG procedure) with treatment group, randomization strata (according to IVRS) and value (s) at the corresponding initial level (s) as effects to compare the effects treatment. The pooled estimate for the mean in both treatment groups, as well as the differences of these estimates, with their corresponding SEs, 95% CI, and p-value (using the SAS MIANALYZE procedure) were provided.

Secondary binary efficacy endpoints were analyzed using the multiple imputation approach for manipulating missing values followed by stratified logistic regression with treatment group as main effect and corresponding baseline value (s) as covariate. , stratified by randomization factors (according to IVRS). Pooled estimates of odds ratio vs. placebo, 95% CI, and p-value (using SAS MIANALYZE procedure) were provided.

Security analysis:

The safety analyzes were descriptive, carried out in the safety population according to the treatment actually received. Safety analyzes focused on the TEAE period defined as the time from the first double-blind IMP dose to 70 days after the last double-blind injection. TEAEs that developed, worsened, or became severe or PCSA occurring after patient inclusion in the open-label extension study (LTS13643) were not considered in the TEAE period. The TEAE period was truncated at the common study deadline.

Results

Study patients

Patient count

Of the 486 randomized patients (323 patients and 163 patients in the alirocumab and placebo groups, respectively), one patient was not treated in the alirocumab group and therefore was not included in the safety population. This patient was also excluded from the TDI population (no LDL-C value within one of the analysis windows up to Week 24 since the patient withdrew consent on Day 1).

Two randomized patients in the alirocumab group were excluded from the mTDI population (one patient was excluded from the TDI population and one patient with no LDL-C value within one of the analysis windows up to Week 24 during the treatment period of effectiveness).

Table 3 - Analysis populations

Placebo Alirocumab 75 Q2W / up to 150 Q2W All

Randomized population 163 (100%) 323 (100%) 486 (100%)

Efficacy populations

Intent to Treat (IDT) 163 (100%) 322 (99.7%) 485 (99.8%)

Modified intention to treat (mTDI) 163 (100%) 321 (99.4%) 484 (99.6%)

Safety population 163 322 485

Note: Patients in the safety population are tabulated based on the treatment actually received (as treated). For the other populations, patients are tabulated according to their randomized treatment.

In the alirocumab group, among 311 patients who received at least one injection after Week 12, 135 (43.4%) patients received automatic dose titration at Week 12 from alirocumab 75 mg Q2W to 150 mg Q2W in a blinded mode.

Study layout

Study readiness, exposure and safety analysis were assessed using all data up to the common study cutoff date (defined as the date of the last patient's Week 52 visit). Therefore, this first stage analysis includes data beyond Week 52 and up to Week 78 or follow-up visit for some patients.

There were a total of 7 (1.4%) randomized patients who completed the 78-week double-blind study treatment period and 424 (87.2%) randomized patients with ongoing treatment at the time of the analysis cut-off date. the first stage. Double-blind IMP was prematurely discontinued before Week 78 for 18 (11.0%) randomized patients in the placebo group and 36 (11.1%) randomized patients in the alirocumab group. The main reasons for discontinuation of study treatment were adverse event and other reasons.

Additionally, among these patients, 34 (10.5%) randomized patients had prematurely discontinued double-blind IMP prior to the Week 52 Visit in the alirocumab group and 15 (9.2%) patients in the placebo group. .

In this first-stage analysis, final results are available for the primary efficacy endpoint at Week 24 and the secondary key efficacy endpoints were assessed at Week 12, Week 24, and Week 52. The criterion primary endpoint was missing for 46 patients at the Week 24 visit for the following reasons: 18 samples were not collected due to premature study discontinuation, 14 were samples outside the analysis time window, 4 samples were missing while the Week 24 visit was made, and 10 samples were taken but the measurement could not be done (lipemia, insufficient quantity, TGs> 400 mg / dL [> 4 , 52 mmol / L], loss of samples, ...).

Demographic characteristics, baseline level, and summary of population characteristics.

Demographic characteristics, disease characteristics, and baseline lipid parameters were similar in the alirocumab group compared to the placebo group (see Table 4). 486 patients with heFH diagnosed by genotyping (39%) or WHO or Simon Broome criteria (61%) were randomized (2: 1) to alirocumab (75 mg Q2W possibly up-titration to 150 mg Q2W) or placebo ( 323 vs. 163, respectively). Half of the randomized population (51%) had a history of at least one coronary heart disease (CHD) or multiple risk factors for CHD that defined these patients as being at very high cardiovascular risk. Demographic characteristics, disease characteristics, and lipid parameters at baseline were similar in the alirocumab group compared to the placebo group. All patients were treated with a statin, 82% receiving a dose defined as a high-intensity statin (atorvastatin 40 to 80 mg daily or rosuvastatin 20 to 40 mg daily) and receiving 57% ezetimibe, in addition to the statin. The mean calculated LDL-C (SD) at baseline was 144.6 (49.7) mg / dL [3.75 (1.29) mmol / L].

Injection exposure was similar across treatment groups with a mean exposure of 59 weeks. In the alirocumab group, among 311 patients who received at least one injection after Week 12, 135 (43.4%) patients received automatic dose titration at Week 12 from alirocumab 75 mg Q2W to 150 mg Q2W in a blinded mode.

Table 4 - Baseline Characteristics of the FH I Patient Population

Table 5 - Disease characteristics and other relevant data at baseline - Randomized population Alirocumab 75

Q2W / up to 150

Placebo Q2W All (N = 163) (N = 323) (N = 486) Type of hypercholesterolemia

Heterozygous familial hypercholesterolemia (HFhe) 163 (100%) 323 (100%) 486 (100%) Non-familial hypercholesterolemia (H no F) 0 0 0

Time since diagnosis of hypercholesterolemia

(years)

Number 163 323 486 Mean (SD) 13.28 (11.38) 12.19 (11.38) 12.55 (11.38) Median 9.43 8.82 9.03 Min: max 0.0: 42 .6 0.0: 60.7 0.0: 60.7

Diagnosis confirmation

By genotyping 62 (38.0%) 129 (39.9%) 191 (39.3%) By WHO / Simon Broomea 101 (62.0%) 193 (59.8%) 294 (60.5%) a for diagnosis of heHF not confirmed by genotyping.

Note: At the time of selection, one patient was enrolled based on clinical criteria with a score of 8 for the WHO criteria. As the clinical score characterized the patient as probable HFH rather than true, genotyping was performed to confirm HFH status but these results are still pending.

Table 6 - Cardiovascular history and breakdown of risk factors

Table 7 - Background TML at Randomization - Randomized Population

Placebo Alirocumab 75 Q2W / up to All 150 Q2W

(N = 163) (N = 323) (N = 486) Any statin 163 (100%) 323 (100%) 486 (100%)

Taking high intensity statin 135 (82.8%) 261 (80.8%) 396 (81.5%) Daily dose of atorvastatin (mg) 64 (39.3%) 113 (35.0%) 177 (36 ,4 %)

10 1 (0.6%) 3 (0.9%) 4 (0.8%)

20 2 (1.2%) 7 (2.2%) 9 (1.9%)

40 23 (14.1%) 23 (7.1%) 46 (9.5%)

80 38 (23.3%) 77 (23.8%) 115 (23.7%)

Other doses 0 3 (0.9%) 3 (0.6%) Rosuvastatin daily dose (mg) 81 (49.7%) 172 (53.3%) 253 (52.1%)

5 4 (2.5%) 7 (2.2%) 11 (2.3%)

10 2 (1.2%) 5 (1.5%) 7 (1.4%)

20 19 (11.7%) 44 (13.6%) 63 (13.0%)

40 55 (33.7%) 116 (35.9%) 171 (35.2%)

Other doses 1 (0.6%) 0 1 (0.2%)

Simvastatin daily dose (mg) 18 (11.0%) 38 (11.8%) 56 (11.5%)

10 2 (1.2%) 2 (0.6%) 4 (0.8%)

20 1 (0.6%) 5 (1.5%) 6 (1.2%)

40 10 (6.1%) 25 (7.7%) 35 (7.2%)

80 3 (1.8%) 6 (1.9%) 9 (1.9%)

Other doses 2 (1.2%) 0 2 (0.4%)

Any TML other than statinase 107 (65.6%) 198 (61.3%) 305 (62.8%) Any TML other than nutraceuticals 105 (64.4%) 192 (59.4%) 297 (61.1 %) Ezetimibe 97 (59.5%) 180 (55.7%) 277 (57.0%) Nutraceuticals 8 (4.9%) 20 (6.2%) 28 (5.8%)

a in combination with statins or not.

The high intensity statin corresponds to atorvastatin 40 to 80 mg daily or rosuvastatin 20 to 40 mg daily.

Table 8 - Baseline lipid efficacy parameters - Quantitative summary in units

conventional - Randomized population

Placebo Alirocumab 75 Q2W / up to 150 Q2W All

(N = 163) (N = 323) (N = 486) Calculated LDL-C (mg / dL)

Number 163 323 486

Mean (SD) 144.4 (46.8) 144.8 (51.1) 144.6 (49.7) Median 138.0 135.0 135.5

Q1: Q3 112.0: 166.0 112.0: 163.0 112.0: 165.0

Min: max 66: 354 39: 384 39: 384

Measured LDL-C (mg / dL)

Number 140 272 412

Mean (SD) 140.0 (43.5) 140.2 (49.7) 140.1 (47.6) Median 135.0 130.5 132.0

Q1: Q3 111.0: 164.0 108.0: 159.5 108.5: 161.0

Min: max 68: 356 37: 378 37: 378

C no HDL (mg / dL)

Number 163 323 486 Placebo Alirocumab 75 Q2W / up to 150 Q2W All (N = 163) (N = 323) (N = 486) Mean (SD) 169.6 (50.6) 170.3 (54.6) 170.1 (53, 3) Median 161.0 158.0 160.0

Q1: Q3 132.0: 195.0 134.0: 198.0 133.0: 196.0

Min: max 78: 390 58: 426 58: 426

Total C (mg / dL)

Number 163 323 486 Mean (SD) 217.6 (50.3) 221.1 (54.3) 219.9 (53.0) Median 210.0 212.0 211.0

Q1: Q3 185.0: 240.0 184.0: 244.0 185.0: 243.0

Min: max 137: 445 123: 482 123: 482

HDL-C (mg / dL)

Number 163 323 486 Mean (SD) 48.0 (14.4) 50.8 (15.7) 49.8 (15.3) Median 45.0 47.0 46.5

Q1: Q3 36.0: 56.0 39.0: 59.0 38.0: 58.0

Min: Max 24: 116 22: 115 22: 116

Fasting TGs (mg / dL)

Number 163 323 486 Mean (SD) 126.5 (62.9) 128.4 (66.7) 127.8 (65.4) Median 111.0 113.0 112.0

Q1: Q3 85.0: 151.0 82.0: 153.0 83.0: 152.0

Min: max 45: 431 35: 566 35: 566

Lipoprotein- (a) (mg / dL)

Number 161 317 478 Mean (SD) 47.2 (51.6) 51.7 (50.2) 50.2 (50.7) Median 23.0 34.0 28.0

Q1: Q3 8.0: 72.0 12.0: 82.0 11.0: 80.0

Min: max 2: 223 2: 229 2: 229

Apo-B (mg / dL)

Placebo Alirocumab 75 Q2W / up to 150 Q2W All (N = 163) (N = 323) (N = 486) Number 161 317 478 Mean (SD) 113.4 (28.5) 114.4 (30.8) 114, 1 (30.0) Median 109.0 108.0 109.0

Q1: Q3 94.0: 128.0 94.0: 130.0 94.0: 129.0

Min: max 64: 249 45: 250 45: 250

Apo-A1 (mg / dL)

Number 161 317 478 Mean (SD) 137.6 (27.2) 142.8 (27.4) 141.1 (27.4) Median 134.0 138.0 137.0

Q1: Q3 121.0: 151.0 124.0: 158.0 122.0: 155.0

Min: max 84: 292 79: 278 79: 292

Apo-B / Apo-A1 (ratio)

Number 161 317 478 Mean (SD) 0.859 (0.292) 0.830 (0.269) 0.839 (0.277) Median 0.810 0.780 0.800

Q1: Q3 0.640: 0.990 0.650: 0.960 0.650: 0.970

Min: max 0.36: 2.42 0.26: 1.84 0.26: 2.42

Total C / HDL-C (ratio)

Number 163 323 486 Mean (SD) 4,907 (1,838) 4,707 (1,756) 4,774 (1,785) Median 4,658 4,321 4,444

Q1: Q3 3,661: 5,658 3,537: 5,649 3,542: 5,649

Min: Max 1.86: 13.64 1.73: 15.14 1.73: 15.14 Note: LDL-C was evaluated as measured by the beta-quantification method.

The LDL-C pool measured was not planned in the initial protocol and was added in a modification. Therefore, the measured LDL-C values are available for fewer patients compared to the calculated LDL-C values.

Dosage and duration

Injection exposure was similar across treatment groups with a mean exposure of 59 weeks.

In the alirocumab group, among the 311 patients who received at least one injection after Week 12, 135 (43.4%) patients received automatic dose titration from 75 mg Q2W to 150 mg Q2W at Week 12 in a blinded mode.

Effectiveness

Primary efficacy endpoint

The TDI analysis includes all calculated LDL-C values collected during treatment and without treatment through Week 52. The analysis of the primary endpoint (percent change in LDL-C calculated from baseline to Week 24) is provided based on a model of MMRM in the IDT population, using estimates of the mean CM at Week 24. Thirty-two (9.9%) patients in the alirocumab group and 14 (8.6%) patients in the The placebo group did not have a calculated LDL-C value at Week 24. These missing values were taken into account by the MMRM model.

Results from the primary endpoint analysis are presented in Table 9, in mmol / L and mg / dL.

Primary efficacy analysis

A statistically significant decrease in the percentage change in LDL-C from baseline to Week 24 was observed in the alirocumab group (mean MC vs. baseline - 48.8%) compared to the placebo group ( mean MC vs. baseline 9.1%) (mean MC difference vs. placebo - 57.9%, p <0.0001). In the alirocumab group, a reduction in LDL-C was observed from baseline from Week 4 and was maintained throughout the study until Week 78 (see Figure 2 and Table 10).

Table 9 - Percentage change from baseline in LDL-C calculated at Week 24: MMRM - IDT Analysis - IDT Population

Placebo Alirocumab 75 Q2W / up to 150 Q2W Calculated LDL cholesterol (N = 163) (N = 322) Baseline level (mmol / L)

Number 163 322

Mean (SD) 3,739 (1,213) 3,748 (1,326) Median 3,574 3,497

Min: max 1.71: 9.17 1.01: 9.95 Initial level (mg / dL)

Number 163 322

Mean (SD) 144.4 (46.8) 144.7 (51.2) Median 138.0 135.0

Min: Max 66: 354 39: 384 Week 24 Percent Change from Baseline (%)

Mean MC (SE) 9.1 (2.2) -48.8 (1.6) Mean difference of MC (SE) vs placebo -57.9 (2.7)

95% CI (-63.3 to -52.6) P-value vs placebo <0.0001 * Note: Least squares (LS) means, standard errors (SE), and p-value taken from MMRM analysis ( mixed effects model with repeated measures). The model includes the fixed categorical effects of treatment group, strata of randomization according to IVRS, moment of time, interaction of treatment by moment of time and strata by moment of time, as well as the continuous fixed covariates of the calculated LDL-C value. baseline and baseline LDL-C value interaction calculated by moment of time

MMRM model and description of the baseline level performed in patients with a baseline value and a post baseline value in at least one of the analysis windows used in the model.

The p-value is followed by a '*' if it is statistically significant according to the fixed hierarchical approach used to ensure a strong control for the overall type I error rate at the 0.05 level.

Table 10 - LDL-C calculated over time - IDT analysis - IDT population

Placebo Alirocumab 75 Q2W / up to 150 Q2W (N = 163) (N = 322)

Change Change in Change Change in LDL-C from percentage from from from percentage from calculated Initial level value initial level Initial level value initial level Mean of MC

(EE) (mmol / L)

Initial level a 3,739 NA NA 3,748 NA NA (0.095) (0.074)

Week 4 3.819 0.074 (0.070) 4.3 (2.1) 1.96 -1.749 (0.050) -46.7 (1.5) (0.070) (0.050)

Week 8 3.805 0.059 (0.073) 3.6 (1.8) 1.986 -1.759 (0.052) -46.4 (1.3) (0.073) (0.052)

Week 12 3.898 0.153 (0.074) 5.7 (2.0) 2.078 -1.668 (0.053) -43.5 (1.4) (0.074) (0.053)

Week 16 3.892 0.147 (0.080) 5.6 (2.1) 1.763 -1.982 (0.057) -51.7 (1.5) (0.080) (0.057)

Week 24 4.029 0.284 (0.084) 9.1 (2.2) 1.846 -1.899 (0.060) -48.8 (1.6) (0.084) (0.060)

Week 36 3.965 0.220 (0.091) 8.5 (2.4) 1.997 -1.748 (0.066) -45.1 (1.8) (0.091) (0.066)

Week 52 4,000 0.255 (0.092) 9.0 (2.6) 1.925 -1.821 (0.066) -47.1 (1.9) (0.092) (0.066)

Week 64 3,947 1,962

(0.086) (0.063)

Week 78 4,082 2,177

(0, 101) (0.073)

Initial level3 144.4 NA NA 144.7 NA NA (3.7) (2.9)

Week 4 147.5 2.9 (2.7) 4.3 (2.1) 77.1 -67.5 (1.9) -46.7 (1.5) (2.7) (1, 9)

Week 8 146.9 2.3 (2.8) 3.6 (1.8) 76.7 -67.9 (2.0) -46.4 (1.3) (2, 8) (2, 0)

Week 12 150.5 5.9 (2.9) 5.7 (2.0) 80.2 -64.4 (2.0) -43.5 (1.4) (2.9) (2, 0)

Week 16 150.3 5.7 (3.1) 5.6 (2.1) 68.1 -76.5 (2.2) -51.7 (1.5) (3.1) (2, two )

Week 24 155.6 11.0 (3.2) 9.1 (2, 2) 71.3 -73.3 (2.3) -48.8 (1.6) (3.2) (2, 3)

Placebo Alirocumab 75 Q2W / up to 150 Q2W (N = 163) (N = 322)

Change Change in Change Change in LDL-C from percentage from from from percentage from calculated Baseline value baseline level Baseline level value baseline level Week 36 153.1 8.5 (3.5) 8.5 (2.4 ) 77.1 -67.5 (2.5) -45.1 (1.8) (3.5) (2.5)

Week 52 154.4 9.8 (3.5) 9.0 (2.6) 74.3 -70.3 (2.6) -47.1 (1.9) (3.5) (2, 6)

Week 64 152.4 75.8

(3.3) (2.4)

Week 78 157.6 84.0

(3.9) (2, 8)

a The baseline level is described using means and standard errors.

Note: Least squares means (LC), standard errors (SE) and p-value taken from MMRM analysis (mixed effects model with repeated measures). The model includes fixed categorical effects of treatment group, strata of randomization according to IVRS, moment of time, interaction of treatment by moment of time, interaction of strata by moment of time, as well as the continuous fixed covariates of LDL-C value. at baseline and the interaction of LDL-C value at baseline by moment of time

MMRM model and description of the baseline level performed in patients with a baseline value and a post baseline value in at least one of the analysis windows used in the model.

Secondary endpoints of key efficacy

Table 11 summarizes the analysis results for the key secondary endpoints in hierarchical order. All key secondary endpoints are statistically significant according to the hierarchical testing procedure.

Table 11

Endpoint Analysis Results Calculated LDL-C p-value - Percentage change from baseline During MC <0.0001 level to Week 24 treatment vs placebo of -58.1%

Calculated LDL-C - Percentage Change from TDI Level Mean Difference from Baseline MC <0.0001 through Week 12 vs. Placebo of -49.2%

Calculated LDL-C - Percentage Change from Baseline During MC <0.0001 Level to Week 12 Treatment vs. Placebo of -49.5%

Apo-B - Percentage Change from Baseline IDT Mean Difference of MC <0.0001 through Week 24 vs. Placebo of -45.8%

Apo-B - Percentage change from baseline During MC mean difference <0.0001 until Week 24 treatment vs placebo of -45.9%

Non-HDL C - Percentage Change from Baseline IDT Mean Difference of MC <0.0001 through Week 24 vs Placebo of -52.4%

Non-HDL C - Percentage Change from Baseline During MC Mean Difference <0.0001 through Week 24 Treatment vs. Placebo of -52.6%

Total C - Percentage change from baseline TDI Mean difference of MC <0.0001 through Week 24 vs placebo of -38.7%

Endpoint Analysis Results P value

Apo-B - Percentage change from baseline TDI Mean difference of MC <0.0001 until Week 12 vs placebo of -37.5%

Non-HDL C - Percentage Change from Baseline IDT Mean Difference of MC <0.0001 through Week 12 vs Placebo of -43.7%

Total C - Percentage change from baseline TDI Mean MC difference <0.0001 through Week 12 vs placebo of -32.5%

Calculated LDL-C - Percentage Change from TDI Level Mean Difference from Baseline MC <0.0001 to Week 52 vs. Placebo of -56.2%

Proportion of patients at very high CV risk who combined TDI estimate for <0.0001 reach calculated LDL-C <70 mg / dL (1.81 mmol / L) or the odds ratio of patients at high CV risk reaching LDL-C vs placebo 155.1

calculated <100 mg / dL (2.59 mmol / L) at Week 24

Proportion of patients at very high CV risk who During pooled estimate for <0.0001 reach calculated LDL-C <70 mg / dL (1.81 mmol / L) or treatment the odds ratio of patients at high CV risk who reach C -LDL vs. placebo of 149.1

calculated <100 mg / dL (2.59 mmol / L) at Week 24

Proportion of patients reaching LDL-C calculated TDI pooled estimate for <0.0001 <70 mg / dL (1.81 mmol / L) at Week 24 Odds Ratio

vs placebo 237.1

Proportion of Patients Reaching LDL-C Calculated During Pooled Estimate for <0, 0001 <70 mg / dL (1.81 mmol / L) at Week 24 Treatment Odds Ratio

versus placebo 237.9

Lp (a) - Percentage change from baseline to IDT pooled estimate for <0, 0001 at Week 24 adjusted mean difference

vs placebo -17.7%

HDL-C - Percentage change from baseline TDI Mean MC difference <0.0001 through Week 24 vs placebo of 8%

Fasting TGs - Percentage change from TDI level pooled estimate for <0.0001 baseline through Week 24 adjusted mean difference

vs placebo of -16.1%

Apolipoprotein A1 - Percentage Change from TDI Mean Difference of MC <0.05 Baseline to Week 24 vs. Placebo of 4.7%

The on-treatment analysis of the percent change in LDL-C from baseline to Week 24 shows results very consistent with the TDI analysis (mean difference in MC vs. placebo of - 58.1% in on-treatment vs. a - 57.9% in the IDT analysis). In fact, few patients had post-treatment LDL-C values (that is, more than 21 days after the last injection) at Week 24: 6 patients (3.7%) in the placebo group and 2 patients (0.6%) in the alirocumab group. A statistically significant decrease in the percent change in LDL-C from baseline to Week 12 (i.e., before possible upward dose adjustment) was observed in the TDI analysis in the alirocumab group (mean MC vs. baseline - 43.5%) compared to placebo group (mean MC vs. baseline 5.7%) (mean MC difference vs. placebo - 49.2%, p <0.0001) .

The key secondary endpoints for Apo B, non-HDL C, total C, Lp (a), HDL-C, and TGs at various time points, as well as the proportion of patients reaching their LDL-C goals and the proportion of patients achieving calculated LDL-C <70 mg / dL at Week 24 were statistically significant according to the hierarchical testing procedure. For the alirocumab group mean LDL-C (SD) at baseline, non-LDL-C, ApoB, and median (IQR) of Lp (a) levels were 144.7 (51.3), 170.3 ( 54.6), 114.3 (30.8) and 34 (12:82) mg / dL respectively. For the placebo group the mean LDL-C (SD) at baseline, non-LDL-C, ApoB, and the median (IQR) of Lp (a) levels were 144.4 (46.8), 169.6 (50.6), 113.4 (28: 5) and 23 (8.72) mg / dL respectively. After 24 weeks, the change in% of mean MC (SE) from baseline to Week 24 for non-LDL C, ApoB Lp (a) levels in the alirocumab group was -42.8%, -41.1% and -25.2%, respectively. The% change from mean MC (SE) from baseline to Week 24 for non-LDL C levels, ApoB Lp (a) for the placebo group it was 9.6%, 4.7%, and -7.5%, respectively. The mean difference of MC versus placebo for non-LDL C, ApoB, and Lp (a) was -52.4%, -45.8%, and 17.7%, respectively.

The proportion of patients at very high cardiovascular (CV) risk who achieve calculated LDL-C <70 mg / dL (1.81 mmol / L) or patients at high CV risk who achieve calculated LDL-C <100 mg / dL (2 , 59 mmol / L) at Week 24 was significantly higher in the alirocumab group than placebo (pooled estimate for the proportion of 72.1% in the alirocumab group vs 2.4% in the placebo group, p <0.0001).

Two consecutive calculated LDL-C values <25 mg / dL (<0.65 mmol / L) were observed in 16 (5.0%) patients. No particular safety concern has been observed in these patients.

Table 12 - Number (%) of patients with 2 consecutive calculated LDL-C <25 mg / dL (<0.65 mmol / L) during the treatment period - Safety population

Placebo Alirocumab 75

Q2W / up to 150 Q2W (N = 163) (N = 322) Patients with 2 consecutive calculated LDL-C values <25 mg / dL 0/163 16/317 (5.0%) 1

Time to first calculated LDL-C value <25 mg / dL (weeks)

two

Number 0 16

Mean (SD) 14.79 (11.37) Median 14.14

Min: max 3.1: 36.1

Patients with 2 consecutive calculated LDL-C values <15 mg / dL 0/163 6/317 (1.9%) 1

Time to first calculated LDL-C value <15 mg / dL

(weeks) 2

Number 0 6

Mean (SD) 18.31 (12.35) Median 20.14

Min: max 4.6: 36.1

The number (n) represents the subset of the total number of patients who met the criteria

The denominator (/ N) within a treatment group is the number of patients for the treatment group who had at least two calculated LDL-C values evaluated at least 21 days apart in the efficacy period.

1 are considered 2 consecutive values if they are separated by at least 21 days

2 First calculated LDL-C value <25 or <15 mg / dL among the first 2 consecutive calculated LDL-C values <25 or <15 mg / dL per patient

Safety Summary Results:

Alirocumab was well tolerated during the treatment period.

Table 13 - Overview of the adverse event profile: Treatment-emergent adverse events - Safety population

Placebo Alirocumab 75 Q2W / up to 150 Q2W

n (%) (N = 163) (N = 322)

Patients with any TEAE 122 (74.8%) 249 (77.3%)

Patients with any treatment-emergent SAE 15 (9.2%) 39 (12.1%)

Patients with any TEAE leading to death 0 4 (1.2%)

Patients with any TEAE leading to permanent treatment interruption 8 (4.9%) 10 (3.1%)

n (%) = number and percentage of patients with at least one TEAE

Overall, the proportions of patients reporting at least one treatment-emergent adverse event (TEAE) (77.3% in the alirocumab group and 74.8% in the placebo group) or at least one TEAE leading to discontinuation permanent (3.1% in the alirocumab group and 4.9% in the placebo group) were similar in both groups. The SOC "musculoskeletal and connective tissue disorders" was reported in 22.4% of patients in the alirocumab group versus 25.2% in the placebo group. The TEAEs most frequently reported in both treatment groups were "injection site reaction" (11.8% vs. 9.8% in the alirocumab group vs. placebo, respectively) and "nasopharyngitis" (9.9% vs 6.7% in the alirocumab group vs placebo, respectively). Among the events of interest, no particular signal was detected for TEAEs related to allergic events, neurological events, neurocognitive disorders, and diabetes. The SOC "benign, malignant and unspecified neoplasms" was observed in 2.8% of patients in the alirocumab group versus 0.6% in the placebo group with no particular clinical pattern in individual events (all these events were reported as not related to IMP by the researcher). TEAEs "validation confirmed cardiovascular events" were reported for 1.9% of patients in the alirocumab group and 1.2% in the placebo group.

Six deaths (1.9%) were reported as unrelated to IMP by the investigator in the alirocumab group versus none in the placebo group: two myocardial infarctions (MI) (one classified as acute MI and one classified as death sudden cardiac), two metastatic cancers (non-small cell lung cancer and pancreatic carcinoma with secondary Trousseau syndrome causing multiple embolic strokes), a colonic pseudo-obstruction following abdominal surgery in one patient, and sudden cardiac death in a patient due to congestive heart failure and coronary artery disease. Both MI patients had multiple risk factors for coronary artery disease. With respect to cancers, the time to the appearance of the first symptoms (approximately 3.5 and 7.5 months after the start of the investigational medicinal product) is not suggestive of a causal role of the investigational medicinal product.

Abnormalities not relevant to PCSA were observed.

Example 3: A randomized, double-blind, placebo-controlled, parallel-group study to evaluate the efficacy and safety of alirocumab in patients with heterozygous familial hypercholesterolemia not adequately controlled by their lipid-modifying therapy

Introduction

The objective of the study was to evaluate the efficacy and safety of alirocumab in improving lipid parameters in patients with heterozygous familial hypercholesterolaemia (HeFH) who have failed to achieve the goal of LDL-C treatment with maximum tolerated statin therapy, with or without additional lipid modifying therapy (TML). Patients who did not reach the goal of maximum tolerated dose of daily statin therapy, with or without another TML, and whose background treatment was maintained throughout the study, were included in this study.

This specific study (Figure 3) was carried out to demonstrate in patients with heFH who were not in their LDL-C target, that alirocumab 75 mg q2w or 75 mg q2w / 150 mg q2w as complementary therapy to statin / -other TML, causes a statistically significant and clinically significant reduction in LDL-C. This non-target LDL-C population with optimized TML represents a higher risk group with a well-identified unmet medical need that can be treated by adding alirocumab to their LDL-C-lowering therapies.

Objectives of the study

The primary objective of the study was to demonstrate the reduction of LDL-C by alirocumab as add-on therapy to stable maximum tolerated daily statin therapy with or without other TML compared to placebo after 24 weeks of treatment in patients with heHF.

The secondary objectives of the study were: 1) to evaluate the effect of alirocumab 75 mg compared to placebo on LDL-C after 12 weeks of treatment; 2) evaluate the effect of alirocumab on other lipid parameters (eg ApoB, non-HDL C, total C, Lp [a], HDL-C, TG levels and ApoA-1 levels); 3) evaluate the long-term effect of alirocumab on LDL-C; 4) evaluate the safety and tolerability of alirocumab; and 5) evaluating the development of anti-alirocumab antibodies.

Study design

It was a randomized, double-blind, placebo-controlled, parallel-group, multinational study in patients with heFH who were not adequately controlled on their TML (ie, stable maximum tolerated daily statin therapy / - other TML). Not adequately controlled was defined as LDL-C> 70 mg / dL (1.81 mmol / L) at the screening visit (Week 2) in patients with a history of documented CVD or LDL-C> 100 mg / dL (2 , 59 mmol / L) at the screening visit (Week -2) in patients with no documented history of CVD. Patients were randomized 2: 1 to receive either alirocumab 75 mg or placebo by SC injection, every 2 weeks, in addition to stable maximum tolerated daily statin therapy (atorvastatin, rosuvastatin, or simvastatin) with or without another TML. Randomization was stratified according to a previous history of either myocardial infarction (MI) or ischemic stroke, and statin therapy (atorvastatin 40 mg to 80 mg daily or rosuvastatin 20 mg to 40 mg daily versus simvastatin whatever the dose daily, atorvastatin less than 40 mg per day, or rosuvastatin less than 20 mg per day).

The study consisted of three periods: a selection period, a treatment period, and a follow-up period.

The selection period was up to 2 weeks, which includes an intermediate visit during which the patient or caregiver was trained to self-inject / inject using a placebo dose.

The double-blind treatment period was 78 weeks. The first injection of study drug was administered at the clinical site on day 1, after the study evaluations were completed, and as soon as possible after the patient was randomized into the study. The patient / caregiver was administered with subsequent injections outside the clinic according to the administration schedule. On days where the clinical study visit coincides with administration, the study drug dose was administered after all study evaluations were performed and all laboratory samples were collected. The last dose of study drug was administered at Week 76. At Week 12, patients randomized to alirocumab, in a blinded mode, or: 1) continued with alirocumab 75 mg every 2 weeks, if LDL-C of the Week 8 was <70 mg / dL (1.81 mmol / L), or 2) dose titration up to alirocumab 150 mg every 2 weeks, if Week 8 LDL-C was> 70 mg / dL (1 , 81 mmol / L).

The follow-up period (if applicable) was 8 weeks after the end of DBTP for patients who did not consent to participate in the open-label extension study, or if they prematurely discontinued study treatment.

Patients were asked to follow a stable diet (equivalent to the Therapeutic Lifestyle Changes diet of the National Cholesterol Education Program's Adult Treatment Panel III [NCEP ATP III TLC] / Appendix 5) from screening to the end of study visit. The daily dose of statin or other TML (if applicable) should remain stable from screening to the end of study visit. Beginning with Week 24, the background TML can be modified under certain conditions as described below. Table 1 of Example 2 is relevant to this example and provides a summary of the TLC diet for high cholesterol.

The central laboratory reported to an independent external physician any patient achieving 2 consecutive calculated LDL-C levels <25 mg / dL (0.65 mmol / L). Patients meeting this criterion were monitored.

Patient selection

The study population consisted of heFH patients who were not adequately controlled with a stable maximum tolerated daily dose of a statin for at least 4 weeks prior to the screening visit (Week -2), with or without another TML.

A patient must have met the following criteria to be eligible for inclusion in the study: 1) HeFH patients * who were not adequately controlled ** with a maximum tolerated daily dose *** of statin with or without another TML, at a stable dose before screening visit (Week -2).

* The diagnosis of heFH must be made either by genotyping or by clinical criteria. For non-genotyped patients, the clinical diagnosis can be based on either the Simon Broome criteria for defined HF or the WHO / Dutch Lipid Network criteria with a score of> 8 points.

Defined familial hypercholesterolemia was defined herein the same as in Example 2. Possible familial hypercholesterolemia was defined herein as in Example 2. WHO criteria (Dutch Lipid Network clinical criteria) for Heterozygous familial hypercholesterolemia (heHF) diagnoses set forth in Table 2 in Example 2 were the same for this example.

** "Not adequately controlled" was defined herein as in Example 2.

ECC Documented Background was defined herein the same as in Example 2.

Risk equivalents of CHD (include 1 or more of the following criteria): 1) Documented peripheral arterial disease (one of the following criteria must be met): A) Current intermittent claudication (lower extremity muscle discomfort caused by exercise that is both reproducible and relieved by rest within 10 minutes) of presumed atherosclerotic origin together with an ankle-brachial index equal to or less than 0.90 in any leg at rest, or B) history of intermittent claudication (muscle discomfort in the exercise-induced lower limb that is both reproducible and relieved by rest within 10 minutes) in conjunction with endovascular procedure or surgery on one or both legs due to atherosclerotic disease, or C) history of critical limb ischemia in conjunction with thrombolysis , endovascular procedure or surgery on one or both legs due to atherosclerotic disease; 2) Documented prior ischemic stroke with focal ischemic neurological deficit that persisted for more than 24 hours, which is considered to be of atherothrombotic origin. CT or MRI should have been done to rule out hemorrhage and nonischemic neurologic disease.

*** "Maximum tolerated dose" was defined herein the same as in Example 2.

Patients who met all the above inclusion criteria were selected by the following exclusion criteria, which are classified into the following three subsections: exclusion criteria related to the study methodology, exclusion criteria related to the active comparator and / or therapies. mandatory background, and exclusion criteria related to current knowledge of alirocumab.

The exclusion criteria related to the study methodology were: 1) patient without a diagnosis of HFhe made either by genotyping or by clinical criteria; 2) LDL-C <70 mg / dL (<1.81 mmol / L) at the screening visit (Week -2) in patients with a documented history of CVD. NOTE: CVD is defined as CHD, ischemic stroke, or peripheral arterial disease as described above; 3) LDL-C <100 mg / dL (<2.59 mmol / L) at the screening visit (Week -2) in patients with no documented history of CVD; 4) are not on a stable dose of TML (including statin) for at least 4 weeks and / or fenofibrate for at least 6 weeks, as applicable, prior to the screening visit (Week -2) and from screening to screening. randomization; 5) currently taking a statin other than simvastatin, atorvastatin or rosuvastatin; 6) simvastatin, atorvastatin or rosuvastatin is not taken daily or is not taken at a recorded dose; 7) daily doses greater than atorvastatin 80 mg, rosuvastatin 40 mg, or simvastatin 40 mg (except patients with simvastatin 80 mg for more than 1 year, who are eligible); 8) use of fibrates, other than fenofibrate within 6 weeks of the screening visit (Week -2) or between screening and randomization visits; 9) use of nutraceutical products or over-the-counter therapies that may affect lipids that have not been in a stable quantity / dose for at least 4 weeks prior to the screening visit (Week -2) or between screening visits and of randomization; 10) use of red yeast rice products within 4 weeks of the screening visit (Week -2), or between screening and randomization visits; 11) patient who has received plasmapheresis treatment within 2 months prior to screening visit (Week -2), or has plans to receive it during the study; 12) Recent MI (within 3 months before screening visit [week -2] or between screening and randomization visits), unstable angina leading to hospitalization, percutaneous coronary intervention (PCI), graft surgery coronary artery bypass (CAD), uncontrolled cardiac arrhythmia, cerebrovascular accident, transient ischemic attack, carotid revascularization, endovascular procedure, or surgical intervention for peripheral vascular disease; 13) plan to undergo scheduled PCI, CABG, carotid or peripheral revascularization during the study; 14) systolic blood pressure> 160 mmHg or diastolic blood pressure> 100 mmHg at screening visit or randomization visit; 15) New York Heart Association (NYHA) history of class III or IV heart failure within the past 12 months; 16) known history of hemorrhagic stroke; 17) age <18 years or legal age of majority at the screening visit (Week -2), whichever is older; 18) patients not previously informed about a low cholesterol diet before the screening visit (Week -2); 19) newly diagnosed diabetes (within 3 calendar months prior to randomization visit [Week 0]) or poorly controlled (hemoglobin A1c [HbA1c]> 9% at screening visit [Week -2]); 20) Presence of any clinically significant uncontrolled endocrine disease known to influence serum lipids or lipoproteins. Note: Patients on thyroid replacement therapy may be enrolled if the dosage has been stable for at least 12 weeks prior to screening and between screening and randomization visits, and the level of thyroid stimulating hormone (TSH) is within the normal range of the central laboratory at the screening visit; 21) history of bariatric surgery within 12 months prior to screening visit (Week -2); 22) unstable weight defined by a variation> 5 kg within 2 months before the screening visit (Week -2); 23) known history of homozygous HF; 24) known history of PCSK9 loss of function (ie, genetic mutation or sequence variation); 25) use of corticosteroids systemic, unless used as replacement therapy for pituitary-adrenal disease with a stable regimen for at least 6 weeks prior to the randomization visit (Week 0). Note: Topical, intra-articular, nasal, inhaled and ophthalmic steroid therapies are not considered 'systemic' and are allowed; 26) use of continuous estrogen or testosterone hormone replacement therapy unless the regimen has been stable in the last 6 weeks prior to the screening visit (Week -2) and with no plans to change the regimen during the study; 27) history of cancer within the past 5 years, except adequately treated basal cell skin cancer, squamous cell skin cancer, or cervical cancer in situ; 28) known history of a positive HIV test; 29) patient who has taken any investigational drug other than alirocumab training placebo kits within 1 month or 5 half-lives, whichever is longer; 30) patient who has previously been treated with at least 1 dose of alirocumab or any other monoclonal anti-PCSK9 antibody in other clinical studies; 31) conditions / situations such as: a) any clinically significant abnormality identified at the time of selection that, in the discretion of the investigator or any collaborating investigator, would prevent the safe termination of the study or restrict the evaluation of endpoints; for example, major systemic diseases, patients with a short life expectancy; or b) deemed by the investigator or any collaborating investigator as inappropriate for this study for any reason, for example: i) deemed unable to meet specific protocol requirements, such as scheduled visits; ii) those considered unable to administer or tolerate long-term injections according to the patient or the investigator; iii) researcher or any collaborating researcher, pharmacist, study coordinator, other study personnel or relatives directly involved in carrying out the protocol, etc .; iv) presence of any other condition (for example, geographic or social), both current and anticipated, that the researcher feels that; 32) laboratory data during the selection period (which does not include randomization labs at Week 0, unless otherwise noted): i) positive test for hepatitis B surface antigen or hepatitis C antibody ( confirmed by reflex test); ii) positive for beta-hCG in serum or urine pregnancy test (including Week 0) in women of childbearing potential; iii) TG> 400 mg / dL (> 4.52 mmol / L) (1 repeat laboratory is allowed); iv) eGFR <30 mL / min / 1.73 m2 according to the equation of the 4-variable MDRD Study (calculated by the central laboratory); v) alanine aminotransferase (ALT) or aspartate aminotransferase (AST)> 3 x upper limit of normal (ULN) (1 repeat laboratory allowed); vi) CPK> 3 x ULN (1 repeat laboratory allowed); vii) TSH <lower limit of normal (LLN) or> ULN (1 repeat laboratory allowed).

Exclusion criteria related to active comparative and / or mandatory background therapies were: 1) all contraindications to background therapies or warnings / precautions for use (where appropriate) as presented in the respective national product labeling.

Exclusion criteria related to current knowledge of alirocumab were: 1) known hypersensitivity to monoclonal antibody or any component of the drug; 2) pregnant or lactating women; 3) women of childbearing potential who are not protected by highly effective method (s) of birth control (as defined in the informed consent document and / or in an attachment to the local protocol) and / or who are unwilling or are unable to take a pregnancy test. Note: Women of childbearing potential must have a confirmed negative pregnancy test at screening and randomization visits. They must use effective contraception for the entire duration of study treatment and for 10 weeks after the last dose of study drug, and agree to repeat the urine pregnancy test at designated visits. The contraceptive methods applied have to meet the criteria for a highly effective method of birth control according to the "Guidance note on non-clinical safety studies for human clinical trials for pharmaceuticals (CPMP / ICH / 286/95)" . Postmenopausal women must be amenorrheic for at least 12 months.

Treatments under study

The study treatment was a single 1 mL SC injection for a 75 mg or 150 mg dose of alirocumab or placebo given in an autoinjector, administered in the abdomen, thigh, or outer area of the upper arm. The first injection of study drug was administered at the clinical site, as soon as possible after the patient was randomized into the study. The patient was monitored at the clinical site for 30 minutes after the first injection. Subsequent injections were administered by the patient / caregiver outside the clinic, according to the administration schedule. On days where the clinical study visit coincided with administration, the study drug dose was administered after all study evaluations were performed and all laboratory samples were collected. Subcutaneous administration of study drug should be administered every 2 weeks at approximately the same time of day (based on patient preference); it was acceptable for management to enter within a window of / - 3 days.

Sterile alirocumab medication was delivered at a concentration of 75 mg / mL or 150 mg / mL in histidine, pH 6.0, polysorbate 20, and sucrose in an autoinjector.

Alirocumab identical placebo was delivered in the same formulation as alirocumab, without the addition of protein, in an autoinjector.

All patients were taking a maximum tolerated stable daily statin (atorvastatin, rosuvastatin, or simvastatin) / - other TML for the entire duration of the study. The statin dose and the dose of another TML (if applicable) must have been stable for the entire duration of the study, from screening to the end of study visit.

During the double-blind treatment period, modification to the background TML prior to Week 24 was allowed only under certain conditions: 1) exceptional circumstances - primary concerns (including, but not limited to, TG alert, below , published by the central laboratory) guarantee said changes, at the discretion of the researcher; or 2) a confirmed TG alert - the patient meets the previously specified TG alert (TG> 500 mg / dL [5.65 mmol / L]).

During the double-blind treatment period, modification to the background TML was allowed after Week 24 only under certain conditions: 1) exceptional circumstances, per investigator discretion; 2) a confirmed TG alert - the patient meets the previously specified TG alert (TG> 500 mg / dL [5.65 mmol / L], or 3) LDL-C increased by at least 25% compared to C -LDL from randomization visit (and there is no other reasonable explanation).

For a laboratory rescue alert of increased LDL-C> 25% compared to LDL-C from the randomization visit on 2 consecutive occasions, the investigator must have ensured that there is no reasonable explanation for the LDL-C control insufficient (such as an alternative medical cause such as the use of corticosteroids, etc.) and in particular that: adherence to the diet was appropriate; background TML compliance was appropriate; and the study treatment was administered as planned. If any of the above could reasonably explain insufficient LDL-C control, the investigator should have emphasized the absolute need to adhere to the treatment and, if necessary, arranged a specific interview with a qualified nutrition professional and emphasized the absolute necessity. adherence to the diet, and performed a blinded LDL-C assessment within 1 to 2 months. Salvage treatment can be started in the event that no reason could be found for LDL-C above the threshold value.

If no reason could be found for LDL-C above the threshold value, or if appropriate action failed to reduce LDL-C below the threshold value, rescue medication may have been introduced. The efficacy of any such changes would be based on the lack of rescue threshold from the blinded lipid test at the next routinely scheduled laboratory draw. Patients per protocol already received a maximum tolerated dose of statin, so upward titration of the statin dose or change would not be an option. For additional LDL-C reduction, the investigator may have considered adding: a cholesterol absorption inhibitor (ezetimibe), or a bile acid binding sequestrant (the resins cholestyramine and colestipol, or colesevelam, a nonabsorbable polymer ). The following lipid modifying agents may also have been considered: fibrate (Note: Caution should be exercised when combining fibrates with other cholesterol-lowering medications such as statins due to the risk of myopathy. When a fibrate is combined with a statin, fenofibrate is the fibrate of choice because it does not affect statin glucuronidation (the only fibrate allowed per protocol was fenofibrate); Nicotinic acid (niacin) (Note: Niacin increases blood glucose, but has been shown to be effective in modifying lipid disorders in people with diabetes if glucose control is maintained).

The study drug dose was increased (dose adjusted upward) from 75 mg to 150 mg SC every 2 weeks, beginning at Week 12, for an individual patient in LDL-C> 70 mg / dL in Week 8 visit.

Patients were randomized to receive either alirocumab or placebo in a 2: 1 ratio, with permuted block randomization. Randomization was stratified by previous history of MI or ischemic stroke (Yes / No) and statin dose ("Yes" as atorvastatin 40 mg to 80 mg daily or rosuvastatin 20 mg to 40 mg daily and "No" as simvastatin whatever the daily dose, atorvastatin less than 40 mg per day or rosuvastatin less than 20 mg per day) as fixed effects; and baseline LDL-C calculated as a covariate.

Concurrent medications must have been kept to a minimum during the study. If deemed necessary for the well-being of the patient and unlikely to interfere with the study drug, concurrent medications (other than those prohibited during the study) may have been administered at the investigator's discretion, at a stable dose (where appropriate). possible).

Nutraceuticals or OTC therapies that could affect lipids were allowed only if they had been used at a stable dose for at least 4 weeks prior to the screening visit, during the screening period, and were maintained for the first 24 weeks of the double-blind treatment period. After the Week 24 visit, modification to these nutraceuticals or OTC therapies was allowed, but generally should have been avoided. Examples of such over-the-counter nutraceuticals or therapies include omega-3 fatty acids at doses <1000 mg, plant stanols such as those found in Benecol, flax seed oil, and psyllium.

Women of childbearing potential must have used effective contraception throughout study treatment, and for 10 weeks after the last dose of study drug.

Concurrent medications prohibited from the initial screening visit to the end of study visit included the following: statins, other than atorvastatin, rosuvastatin, or simvastatin; fibrates, other than fenofibrate; and red yeast rice products.

Study endpoints

Baseline characteristics included standard demographics (eg, age, race, weight, height, etc.), disease characteristics including personal history, and medication history for each patient.

The primary efficacy endpoint was the percent change in LDL-C calculated from baseline to Week 24, which was defined as: 100 x (LDL-C value calculated at Week 24 - LDL-C value calculated at baseline) / LDL-C value calculated at baseline. The LDL-C value calculated at baseline was the last LDL-C level obtained prior to the first dose of study drug. The calculated LDL-C at Week 24 was the LDL-C level obtained within the Week 24 analysis window and during the primary efficacy period. The primary efficacy period was defined as the time from the first injection of the double-blind study drug to 21 days after the last injection of the double-blind study drug or to the upper limit of the Week 24 analysis window. , whichever comes first.

The secondary key efficacy endpoints were: 1) percent change in LDL-C calculated from baseline to Week 12: similar definition and rules as for the primary efficacy endpoint, except that C- LDL calculated at Week 12 was the level of LDL-C obtained within the Week 12 analysis window and during the 12 week efficacy period. The 12-week efficacy period was defined as the time from the first injection of double-blind study drug to contact with the resupply IVRS at Visit 6 or up to 21 days after the last injection of study drug. , whichever comes first. The blood sample collected on the day of contact with the IVRS for re-supply of Visit 6 will be considered as before the assessment; 2) the percentage change in ApoB from baseline to Week 24. Same definition and rules as for primary endpoint; 3) percent change in non-HDL C from baseline to Week 24. Same definition and rules as for primary endpoint; 4) the percent change in total C from baseline to Week 24. Same definition and rules as for primary endpoint; 5) percentage change in ApoB from baseline to Week 12. Same definition and rules as for percent change in LDL-C calculated from baseline to Week 12; 6) the percent change in non-HDL-C from baseline to Week 12. Same definition and rules as for percent change in LDL-C calculated from baseline to Week 12; 7) the percent change in total C from baseline to Week 12. Same definition and rules as for percent change in LDL-C calculated from baseline to Week 12; 8) the percent change in LDL-C calculated from baseline to Week 52. Definitions and rules are similar to those used for the primary endpoint substituting Week 24 for Week 52; 9) the proportion of patients who reach the goal of LDL-C at Week 24, that is, LDL-C <70 mg / dL (1.81 mmol / L) with previous CVD or <100 mg / dL ( 2.59 mmol / L) for patients without previous CVD, defined as: (number of patients whose calculated LDL-C value at Week 24 reached the LDL-C target / number of patients in [modified intention to treat (mTDI population )] * 100, using the definition and rules used for the primary endpoint: 10) the proportion of patients achieving LDL-C <70 mg / dL (1.81 mmol / L) at Week 24; 11) the percentage change in Lp (a) from baseline to Week 24. Same definition and rules as for primary endpoint; 12) the percent change in HDL-C from baseline to Week 24. Same definition and rules as for primary endpoint; 13) percent change in HDL-C from baseline to Week 12. Same definition and rules as for percent change in LDL-C calculated from baseline to Week 12; 14) the percentage change in Lp (a) from baseline to Week 12. Same definition and rules as for the percent change in LDL-C calculated from baseline to Week 12; 15) percent change in fasting TG from baseline to Week 24. Same definition and rules as for primary endpoint; 16) percentage change in fasting TG from baseline to Week 12. Same definition and rules as for percent change in LDL-C calculated from baseline to Week 12; 17) the percentage change in ApoA-1 from baseline to Week 24. Same definition and rules as for primary endpoint; 18) the percentage change in ApoA-1 from baseline to Week 12. Same definition and rules as for percentage change in LDL-C calculated from baseline to Week 12.

Other secondary efficacy endpoints were: 1) the percentage change in LDL-C calculated from baseline to Week 78. Definitions and rules are similar to those used for the primary endpoint substituting week 24 for Week 78; 2) the proportion of patients who reach the LDL-C goal at Weeks 12, 52, and 78, that is, LDL-C <70 mg / dL (1.81 mmol / L) in case of previous CVD or <100 mg / dL (2.59 mmol / L) for patients without previous CVD; 3) the proportion of patients achieving LDL-C <100 mg / dL (2.59 mmol / L) at Week 24; 4) the proportion of patients achieving LDL-C <100 mg / dL (2.59 mmol / L) at Week 12; 5) the proportion of patients achieving LDL-C <70 mg / dL (1.81 mmol / L) at Week 12; 6) the absolute change in calculated LDL-C (mg / dL and mmol / L) from baseline to weeks 12, 24, 52, and 78; 7) the percentage change in ApoB, non-HDL C, total C, Lp (a), HDL-C, fasting TG, and ApoA-1 from baseline to weeks 52 and 78; 8) the change in the ApoB / ApoA-1 ratio from baseline to weeks 12, 24, 52 and 78; 9) the proportion of patients with ApoB <80 mg / dL (0.8 g / L) at Weeks 12, 24, 52 and 78; 10) the proportion of patients with non-HDL C <100 mg / dL at Weeks 12, 24, 52 and 78; 11) the proportion of patients with calculated LDL-C <70 mg / dL (1.81 mmol / L) and / or> 50% reduction in calculated LDL-C (if calculated LDL-C> 70 mg / dL [1 , 81 mmol / L]) at Weeks 12, 24, 52 and 78.

Other endpoints were: 1) anti-alirocumab antibody status (positive / negative) and titers assessed throughout the study; 2) the percentage change in high sensitivity C-reactive protein (hs-CRP) from baseline to weeks 24, 52, and 78; 3) the absolute change in HbA1c (%) from baseline to weeks 24, 52, and 78; and 4) response for each EQ-5D point, index score, and change in index score from baseline to week 52.

Study visits

The following visits were scheduled:

On visit 1 / Selection / Day -14 to -8; Visit 2 / Selection / Day -7 (+/- 3 days); Visit 3 / Initial level / Week 0 / Day 1; Visit 4 / Week 4 / Day 29 (+/- 7 days); Visit 6 / Week 12 / Day 85 (+/- 3 days); Visit 7 / Week 16 / Day 113 (+/- 7 days); Visit 8 / Week 24 / Day 169 (+/- 3 days) / Primary endpoint evaluation; Visit 9 / Week 36 / Day 253 (+/- 7 days); Visit 10 / Week 52 / Day 365 (+/- 5 days); Visit 11 / Week 64 / Day 449 (+/- 7 days); Visit 12 / Week 78 / Day 547 (+/- 5 days); and the End of study / Visit 13 / Week 86 / Day 603 (+/- 7 days).

Medical / surgical history, medication history, demographic characteristics, height, Hepatitis B surface antigen, and serum pregnancy test were performed in order to determine study eligibility or characterize the population at baseline.

All laboratory samples were collected prior to administration of the study drug dose.

Blood samples for lipid panels should be collected in the morning, fasting (i.e. overnight for at least 10 hours of fasting, water only, and abstaining from smoking) for all clinic visits. It was recommended not to consume alcohol within 48 hours and not to do intense physical exercise within 24 hours prior to blood sampling. Note: if the patient was not fasting, the blood sample for lipids was not collected and a new appointment was scheduled the day after (or as soon as possible on that date), with a reminder that the patient was fasting .

Sample size and power considerations

A total sample size of 45 patients (30 on alirocumab and 15 on placebo) will have 95% power to detect a difference in the mean percentage change in LDL-C of 30% with a two-sided significance level of 0.05; assuming a common standard deviation of 25% and that all 45 patients have an assessable primary endpoint.

To meet regulatory requirements through the program, the sample size increased to 126 alirocumab patients, in an attempt to understand safety in a larger population. To have at least 126 alirocumab patients treated for 12 months in this study, and assuming a dropout rate of 10% during the first 3-month period and a dropout rate of 20% during the remaining 9-month period, we increased the total sample size and was rounded up to 250 patients, with a 2: 1 randomization ratio (alirocumab: 167, placebo: 83).

Analysis populations

Intention-to-treat population

The randomized population included all randomized patients, and was analyzed according to the treatment assigned by randomization.

The TDI population (also known as the complete analysis population [PCA]) was defined as all randomized patients who had an evaluable primary endpoint. The endpoint was evaluable when the following two conditions were met: 1) availability of an LDL-C value calculated at baseline; and 2) availability of at least 1 value of LDL-C calculated within 1 of the analysis windows through Week 24.

Patients in the IDT population were analyzed according to randomized treatment group (ie, treatment group as randomized).

Intent to treat modified

The mTDI population was defined as any randomized population that took at least 1 dose or part of a dose of study drug and had an assessable primary endpoint. The endpoint was considered evaluable (ie, efficacy treatment period) when both of the following conditions were met: 1) availability of a baseline calculated LDL-C value; and 2) availability of at least 1 calculated LDL-C value during the efficacy treatment period and within one of the analysis windows up to Week 24. The efficacy treatment period is defined as the time from the first injection of the double-blind study drug up to 21 days after the last injection of the double-blind study drug.

Patients in the mTDI population were analyzed according to the treatment group assigned by randomization.

Safety analysis population

The safety population considered for safety analyzes was the randomized population that received at least 1 dose or part of a dose of study drug. Patients were analyzed according to the treatment actually received (ie, treatment group as treated, placebo or alirocumab).

Results

Description of the study populations

A total of 249 patients (82 to the placebo group and 167 to the alirocumab group) were randomized in this study. One patient was randomized to the placebo group, but received no study treatment due to the reason for withdrawing consent before receiving the first IMP injection. Therefore, the patient was excluded from the safety population. Two patients were excluded from the randomized patients (one in the above placebo group and one in the alirocumab group) from the TDI and TDI populations due to the lack of post-LDL-C assessments at baseline.

Table 14: Analysis populations

Placebo Alirocumab 75 Q2W / up to 150 Q2W All

(N = 82) (N = 167) (N = 249)

Randomized population 82 (100%) 167 (100%) 249 (100%)

Efficacy population:

Intention to Treat (IDT) 81 (98.8%) 166 (99.4%) 247 (99.2%)

Modified intention to treat (mTDI) 81 (98.8%) 166 (99.4%) 247 (99.2%)

Quality of life population 80 (97.6%) 164 (98.2%) 244 (98.0%)

Anti-alirocumab antibody population 77 (93.9%) 166 (99.4%) 243 (97.6%)

Safety population 81 (98.8%) 167 (100%) 248 (99.6%)

Note: Patients in the safety and anti-alirocumab antibody populations are tabulated according to the treatment actually received (as treated). For the other populations, patients are tabulated according to their randomized treatment

In the alirocumab group, among the 158 patients who received at least one injection after Week 12, 61 (38.6%) patients received automatic dose titration at Week 12 in a blinded mode from alirocumab 75 mg Q2W up to 150 mg Q2W.

Subject dispositions

Regarding the first stage analysis data deadline, the patient status for the 249 randomized patients is presented below: 1) 0 (0.0%) patients completed the 78-week double-blind treatment period , due to patients in treatment who have not yet made it to the Week 78 visit; 2) 234 (94.0%) patients were still in treatment: 78 (95.1%) in the placebo group and 156 (93.4%) in the alirocumab group; 3) 9 (3.6%) randomized and treated patients prematurely discontinued study treatments before Week 24: 1 (1.2%) in the placebo group and 8 (4.8%) in the alirocumab group . 4 (1.6%) patients terminated study treatments prematurely due to adverse events: 0 in the placebo group vs 4 (2.4%) in the alirocumab group. 3 (1.2%) patients prematurely terminated treatments in study due to poor protocol compliance: 1 (1.2%) in the placebo group and 2 (1.2%) in the alirocumab group. 2 (0.8%) patients terminated study treatments prematurely due to various other reasons: 0 in the placebo group vs 2 (1.2%) in the alirocumab group; 4) 13 (5.2%) randomized and treated patients prematurely discontinued study treatments before Week 52: 2 (2.4%) in the placebo group and 11 (6.6%) in the alirocumab group . 5 (2.0%) patients terminated study treatments prematurely due to adverse events: 0 in the placebo group vs 5 (3.0%) in the alirocumab group.

3 (1.2%) patients prematurely terminated study treatments due to poor protocol compliance: 1 (1.2%) in the placebo group and 2 (1.2%) in the alirocumab group. 5 (0.8%) patients terminated study treatments prematurely due to various other reasons: 1 (1.2%) in the placebo group and 4 (2.4%) in the alirocumab group; 5) 14 (5.6%) patients prematurely terminated study treatments before completing the 78-week treatment period: 3 (3.7%) in the placebo group and 11 (6.6%) in the group with alirocumab. 6 (2.4%) patients terminated study treatments prematurely due to adverse events: 1 (1.2%) in the placebo group and 5 (3.0%) in the alirocumab group. 3 (1.2%) patients prematurely terminated study treatments due to poor protocol compliance: 1 (1.2%) in the placebo group and 2 (1.2%) in the alirocumab group. 5 (2.0%) patients terminated study treatments prematurely for various other reasons: 1 (1.2%) in the placebo group and 4 (2.4%) in the alirocumab group.

The following table provides the availability of LDL-C values over time. At Week 24, the primary efficacy endpoint was available for 78 (96.3%) patients in the placebo group and 157 (94.5%) in the alirocumab group. There were 77 (95.1%) evaluations during treatment and 1 (1.2%) evaluation without treatment in the placebo group, as compared to 155 (93.4%) evaluations during treatment and 2 (1.2 %) evaluations without treatment in the alirocumab group. At Week 52, the key secondary efficacy endpoint was available for 78 (96.3%) patients in the placebo group and 158 (95.2%) patients in the alirocumab group.

Table 15: Availability of LDL-C calculated over time - IDT population

Placebo Alirocumab 75 Q2W / up to 150 Q2W

LDL-C (N = 81) (N = 166)

calculated

Value during the Post Value Value during the Post Value Value treatment missing treatment treatment missing treatment

WEEK 4 79 (97.5%) 0 2 162 (97.6%) 0 4 (2.4%)

(2.5%)

WEEK 8 79 (97.5%) 0 2 156 (94.0%) 0 10 (6.0%)

(2.5%)

WEEK 12 76 (93.8%) 0 5 151 (91.0%) 1 (0.6%) 14 (8.4%)

(6, 2%)

WEEK 16 77 (95.1%) 0 4 149 (89.8%) 3 (1.8%) 14 (8.4%)

(4.9%)

WEEK 24 77 (95.1%) 1 (1, 2%) 3 155 (93.4%) 2 (1, 2%) 9 (5.4%)

(3.7%)

WEEK 36 73 (90.1%) 0 8 153 (92.2%) 2 (1, 2%) 11 (6, 6%)

(9.9%)

WEEK 52 78 (96.3%) 0 3 155 (93.4%) 3 (1.8%) 8 (4.8%)

(3.7%)

A value was obtained during treatment after the first injection of the study treatment and within 21 days after the last injection of the study treatment.

A post-treatment value was obtained more than 21 days after the last injection of the study treatment.

Primary endpoint was missing for 12 (4.9%) patients at Week 24. At the Week 24 visit, the reasons for missing values were as follows: 1) 4 subject with no samples obtained due to study discontinuation early; 2) 2 subjects were still on treatment, but no LDL-C was made at Week 24; 3) 6 samples were obtained at Week 24, but no LDL-C could be calculated (5 with TGs> 400 mg / dL and measured LDL-C reported, 1 with> 400 mg / dL but no LDL-C reported measured).

Demographic and baseline characteristics

Overall, demographic characteristics, baseline disease characteristics, baseline efficacy lipid parameters, history of TML, and background TML use were homogeneous between patients randomized to the alirocumab group and randomized patients. to the placebo group (see Table 16). In particular, the mean baseline LDL-C in the alirocumab group was 134.6 mg / dL (SD = 41.1 mg / dL) compared to that in the placebo group which was 134.0 mg / dL. (SD = 41.4 mg / dL) with a global mean of 134.4 mg / dL (SD = 41.1 mg / dL). A possibly notable exception is the observed difference in BMI at baseline, with a mean BMI of 28.6 kg / m2 (SD = 4.6 kg / m2) in the alirocumab group compared to 27.7 kg / m2. (SD = 4.7 kg / m2) in the placebo group.

Table 16. Baseline Characteristics of the FH II Patient Population

Table 17: Characteristics of the disease and other relevant data at baseline - Randomized population

Placebo Alirocumab 75 All p-value (N = 82) Q2W / up to 150 Q2W (N = 249) vs (N = 167) placebo

Type of hypercholesterolemia

Familial hypercholesterolemia 82 (100%) 167 (100%) 249 (100%) heterozygous (HFhe)

Unfamilial hypercholesterolemia (H no F) 0 0 0

Time since diagnosis of

hypercholesterolemia (years)

Placebo Alirocumab 75 All P-value (N = 82) Q2W / up to 150 Q2W (N = 249) vs (N = 167) placebo Number 82 167 249 0.4938 Mean (SD) 12.7 (8, 8) 12 , 9 (7.9) 12, 8 (8, 2)

Median 10.8 12.3 11.5

Min: max 0: 42 0: 40 0: 42

Diagnosis confirmation *

By genotyping 67 (81.7%) 117 (70.1%) 184

(73.9%)

By WHO / Simon Broome 18 (22, 0%) 52 (31.1%) 70 (28.1%) Definite / Certain 18 (22, 0%) 52 (31.1%) 70 (28.1%)

* The diagnosis of HeHF can be confirmed by genotyping as well as WHO or Simon Broome criteria. Note: P-values comparing baseline data between treatment groups are provided for descriptive purposes, as a selection tool, using Fisher's exact test for qualitative data and the one-sided asymptotic ANOVA test for Wilcoxon scores ( Kruskal-Wallis test) for continuous data.

Table 18: Background TML at Randomization - Randomized Population

Placebo Alirocumab 75 All (N = 82) Q2W / up to 150 Q2W (N = 249) p-value vs (N = 167) placebo Any statin 82 (100%) 167 (100%) 249 (100%) Taking statin from high intensity 72 (87.8%) 144 (86.2%) 216 (86.7%) 0.8434

Atorvastatin daily dose (mg)

10 2 (2.4%) 2 (1.2%) 4 (1.6%) 20 0 8 (4.8%) 8 (3.2%)

40 13 (15.9%) 27 (16.2%) 40 (16.1%)

80 16 (19.5%) 28 (16.8%) 44 (17.7%)

Other doses 1 (1.2%) 0 1 (0.4%)

Rosuvastatin Daily Dose (mg)

5 1 (1.2%) 1 (0.6%) 2 (0.8%) 10 2 (2.4%) 4 (2.4%) 6 (2.4%) 20 8 (9.8 %) 30 (18.0%) 38 (15.3%)

40 33 (40.2%) 59 (35.3%) 92 (36.9%)

Other doses 1 (1.2%) 1 (0.6%) 2 (0.8%)

Simvastatin daily dose (mg)

10 1 (1.2%) 0 1 (0.4%) 20 1 (1.2%) 3 (1.8%) 4 (1.6%)

40 0 3 (1, 8%) 3 (1, 2%)

80 3 (3.7%) 1 (0.6%) 4 (1.6%)

Other doses 0 0 0

Any TML other than statins * 57 (69.5%) 117 (70.1%) 174 (69.9%) 1.0000 Any TML other than 54 (65.9%) 115 (68.9%) 169 ( 67.9%) nutraceuticals

Ezetimibe 53 (64.6%) 112 (67.1%) 165 (66.3%) Nutraceuticals 7 (8.5%) 8 (4.8%) 15 (6.0%) Note: Values of p comparing data at baseline between treatment groups for descriptive purposes, as a selection tool, using Fisher's exact test.

* in combination with statins or not.

Table 19 - Cardiovascular history and breakdown of risk factors

Table 20: Baseline Lipid Efficacy Parameters - Quantitative Summary in Conventional Units - Randomized Population

Placebo Alirocumab 75 Q2W / up to 150 Q2W All p-value (N = 82) (N = 167) (N = 249) vs placebo calculated LDL-C

(mg / dL)

Number 82 167 249 0.8507 Mean (SD) 134.0 (41.4) 134.6 (41.1) 134.4 (41.1) Median 126.0 128.0 126.0

Q1: Q3 109.0: 151.0 107.0: 154.0 108.0: 151.0

Min: Max 74: 295 58: 303 58: 303

Measured LDL-C (mg / dL)

Number 70 149 219 0.6375 Mean (SD) 130.2 (36.6) 132.6 (40.6) 131.8 (39.3) Median 125.5 126.0 126.0

Q1: Q3 104.0: 145.0 104.0: 149.0 104.0: 147.0

Min: Max 71: 249 49: 310 49: 310

HDL-C (mg / dL)

Number 82 167 249 0.4437 Mean (SD) 54.2 (15.7) 52.6 (15.7) 53.1 (15.7)

Median 51.0 50.0 51.0

Q1: Q3 42.0: 63.0 42.0: 61.0 42.0: 62.0

Min: Max 25: 103 24: 110 24: 110

Total C (mg / dL)

Number 82 167 249 0.9589 Mean (SD) 211.7 (45.6) 211.6 (45.8) 211.6 (45.6) Median 200, 0 205.0 202, 0

Q1: Q3 179.0: 237.0 178.0: 242.0 179.0: 239.0

Min: Max 133: 376 123: 391 123: 391

C no HDL (mg / dL)

Number 82 167 249 0.8208 Mean (SD) 157.5 (43.7) 159.0 (44.8) 158.5 (44.4) Median 150.5 147.0 149.0

Q1: Q3 129.0: 170.0 127.0: 181.0 127.0: 177.0

Placebo Alirocumab 75 Q2W / up to 150 Q2W All p-value (N = 82) (N = 167) (N = 249) vs placebo Min: max 93: 320 76: 326 76: 326

Fasting TGs (mg / dL)

Number 82 167 249 0.6593

Mean (SD) 116.6 (56.8) 123.2 (69.3) 121.0 (65.4)

Median 100.5 105.0 104.0

Q1: Q3 81.0: 136.0 81.0: 144.0 81.0: 141.0

Min: Max 47: 366 46: 581 46: 581

Apo-B (mg / dL)

Number 81 167 248 0.9533

Mean (SD) 107.7 (23.9) 107.9 (27.4) 107.9 (26.3)

Median 103.0 102, 0 102, 0

Q1: Q3 91.0: 116.0 91.0: 122.0 91.0: 121.0

Min: Max 74: 187 57: 208 57: 208

Apo-A1 (mg / dL)

Number 81 167 248 0.3472

Mean (SD) 148.9 (29.6) 146.3 (29.4) 147.2 (29.4)

Median 150.0 142.0 144.5

Q1: Q3 129.0: 166.0 127.0: 160.0 128.0: 162.5

Min: max 82: 223 90: 252 82: 252

Apo-B / Apo-A1 (ratio)

Number 81 167 248 0.7518

Mean (SD) 0.8 (0, 2) 0.8 (0, 2) 0.8 (0, 2)

Median 0.7 0.7 0.7

Q1: Q3 0.6: 0.8 0.6: 0.9 0.6: 0.9

Min: max 0: 1 0: 2 0: 2

Lipoprotein- (a) (mg / dL)

Number 81 167 248 0.9910

Mean (SD) 50.9 (59.7) 49.8 (69.2) 50.2 (66.1)

Placebo Alirocumab 75 Q2W / up to 150 Q2W All p-value (N = 82) (N = 167) (N = 249) vs placebo

Median 21, 0 22, 0 22, 0

Q1: Q3 7.0: 76.0 8.0: 70.0 7.5: 75.0

Min: max 2: 232 2: 555 2: 555

Total C / HDL-C (ratio)

Number 82 167 249 0.6572

Mean (SD) 4.2 (1.3) 4.3 (1.5) 4.3 (1.5)

Median 3.9 3.9 3.9

Q1: Q3 3.3: 4.8 3.3: 5.0 3.3: 4.9

Min: Max 2: 9 2: 11 2: 11

Note: P-values comparing baseline data between treatment groups are provided for descriptive purposes, as a selection tool, using the one-sided asymptotic ANOVA test for Wilcoxon scores (Kruskal-Wallis test).

The collection of LDL-C measured in the initial protocol was not planned and was added in a modification. Therefore, the measured LDL-C values are available for fewer patients compared to the calculated LDL-C values.

Degree of exposure

Injection exposure was similar across the treatment groups with a mean exposure of approximately 58-60 weeks. Alirocumab-treated patients were exposed for 2-75.9 weeks and placebo-treated patients for 11.6-75.7 weeks. The majority (93.5%: 97.5%, alirocumab: placebo, respectively) of the patients were treated for more than 52 weeks.

In the alirocumab group, among the 158 patients who received at least one injection after Week 12, 61 (38.6%) patients received automatic up-titration in a blinded mode at Week 12 from alirocumab 75 mg Q2W up to 150 mg Q2W. 26 patients were not given the opportunity for up-titration at Week 12 due to missing LDL-C values at Week 8 at the time of the up-titration decision. Of the 26 patients for whom the Week 8 LDL-C value was missing, 4 alirocumab patients should have had their dose adjusted upward based on the now available Week 8 LDL-C data. The remaining patients were either in the placebo treatment group, or the Week 8 LDL-C visits for the alirocumab patients were below the LDL-C cut-off <70 mg / dL for upward dose adjustment. .

Efficacy analysis

Primary efficacy analysis in the IDT population

Primary endpoint analysis (percent change in LDL-C calculated from baseline to Week 24) is provided based on an MMRM model over the IDT population, using estimates of CM means at Week 24. This The repeated measures approach includes all LDL-C values collected during treatment and without treatment up to Week 52. At Week 24, 3 (3.7%) patients in the placebo group and 9 (5.4%) patients in the alirocumab group did not have a calculated LDL-C value (Table 15). These missing values were taken into account by the MMRM model.

The primary efficacy analysis showed a statistically significant decrease in the percentage change from baseline LDL-C calculated at Week 24 in the TDI analysis for the alirocumab treatment group (mean MC = -48.7% ) compared to placebo (mean MC = 2.8%). The mean MC difference between the alirocumab treatment group and the placebo treatment groups is -51.4% (p <0.0001). 81.4% of HeFH patients in the alirocumab group achieved LDL-C targets at 24 weeks, compared with 11.3% for the placebo group.

Table 21: Percentage change from baseline in LDL-C calculated at Week 24 (IDT analysis):

MMRM Analysis - IDT Population

Calculated LDL cholesterol ^Placebo Alirocumab 75 Q2W / up to 150 _{(N = 81)} Q2W (N = 166)

Initial level (mmol / L)

Number 81 166

Mean (SD) 3,470 (1,078) 3,486 (1,069)

Median 3,263 3,289

Min: Max 1.92: 7.64 1.50: 7.85

Initial level (mg / dL)

Number 81 166

Mean (SD) 134.0 (41.6) 134.6 (41.3)

Median 126.0 127.0

Min: Max 74: 295 58: 303

Percentage change in Week 24 from level

initial (%)

LS mean (SE) 2, 8 (2, 8) -48.7 (1.9)

Mean difference of MC (SE) vs placebo -51.4 (3.4)

95% CI (-58.1 to -44.8)

P-value vs placebo <.0001

Note: Least squares means (LC), standard errors (SE) and p-value taken from MMRM analysis (mixed effects model with repeated measures). The model includes the fixed categorical effects of treatment group, randomization strata according to IVRS, moment of time, interaction of treatment by moment of time, interaction of strata by moment of time, as well as the continuous fixed covariates of the LDL-C value. calculated at the initial level and the interaction of value at the initial level per moment of time.

MMRM model and description of the baseline level performed in patients with a baseline value and a post baseline value in at least one of the analysis windows used in the model.

The p-value is followed by a '*' if it is statistically significant according to the fixed hierarchical approach used to ensure a strong control for the overall type 1 error rate at the 0.05 level.

LDL-C calculated over time

Figure 4 is a graph showing the mean percent change in MC (+/- SE) of LDL-C from baseline over time for the IDT population. Note: Least squares means (LC) and standard errors (SE) taken from MMRM analysis (mixed effects model with repeated measures).

The model includes the fixed categorical effects of the treatment group, point in time, treatment interaction by time point, as well as the continuous fixed covariates of the baseline LDL-C value and the LDL-C interaction at the baseline level. initial by moment of time.

Table 22: LDL-C calculated over time - IDT population

Placebo Alirocumab 75 Q2W / up to 150 Q2W (N = 81) (N = 166)

Calculated LDL-C

Change Change in Change in Value from percentage Initial level change percentage from level Value from level

baseline from baseline baseline mean MC (SE) (mmol / L)

Initial level 3,470 NA NA 3,486 NA NA

(0, 120) (0.083)

Week 4 3.485 0.004 (0.077) 1.1 (2.0) 1.924 -1.56 (0.054) -45.2 (1.4)

(0.077) (0.054)

Week 8 3.561 0.081 (0.090) 3.3 (2.4) 1.913 -1.57 (0.063) -45.3 (1.7)

(0.090) (0.063)

Week 12 3.585 0.104 (0.097) 4.6 (2.6) 1.960 -1.52 (0.068) -43.8 (1.8)

(0.097) (0.068)

Week 16 3.508 0.028 (0.11) 2.4 (2.7) 1.649 -1.83 (0.071) -51.9 (1.9)

(0, 101) (0.071)

Week 24 3,537 0.057 (0.103) 2, 8 (2, 8) 1,754 -1.73 (0.072) -48.7 (1.9)

(0.103) (0.072)

Week 36 3.603 0.122 (0.117) 5.1 (3.2) 1.788 -1.69 (0.081) -48.0 (2.2)

(0.117) (0.081)

Week 52 3.718 0.237 (0.125) 8.4 (3.3) 1.708 -1.77 (0.088) -50.3 (2.3)

(0.125) (0.088)

Week 64 3,601 1,657

(0.107) (0.075)

Week 78 3,574 1,806

(0.109) (0.076)

Mean MC (SE) (mg / dL)

Initial level 134.0 NA NA 134.6 NA NA

(4.6) (3.2)

Week 4 134.6 0.2 (3.0) 1.1 (2.0) 74.3 -60.1 (2.1) -45.2 (1.4)

(3,0) (2, 1)

Week 8 137.5 3.1 (3.5) 3.3 (2.4) 73.9 -60.5 (2.4) -45.3 (1.7)

(3.5) (2.4)

Week 12 138.4 4.0 (3.7) 4.6 (2.6) 75.7 -58.7 (2.6) -43.8 (1.8)

(3.7) (2, 6)

Week 16 135.5 1.1 (3.9) 2.4 (2.7) 63.7 -70.7 (2.7) -51.9 (1.9)

(3.9) (2.7)

Placebo Alirocumab 75 Q2W / up to 150 Q2W

(N = 81) (N = 166)

Calculated LDL-C

Change Change in Change in Value from Percentage Change

Value from initial level percentage from initial level initial level from initial

Week 24 136.6 2.2 (4.0) 2, 8 (2, 8) 67.7 -66.7 (2.8) -48.7 (1.9)

(4,0) (2, 8)

Week 36 139.1 4.7 (4.5) 5.1 (3.2) 69.0 -65.3 (3.1) -48.0 (2.2)

(4.5) (3.1)

Week 52 143.6 9.2 (4.8) 8.4 (3.3) 65.9 -68.4 (3.4) -50.3 (2.3)

(4.8) (3.4)

Week 64 139.0 64.0

(4.1) (2.9)

Week 78 138.0 69.7

(4.2) (2.9)

* Baseline is described using means and standard errors.

Note: Least squares means (LC), standard errors (SE) and p-value taken from MMRM analysis (mixed effects model with repeated measures). The model includes the fixed categorical effects of treatment group, randomization strata according to IVRS, moment of time, interaction of treatment by moment of time, interaction of strata by moment of time, as well as the continuous fixed covariates of the LDL-C value. at baseline and the interaction of LDL-C value at baseline by moment of time.

MMRM model and description of the baseline level performed in patients with a baseline value and a post baseline value in at least one of the analysis windows used in the model.

Sensitivity to severe GCP breach

There were no seriously GCP non-compliant sites in this study.

Key secondary efficacy analysis

The following table summarizes the analysis results for all the key secondary endpoints in rank order for the statistical test at the 0.05 level of significance. This study has achieved statistically significant effects in favor of alirocumab-treated patients for all but the last in the hierarchy (i.e. Apo A-1 - Percentage change from baseline to Week 12) of the secondary criteria effectiveness assessment key.

For clarification, IDT analysis is defined for patients in the IDT population and includes all endpoint assessments in one analysis window, regardless of study treatment administration status (i.e., includes post-treatment assessments). During treatment analysis is defined for patients in the mTDI population and includes all endpoint assessments from the first injection of double-blind study drug to the day of the last 21 day injection (i.e., includes assessments at the efficacy treatment period).

Table 23: Summary of Secondary Key Efficacy Endpoints

Endpoint / Analysis Comparison Result Result Placebo value alirocumab p

1. LDL-C in WK24 - IDT analysis Mean of MC: Mean of MC: - Dif: -51.4% <0, 0001

2, 8% 48.7%

2. LDL-C in WK 24 - analysis Mean of MC: Mean of MC: - Dif: -52.2% <0, 0001 during treatment 2.7% 49.4%

Endpoint / Analysis Comparison Result Result Placebo value alirocumab p 3. LDL-C in WK12 - TDI analysis Mean MC: Mean MC: - Dif: -48.4% <0, 0001

4.6% 43.8%

4. LDL-C in WK12 - analysis Mean of MC: Mean of MC: - Dif: -48.8% <0, 0001 during treatment 4.6% 44.2%

5. Apo B in WK24 - IDT analysis Mean of MC: - Mean of MC: - Dif: -39.3% <0, 0001

3.5% 42.8%

6. Apo B in WK24 - analysis Mean of MC: - Mean of MC: - Dif: -39.8% <0, 0001 during treatment 3.5% 43.2%

7. C not HDL in WK24 - analysis Mean of MC: Mean of MC: - Dif: -45.7% <0, 0001 IDT 3.1% 42.6%

8. C not HDL in WK24 - analysis Mean of MC: Mean of MC: - Dif: -46.4% <0, 0001 during treatment 3.1% 43.2%

9. Total cholesterol in WK24 - MC mean: MC mean: - Diff: -32.8% <0, 0001 TDI analysis 2.1% 30.6%

10. Apo B in WK12 - analysis Mean of MC: - Mean of MC: - Dif: -34.5% <0, 0001 IDT 0.9% 35.4%

11. C non-HDL in WK12 - analysis Mean of MC: Mean of MC: - Dif: -42.0% <0, 0001 IDT 4.1% 37.9%

12. Total cholesterol in WK12 - MC mean: MC mean: - Diff: -29.9% <0, 0001 TDI analysis 3.4% 26.6%

13. LDL-C in WK52 - analysis Mean of MC: Mean of MC: - Dif: -58.8% <0, 0001 IDT 8.4% 50.3%

14. Very high CV LDL-C <Proportion = 11.3% Proportion == 81.4% Ratio of <0, 0001 70 mg / dL or High CV-LDL-C <odds = 52.2

100 mg / dL in WK24 - analysis

IDT

15. Very high CV LDL-C <Proportion = 11.6% Proportion = 82.1% Ratio of <0, 0001 70 mg / dL or high CV-LDL-C <odds = 53.3

100 mg / dL in WK24 - analysis

during treatment

16. LDL-C <70 mg / dL in WK24 Ratio = 1.2% Ratio = 68.2% Ratio of <0, 0001 - IDT probability analysis = 239.7

17. LDL-C <70 mg / dL in WK24 Ratio = 1.3% Ratio = 68.8% Ratio of <0, 0001 - analysis during treatment odds = 240.6

18. Lp (a) in WK24 - IDT analysis Mean of MC: - Mean of MC: - Dif: -20.3% <0, 0001

10.0% 30.3%

19. HDL-C in WK24 - analysis Mean of MC: - Mean of MC: 6.0% Diff: 6.8% 0.0009 IDT 0.8%

20. Fasting triglycerides in MC mean: MC mean: - Dif: -10.9% 0.0017 WK24 - TDI analysis 0.4% 10.5%

19. Apo A-1 in WK24 - analysis Mean of MC: - Mean of MC: 2.8% Dif: 4.4% 0.0062 IDT 1.6%

20. Lp (a) in WK12 - IDT analysis Mean of MC: - Mean of MC: - Diff: -19.1% <0, 0001

5.6% 24.7%

Endpoint / Analysis Comparison Result Result Placebo value alirocumab p

21. HDL-C in WK12 - analysis Mean of MC: Mean of MC: 6.0% Diff: 4.3% 0.0147 IDT 1.7%

22. Fasting triglycerides in MC mean: MC mean: - Dif: -8.9% 0.0258 WK12 - TDI analysis 0.9% 8, 0%

Apo A-1 in WK12 - IDT analysis Mean of MC: - Mean of MC: 0.4% Diff: 2.3% 0.1475

1.9%

Hierarchical test completed

All key secondary efficacy endpoints, except the percentage change in Apo A-1 from baseline to Week 12 in the IDT population, achieved statistically significant effects in favor of patients treated with alirocumab according to the procedure of hierarchical test.

Key secondary efficacy analysis for percent change from baseline calculated LDL-C to week 24 in mTDI population (analysis during treatment) showed results consistent with TDI analysis with statistically significant decrease in calculated LDL-C in the alirocumab treatment group (mean MC = -49.4%) compared to placebo (mean MC = 2.7%). The mean MC treatment difference between alirocumab-treated patients and placebo-treated patients is -52.2% (p <0.0001). In fact, few LDL-C values were collected from post-treatment patients (i.e., more than 21 days after the last injection) at Week 24: 1 patient (1.2%) in the placebo group and 2 patients (1.2%) in the alirocumab group.

The decrease in the percentage change in Apo A-1 from baseline to Week 24 in the IDT analysis was not statistically significant: mean MC vs. baseline was 0.4% in the alirocumab and -1 group , 9% in the placebo group (mean difference of MC vs placebo 2.3%, p = 0.1475).

LDL-C calculated over time (includes observed data)

Figure 5 is a graph showing the mean percent change of MC (+/- SE) of LDL-C from baseline during the efficacy treatment period with time for the mTDI population.

Summary

In general, demographic characteristics, baseline disease characteristics, baseline efficacy lipid parameters, history of TML, and background TML use were comparable between patients randomized to the alirocumab group and patients randomized to the group with placebo. In particular, the mean baseline (SD) LDL-C in the alirocumab group was 134.6 (41.1) mg / dL compared to that in the placebo group which was 134.0 (41.4) mg / dL.

The primary efficacy endpoint and all secondary endpoints, except for percentage change in Apo A-1 from baseline to Week 12 in the IDT population (IDT analysis), achieved a statistically significant benefit in favor of patients treated with alirocumab according to the hierarchical testing procedure.

Summary safety results

A total of 248 patients were randomized and received at least a partial dose of study treatment (safety population). Below is a high-level summary of adverse events and events of interest.

Table 24: Adverse Event Profile Overview: Treatment-Emergent Adverse Events - Safety Population

Placebo Alirocumab 75 Q2W / up to 150 (N = 81) Q2W (N = 167)

Patients with any TEAE 62 (76.5%) 117 (70.1%)

Patients with any treatment-emergent SAE 7 (8, 6%) 10 (6, 0%)

Patients with any TEAE leading to death 0 0

Patients with any TEAE leading to permanent 1 (1.2%) 5 (3.0%) interruption of treatment

TEAE: Treatment Emergent Adverse Event, SAE: Serious Adverse Event

n (%) = number and percentage of patients with at least one TEAE

Treatment-emergent SAEs occurred in a total of 17 patients, specifically 10 (6.0%) patients in the alirocumab treatment group and 7 (8.6%) patients in the placebo treatment group. There were no more than 2 reports in any SOC for any treatment group and no individual SAE was reported more than once in any treatment group.

No patient deaths were reported at the time of this first stage analysis.

A total of 6 patients prematurely discontinued study treatment due to TEAE. Specifically, 5 (3.0%) patients in the alirocumab treatment group discontinued treatment prematurely due to rectal adenocarcinoma, diarrhea, nausea, angioedema, asthenia, and elevated alanine aminotransferase. One (1.2%) patient in the placebo treatment group discontinued due to syncope.

TEAEs occurred in 117 (70.1%) patients in the alirocumab treatment group and 62 (76.5%) patients in the placebo treatment group. TEAEs that occurred in> 5% of patients in any treatment group are: injection site reaction (10.8% vs 7.4% in the alirocumab and placebo groups, respectively), headache (8.4 % vs. 8.6% in the alirocumab and placebo group, respectively), myalgia (6.0% vs. 6.2% in the alirocumab and placebo group, respectively), and diarrhea (5.4% vs. 1.2% in the alirocumab and placebo groups, respectively).

For TEAEs of special interest (AESIs), the results are presented by groupings of pre-defined SMQ preferred terms.

Treatment-emergent injection site reactions (ISRs) occurred in 18 (10.8%) patients in the alirocumab treatment group and 6 (7.4%) patients in the placebo treatment group. None of the AEs were serious.

General allergic TEAEs, identified by the MedDRA SMQ of "hypersensitivity" occurred in 17 (10.2%) patients in the alirocumab treatment group and 6 (7.4%) patients in the placebo treatment group. None of the AEs were serious.

Treatment-emergent neurological disorders occurred in 7 (4.2%) patients in the alirocumab treatment group and 2 (2.5%) patients in the placebo treatment group. In the alirocumab group, the PTs were: hypoaesthesia in 4 (2.4%) patients, paresthesia in 2 (1.2%) and balance disorder in 1 (0.6%). None of the AEs were serious.

Treatment-emergent neurocognitive disorders occurred in 0 (0.0%) patients in the alirocumab treatment group and 1 (1.2%) patients in the placebo treatment group. The AE was not serious.

A total of 9 (5.4%) patients in the alirocumab treatment group and 0 (0.0%) patients in the placebo treatment group had 2 consecutive calculated LDL-C measurements less than 25 mg / dL. For patients with 2 consecutive calculated LDL-C measurements less than 25 mg / dL, TEAEs occurred in 3 (33.3%) patients on alirocumab treatment. The PTs were: influenza, influenza-like illness and nasopharyngitis. None of these AEs were serious, nor were they AESIs.

conclusion

The following conclusions can be drawn from this early review of the study data: 1) the study reached the primary efficacy endpoint with a statistically significant reduction in calculated LDL-C in alirocumab-treated patients; 2) this study also met all secondary key efficacy endpoints except the last endpoint (Apo A-1 at Week 12 in the IDT population (IDT analysis)); and 3) based on the data available at the time of this first-stage analysis, subcutaneous administration of alirocumab to patients with heterozygous familial hypercholesterolemia and an LDL-C> 70 mg / dL or LDL-C> 100 mg / dL, depending on the history of MI or stroke at baseline, it was generally safe and well tolerated.

Summary of pooled data from studies F H I and F H II

From the pooled times of studies FH I and FH II the following conclusions can be drawn: 1) self-administered alirocumab produced significantly greater reductions in LDL-C versus placebo after 24 weeks (mean MC difference 51.4-57.9%); 2) most patients (> 70%) reached their LDL-C goals at Week 24; 3) LDL-C reductions of 47.1-50.3% at Week 52 were achieved with alirocumab; 4) mean LDL-C levels of 1.7-1.9 mmol / L (65.9-74.3 mg / dL) at Week 52 were achieved with alirocumab; 5) approximately 50% of patients did not require upward titration until alirocumab 150 mg Q2W, suggesting that 75 mg Q2W may be sufficient for many patients; and 6) TEAEs occurred at a similar frequency in the alirocumab and placebo arms.

Specifically, the combined data from studies FH I and FH II show that alirocumab produced a significant reduction in LDL-C at Week 24 relative to placebo. The% change in mean MC (SE) from baseline at Week 24 was -48.8% for the alirocumab group (N = 488), compared to 7.1% for the placebo group ( N = 244). The mean difference of MC (SE) versus placebo was -55.8% (2.1) (P <0.0001). Additionally, only 42% of alirocumab patients required upward dose adjustment at Week 12 to the 150 mg Q2W dose.

The mean calculated LDL-C values of MC (SE) versus time for the ODYSSEY FH I and FH II studies are shown in Figure 8. The values indicated in the graph are the mean% change in MC from baseline. through Week 24 and Week 52. Figure 9 is a graph showing the mean calculated LDL-C values of MC (EE) versus time for the ODYSSEY FH I and FH II studies. The values indicated below the graph are the numbers of patients analyzed at the various moments in time.

Among patients who received double-blind treatment for at least 12 weeks, 176/311 (56.6%) in FH I and 97/158 (61.4%) in FH II had LDL-C levels <1 , 8 mmol / L at Week 8 and they were maintained with alirocumab 75 mg Q2W. LDL-C levels were stable over time in these patients (Figure 10). For patients in FH I who received dose escalation to 150 mg Q2W, mean LDL-C levels were 2.7 mmol / L (104.3 mg / dL) at Week 12 and 2.0 mmol / L (78 , 5 mg / dL) at Week 24. The corresponding values in FH II were 2.6 mmol / L (98.6 mg / dL) at Week 12 and 1.9 mmol / L (71.8 mg / dL ) in Week 24.

Subgroup analyzes of the primary efficacy endpoint showed a consistent reduction in LDL-C calculated across a range of baseline and demographic characteristics (Figure 11). The percentage reduction in LDL-C (alirocumab vs placebo) was 60.1% in men and 50.6% in women (pooled data for FH I and FH II), with a p value for interaction of 0.0267 . In individual studies, LDL-C reductions (versus placebo) were 62.6% for men and 51.9% for women in FH I, and 53.5% for men and 49.2% for women in FH II.

A summary of pooled interim safety data from Studies FH I and FH II is set out in Table 25A. All data were collected up to the patient's last visit at Week 52. The percentage of patients who had ASDs, severe AEs, and ASDs leading to treatment discontinuation were comparable between treatment groups in individual studies (Table 25B). A higher proportion of patients had injection site reactions in the alirocumab versus placebo groups in FH I (12.4% versus 11.0%) and FH II (11.4% versus 7.4%). The majority of injection site reactions were classified as mild in intensity. No injection site reaction led to the discontinuation of the study drug. None of the neurological or allergic events reported (Table 3) were serious. Pruritus was reported in two (0.6%) and three (1.8%) patients treated with alirocumab in FH I and II, respectively, and one patient treated with placebo in each study (0.6% and 1.2% , respectively). Few neurocognitive events were reported with alirocumab (2 [0.6%] in FH I, none in FH II) or placebo (2 [1.2%] in FH I, 1 [1.2%] in FH II; Table 3). In FH I and FH II, respectively, 85.8% and 91.6% of alirocumab-treated patients (87.7% and 90.1% placebo) received study treatment for> 76 weeks.

Table 25A: Interim Safety Analysis (Pooled Data from Studies FH I and FH II)

Table 25B: Final Safety Analysis (Pooled Data from Studies FH I and FH II)

Example 4: A randomized, double-blind, placebo-controlled, parallel-group study to evaluate the efficacy and safety of alirocumab in patients with heterozygous familial hypercholesterolaemia and LDL-C greater than or equal to 160 mg / dl with its modifying therapy. lipids

Introduction

This study included patients with heterozygous familial hypercholesterolaemia (heHF) with or without a history of documented MI or ischemic stroke.

The objective of the present study was to evaluate the efficacy and safety of alirocumab in patients with HeFH whose LDL-C level was greater than or equal to 160 mg / dL (4.14 mmol / L) with maximum tolerated statin therapy with or without TML. additional.

This specific study (Figure 6) was carried out to demonstrate in patients with heHF, with LDL-C greater than or equal to 160 mg / dL, that alirocumab 150 mg Q2W as complementary therapy to statin / - other TML causes a reduction in LDL-C statistically significant and clinically significant. This population with such a high level of LDL-C despite an optimized TML represents a higher risk group with a well-identified unmet medical need that can be treated by adding alirocumab to their LDL-C-lowering therapy.

Objectives of the study

The primary objective of the study was to demonstrate the reduction of LDL-C by alirocumab as add-on therapy to stable maximum tolerated daily statin therapy with or without another TML compared to placebo after 24 weeks of treatment in patients with heterozygous familial hypercholesterolaemia (HFhe ) and LDL-C greater than or equal to 160 mg / dL (4.14 mmol / L).

The secondary objectives were: 1) to evaluate the effect of alirocumab in comparison with placebo on LDL-C after 12 weeks of treatment; 2) evaluate the effect of alirocumab on other lipid parameters (ie Apo B, non-HDL C, total C, Lp (a), HDL-C, TG and Apo A-1 levels); 3) evaluate the long-term effect of alirocumab on LDL-C; 4) evaluate the safety and tolerability of alirocumab; 5) evaluate the development of antialirocumab antibodies.

Study design

This was a randomized, double-blind, placebo-controlled, parallel group, unbalanced (2: 1, alirocumab: placebo), multicenter, multinational study to evaluate the efficacy and safety of alirocumab in patients with heterozygous familial hypercholesterolaemia ( HFhe) and LDL-C greater than or equal to 160 mg / dL with or without your TML (i.e. stable maximum tolerated daily statin therapy / - other TML). Randomization was stratified according to previous history of myocardial infarction (MI) or ischemic stroke [Yes / No], and treatment with statins (atorvastatin 40 to 80 mg daily or rosuvastatin 20 to 40 mg daily versus simvastatin whatever the daily dose, atorvastatin less than 40 mg per day or rosuvastatin less than 20 mg per day). After randomization, patients received double-blind study treatment (or alirocumab or placebo) every 2 weeks for a period of 78 weeks, in addition to stable maximum tolerated daily statin therapy / - other TML.

After the completion of the 18-month double-blind treatment period, all patients who successfully completed the ODYSSEY High FH study had the opportunity to participate in an open-label extension study. Therefore, all patients will receive alirocumab at entry into the open-label extension study regardless of the study treatment received during the 18-month double-blind treatment period.

The study consisted of 3 periods: screening, double-blind treatment, and follow-up.

The screening period was up to 3 weeks in length that includes an intermediate visit during which the patient (or other designated person, such as spouse, relative, etc.) was trained to self-inject / inject with placebo for alirocumab. Eligibility assessments were performed to allow for randomization of patients in the study.

The double-blind treatment period was an 18-month randomized double-blind study treatment period. The first injection during the double-blind period was made at the site on the day of randomization (Week 0 [D1] - V3) and as soon as possible after the call to the IVRS / IWRS for study randomization. Subsequent injections were made by the patient (self-injection) or another designated person (such as spouse, relative, etc.) at a location preferred by the patient (home ...). Patients randomized to alirocumab received a 150 mg dose of IMP from randomization (V3) through Week 76 (i.e. Weeks 0, 2, 4, 6, 8 ... to 76).

The follow-up period (if applicable) was a period of 8 weeks after the end of DBTP for patients who did not consent to participate in the open-label extension study or prematurely discontinued study treatment.

Laboratory measurement of lipid parameters was performed by a central laboratory during the study.

Patients who achieved 2 consecutive calculated LDL-C levels <25 mg / dL (0.65 mmol / L) during the study were monitored and managed.

The statin and other TML (if applicable) must have been stable (including dose) during the first 24 weeks of DBTP, except in exceptional circumstances where primary concerns (including, but not limited to, TG alert reported by the central laboratory) guarantee such changes, at the discretion of the investigator. In Week 24 and beyond, the background TML was modified only under certain conditions as described below.

Patients must have been on a stable diet (NCEP-ATPIII TLC diet or equivalent) for the entire duration of the study since screening, as described above in Example 2 (see Table 1). The dietitian or appropriately trained site staff reviewed the patient's diet at the screening visit and periodically throughout the study.

Study duration included a screening period of up to 3 weeks, a 78-week DBTP for efficacy and safety assessment, and an 8-week post-treatment follow-up period after the last DBTP visit for patients who did not give the consent to participate in the open-label extension study or if they prematurely discontinued study treatment. Thus, the maximum study duration per patient was approximately 89 weeks (ie, 20 months) (up to 3 weeks selection 78 weeks double-blind treatment 8 weeks follow-up). End of study per patient was the last planned protocol visit or resolution / stabilization of all SAEs, and AESI, whichever comes first.

Patient selection

The inclusion criteria were: 1) patients with heterozygous familial hypercholesterolaemia (heFH) * who were not adequately controlled with a maximum tolerated daily dose of statin, ** with or without other lipid-modifying therapy (TML) at stable doses before screening visit (Week -3).

The diagnosis of heFH must have been made either by genotyping or by clinical criteria. For non-genotyped patients, the clinical diagnosis may have been based on either the Simon Broome criteria with a criterion for defined HF or the WHO / Dutch Lipid Network criteria with a score> 8 points. See the criteria described above in Example 2.

Definition of maximum tolerated dose: any of the following were acceptable): 1) rosuvastatin 20 mg or 40 mg daily; 2) atorvastatin 40 mg or 80 mg daily; 3) simvastatin 80 mg daily (if already taking the dose for> 1 year - see exclusion criterion E 06); or 4) patients unable to be on any of the above statin doses must have been treated with the daily dose of atorvastatin, rosuvastatin, or simvastatin that was deemed appropriate for the patient based on the investigator's discretion or concerns. Some examples of acceptable reasons for a patient to take a lower dose of statins include, but are not limited to: adverse effects at any dose, advanced age, low body mass index, regional practices, local drug label, concurrent medications such as intolerance to glucose / impaired fasting glucose.

Patients who met all the above inclusion criteria were selected by the following exclusion criteria, which are classified and numbered in the following 3 subsections: exclusion criteria related to study methodology, exclusion criteria related to background therapies, and exclusion criteria related to alirocumab.

The exclusion criteria related to the study methodology were: 1) patient without a diagnosis of HFhe made either by genotyping or clinical criteria; 2) LDL-C <160 mg / dL (<4.14 mmol / L) at the screening visit (Week -3); 3) are not on a stable dose of TML (including statin) for at least 4 weeks and / or fenofibrate for at least 6 weeks as applicable, prior to the screening visit (Week -3) or from screening to randomization ; 4) he was currently taking a statin other than simvastatin, atorvastatin, or rosuvastatin; 5) simvastatin, atorvastatin, or rosuvastatin is not taken daily or is not taken at a recorded dose; 6) daily doses higher than atorvastatin 80 mg, rosuvastatin 40 mg or simvastatin 40 mg (except for patients with simvastatin 80 mg for more than one year, who are eligible); 7) use of fibrates, other than fenofibrate within 6 weeks of the screening visit (Week -3) or between screening and randomization visits; 8) use of nutraceutical products or over-the-counter therapies that may affect lipids that have not been in a stable quantity / dose for at least 4 weeks prior to the screening visit (Week -3) or between screening visits and of randomization; 9) use of red yeast rice products within 4 weeks of the screening visit (Week -3) or between screening and randomization visits; 10) patient who has received plasmapheresis treatment within 2 months prior to screening visit (Week -3), or plans to receive it during the study; 11) Recent MI (within 3 months before screening visit [Week -3] or between screening and randomization visits), unstable angina leading to hospitalization, percutaneous coronary intervention (PCI), graft surgery of coronary artery bypass (CABG), uncontrolled cardiac arrhythmia, cerebrovascular accident, transient ischemic attack (TIA), carotid revascularization, endovascular procedure or surgical intervention for peripheral vascular disease; 12) plan to undergo scheduled PCI, CABG, carotid or peripheral revascularization during the study; 13) systolic blood pressure> 160mmHg or diastolic blood pressure> 100mmHg at screening visit or randomization visit; 14) New York Heart Association (NYHA) history of class III or IV heart failure within the last 12 months; 15) known history of hemorrhagic stroke; 16) age <18 years or legal age of majority at the screening visit (Week -3), whichever is greater; 17) patients not previously informed about a low cholesterol diet before the screening visit (Week -3); 18) newly diagnosed diabetes (within 3 calendar months prior to randomization visit [Week 0]) or poorly controlled (HbA1c> 9% at screening visit [Week -3]); 19) Presence of any clinically significant uncontrolled endocrine disease known to influence serum lipids or lipoproteins. Note: Patients on thyroid replacement therapy may be enrolled if the dosage has been stable for at least 12 weeks prior to screening and between screening and randomization visits, and the TSH level is within the normal central laboratory range in the screening visit; 20) history of bariatric surgery within 12 months prior to screening visit (Week -3); 21) unstable weight defined by a variation> 5 kg within 2 months before the screening visit (Week -3); 22) known history of homozygous HF; 23) known history of PCSK9 loss of function (ie, genetic mutation or sequence variation); 24) use of systemic corticosteroids, unless used as replacement therapy for pituitary-adrenal disease with a stable regimen for at least 6 weeks prior to the randomization visit (Week 0). Note: Topical, intra-articular, nasal, inhaled and ophthalmic steroid therapies are not considered "systemic" and are allowed; 25) use of continuous estrogen or testosterone hormone replacement therapy unless the regimen has been stable in the last 6 weeks prior to the screening visit (Week -2) and with no plans to change the regimen during the study; 26) History of cancer within the last 5 years, except adequately treated basal cell skin cancer, squamous cell skin cancer, or cervical cancer in situ; 27) known history of a positive HIV test; 28) patient who has taken any investigational drug other than alirocumab training placebo kits within 1 month or 5 half-lives, whichever is longer; 29) patient who has previously been treated with at least one dose of alirocumab or any other monoclonal anti-PCSK9 antibody in other clinical trials; 30) patient who withdraws consent during the selection period (patient who does not wish to continue or does not return); 31) conditions / situations such as any clinically significant abnormalities identified at the time of selection that, in the discretion of the investigator or any collaborating investigator, would prevent the safe termination of the study or restrict the evaluation of endpoints; for example, major systemic diseases, patients with a short life expectancy considered by the investigator or any collaborating investigator as inappropriate for this study for any reason, for example: a) considered unable to meet specific protocol requirements, such as scheduled visits; b) considered unable to administer or tolerate long-term injections according to the patient or the investigator; c) researcher or any collaborating researcher, pharmacist, study coordinator, other study personnel or relatives directly involved in carrying out the protocol, etc .; d) presence of any other current or anticipated condition (for example, geographic or social ...) that the investigator feels would restrict or limit the patient's participation during the duration of the study; 32) laboratory data during the selection period (which does not include randomization laboratories at Week 0): a) positive test for hepatitis B surface antigen or hepatitis C antibody; b) positive for beta-hCG in serum or urine pregnancy test (including Week 0) in women of childbearing potential; c) triglycerides> 400 mg / dL (> 4.52 mmol / L) (1 repeat laboratory is allowed); d) eGFR <30 mL / min / 1.73 m2 according to the equation of the MDRD Study of 4 variables (calculated by the central laboratory); e) ALT or AST> 3 x ULN (1 repeat laboratory allowed); f) CPK> 3 x ULN (1 repeat laboratory is allowed); g) TSH <lower limit of normal (LLN) or> upper limit of normal (ULN) (1 repeat laboratory allowed).

Exclusion criteria related to background therapies were: 1) all contraindications to background therapies or warnings / precautions for use (where appropriate) as presented in the respective national product labeling.

Exclusion criteria related to alirocumab were: 1) known hypersensitivity to monoclonal antibody or any component of the drug; 2) pregnant or lactating women; 3) women of childbearing potential who are not protected by highly effective method (s) of birth control (as defined in the informed consent document and / or in an attachment to the local protocol) and / or who are unwilling or are unable to take a pregnancy test. Note: Women of childbearing potential must have had a confirmed negative pregnancy test at screening and randomization visits. They must have used effective contraception for the entire duration of study treatment, and for 10 weeks after the last IMP intake, and agree to repeat the urine pregnancy test at designated visits. The contraceptive methods applied had to meet the criteria of a highly effective method of birth control according to the "Guidance note on non-clinical safety studies for human clinical trials and the marketing authorization of pharmaceutical products (CPMP / ICH / 286/95) ". Postmenopausal women must have been amenorrheic for at least 12 months.

Treatments under study

The sterile drug alirocumab was supplied at a concentration of 150 mg / mL in histidine, pH 6.0, polysorbate 20 and sucrose. The drug was delivered as a 1 mL volume in an autoinjector.

Sterile placebo for alirocumab was prepared in the same formulation as alirocumab without the addition of protein as 1 mL volume in an autoinjector.

During the double-blind treatment period, alirocumab or placebo was administered subcutaneously every 2 weeks, starting at Week 0 continuing until the last injection (Week 76) 2 weeks before the end of the double-blind treatment period. If the injection was scheduled to take place on the same date as the study visit, then the IMP must have been administered after the blood sampling had been completed.

The IMP should ideally have been administered subcutaneously every 2 weeks at approximately the same time of day; however, it was acceptable to have a window period of ± 3 days. Time of day was based on patient preference.

The following drug classes were identified as non-investigational specialty pharmaceuticals (NIMPs) because the medication was either background therapy or possible rescue medication: statins (rosuvastatin, atorvastatin, simvastatin); cholesterol absorption inhibitors (ezetimibe); bile acid binding sequestrants (such as cholestyramine, colestipol, colesevelam); nicotinic acid; fenofibrate; omega-3 fatty acids (> 1000 mg per day).

Patients who reached 2 consecutive calculated LDL-C <25 mg / dL (0.65 mmol / L) were monitored.

Additional sample (s) were taken from patients who had a titer of 240 or higher for antialirocumab antibody at the follow-up visit, 6 to 12 months after the last dose, and approximately every 3 years thereafter. 6 months until the title went back below 240.

Patients were randomized to receive either placebo or alirocumab during the double-blind study treatment period using a 1: 2 ratio, with permuted block randomization. Randomization was stratified according to previous history of myocardial infarction (MI) or ischemic stroke [Yes / No], and statin treatment (atorvastatin 40 to 80 mg daily or rosuvastatin 20 to 40 mg daily versus simvastatin whatever the daily dose, atorvastatin less than 40 mg per day or rosuvastatin less than 20 mg per day).

Concurrent medication was any treatment received by the patient concurrent with the study (up to the follow-up visit). Concurrent medications had to be kept to a minimum during the study. However, if these were considered necessary for the well-being of the patient and were unlikely to interfere with IMP, they could be administered at the investigator's discretion, at a stable dose (where possible). In addition to the specific information regarding concurrent medications provided in this section, any other concurrent medication (s) will be permitted. If the patient had an LDL-C> or equal to 160 mg / dL (4.14 mmol / L) at the screening visit (Week -3) and was treated with a statin alone, that is, without additional TML, the Investigator had to report the patient's reason for not being on a second TML. For background TML, which includes statins, sites must have followed the national product label for safety monitoring and patient management.

Nutraceuticals or OTC therapies that could affect lipids were allowed only if they had been used at a stable dose for at least 4 weeks prior to the screening visit, during the screening period, and were maintained for the first 24 weeks of the double-blind treatment period. After the Week 24 visit, modification to these nutraceuticals or OTC therapies was allowed, but generally should have been avoided. Examples of such nutraceutical products or OTC therapies included omega-3 fatty acids at doses <1000 mg, plant stanols such as those found in Benecol, flax seed oil, and psyllium.

Patients must have been on stable maximum tolerated daily recorded statin doses with another TML for at least 4 weeks (6 weeks for fenofibrate) prior to the screening visit. During the study, patients should have remained on these stable maximum tolerated daily recorded statin doses with another TML. The lipid profile values obtained after randomization were blinded. However, the sites were aware of the triglyceride alert as well as the salvage threshold of the LDL-C value to make decisions about the patient's background TML. From the screening visit (Week -3) to Week 24 of the double-blind treatment period, the background TML should not have been changed. No dose adjustment, interruption, or initiation of other statins or other TMLs should have occurred during this time, except for exceptional circumstances where major concerns (including, but not limited to, central laboratory-reported triglyceride alert) guaranteed such changes, at the discretion of the researcher.

For a triglyceride alert (TG> 500 mg / dL (5.65 mmol / L)) that was confirmed by retest, the investigator would conduct investigations, manage the patient, and modify the background TML at their discretion. doctor.

For an LDL-C salvage notification at the Week 24 visit and beyond, that is, an increase in LDL-C> 25% compared to the LDL-C randomization visit on two consecutive occasions, the investigator must have guaranteed that there was no reasonable explanation for insufficient LDL-C control (such as an alternative medical cause such as the use of corticosteroids, etc.) and in particular that: adherence to the diet was appropriate; background TML compliance was appropriate; and the study treatment was administered as planned. If any of the above could have reasonably explained the insufficient LDL-C control, the investigator should have taken appropriate action, i.e., emphasizing the absolute need to adhere to treatment, if a specific interview with a qualified nutrition professional needs to be arranged and emphasizing the absolute need to adhere to the diet, and performed a blinded LDL-C assessment within 1 to 2 months. If none of the reasons listed above could be found, or if the appropriate action failed to lower LDL-C below the alert value, rescue medication may have been introduced. The efficacy of any such changes was made based on the absence of LDL-C salvage notification sent in the blinded lipid test at the next scheduled lab draw.

If the reason for LDL-C above the threshold value could not be found, or if the appropriate action failed to reduce LDL-C below the threshold value, rescue medication may have been introduced. The efficacy of any such changes would be made based on the lack of rescue threshold of the blinded lipid test at the next routinely scheduled lab draw. The patients per protocol already received a maximum tolerated dose of statin, so upward adjustment of the statin dose or change was not an option. For additional LDL-C reduction, the investigator may have considered: a cholesterol absorption inhibitor (ezetimibe), or a bile acid binding sequestrant (the resins cholestyramine and colestipol, or colesevelam, a nonabsorbable polymer) . The following lipid modifying agents may also have been considered: fibrate (Note: Caution should be exercised when combining fibrates with other cholesterol-lowering medications such as statins due to the risk of myopathy. When a fibrate is combined with a statin, fenofibrate is the fibrate of choice because it does not affect statin glucuronidation); the only fibrate allowed by protocol was fenofibrate; Nicotinic acid (niacin) (Note: Niacin increases blood glucose, but has been shown to be effective in modifying lipid disorders in people with diabetes if glucose control is maintained).

In summary, the background TML should not have changed from selection to follow-up visit. However, until Week 24, if a confirmed TG alert was reached or if there was overwhelming clinical concern (at the discretion of the investigator), then background TML modification was allowed. From Week 24 onwards, if a confirmed TG alert was reached, or if a salvage threshold for LDL-C was obtained (and there is no other reasonable explanation), or if there was an overwhelming clinical concern (at the discretion of the investigator) , then a modification of the background TML was allowed.

Women of childbearing potential were required to take effective contraception throughout study treatment and for 10 weeks after the last IMP injection (ie, follow-up visit).

Concurrent medications prohibited from the initial screening visit to the follow-up visit included the following: statins other than simvastatin, atorvastatin, and rosuvastatin; fibrates, other than fenofibrate; and red yeast rice products.

Study endpoints

The primary efficacy endpoint was the percent change in LDL-C calculated from baseline to Week 24, which was defined as: 100 x (LDL-C value calculated at Week 24 - LDL-C value calculated at baseline) / LDL-C value calculated at baseline. The LDL-C value calculated at baseline was the last LDL-C level obtained before the first double-blind IMP injection. The calculated LDL-C at Week 24 was the LDL-C level obtained within the Week 24 analysis window and during the primary efficacy period. The primary efficacy period was defined as the time from the first double-blind IMP injection to 21 days after the last double-blind IMP injection or to the upper limit of the Week 24 analysis window, whichever is. First. All calculated LDL-C values (scheduled or unscheduled, fasting or non-fasting) can be used to provide a value for the primary efficacy endpoint if appropriate as defined above.

The key secondary efficacy endpoints were: 1) The percent change in LDL-C calculated from baseline to Week 12: similar definition and rules as for the primary efficacy endpoint, except that C- LDL calculated at Week 12 was the level of LDL-C obtained within the Week 12 analysis window and during the 12 week efficacy period. The 12-week efficacy period was defined as the time from the first double-blind IMP injection to contact with the IVRS re-supply at Visit 6 or up to 21 days after the last double-blind IMP injection, whichever comes first. The blood sample collected on the day of contact with the IVRS re-supply of Visit 6 was considered as before the assessment; 2) the percentage change in Apo B from baseline to Week 24. Same definition and rules as for primary endpoint; 3) percent change in non-HDL C from baseline to Week 24. Same definition and rules as for primary endpoint; 4) the percent change in total C from baseline to Week 24. Same definition and rules as for primary endpoint; 5) percentage change in Apo B from baseline to Week 12. Same definition and rules as for percent change in LDL-C calculated from baseline to Week 12; 6) the percent change in non-HDL-C from baseline to Week 12. Same definition and rules as for percent change in LDL-C calculated from baseline to Week 12; 7) the percent change in total C from baseline to Week 12. Same definition and rules as for percent change in LDL-C calculated from baseline to Week 12; 8) the percent change in LDL-C calculated from baseline through Week 52. Definitions and rules were similar to those used for the primary endpoint substituting Week 24 for Week 52. The 52-week efficacy period was defined as the time from the first double-blind IMP to 21 days after the last injection double-blind IMP, or up to the upper limit of the Week 52 analysis window, whichever comes first; 9) the proportion of patients who reach the goal of LDL-C at Week 24, that is, LDL-C <70 mg / dL (1.81 mmol / L) in case of previous CVD or <100 mg / dL ( 2.59 mmol / L) for patients without CVD, it was defined as: (number of patients whose calculated LDL-C value at Week 24 reached the LDL-C target / number of patients in the mTDI population) * 100, using the definition and rules used for the primary endpoint; 10) the percentage change in Lp (a) from baseline to Week 24. Same definition and rules as for primary endpoint; 11) the percent change in HDL-C from baseline to Week 24. Same definition and rules as for primary endpoint; 12) percent change in HDL-C from baseline to Week 12. Same definition and rules as for percent change in LDL-C calculated from baseline to Week 12; 13) the percent change in Lp (a) from baseline to Week 12. Same definition and rules as for percent change in LDL-C calculated from baseline to Week 12; 14) percent change in fasting TG from baseline to Week 24. Same definition and rules as for primary endpoint; 15) percentage change in fasting TG from baseline to Week 12. Same definition and rules as for percent change in LDL-C calculated from baseline to Week 12; 16) the percentage change in Apo A-1 from baseline to Week 24. Same definition and rules as for primary endpoint; 17) the percentage change in Apo A-1 from baseline to Week 12. Same definition and rules as for percent change in LDL-C calculated from baseline to Week 12.

Other secondary efficacy endpoints were: 1) percent change in LDL-C calculated from baseline to Week 78: Definitions and rules were similar to those used for the primary endpoint substituting Week 24 for Week 78; 2) the proportion of patients who reach the LDL-C goal at Weeks 12, 52, and 78, that is, LDL-C <70 mg / dL (1.81 mmol / L) in case of previous CVD or <100 mg / dL (2.59 mmol / L) for patients without previous CVD; 3) the proportion of patients achieving LDL C <100 mg / dL (2.59 mmol / L) at Week 24; 4) the proportion of patients achieving LDL-C <100 mg / dL (2.59 mmol / L) at Week 12; 5) the proportion of patients achieving LDL-C <70 mg / dL (1.81 mmol / L) at Week 24; 6) the proportion of patients achieving LDL-C <70 mg / dL (1.81 mmol / L) at Week 12; 7) the absolute change in calculated LDL-C (mg / dL and mmol / L) from baseline through Weeks 12, 24, 52, and 78; 8) the percentage change in Apo B, non-HDL C, total C, Lp (a), HDL-C, fasting TG, and Apo A-1 from baseline to Week 52 and 78; 9) the change in Apo B / Apo A-1 ratio from baseline to Weeks 12, 24, 52 and 78; 10) the proportion of patients with Apo B <80 mg / dL (0.8 g / L) at Weeks 12, 24, 52 and 78; 11) the proportion of patients with non-HDL C <100 mg / dL at Weeks 12, 24, 52 and 78; 12) the proportion of patients with calculated LDL-C <70 mg / dL (1.81 mmol / L) and / or > 50% reduction in calculated L-C-L (if calculated LDL-C > 70 mg / dL [ 1.81 mmol / L]) at Weeks 12, 24, 52 and 78.

Total C, HDL-C, TG, Apo B, Apo A-1, and Lp (a) were directly measured by the central laboratory. LDL-C was calculated using Friedewald's formula at all visits (except Week -1 and the follow-up visit). If TG values exceeded 400 mg / dL (4.52 mmol / L), then the central laboratory reflexively measured (using the beta quantification method) LDL-C instead of calculating it. Non-HDL C was calculated by subtracting HDL-C from total C. The Apo B / Apo A-1 ratio was calculated.

Clinical laboratory data consisted of urinalysis, hematology (number of red blood cells, hemoglobin, red blood cell distribution width (RDW), reticulocyte count, hematocrit, platelets, number of leukocytes with leukocyte formula), standard chemical analysis ( glucose, sodium, potassium, chloride, bicarbonate, calcium, phosphorus, urea nitrogen, creatinine, uric acid, total protein, LDH, albumin, Y- glutamyltransferase [yGT]), hepatitis C antibody, liver panel (ALT, AST, Al P and total bilirubin) and CPK.

The vital signs included: heart rate, systolic and diastolic blood pressure while sitting.

Other endpoints included: anti-alirocumab antibody evaluations, hs-CRP, HbA1c, EQ-5D Questionnaire, and pharmacogenetic samples.

Anti-alirocumab antibodies included antibody status (positive / negative) and antibody titers. Serum samples were collected periodically throughout the study for the determination of antialirocumab antibodies. The first sample scheduled at the randomization visit was obtained prior to IMP injection (predose). Additional sample (s) were taken from patients who had a titer of 240 or higher for anti-alirocumab antibodies at the follow-up visit 6 to 12 months after the last dose, and approximately every 3 to 6 months until the titer dropped below 240. Samples for anti-alirocumab antibodies were tested using a non-quantitative validated titer-based double capture immunoassay. It involved an initial screening, a confirmatory assay based on drug specificity, and a measurement of the anti-alirocumab antibody titer in the sample. The lower limit of detection was approximately 1.5 ng / mL. Samples that tested positive for ADA were evaluated for neutralizing antibodies using a competitive, non-quantitative, validated ligand binding assay. The lower limit of detection based on a monoclonal positive control neutralizing antibody was 390 ng / mL.

The percent change in hs-CRP from baseline through Week 24, 52, and 78.

The absolute change in HbA1c (%) from baseline through Week 24, 52, and 78.

EQ-5D is a standardized measure of health status developed by the EuroQol Group to provide a simple generic measure of health for clinical and economic assessment. EQ-5D, as a health-related quality of life measure, defines health in terms of 5 dimensions: mobility, self-care, usual activities, pain / discomfort, anxiety / depression. Each dimension can adopt one of three responses (3 ordinal levels of severity): 'no problem' (1), "some problems" (2), "serious problems" (3). General health status was defined as a 5-digit number. The health states defined by the 5-dimensional classification can be converted into corresponding index scores that quantify health status, where 0 represents 'death' and 1 represents 'perfect health'.

Study procedures

For all visits after Day 1 / Week 0 (randomization visit), a time period of a certain number of days was allowed. The window period for visits at Weeks 12 and 24 was ± 3 days, at Week 52 and 78 it was ± 5 days, and for all other site visits it was ± 7 days during the double-blind treatment period. , and follow-up period. A window period of 3 days was allowed for the randomization visit (Day 1 / Week 0) and ± 7 days for the injection training visit at screening (Week -1).

Blood sampling for determination of lipid parameters (i.e. total C, LDL-C, HDL-C, TG, non-HDL C, Apo B, Apo A-1, Apo B / Apo A-1 ratio, Lp [a]) should have been performed in the morning, fasting (ie overnight, at least 10-12 hours of fasting and abstaining from smoking) for all site visits throughout the study. It was recommended not to consume alcohol within 48 hours and not to do intense physical exercise within 24 hours prior to blood sampling. Note: If the patient was not fasting, the blood sample was not collected and the patient was given a new appointment the day after (or as soon as possible on that date) with instructions to be fasting (see conditions above).

Only patients who met the inclusion criteria were selected. The screening period took place up to 3 weeks or 21 days (and as soon as possible after receipt of the laboratory eligibility criteria) prior to the randomization visit / Day 1. The first screening visit (Week -3 ) took place 21 to 8 days before the randomization visit. If another designated person was planned to administer the injections to the patient during the study, then this person must have been present at the injection training visit (Week -1).

The following visits were scheduled: Selection visit (Visit 1 / Week -3 / Day -21 to -8); Selection (Visit 2 / Week -1 / Day -7 ± 7); Randomization visit (Visit 3 / Week 0 / Day 1 3); Visit 4 / Week 4 / Day 29 ± 7); Visit 5 / Week 8 / Day 57 ± 7); Visit 6 / Week 12 / Day 85 ± 3; Visit 7 / Week 16 / Day 113 ± 7): Visit 8 / Week 24 / Day 169 ± 3; Visit 9 / Week 36 / Day 253 ± 7; Visit 10 / Week 52 / Month 12 / Day 365 ± 5; Visit 11 / Week 64 / Day 449 ± 7; Visit 12 / Week 78 / Month 18 / Day 547 ± 5 (end of treatment visit); and Visit 13 / Week 86 / Day 603 ± 7 (follow-up visit).

Security

The safety events monitored were the occurrence of treatment emergent adverse events (TEAEs) reported by the patient or observed by the investigator, serious adverse events (SAEs), TEAEs leading to treatment discontinuation, AEs of special concern (local reactions injection site, allergic events, selected neurological events and cardiovascular events with validation result), manifestation of PCSA (potentially clinically significant abnormalities) in laboratory parameters, specific tests for diabetes or impaired glucose control, and patients with 2 C- Consecutive LDL <25 mg / dL.

Statistical methods

Determination of sample size:

A total sample size of 45 patients (30 on alirocumab and 15 on placebo) had 95% power to detect a difference in mean percent change in LDL-C of 30% with a two-sided significance level of 0.05 and assuming a common standard deviation of 25%, and all of these 45 patients having an assessable primary endpoint. A final total sample size of 105 patients with a 2: 1 randomization ratio (alirocumab 70: placebo 35) has been selected to provide at least 50 patients exposed to alirocumab for 12 months in the first stage analysis and assuming a rate of dropouts of 10% compared to the first period of 3 months and a dropout rate of 20% compared to the first period of 3-12 months.

Exact time of analysis:

The first-stage analyzes include efficacy endpoints through Week 52 (final efficacy analysis) and interim safety analysis, which was performed on all safety data up to the common study deadline (Weekly visit 52 of the last patient). Analysis of lipid data beyond Week 52 was descriptive. The results of the first stage analyzes are presented in this document.

The second (final) stage analysis will be performed at the end of the study and will consist of the final analysis of the efficacy endpoints up to Week 78 and the final safety analyzes.

Analysis populations:

The primary efficacy analysis population was the intention-to-treat (IDT) population, defined as all randomized patients who had a primary evaluable efficacy endpoint, that is, those with a calculated LDL-C value at the level baseline available, and at least one calculated LDL-C value available within one of the analysis windows through Week 24 (including all calculated LDL-C during treatment and without treatment).

The secondary efficacy analysis population was the modified intention-to-treat (mTDI) population, defined as all randomized patients who took at least one dose or part of a dose of the double-blind investigational pharmaceutical specialty (IMP) and who had a calculated LDL-C value available at baseline and at least one within one of the analysis windows through Week 24 during the efficacy treatment period. The efficacy treatment period was defined as the time from the first double-blind IMP administration to 21 days after the last double-blind injection.

The safety population included all randomized patients who received at least one dose or part of a dose of the double-blind IMP.

Efficacy analysis:

Primary efficacy endpoint analyzes were performed using an IDT approach (based on the IDT population defined above), including all lipid data, regardless of whether the patient was continuing therapy or not. This corresponds to IDT estimates, defined for primary and secondary key endpoints. In addition, analyzes were also performed using an during treatment approach (based on the mTDI population defined above), including lipid data collected during the efficacy treatment period. This corresponds to estimates during treatment of key secondary endpoints.

The IDT approach analyzed all patients, regardless of their compliance with treatment; assessed the benefit of the treatment strategy and reflected the effect as far as possible in a patient population. The during treatment approach looked at the effect of treatment, restricted to the period during which patients actually received the treatment. It evaluated the benefit that a treatment would achieve in patients who comply with the treatment until the moment of time considered.

Efficacy analyzes were performed by treatment as randomized.

All measurements, scheduled or unscheduled, fasting or non-fasting, were assigned to analysis windows to provide an evaluation for time points from Week 4 to Week 78.

Regarding the primary efficacy analysis (TDI approach), the percentage change in LDL-C calculated from baseline to Week 24 was analyzed using a mixed effects model with repeated measures (MMRM) approach. All available post-baseline data from the Week 4 through Week 52 analysis windows were used and the MMRM accounted for missing data. The model included the fixed categorical effects of treatment group (placebo vs alirocumab), randomization strata (according to IVRS), moment of time (Week 4 to Week 52), interaction of treatment by moment of time and interaction of strata by moment time, as well as the continuous fixed covariates of baseline LDL-C value and baseline value interaction by time. This model provided estimates of least squares means (MC means) adjusted to baseline at Week 24 for both treatment groups with their corresponding 95% confidence interval. To compare the alirocumab to the placebo group, an appropriate instruction was used to test for differences in these estimates at the 5% alpha level.

A hierarchical procedure has been defined to test secondary key endpoints while controlling for multiplicity (using the previous order of secondary key endpoints). The first key secondary endpoint was the percent change in LDL-C calculated from baseline to Week 24 using an on-treatment approach.

Continuous secondary variables anticipated to have a normal distribution (ie, lipids other than TGs and Lp (a)) were analyzed using the same MMRM model as for the primary endpoint. Continuous endpoints that were anticipated to have a non-normal distribution (i.e. TGs and Lp (a)) were analyzed. using multiple imputation approach for missing value manipulation followed by robust regression model with endpoint of interest as response variable using M estimation (using SAS ROBUSTREG procedure) with treatment group, randomization strata (according to IVRS) and value (s) at the corresponding baseline level (s) as effects to compare treatment effects. The pooled estimate for the mean in both treatment groups, as well as the differences of these estimates, with their corresponding SEs, 95% CI, and p-value (using the SAS MIANALYZE procedure) were provided.

Binary secondary efficacy endpoints were analyzed using the multiple imputation approach for manipulating missing values followed by stratified logistic regression with treatment group as main effect and corresponding baseline value (s) as covariate. , stratified by randomization factors (according to IVRS). Pooled estimates of odds ratio vs. placebo, 95% CI, and p-value (using SAS MIANALYZE procedure) were provided.

Security analysis:

The safety analyzes were descriptive, carried out in the safety population according to the treatment actually received. Safety analyzes focused on the TEAE period defined as the time from the first double-blind dose to 70 days after the last double-blind injection. TEAEs that developed, worsened, or became severe or PCSA occurring after patient inclusion in the open-label extension study (LTS13643) were not considered in the TEAE period. The TEAE period was truncated at the common study deadline.

Results

Study patients

Patient count

Of the 107 randomized patients (72 and 35 patients in the alirocumab and placebo groups, respectively), one patient in the alirocumab group had no baseline calculated LDL-C value and therefore was not included in the IDT and TMDI populations.

Table 26 - Analysis populations

Placebo Alirocumab 150 Q2W All

Randomized population 35 (100%) 72 (100%) 107 (100%)

Efficacy populations

Intention to Treat (IDT) 35 (100%) 71 (98.6%) 106 (99.1%)

Modified intention to treat (mTDI) 35 (100%) 71 (98.6%) 106 (99.1%)

Safety population 35 72 107

Note: Patients in the safety population are tabulated based on the treatment actually received (as treated).

For the other populations, patients are tabulated according to their randomized treatment.

Study layout

Study readiness, exposure and safety analysis were assessed using all data up to the common study cutoff date (defined as the date of the last patient's Week 52 visit). Therefore, this first stage analysis includes efficacy data up to Week 52 and safety data beyond Week 52 and up to Week 78 or follow-up visit for some patients. The patient arrangement is shown in Figure 12.

In this study, one site with 7 randomized patients and a second site with 6 randomized patients with severe GCP non-compliance were identified, and the sites were closed. For the first closed site, one of the key findings was related to IMP injections, which were reported by some patients they had received while the corresponding kits were discovered in the fridge. The reporting of these injections was corrected in the database but other problems about the injections could not be excluded. For the second site, constant concerns with study conduct and associated study-related documentation during routine monitoring.

Among these 13 patients, one was still ongoing at the deadline, one discontinued due to an adverse event, one patient moved, 3 patients discontinued due to poor protocol compliance, and 7 patients discontinued due to the decision to close the site.

There were a total of 10 (9.3%) randomized patients who completed the 78-week double-blind study treatment period and 76 (71.0%) randomized patients with ongoing treatment at the time of the analysis cut-off date. the first stage. Double-blind IMP was discontinued prematurely before Week 78 for 6 (17.1%) patients in the placebo group and 15 (20.8%) patients in the alirocumab group. All of these patients actually discontinued prematurely before Week 52. The main reasons for discontinuation of study treatment were 'other reasons', poor compliance, and adverse events. These 'other reasons' included the 7 patients who discontinued due to the site closure decision as mentioned above, 1 patient withdrawal not otherwise specified, 1 patient withdrawal due to independently obtained cholesterol results, and 1 patient he moved.

In this first-stage analysis, final results are available for primary efficacy endpoints at Week 24 and secondary key efficacy endpoints were assessed at Week 12, Week 24, and Week 52. The table below provides the availability of LDL-C over time. At Week 24, the primary efficacy endpoint was available for 33 (94.3%) in the placebo group and 63 (88.7%) in the alirocumab group.

Table 27 - Availability of LDL-C calculated over time - IDT population

Placebo Alirocumab 150 Q2W

(N = 35) (N = 71)

LDL-C Value during post value Value during post value Value calculated treatment missing treatment treatment missing treatment

Week 4 31 (88.6%) 0 4 (11.4%) 67 (94.4%) 0 4 (5.6%)

Week 8 34 (97.1%) 0 1 (2.9%) 66 (93.0%) 0 5 (7.0%)

Week 12 33 (94.3%) 0 2 (5.7%) 68 (95.8%) 0 3 (4.2%)

Week 16 28 (80.0%) 0 7 (20.0%) 66 (93.0%) 0 5 (7.0%)

Week 24 33 (94.3%) 0 2 (5.7%) 62 (87.3%) 1 (1.4%) 8 (11.3%)

Week 36 30 (85.7%) 1 (2.9%) 4 (11.4%) 60 (84.5%) 3 (4.2%) 8 (11.3%)

Week 52 27 (77.1%) 0 8 (22.9%) 52 (73.2%) 2 (2.8%) 17 (23.9%)

The primary endpoint was missing for 10 patients at Week 24 (2 and 8 patients in the placebo and alirocumab groups, respectively). At the Week 24 visit (as monitored by CRF), the reasons for loss were as follows: 3 samples were not taken due to premature study interruption; 3 samples were taken outside the analysis time window; 2 samples were not made because the visit was not made in Week 24; and 2 samples available, but the measurement could not be made (lipemia, insufficient quantity, TGs> 400 mg / dL [> 4.52 mmol / L], loss of sample, etc.).

The higher number of missing data in Week 52 is primarily due to the decision to shut down the two sites due to severe GCP breach.

The LDL-C endpoint was missing at Week 52 for 25 of the 106 patients. The reasons for missing results were as follows: 17 samples were not taken due to discontinuation of the study prematurely including 11 patients from the two closed sites; 3 samples were taken outside the analysis time window; 1 sample was not done because Week 52 was not done; 1 sample missing while on the Week 52 visit; and 3 samples available but the measurement could not be made (TGs> 400 mg / dL [> 4.52 mmol / L] and hemolysis).

Demographic and baseline characteristics

Summary of the characteristics of the population:

107 HFhe patients diagnosed by genotyping (17.8%) and WHO / Dutch Lipid Network criteria (score> 8 points) or Simon Broome criteria for defined HF (82.2%) were randomized 2: 1 with respect to alirocumab (150 mg Q2W) or placebo.

Demographic characteristics, disease characteristics, and lipid parameters at baseline were generally similar in the alirocumab group compared to the placebo group: heFH diagnosis by genotyping in the alirocumab group (19,4 %) versus placebo (14.3%); HeFH diagnosis through clinical criteria in the group with alirocumab (80.6%) compared to placebo (85.7%); a mean age (SD) in the alirocumab group of 49.8 years (14.2) versus a mean age of the placebo group of 52.1 years (11.2); percentage of white race in the group with alirocumab (88.9%) versus with placebo (85.7%); and a mean BMI (SD) in the alirocumab group of 28.8 kg / m2 (5.2) versus a mean BMI in the placebo group of 28.9 kg / m2 (4.2). Some imbalances were observed due to the small sample size of the study: a higher proportion of female patients in the alirocumab group (51.4%) versus the placebo group (37.1%); more recent diagnosis of hypercholesterolemia in the alirocumab group (median 9.8 years) versus the placebo group (median 17.4 years); a lower proportion of patients considered to be at very high CV risk in the alirocumab group (52.8%) than in the placebo group (65.7%), mainly caused by personal history of the coronary revascularization procedure; and a smaller proportion of patients received ezetimibe at randomization in the alirocumab group (19.4%) than in the placebo group (34.3%). The cardiovascular history and risk factors of the patients in both the alirocumab and placebo groups are shown in Table 28.

Table 28 - Cardiovascular history and risk factors

Placebo Alirocumab 150 All Q2W

(N = 35) (N = 72) (N = 107)

All patients on maximal statin background therapy

tolerated ± other THL

History of CHD,% (n) 43.1% (31) 62.9% (22) 49.5% (53)

Acute MI,% (n) 22.1% (16) 22.9% (8) 22.4% (24)

Silent MI,% (n) 1.4% (1) 0 0.9% (1)

Unstable angina,% (n) 9.7% (7) 17.1% (6) 12.1% (13)

Coronary revascularization procedures,% (n) 15.3% (11) 40.0% (14) 23.4% (25)

Other clinically significant CCT,% (n) 27.8% (20) 28.6% (10) 28.0% (30)

Current smoker,% (n) 16.7% (12) 25.7% (9) 19.6% (21)

Hypertension,% (n) 55.6% (40) 60.0% (21) 57.0% (61)

Type 2 diabetes,% (n) 12.5% (9) 17.1% (6) 14.0% (15)

At randomization, all patients were treated with a statin, receiving 72.9% of the high-intensity statin (atorvastatin 40 to 80 mg daily or rosuvastatin 20 to 40 mg daily) and receiving 6.5% simvastatin 80 mg. In addition to the statin, 19.4% and 34.3% of patients were receiving ezetimibe in the alirocumab and placebo groups, respectively. Table 30 shows the background lipid modifying therapies (TMLs) of the populations treated with alirocumab and placebo at randomization, as well as the total randomized population.

Table 31 shows the baseline lipid efficacy parameters for the alirocumab and placebo treated populations, as well as the total randomized population. The mean calculated LDL-C (SD) at baseline was 197.8 (53.4) mg / dL (5.123 (1.38) mmol / L). Mean non-HDL C (SD) at baseline was 226.4 (55.3) mg / dL. Mean total C (d E) at baseline was 274.4 (54.0) mg / dL. Mean HDL-C (SD) at baseline was 48.1 (13.3) mg / dL. The mean MC (SD) ratio of total C / HDL-C at baseline was 6.135 (2.119). Mean fasting triglycerides (SD) (TGs) at baseline were 149.8 (86.6) mg / dL. The mean lipoprotein- (a) (SD) at baseline was 41.2 (46.6) mg / dL. Apo-B mean (SD) at baseline was 140.9 (31.0) mg / dL. Apo-A1 mean (SD) at baseline was 137.5 (23.3) mg / dL. The mean Apo-B / Apo-A1 ratio (SD) at baseline was 1.061 (0.323) mg / dL.

Exposure to injections was similar across treatment groups with a mean exposure of 60.7 weeks in the placebo group and 58.3 weeks in the alirocumab group.

Table 29 - Characteristics of the disease and other relevant data from baseline - Randomized population Placebo Alirocumab 150 All Q2W

(N = 35) (N = 72) (N = 107) Type of hypercholesterolemia

Heterozygous familial hypercholesterolemia (HFhe) 35 (100%) 72 (100%) 107 (100%) Non-familial hypercholesterolemia (non-FH) 0 0 0

Time since diagnosis of hypercholesterolemia (years)

Number 35 72 107 Mean (SD) 16.41 (12.62) 11.48 (10.48) 13.09 (11.41) Median 17.42 9.76 11.70 Min: max 0.0: 42 .8 0.0: 39.9 0.0: 42.8

Diagnosis confirmation

By genotyping 5 (14.3%) 14 (19.4%) 19 (17.8%) By WHO / Simon Broomea 30 (85.7%) 58 (80.6%) 88 (82.2%) a for diagnosis of heHF not confirmed by genotyping.

Table 30 - Background TML at Randomization - Randomized Population

Placebo Alirocumab 150 Q2W All (N = 35) (N = 72) (N = 107) Any statin 35 (100%) 72 (100%) 107 (100%)

Taking high-dose statin 28 (80.0%) 57 (79.2%) 85 (79.4%) Taking high-intensity statin 25 (71.4%) 53 (73.6%) 78 (72.9 %) Atorvastatin daily dose (mg) 10 (28.6%) 22 (30.6%) 32 (29.9%) 10 0 0 0

20 0 0 0

40 3 (8.6%) 10 (13.9%) 13 (12.1%)

80 7 (20.0%) 11 (15.3%) 18 (16.8%) Placebo Alirocumab 150 Q2W All (N = 35) (N = 72) (N = 107) Other doses 0 1 (1.4%) 1 (0.9%) Rosuvastatin daily dose (mg) 16 (45.7% ) 33 (45.8%) 49 (45.8%)

5 1 (2.9%) 2 (2.8%) 3 (2.8%)

10 0 0 0

20 3 (8.6%) 8 (11.1%) 11 (10.3%)

40 12 (34.3%) 23 (31.9%) 35 (32.7%)

Other doses 0 0 0

Simvastatin daily dose (mg) 10 (28.6%) 19 (26.4%) 29 (27.1%)

10 1 (2.9%) 4 (5.6%) 5 (4.7%)

20 1 (2.9%) 2 (2.8%) 3 (2.8%)

40 5 (14.3%) 9 (12.5%) 14 (13.1%)

80 3 (8.6%) 4 (5.6%) 7 (6.5%)

Other doses 0 0 0

Any TML other than statins3 13 (37.1%) 16 (22.2%) 29 (27.1%) Any TML other than nutraceuticals 12 (34.3%) 16 (22.2%) 28 (26.2 %) Ezetimibe 12 (34.3%) 14 (19.4%) 26 (24.3%) Nutraceuticals 1 (2.9%) 0 1 (0.9%) a in combination with statins or not.

The high intensity statin corresponds to atorvastatin 40 to 80 mg daily or rosuvastatin 20 to 40 mg daily. The high-dose statin corresponds to atorvastatin 40 to 80 mg daily, rosuvastatin 20 to 40 mg daily, or simvastatin 80 mg daily.

Table 31 - Baseline lipid efficacy parameters - Quantitative summary in conventional units - Randomized population

Placebo Alirocumab 150 Q2W All (N = 35) (N = 72) (N = 107) Calculated LDL-C (mg / dL)

Number 35 71 106 Mean (SD) 201.0 (43.4) 196.3 (57.9) 197.8 (53.4) Median 201.0 180.0 181.0

Q1: Q3 166.0: 240.0 165.0: 224.0 165.0: 224.0

Min: Max 137: 279 89: 402 89: 402 Placebo Alirocumab 150 Q2W All (N = 35) (N = 72) (N = 107) C no HDL (mg / dL)

Number 35 72 107 Mean (SD) 231.5 (47.6) 223.9 (58.8) 226.4 (55.3) Median 226.0 204.0 209.0

Q1: Q3 194.0: 274.0 189.5: 251.0 191.0: 260.0

Min: max 153: 326 117: 419 117: 419

Total C (mg / dL)

Number 35 72 107 Mean (SD) 276.4 (46.8) 273.5 (57.5) 274.4 (54.0) Median 272.0 256.0 259.0

Q1: Q3 237.0: 313.0 242.5: 300.5 241.0: 310.0

Min: max 202: 364 171: 458 171: 458

HDL-C (mg / dL)

Number 35 72 107 Mean (SD) 44.9 (11.3) 49.6 (14.0) 48.1 (13.3) Median 42.0 45.5 45.0

Q1: Q3 39.0: 51.0 39.5: 57.5 39.0: 55.0

Min: max 24: 72 28: 84 24: 84

Fasting TGs (mg / dL)

Number 35 72 107 Mean (SD) 156.3 (89.3) 146.6 (85.6) 149.8 (86.6) Median 122.0 131.5 129.0

Q1: Q3 95.0: 193.0 87.5: 160.5 94.0: 171.0

Min: Max 62: 455 40: 512 40: 512

Lipoprotein- (a) (mg / dL)

Number 34 71 105 Mean (SD) 46.2 (50.3) 38.8 (44.9) 41.2 (46.6) Median 30.0 22.0 26.0

Q1: Q3 11.0: 42.0 8.0: 50.0 10.0: 48.0

Min: Max 2: 201 2: 189 2: 201 Placebo Alirocumab 150 Q2W All (N = 35) (N = 72) (N = 107)

Apo-B (mg / dL)

Number 34 71 105 Mean (SD) 146.6 (28.3) 138.2 (32.0) 140.9 (31.0) Median 143.0 130.0 134.0

Q1: Q3 121.0: 173.0 118.0: 154.0 119.0: 158.0

Min: Max 99: 208 81: 255 81: 255

Apo-A1 (mg / dL)

Number 34 71 105 Mean (SD) 131.5 (19.2) 140.3 (24.6) 137.5 (23.3) Median 127.5 137.0 134.0

Q1: Q3 120.0: 142.0 122.0: 155.0 122.0: 151.0

Min: Max 97: 181 97: 211 97: 211

Apo-B / Apo-A1 (ratio)

Number 34 71 105 Mean (SD) 1.141 (0.287) 1.023 (0.334) 1.061 (0.323) Median 1.170 0.950 1.020

Q1: Q3 0.900: 1.300 0.800: 1.170 0.850: 1.230

Min: Max 0.58: 1.86 0.49: 2.32 0.49: 232

Total C / HDL-C (ratio)

Number 35 72 107 Mean (SD) 6,540 (1,986) 5,938 (2,167) 6,135 (2,119) Median 6,417 5,647 5,863

Q1: Q3 4,936: 7,600 4,399: 6,878 4,545: 7,370

Min: Max 3.29: 11.19 2.92: 13.48 2.92: 13.48

Dosage and duration

Exposure to injections was similar across the treatment groups with a mean exposure of 60.7 weeks in the placebo group and 58.3 weeks in the alirocumab group. The duration of exposure for injection could not be calculated for 1 patient in the alirocumab group due to the unknown date of the last injection.

Effectiveness

Primary efficacy endpoint

The TDI analysis includes all calculated LDL-C values collected during treatment and without treatment through Week 52. The analysis of the primary endpoint (percent change in LDL-C calculated from baseline to Week 24) is provided based on a model of MMRM in the IDT population, using estimates of the mean CM at Week 24. Nine (11.3%) patients in the alirocumab group and 2 (5.7%) patients in the group with placebo did not have a calculated LDL-C value at Week 24. These missing values were accounted for by the MMRM model.

The results of the primary endpoint analysis are presented in Table 32, in mmol / L and mg / dL. Primary efficacy analysis

A statistically significant decrease in the percent change in LDL-C from baseline to Week 24 was observed in the alirocumab group (mean MC vs. baseline -45.7%) compared to the placebo group ( mean MC vs baseline -6.6%) (mean MC difference vs placebo (SE) -39.1% (6.0%), p <0.0001) (see Table 31). This represents an absolute reduction (SD) of -90.8 (6.7) mg / dL in the alirocumab group and -15.5 (9.5) mg / dL in the placebo group (see Table 33). . The percentage change from baseline to Week 24 in LDL-C by individual patients is shown in Figure 13. All patients were on a background statin (at the maximum tolerated level). A subset of patients also received additional lipid-lowering therapy.

In the alirocumab group, LDL-C reduction was observed from baseline from Week 4 to Week 52 (see Figure 7, Figure 14 and Table 33). A slight decrease in LDL-C reduction was observed over time in the alirocumab group (mean MC vs. baseline at Week 52 of -42.1 vs. -45.7 at Week 24), although overall the amount of decrease remained the same (75 mg / dL; see Figure 14). In addition, significant numbers of alirocumab patients achieved LDL-C levels of <100 mg / dL (57% vs 11% of placebo patients) and <70 mg / dL (<1.81 mmol / L; 32% versus 3% placebo patients) at Week 24 despite baseline LDL-C levels of> 190 mg / dL (mean (SD) baseline LDL-C calculated 196.3 (57 .9) mg / dL for the alirocumab group; 201 (43.4) mg / dL for the placebo group). At Week 12, 31.0% of patients in the alirocumab group (vs. 0.0% of the placebo group; TDI analysis) achieved calculated LDL-C levels of <70 mg / dL (<1.81 mmol / L). Similarly, at Week 52, 31.0% of patients in the alirocumab group (vs. 5.7% of the placebo group; TDI analysis) achieved calculated LDL-C levels of <70 mg / dL (<1 , 81 mmol / L).

A sensitivity analysis of the primary efficacy endpoint was performed excluding 13 patients from 2 sites with severe GCP noncompliance. The decrease in percent change in LDL-C from baseline to Week 24 was still statistically significant in the alirocumab group (mean MC vs. baseline -50.3%) compared to the placebo group ( mean MC vs. baseline -2.3%) (mean MC difference vs. placebo (SE) -48.0% (5.8%), p <0.0001) (see Table 34).

Table 32 - Percentage change from baseline in LDL-C calculated at Week 24: MMRM - IDT Analysis - IDT Population

Placebo Alirocumab 150 Q2W Calculated LDL cholesterol (N = 35) (N = 71) Baseline level (mmol / L)

Number 35 71

Mean (SD) 5,205 (1,125) 5,083 (1,499) Median 5,206 4,662

Min: Max 3.55: 7.23 2.31: 10.41 Initial level (mg / dL)

Number 35 71

Mean (SD) 201.0 (43.4) 196.3 (57.9) Median 201.0 180.0

Min: Max 137: 279 89: 402 Placebo Alirocumab 150 Q2W Calculated LDL cholesterol (N = 35) (N = 71)

Week 24 percent change from baseline (%)

Mean MC (SE) -6.6 (4.9) -45.7 (3.5) Mean difference of MC (SE) vs placebo -39.1 (6.0) 95% CI (-51 , 1 to -27.1) P-value vs placebo <0.0001 * Note: Least squares (LS) means, standard errors (SE), and p-values taken from MMRM analysis (mixed effects model with repeated measures ). The model includes the fixed categorical effects of treatment group, randomization strata according to IVRS, moment of time, interaction of treatment by moment of time and strata by moment of time, as well as the continuous fixed covariates of the calculated LDL-C value. baseline and baseline LDL-C value interaction calculated by moment of time

MMRM model and description of the baseline level performed in patients with a baseline value and a post baseline value in at least one of the analysis windows used in the model.

The p-value is followed by a '*' if it is statistically significant according to the fixed hierarchical approach used to ensure a strong control for the overall type I error rate at the 0.05 level.

Table 33 - LDL-C calculated over time - IDT analysis - IDT population

Placebo Alirocumab 150 Q2W (N = 35) (N = 71)

Change in Percentage Change LDL-C Percentage Change Change from from level from level from calculated Initial initial level value Initial initial level value Mean of MC

(EE) (mmol / L)

Initial level at 5,205 NA NA 5,083 NA NA

(0.190) (0.178)

Week 4 4.537 -0.586 (0.221) -11.5 (4.1) 2.522 -2.601 (0.154) -52.9 (2.8)

(0.221) (0.154)

Week 8 4.435 -0.688 (0.229) -12.4 (4.3) 2.647 -2.477 (0.161) -48.6 (3.1)

(0.229) (0.161)

Week 12 4.702 -0.422 (0.234) -6.6 (4.6) 2.692 -2.432 (0.164) -46.9 (3.2)

(0.234) (0.164)

Week 16 4.779 -0.344 (0.235) -6.1 (4.8) 2.633 -2.490 (0.161) -48.0 (3.3)

(0.235) (0.161)

Week 24 4.722 -0.401 (0.246) -6.6 (4.9) 2.771 -2.352 (0.174) -45.7 (3.5)

(0.246) (0.174)

Week 36 4.666 -0.457 (0.251) -8.9 (5.0) 2.832 -2.292 (0.176) -44.0 (3.5)

(0.251) (0.176)

Week 52 4.862 -0.262 (0.275) -3.0 (5.9) 2.921 -2.202 (0.197) -42.1 (4.2)

(0.275) (0.197)

Placebo Alirocumab 150 Q2W (N = 35) (N = 71)

Change in percent change LDL-C percent change Change from from level from level from calculated Baseline baseline value Baseline baseline value Week 78 1.2 (6.4) -37.9 (4.5)

MC mean

(EE) (mg / dL)

Initial level at 201.0 (7.3) NA NA 196.3 NA NA (6.9)

Week 4 175.2 -22.6 (8.5) -11.5 (4.1) 97.4 -100.4 (5.9) -52.9 (2.8)

(8.5) (5.9)

Week 8 171.2 -26.6 (8.8) -12.4 (4.3) 102.2 -95.6 (6.2) -48.6 (3.1)

(8.8) (6.2)

Week 12 181.5 -16.3 (9.0) -6.6 (4.6) 103.9 -93.9 (6.3) -46.9 (3.2)

(9.0) (6.3)

Week 16 184.5 -13.3 (9.1) -6.1 (4.8) 101.7 -96.1 (6.2) -48.0 (3.3)

(9.1) (6.2)

Week 24 182.3 -15.5 (9.5) -6.6 (4.9) 107.0 -90.8 (6.7) -45.7 (3.5)

(9.5) (6.7)

Week 36 180.2 -17.7 (9.7) -8.9 (5.0) 109.3 -88.5 (6.8) -44.0 (3.5)

(9.7) (6.8)

Week 52 187.7 -10.1 (10.6) -3.0 (5.9) 112.8 -85.0 (7.6) -42.1 (4.2)

(10.6) (7.6)

a The baseline level is described using means and standard errors.

Note: Least squares means (LC), standard errors (SE) and p-value taken from MMRM analysis (mixed effects model with repeated measures). The model includes fixed categorical effects of treatment group, strata of randomization according to IVRS, moment of time, interaction of treatment by moment of time, interaction of strata by moment of time, as well as the continuous fixed covariates of LDL-C value. at baseline and the interaction of LDL-C value at baseline by moment of time. MMRM model and description of the initial level performed in patients with a value at the initial level and a post-initial level value in at least one of the analysis windows used in the model.

Sensitivity analysis of the primary endpoint

Table 34 - Percentage change from baseline in LDL-C calculated at Week 24: MMRM - IDT analysis - IDT population excluding sites with severe GCP noncompliance

Placebo Alirocumab 150 Q2W Calculated LDL cholesterol (N = 31) (N = 62) Baseline level (mmol / L)

Number 31 62 Mean (SD) 5,310 (1,146) 5,101 (1,460) Median 5,258 4,675 Min: Max 3.55: 7.23 2.31: 10.41 Placebo Alirocumab 150 Q2W Calculated LDL cholesterol (N = 31) (N = 62) Baseline level (mg / dL)

Number 31 62 Mean (SD) 205.0 (44.2) 197.0 (56.4) Median 203.0 180.5 Min: Max 137: 279 89: 402

Percentage change at Week 24 from baseline (%)

Mean MC (SE) -2.3 (4.7) -50.3 (3.3) Mean difference MC (SE) vs placebo -48.0 (5.8) 95% CI (-59 .4 to -36.6) P-value vs placebo <0.0001 Note: Least squares means (LC), standard errors (SE) and p-value taken from MMRM analysis (mixed effects model with repeated measures) . The model includes the fixed categorical effects of treatment group, randomization strata according to IVRS, moment of time, interaction of treatment by moment of time and strata by moment of time, as well as the continuous fixed covariates of the calculated LDL-C value. baseline and baseline LDL-C value interaction calculated by moment of time

MMRM model and description of the baseline level performed in patients with a baseline value and a post baseline value in at least one of the analysis windows used in the model.

The p-value is not adjusted for multiplicity and is provided for descriptive purposes only

Note: Sites N ° 643-710 and N °. 840-743 were excluded from the analysis

Secondary endpoints of key efficacy

The following table summarizes the analysis results for the secondary key endpoints in the hierarchical order. All key secondary endpoints are statistically significant by the included hierarchical testing procedure up to the endpoint of Lp (a) at Week 24 (TDI estimate). Statistical significance for HDL-C was not reached at Week 24 (TDI estimate) and thus the testing procedure was stopped, only p-values for this endpoint were provided for descriptive purposes.

Table 35

Endpoint Analysis Results Calculated LDL-C p-value - Percentage change from baseline During MC <0.0001 level to Week 24 treatment vs placebo of -38.9%

Calculated LDL-C - Percentage Change from TDI Level Mean Difference from Baseline MC <0.0001 through Week 12 vs. Placebo of -40.3%

Calculated LDL-C - Percentage Change from Baseline During MC <0.0001 Mean Difference to Week 12 Treatment vs. Placebo of -40.3%

Apo-B - Percentage change from baseline TDI Mean MC difference <0.0001 through Week 24 vs placebo of -30.3%

Apo-B - Percentage change from baseline During MC mean difference <0.0001 through Week 24 treatment vs placebo of -30.2%

Endpoint Analysis Results P value

Non-HDL C - Percentage change from baseline TDI Mean MC difference <0.0001 through Week 24 vs placebo of -35.8%

Non-HDL C - Percentage change from baseline During MC mean difference <0.0001 through Week 24 treatment vs placebo of -35.5%

Total C - Percentage change from baseline TDI Mean MC difference <0.0001 through Week 24 vs placebo of -28.4%

Apo-B - Percentage change from baseline TDI Mean MC difference <0.0001 through Week 12 vs placebo of -30.2%

Non-HDL C - Percentage change from baseline IDT Mean MC difference <0.0001 through Week 12 vs placebo of -34.5%

Total C - Percentage change from baseline TDI Mean difference of MC <0.0001 through Week 12 vs placebo of -27.8%

Calculated LDL-C - Percent Change from TDI Level Baseline MC <0.0001 Difference to Week 52 vs. Placebo of -39.1%

Proportion of patients at very high CV risk who combined TDI estimate for 0.0016 reach calculated LDL-C <70 mg / dL (1.81 mmol / L) or the odds ratio of patients at high CV risk reaching LDL-C vs. a placebo of 11.7

calculated <100 mg / dL (2.59 mmol / L) at Week 24

Proportion of patients at very high CV risk who during the pooled estimate for 0.0014 reach calculated LDL-C <70 mg / dL (1.81 mmol / L) or treatment the odds ratio of patients at high CV risk who reach C- LDL versus placebo of 11.9

calculated <100 mg / dL (2.59 mmol / L) at Week 24

Lp (a) - Percentage change from baseline to IDI pooled estimate for 0.0164 at Week 24 adjusted mean difference

versus placebo -14.8%

HDL-C - Percentage change from baseline TDI Mean difference of MC 0.2745 through Week 24 vs placebo 3.7%

Fasting TGs - Percentage change from TDI level pooled estimate for baseline 0.1386 through Week 24 adjusted mean difference

vs. placebo -8.7%

The analysis during treatment of the percentage change in LDL-C from baseline to Week 24 shows results very consistent with the TDI analysis (mean difference of MC vs placebo of -38.9% in analysis during treatment vs to -39.1% in the TDI analysis). In fact, only 3 patients (2 on placebo and 1 on alirocumab) had post-treatment LDL-C values (that is, more than 21 days after the last injection) at Week 24.

Key secondary endpoints including Apo B, Non-HDL C, Total C, Lp (a) at various time points, as well as the proportion of patients reaching their LDL-C goals at Week 24, were statistically significant. according to the hierarchical test procedure. Significant reductions in non-HDL C, Apo B, and Lp (a) levels were observed at Week 24. The mean percent change in MC of alirocumab versus placebo from baseline to Week 24 was -41.9 versus a -6.2 for non-HDL C (p <0.0001), -39.0 vs -8.7 for Apo B (p <0.0001) and -23.5 vs -8 .7 for Lp (a) (p value = 0.0164).

The proportion of patients at very high cardiovascular (CV) risk who achieve calculated LDL-C <70 mg / dL (1.81 mmol / L) or patients at high CV risk who achieve calculated LDL-C <100 mg / dL (2 , 59 mmol / L) at Week 24 was significantly higher in the alirocumab group than placebo (pooled estimate for the proportion of 41.0% in the alirocumab group vs 5.7% in the placebo group, p = 0.0016).

The analyzes performed with the treatment approach were consistent with these analyzes.

Differences in percentage change in fasting HDL-C and TG from baseline to Week 24 in the IDT analysis were not statistically significant: HDL-C at Week 24: mean MC vs. baseline was + 7.5% in the alirocumab group and 3.9% in the placebo group (mean difference of MC vs. placebo of 3.7%, p = 0.2745); and fasting TGs at Week 24: mean MC vs. baseline was -10.5% in the alirocumab group and -1.1% in the placebo group (mean MC vs. placebo difference of -9, 4%, p = 0.1299).

Four (5.6%) patients had two consecutive calculated LDL-C values <25 mg / dL. No particular safety concern has been observed in these patients.

Summary of safety results:

The proportion of patients who had a treatment emergent adverse event (TEAE) was lower in the alirocumab group (61.1%) compared to the placebo group (71.4%) in the present study. The proportion of patients who had a severe TEAE was similar between treatment groups. A similar proportion of patients had TEAEs leading to treatment discontinuation (1 patient (2.9%) and 3 patients (4.2%) in the placebo and alirocumab groups, respectively). These results are in agreement with the proportion of patients who have had TEAEs in previous placebo-controlled phase 2/3 alirocumab studies (results of 2476 and 1276 patients in the alirocumab and placebo groups, respectively). Specifically, in this study, TEAEs were 75.8% vs. 76.4%, treatment-emergent SAEs were 13.7% vs. 14.3%, TEAEs leading to death were 0.5% vs. 0.9%, and TEAEs leading to discontinuation were 5.3% versus 5.1%, for alirocumab versus placebo groups, respectively.

The most frequently reported SOCs (and PTs) in both treatment groups of the present study were: "infections and infestations": 40.3% in the alirocumab group vs 34.3% in the placebo group (with influenza reported in 11.1% vs 2.9% and urinary tract infection in 6.9% vs 0 in the group with alirocumab vs placebo respectively); "cardiac disorders": 12.5% in the alirocumab group versus no cases in the placebo group. Among the events submitted for validation, the events were confirmed for 6 patients presenting: 4 MI, 1 heart failure requiring hospitalization, and 5 coronary revascularization procedures caused by ischemia; "nervous system disorders": 11.1% in the alirocumab group vs 8.6% in the placebo group (with reported headache in 5.6% vs 0 and dizziness 4.2% vs 0 in the group with alirocumab vs placebo respectively); and "musculoskeletal and connective tissue disorders": 16.7% in the alirocumab group versus 28.6% in the placebo group. No deaths were reported during the study in either group.

SAEs were reported by 11.1% patients in the alirocumab group and 11.4% in the placebo group. There is no particular clinical pattern among the preferred SAEs terms that were reported individually. The most frequently reported SOC (System Organ Class) for SAEs is "cardiac disorders".

Seven patients, 6 (8.3%) in the alirocumab group and 1 (2.9%) in the placebo group had a local injection site reaction emerging from treatment. These events were of mild intensity except one of moderate intensity. Two patients, one (1.4%) in the alirocumab group and one (2.9%) in the placebo group, had neurocognitive disorders. Four patients, three (4.2%) in the alirocumab group and one (2.9%) in the placebo group, had ALT> 3 x ULN. Two patients out of 71 analyzed (2.8%, compared with 0 patients in the placebo group) had a creatine kinase level> 3 x ULN. None of the events were serious or led to treatment discontinuation. TEAEs occurring in the alirocumab and placebo patient groups up to the last patient visit at Week 52 were collected and classified in Table 36.

Table 36 - TEAE safety analysis up to week 52.

Placebo Alirocumab 150 Q2W

(N = 35) (N = 72)

% (n) of patients

All patients with maximum tolerated statin background therapy ± other LTL

Nasopharyngitis 11.4% (4) 11.1% (8)

Flu 2.9% (1) 11.1% (8)

Injection site reaction 2.9% (1) 8.3% (6)

Urinary tract infection 0 6.9% (5)

Diarrhea 8.6% (3) 5.6% (4) Placebo Alirocumab 150 Q2W (N = 35) (N = 72) Sinusitis 5.7% (2) 5.6% (4) Bronchitis 2.9% (1) 5.6% (4) Headache 0 5.6% (4) Fatigue 0 5.6% (4) Myalgia 8.6% (3) 4.2% (3) Nausea 5.7% (2) 1.4% (1) Vertigo 5.7% (2) 1.4% (1) Dyspepsia 5.7% (2) 0 Elevated uric acid in blood 5.7% (2) 0 Rheumatoid arthritis 5.7% (2) 0

Among the events of interest, no particular signal was detected for TEAE related to neurological events, general allergic events and diabetes.

No relevant abnormality was observed for PCSA.

Conclusion:

The following conclusions can be drawn regarding patients with heHF and high baseline LDL-C levels despite maximum tolerated statin with or without another THL from the ODYSSEY HIGH FH study: 1) self-administered alirocumab produced reductions in LDL-C significantly higher compared to placebo after 24 weeks, with the mean absolute decrease from baseline in LDL-C -90.8 mg / dL at Week 24 with alirocumab compared to -15.5 mg / dL with placebo , and resulting LDL-C levels of 107 mg / dL with alirocumab at Week 24 versus 182 mg / dL with placebo; 2) 32% of patients with alirocumab achieved LDL-C <70 mg / dL despite baseline LDL-C> 190 mg / dL; 3) 57% of patients with alirocumab reached LDL-C <100 mg / dL at Week 24; 4) alirocumab was generally well tolerated and TEAEs occurred at a similar frequency in the alirocumab and placebo arms.

Example 5: Efficacy and safety of the monoclonal antibody against PCSK9 alirocumab versus placebo in 1254 patients with heterozygous familial hypercholesterolemia (HFhe): analysis up to 78 weeks of four ODYSSEY trials

Background:

Previous studies have shown that only -20% of patients with heterozygous familial hypercholesterolemia (heFH) treated with lipid-lowering therapies (LTHs) achieved predefined target LDL-C levels of <2.5 mmol / L [97 mg / dL]. The efficacy and safety of alirocumab versus placebo was studied in 1,254 heFH patients with maximally tolerated statin ± other THL from four 18-month placebo-controlled ODYSSEY trials (FH I, FH II, HIGH FH, LONG TERM). This represents the single largest set of HeFH patients studied in a phase 3 clinical trial program. A description of the LONG TERM study is set forth in Robinson et al., (2015) NEJM 372: 16 pp 1489-99.

Methods:

Data were pooled by initial alirocumab dose. In FH I / II, patients with LDL-C levels> 1.81 / 2.59 mmol / L [70/100 mg / dL], depending on CV risk, received placebo (N = 244) or alirocumab 75 mg Q2W (N = 488); alirocumab dose increased to 150 mg Q2W at Week 12 if LDL-C at Week 8> 1.81 mmol / L [70 mg / dL] (41.8% of patients). Separately, data for HIGH FH (LDL-C> 4.14 mmol / L [160 mg / dL]) and the LONG TERM subset of patients with heFH (LDL-C> 1.81 mmol / L [70 mg / dL]), where patients received placebo (N = 180) or alirocumab 150 mg Q2W (N = 342). All doses were 1 mL subcutaneous (SC) injections. Data were pooled for change in LDL-C from baseline through Week 52.

Results:

Baseline LDL-C levels and changes from baseline with treatment are shown in Table 37. Compared with placebo, alirocumab reduced LDL-C by 49% and 61% (p <0.0001) in Week 12 for the 75 and 150 mg Q2W doses, respectively. At Week 24, the LDL-C reductions with alirocumab versus placebo were 56% (alirocumab 75 mg Q2W with a possible dose increase at Week 12) and 59% (alirocumab 150 mg Q2W), respectively (p <0 , 0001). For both dose regimens, despite high baseline LDL-C levels, mean MC LDL-C levels of ~ 2 mmol / L [77 mg / dL] were achieved at Week 12 (Table 37 ), with reductions maintained through Week 52. Additional beneficial effects were seen in other parameters including non-HDL C and Apo B.

In individual studies to date, overall, similar rates of treatment emergent adverse events (TEAEs) have been observed in alirocumab and placebo-treated patients. Across placebo-controlled studies in the ODYSSEY Program (both patients with and without HeFH), TEAEs (preferred terms) reported in> 5% of patients on alirocumab or placebo include nasopharyngitis (11.3% and 11.1 % of patients treated with alirocumab and placebo, respectively), upper respiratory tract infection (u Ri) (6.1% vs 7.0%), injection site reaction (6.7% vs 4, 8%), flu (5.7% vs. 4.6%), headache (4.8% vs. 5.2%) and arthralgia (4.0% vs. 5.5%).

Table 37: Mean Least Squares (LS) (SE) calculated LDL-C at Week 12 (W12), Week 24 (W24) and Week 52 (W52) (intention-to-treat analysis)

Conclusions:

Claims (11)

1. A proprotein convertase subtilisin / kexin type 9 inhibitor (PCSK9) for use in the treatment of hypercholesterolemia in a patient with heterozygous familial hypercholesterolemia (HeFH) who has a serum LDL-C concentration greater than or equal to 70 mg / dL despite taking a stable daily dose of statin for at least 4 weeks, and a history of documented cardiovascular disease, wherein the PCSK9 inhibitor is an antibody or antigen-binding fragment thereof that specifically binds to human PCSK9 and comprises an HCVR and an LCVR having the amino acid sequences of SEQ ID NOs: 1 and 6, respectively. 2. The PCSK9 inhibitor for use of claim 1, wherein the diagnosis of heHF is made either by genotyping or clinical criteria, and wherein the clinical criteria is or the Simon Broome Register Diagnostic Criteria for Heterozygous Familial Hypercholesterolemia, or the WHO / Dutch Lipid Network criteria with a score> 8. The PCSK9 inhibitor for the use of any one of claims 1-2, wherein the antibody or antigen-binding fragment thereof that specifically binds PCSK9 is administered to the patient at a dose of 75 mg at a frequency once every two weeks. 4. The PCSK9 inhibitor for use of claim 3, wherein: (a) the 75 mg dose is maintained if the patient's LDL-C measured after five or more doses is less than 70 mg / dL; and (b) the 75 mg dose is discontinued if the patient's LDL-C measured after five or more doses remains greater than or equal to 70 mg / dL, and the antibody or antigen-binding fragment thereof that specifically binds PCSK9 is subsequently administered to the patient at a dose of 150 mg at a frequency of once every two weeks. 5. The PCSK9 inhibitor for the use of any one of claims 1 or 2, wherein the antibody or antigen-binding fragment thereof that specifically binds PCSK9 is administered to the patient at a dose of 150 mg at a frequency once every two weeks. 6. The PCSK9 inhibitor for the use of any one of claims 1-5, wherein the PCSK9 inhibitor is administered to the patient in combination with the daily dose of stable statin. 7. The PCSK9 inhibitor for use of any one of claims 1-6, wherein the stable daily dose of statin comprises a daily dose of 40 mg to 80 mg of atorvastatin, a daily dose of 20 mg to 40 mg of rosuvastatin, or a daily dose of 80 mg simvastatin. 8. The PCSK9 inhibitor for use of claim 6 or 7, wherein the PCSK9 inhibitor is administered to the patient in further combination with at least one other lipid-lowering therapy. The PCSK9 inhibitor for the use of any one of claims 1-8, wherein the patient has one or more of the indications selected from the group consisting of type 2 diabetes, high blood pressure, myocardial infarction and stroke. ischemic. The PCSK9 inhibitor for use of claim 9, wherein the patient has type 2 diabetes. The PCSK9 inhibitor for the use of any one of claims 1-10, wherein the PCSK9 inhibitor improves at least one parameter associated with hypercholesterolemia selected from the group consisting of: (a) reduction of the patient's low-density lipoprotein cholesterol (LDL-C) by at least 40%; (b) reduction of the patient's apolipoprotein B (ApoB) by at least 30%; (c) reduction of the patient's non-high-density lipoprotein (non-HDL C) cholesterol by at least 40%; (d) reduction of the patient's total cholesterol by at least 20%; (e) increase in the patient's high-density lipoprotein cholesterol (HDL-C) by at least 3%; (f) reduction of the patient's triglycerides by at least 5%; (g) reduction of the patient's lipoprotein a (Lp (a)) by at least 20%; and (h) Increase in the patient's apolipoprotein A-1 by at least 1%.