Patent Description:
Diverse diseases or illnesses may have common, partially non-specific symptoms that can range from unpleasant to unbearable for the individual suffering from therefrom. Quite often individuals experiencing more than one symptom need to take several drugs to experience alleviation of these symptoms. There is an ongoing need for new forms of therapy or prevention of symptoms associated with many different underlying diseases or illnesses. In particular, it would be helpful to provide medicaments or drugs that can be used in the therapy or prevention of more than one symptom associated with an underlying disease or illness. The present invention addresses this need.

The peptide adrenomedullin (ADM) was described for the first time in <NUM> (<NPL>) as a novel hypotensive peptide comprising <NUM> amino acids, which had been isolated from a human pheochromocytome. In the same year, cDNA coding for a precursor peptide comprising <NUM> amino acids and the complete amino acid sequence of this precursor peptide were also described. The precursor peptide, which comprises, inter alia, a signal sequence of <NUM> amino acids at the N-terminus, is referred to as "preproadrenomedullin" (pre-proADM). In the present description, all amino acid positions specified usually relate to the pre-proADM which comprises the <NUM> amino acids. The peptide adrenomedullin (ADM) is a peptide which comprises <NUM> amino acids (SEQ ID NO: <NUM>) and which comprises the amino acids <NUM> to <NUM> of pre-proADM, from which it is formed by proteolytic cleavage. To, date, only a few fragments of the peptide fragments formed in the cleavage of the pre-proADM have been more exactly characterized, in particular the physiologically active peptides adrenomedullin (ADM) and "PAMP", a peptide comprising <NUM> amino acids (<NUM>-<NUM>) which follows the <NUM> amino acids of the signal peptide in pre-proADM. The discovery and characterization of ADM in <NUM> triggered intensive research activity, the results of which have been summarized in various review articles, in the context of the present description, reference being made in particular to the articles to be found in an issue of "Peptides" devoted to ADM in particular (<NPL>) and (<NPL>). A further review is (<NPL>). In the scientific investigations to date, it has been found, inter alia, that ADM may be regarded as a poly-functional regulatory peptide. It is released into the circulation in an inactive form extended by glycine (<NPL>). There is also a binding protein (<NPL>) which is specific for ADM and probably likewise modulates the effect of ADM. Those physiological effects of ADM as well as of PAMP which are of primary importance in the investigations to date were the effects influencing blood pressure.

ADM is an effective vasodilator, and it is possible to associate the hypotensive effect with the particular peptide segments in the C-terminal part of ADM. It has furthermore been found that the above-mentioned further physiologically active peptide PAMP formed from pre-proADM likewise exhibits a hypotensive effect, even if it appears to have an action mechanism differing from that of ADM.

It has furthermore been found that the concentrations of ADM, which can be measured in the circulation and other biological liquids are, in a number of pathological states, significantly above the concentrations to be found in healthy control persons. Thus, the ADM level in patients with congestive heart failure, myocardial infarction, kidney diseases, hypertensive disorders, diabetes mellitus, in the acute phase of shock and in sepsis and septic shock are significantly increased, although to different extents. The PAMP concentrations are also increased in some of said pathological states, but the plasma levels are lower relative to ADM ((Eto, T. It is furthermore known that unusually high concentrations of ADM are to be observed in sepsis, and the highest concentrations in septic shock (cf. , "supra) and (<NPL>; <NPL> ):<NPL> ;<NPL>; and <NPL>).

There is only limited knowledge about the role of adrenomedullin in migraine. As adrenomedullin has vasodilatory effects within the cerebral circulation, it is hypothesized to be involved in migraine. On the one hand, the administration of human ADM intravenous infusion to patients suffering from migraine did not induce significantly more headache or migraine compared with placebo (<NPL>). Moreover, Akcali et al. reported low plasma adrenomedullin levels in the natural course of migraine patients (<NPL>). On the other hand, it has been stated by Kis et al. that there is a need for the development of novel, potent, specific and possibly non-peptide receptor antagonists as potential therapeutic tools for the suppression of ADM-mediated proliferative effects as anti-migraine drugs (<NPL>). <CIT> discloses murine and humanized anti adrenomedullin antibodies which reduce mortality in a mouse model of sepsis.

Before describing the invention in detail, it is deemed expedient to provide definitions for certain technical terms used throughout the description. Although the present invention will be described with respect to particular embodiments, this description is not to be construed in a limiting sense. Before describing in detail exemplary embodiments of the present invention, definitions important for understanding the present invention are given.

As used in this specification and in the appended claims, the singular forms of "a" and "an" also include the respective plurals unless the context clearly dictates otherwise.

In the context of the present invention, the terms "about" and "approximately" denote an interval of accuracy that a person skilled in the art will understand to still ensure the technical effect of the feature in question. The term typically indicates a deviation from the indicated numerical value of ±<NUM> %, preferably ±<NUM> %, more preferably ±<NUM> %, and even more preferably ±<NUM> %.

It is to be understood that the term "comprising" is not limiting. For the purposes of the present invention the term "consisting" of is considered to be a preferred embodiment of the term "comprising" of. If hereinafter a group is defined to comprise at least a certain number of embodiments, this is meant to also encompass a group, which preferably consists of these embodiments only.

It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the present invention that will be limited only by the appended claims.

As used herein, the term "nausea" refers to a sensation of unease and discomfort in the upper stomach with an involuntary urge to vomit (<NPL>). It may precede vomiting, but a person can have nausea without vomiting. When prolonged, it is a debilitating symptom. Nausea is a non-specific symptom, which means that it has many possible causes. Some common causes of nausea are motion sickness, dizziness, migraine, fainting, low blood sugar, gastroenteritis (stomach infection) or food poisoning. Nausea is a side effect of many medications including chemotherapy, or morning sickness in early pregnancy. Nausea may also be caused by anxiety, disgust and depression.

As used herein, the term "headache" is the symptom of pain anywhere in the region of the head or neck. It occurs in migraines (sharp or throbbing pains), tension-type headaches, and cluster headaches (<NPL>). There is also an increased risk of depression in those with severe headaches. Headaches can occur as a result of many conditions whether serious or not. There are a number of different classification systems for headaches. It is well-recognized that causes of headaches may include fatigue, sleep deprivation, stress, effects of medications, the effects of recreational drugs, viral infections, loud noises, common colds, head injury, rapid ingestion of a very cold food or beverage, and dental or sinus issues.

Headaches are broadly classified as "primary" or "secondary" (O<NPL>). Primary headaches are benign, recurrent headaches not caused by underlying disease or structural problems. For example, migraine is a type of primary headache. While primary headaches may cause significant daily pain and disability, they are not dangerous. The four categories of primary headaches are: migraine, tension-type headache (TTH) cluster headache and other trigeminal autonomic cephalalgias, and other primary headaches.

Secondary headaches, which are of organic, metabolic or drug-induced origin, are caused by an underlying disease, like an infection, head injury, vascular disorders, brain bleed or tumors. Secondary headaches can be harmless or dangerous. A migraine is a primary headache disorder characterized by recurrent headaches that are moderate to severe (for review see: <NPL>). Typically, the headaches affect one half of the head, are pulsating in nature, and last from two to <NUM> hours. Associated symptoms may include nausea, vomiting, and sensitivity to light, sound, or smell. The pain is generally made worse by physical activity. Up to one-third of people have an aura: typically a short period of visual disturbance, which signals that the headache will soon occur. Occasionally, an aura can occur with little or no headache following it.

As used herein, the terms "muscle aches" or "muscle pain", also termed "myalgia", refer to a symptom of many diseases and disorders (for review see: <NPL>). The most common causes are the overuse or overstretching of a muscle or group of muscles. Myalgia without a traumatic history is often due to viral infections. Longer-term myalgias may be indicative of a metabolic myopathy, some nutritional deficiencies or chronic fatigue syndrome.

As used herein, the term "back pain" refers to painful sensations in the any part of the back. Episodes of back pain may be acute, sub-acute, or chronic depending on the duration. The pain may be characterized as a dull ache, shooting or piercing pain, or a burning sensation. The pain may radiate into the arms and hands as well as the legs or feet, and may include paresthesia (tingling with no apparent cause), weakness or numbness in the legs and arms. The anatomic classification of back pain follows the segments of the spine: neck pain (cervical), middle back pain (thoracic), lower back pain (lumbar) or coccydynia (tailbone or sacral pain) with the lumbar vertebrae area most common for pain. The pain may originate from the muscles, nerves, bones, joints or other structures in the vertebral column (spine), however, internal structures such as the gallbladder and pancreas may also cause referred pain in the back (<NPL>).

As used herein, the term "shivering" (also called "shuddering") is a bodily function in response to early hypothermia or just feeling cold in warm-blooded animals. When the core body temperature drops, the shivering reflex is triggered to maintain homeostasis. Skeletal muscles begin to shake in small movements, creating warmth by expending energy. Shivering can also be a response to a fever, as a person may feel cold. During fever the hypothalamic set point for temperature is raised. The increased set point causes the body temperature to rise (pyrexia), but also makes the patient feel cold until the new set point is reached. Severe chills with violent shivering are called rigors. Rigors occur because the patient's body is shivering in a physiological attempt to increase body temperature to the new set point. Shivering can also appear after surgery, known as post-anesthetic shivering.

As used herein, the term "vomiting", also known as emesis and throwing up, among other terms, is the involuntary, forceful expulsion of the contents of one's stomach through the mouth and sometimes the nose (<NPL>). Vomiting can be caused by a wide variety of conditions; it may present as a specific response to ailments like gastritis or poisoning, or as a non-specific sequela of disorders ranging from brain tumors and elevated intracranial pressure to overexposure to ionizing radiation.

As used herein, the symptoms of illnesses or diseases in a patient in need of therapy and/or prevention of such symptoms are selected from the groups of disease indications comprising inflammatory conditions, autoimmune diseases, metabolic diseases, brain diseases, cardiovascular diseases and drug-induced diseases.

Below, the types of symptoms and illnesses associated herewith are provided in form of nonlimiting lists. It is noted that the herein described therapy or prevention may be directed to more than one type of symptom. Further, it is noted that a given medical indication, illness, or disease may be associated with more than one of the symptoms.

The symptom "nausea" may be associated with illnesses inside the abdomen, e.g. obstructing disorders (for example pyloric obstruction, small bowel obstruction, colonic obstruction, superior mesenteric artery syndrome), enteric infections (for example viral or bacterial infection), inflammatory diseases (such as cholecystitis, pancreatitis, appendicitis, hepatitis), sensorimotor dysfunction (for example, gastroparesis, intestinal pseudo-obstruction, gastroesophageal reflux disease, chronic idiopathic nausea, functional vomiting, cyclic vomiting syndrome, rumination syndrome) or biliary colic; illnesses outside the abdomen, e.g. cardiopulmonary disorder (such as cardiomyopathy, myocardial infarction), inner-ear diseases (such as motion sickness, labyrinthitis, malignancies), intracerebral disorders (for example hemorrhage, abscess, hydrocephalus, malignancies), psychiatric illnesses (for example anorexia and bulimia nervosa, depression), post-operative vomiting, nausea associated with medications and drugs (for example chemotherapy and biologics therapy, antibiotics, anti-arrhythmics, digoxin, oral hypoglycemic medications, oral contraceptives), nausea associated with endocrine/metabolic diseases (such as pregnancy, uremia, ketoacidosis, thyroid and parathyroid disease, adrenal insufficiency), nausea due to intoxication because of liver failure, alcohol abuse, etc..

The symptom "back pain" may be associated with illnesses due to inflammation, especially in the acute phase, which typically lasts from two weeks to three months, and may be associated with lumbago, trauma, injury, infections, cancer (especially cancers known to spread to the spine like breast, lung and prostate cancer), etc..

The symptom "myalgia" or "muscle pain" may be associated with injury or trauma, including sprains, hematoma, or overuse, wherein a muscle was used too much, too often, including protecting a separate injury, chronic tension, muscle pain due to rhabdomyolysis, associated with, e.g., viral infections, compression injury, drug-related, e.g., due to fibrates and statins, ACE inhibitors, cocaine, some anti-retroviral drugs, severe potassium deficiency, fibromyalgia, Ehlers-Danlos syndrome, auto-immune disorders (including for example mixed connective tissue disease, Systemic lupus erythematosus, polymyalgia rheumatic, Myositis, such as polymyositis, dermatomyositis, and inclusion body myositis, multiple sclerosis, Myalgic Encephalomyelitis (chronic fatigue syndrome), Familial Mediterranean fever, Polyarteritis nodosa, Devic's disease, Morphea, Sarcoidosis), metabolic diseases (such as carnitine palmitoyltransferase II deficiency, Conn's syndrome, Adrenal insufficiency, Hyperthyroidism, Hypothyroidism, Diabetes, Hypogonadism, , as well as Channelopathy, Stickler Syndrome, Hypokalemia, Hypotonia (Low Muscle Tone), Exercise intolerance, Mastocytosis, Peripheral neuropathy, Eosinophilia myalgia syndrome, Barcoo Fever, herpes, hemochromatosis also known as iron overload disorder, delayed onset muscle soreness, AIDS, HIV infections, tumor-induced osteomalacia, hypovitaminosis D, myocardial infarction.

The symptom "headache" may be associated with primary headaches. <NUM>% of all headaches are primary headaches. Primary headaches usually first start when people are between <NUM> and <NUM> years old. The most common types of primary headaches are migraines and tension-type headaches. They have different characteristics. Migraines typically present with pulsing head pain, nausea, photophobia (sensitivity to light) and phonophobia (sensitivity to sound).

Tension-type headaches usually present with non-pulsing "bandlike" pressure on both sides of the head, not accompanied by other symptoms. Other very rare types of primary headaches include: cluster headaches: short episodes (<NUM>-<NUM> minutes) of severe pain, usually around one eye, with autonomic symptoms (tearing, red eye, nasal congestion) which occur at the same time every day. Cluster headaches can be treated with triptans and prevented with prednisone, ergotamine or lithium; trigeminal neuralgia or occipital neuralgia characterized by shooting face pain, hemicrania continua, i.e., continuous unilateral pain with episodes of severe pain; primary stabbing headaches, e.g., recurrent episodes of stabbing "ice pick pain" or "jabs and jolts" for <NUM> second to several minutes without autonomic symptoms (tearing, red eye, nasal congestion); Primary cough headache that starts suddenly and lasts for several minutes after coughing, sneezing or straining (anything that may increase pressure in the head). Serious causes (see secondary headaches red flag section) must be ruled out before a diagnosis of "benign" primary cough headache can be made; primary exertional headache characterized by throbbing, pulsatile pain which starts during or after exercising, lasting for <NUM> minutes to <NUM> hours. The mechanism behind these headaches is unclear, possibly due to straining causing veins in the head to dilate, causing pain; primary sex headache characterized by a dull, bilateral headache that starts during sexual activity and becomes much worse during orgasm; hypnic headache; secondary headaches that may be caused by problems elsewhere in the head or neck. Some of these are not harmful, such as cervicogenic headache (pain arising from the neck muscles), medication overuse headache in those using excessive painkillers for headaches, meningitis characterized by inflammation of the meninges which presents with fever and meningismus, or stiff neck; bleeding inside the brain (intracranial hemorrhage); subarachnoid hemorrhage (acute, severe headache, stiff neck without fever); ruptured aneurysm, arteriovenous malformation, intraparenchymal hemorrhage; temporal arteritis, i.e., an inflammatory disease of arteries common in the elderly (average age <NUM>), polymyalgia rheumatic; acute closed angle glaucoma (increased pressure in the eyeball); post-ictal headaches that happen after a convulsion or other type of seizure, as part of the period after the seizure (the post-ictal state); gastrointestinal disorders may cause headaches, including Helicobacter pylori infection, celiac disease, non-celiac gluten sensitivity, irritable bowel syndrome, inflammatory bowel disease, gastroparesis, and hepatobiliary disorders. The treatment of the gastrointestinal disorders may lead to a remission or improvement of headaches.

The term headache is defined as primary or secondary headache. Primary headache is defined as migraine, tension-type headache (TTH), cluster headache and other trigeminal autonomic cephalalgias, and other primary headaches.

Diagnosis or assessment of headache is well-established in the art. Assessment may be performed based on subjective measures, such as patient characterization of symptoms. For example, migraine may be diagnosed based on the following criteria: <NUM>) episodic attacks of headache lasting <NUM> to <NUM> hours; <NUM>) with two of the following symptoms: unilateral pain, throbbing, aggravation on movement, and pain of moderate or severe intensity; and <NUM>) one of the following symptoms: nausea or vomiting, and photophobia or phonophobia (<NPL>).

The symptom "shivering" may be associated with fever, cold sensitivity, menopause, panic attack, anxiety, bacterial infection, rickettsial infection, viral infection, drug withdrawal, etc..

The symptom "vomiting" may be associated with gastritis (inflammation of the gastric wall), gastroenteritis, gastroesophageal reflux disease, celiac disease, non-celiac gluten sensitivity, pyloric stenosis, bowel obstruction, overeating, acute abdomen and/or peritonitis, ileus, food allergies (often in conjunction with hives or swelling); cholecystitis, pancreatitis, appendicitis, hepatitis; food poisoning, allergic reaction to cow's milk proteins, e.g., milk allergy or lactose intolerance; motion sickness, Ménière's disease, concussion, cerebral hemorrhage, migraine, brain tumors, benign intracranial hypertension and hydrocephalus, metabolic disturbances such as hypercalcemia, Uremia adrenal insufficiency, hypoglycemia, hyperglycemia, drug reaction, alcohol intoxication, opioid uptake, selective serotonin reuptake inhibitors; use of chemotherapy drugs; gastric inflammation caused by a range of viruses and bacteria, e.g., norovirus, influenza; or psychiatric/behavioral illnesses such as bulimia nervosa and purge disorder.

Below, embodiments of the invention are provided. It is noted that generally embodiments can be combined with any other embodiment of the same category (product, process, use, method).

Subject matter of the present invention is an anti-adrenomedullin (ADM) antibody or an anti-adrenomedullin antibody fragment for use in therapy or prevention of symptoms of illness in a subject in need thereof, wherein the subject is human, and.

The antibody or fragment binds to mature ADM, e.g., to ADM of amino acids <NUM> to <NUM> (SEQ ID NO: <NUM>), in particular to N-terminal ADM (SEQ ID NO: <NUM>), as detailed below.

In a further embodiment, the illness is migraine.

The sample is selected from the group comprising whole blood, plasma, and serum.

Mean arterial pressure (MAP) is defined as the average pressure in a patient's arteries during one cardiac cycle. It is considered as indicator of perfusion to vital organs. It is believed that a MAP that is greater than <NUM> mmHg is enough to sustain the organs of the average person. In the context of the present invention the term "mean arterial pressure is decreased" means a MAP below <NUM> mmHg.

The mature adrenomedullin peptide is an amidated peptide (ADM-NH<NUM>), which comprises <NUM> amino acids (SEQ ID No: <NUM>) and which comprises the amino acids <NUM> to <NUM> of pre-proADM <NUM>-<NUM>, from which it is formed by proteolytic cleavage. Mature ADM, bio-ADM and ADM-NH<NUM> is used synonymously throughout this application and is a molecule according to SEQ ID No.: <NUM>.

The N-terminal part (aa <NUM>-<NUM>) of ADM has the sequence YRQSMNNFQGLRSFGCRFGTC (SEQ ID No. <NUM>).

Another embodiment of the invention relates to an anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering , in a subject in need thereof according to any of the preceding embodiments, wherein said antibody or fragment exhibits a binding affinity to ADM of at least <NUM>-<NUM> M by label-free surface plasmon resonance using a Biacore <NUM> system.

The anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject in need thereof according to any of the preceding embodiments, is not ADM-binding-Protein-<NUM> (complement factor H).

Another embodiment of the invention relates to an anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject in need thereof according to any of the preceding embodiments, wherein said antibody or fragment recognizes and binds to the N-terminal end (aa <NUM>) of ADM.

Another embodiment of the invention relates to an anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject in need thereof according to any of the preceding embodiments, wherein said antibody or fragment is an ADM stabilizing antibody or fragment that enhances the half-life (t<NUM>/<NUM> half retention time) of ADM in serum, blood, plasma at least <NUM> %, preferably at least, <NUM> %, more preferably > <NUM> %, most preferably > <NUM> %.

Another embodiment of the invention relates to an anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject in need thereof according to any of the preceding embodiments, wherein said antibody or fragment blocks the bioactivity of ADM not more than <NUM> %, preferably not more than <NUM>% using hADM <NUM>-<NUM> as a reference antagonist in CHO-K1 cells expressing human recombinant ADM receptor.

Another embodiment of the invention relates to an anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering according to any of the preceding embodiments, wherein said subject in need thereof, is selected from the group of subjects suffering from a disease selected from the group comprising migraine, viral infections, drug-induced symptoms of disease (e.g., chemotherapy-induced, or induced by treatment with biologicals, antibiotics, anti-viral compounds, etc.). In a particular embodiment, the disease or illness (these terms may be used interchangeably) is migraine.

Another embodiment of the invention relates to an anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject according to any of the preceding embodiments, wherein said subjects undergoes cancer therapy (chemotherapy).

Chemotherapy is a category of cancer treatment that uses one or more anti-cancer drugs (chemotherapeutic agents) as part of a standardized chemotherapy regimen. Chemotherapeutic drugs may include the following drug categories: alkylating agents, kinase inhibitors, vinca alkaloids, anthracyclines, antimetabolites, aromatase inhibitors, topoisomerase inhibitors, engineered anti-cancer agents such as monoclonal antibodies, cytokines, gene therapy vectors, antisense and peptide molecules.

The anti-ADM antibody or an anti-ADM antibody fragment for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering, in a subject according to any of the preceding embodiments is a humanized monoclonal antibody or fragment that binds to the N-terminal region (aa <NUM>-<NUM>) of ADM (SEQ ID No. <NUM>) or an antibody fragment thereof wherein the heavy chain comprises the sequences:.

and wherein the light chain comprises the sequences:.

Another embodiment of the invention relates to a humanized monoclonal antibody or fragment that binds to ADM or an antibody fragment thereof as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject according to any of the preceding embodiments, wherein said antibody or fragment comprises a sequence selected from the group comprising as a VH region:.

and comprises the following sequence as a VL region:.

Also disclosed is a humanized monoclonal antibody or fragment that binds to ADM or an antibody fragment thereof as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject according to any of the preceding embodiments, wherein said antibody or fragment comprises the following sequence as a heavy chain:
SEQ ID NO: <NUM>
<IMG>
and comprises the following sequence as a light chain:
SEQ ID NO: <NUM>
<IMG>.

The antibody comprises the following sequence as a heavy chain:
SEQ ID NO: <NUM>
<IMG>
or a sequence that is > <NUM>% identical to it, preferably > <NUM>%, preferably > <NUM>% and comprises the following sequence as a light chain:
SEQ ID NO: <NUM>
<IMG>
or a sequence that is > <NUM>% identical to it, preferably > <NUM>%, preferably > <NUM>%.

To assess the identity between two amino acid sequences, a pairwise alignment is performed. Identity defines the percentage of amino acids with a direct match in the alignment.

Pharmaceutical composition comprising an antibody according as above outlined.

Another embodiment of the invention relates to an anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject in need thereof according to any of the preceding embodiments to be used in combination with known medicaments against nausea.

Another embodiment of the invention relates to an anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject in need thereof according to any of the preceding embodiments to be used in combination with known medicaments against headache.

Another embodiment of the invention relates to an anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject in need thereof according to any of the preceding embodiments to be used in combination with known medicaments against myalgia.

Another embodiment of the invention relates to an anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject in need thereof according to any of the preceding embodiments to be used in combination with known medicaments against shivering.

Another embodiment of the invention relates to an anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject in need thereof according to any of the preceding embodiments to be used in combination with known medicaments against vomiting.

Another embodiment of the invention relates to an anti-ADM antibody or an anti-ADM antibody fragment as defined herein for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering in a subject in need thereof according to any of the preceding embodiments to be used in combination with known medicaments against back pain.

Another embodiment of the invention relates to a pharmaceutical formulation for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering as defined herein in a subject in need thereof comprising an antibody or fragment according to any of the preceding embodiments.

Another embodiment of the invention relates to a pharmaceutical formulation for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering as defined herein in a subject in need thereof according to the preceding embodiment, wherein said pharmaceutical formulation is a solution; preferably a ready-to-use solution.

Another embodiment of the invention relates to a pharmaceutical formulation for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering as defined herein in a subject according to the preceding embodiment, wherein said pharmaceutical formulation is in a freeze-dried state.

Another embodiment of the invention relates to a pharmaceutical formulation for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering as defined herein in a subject in need thereof according to any of the preceding embodiments relating to such formulations, wherein said pharmaceutical formulation is administered intra-muscular.

Another embodiment of the invention relates to a pharmaceutical formulation for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering as defined herein in a subject in need thereof according to any of the preceding embodiments relating to such formulations, wherein said pharmaceutical formulation is administered intra-vascular.

Another embodiment of the invention relates to a pharmaceutical formulation for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering as defined herein in a subject according to the preceding embodiment, wherein said pharmaceutical formulation is administered via infusion.

Another embodiment of the invention relates to a pharmaceutical formulation for use in therapy or prevention of symptoms of illness selected from the group of nausea, headache, muscle aches, back pain, shivering as defined herein in a subject according to any of the according to any of the preceding embodiments relating to such formulations, wherein said pharmaceutical formulation is to be administered systemically.

As used herein, the definition of the "illness score" relates to LPS-induced clinical symptoms including nausea, headache, muscle aches, back pain, shivering, and vomiting. Those symptoms were combined into one score (= Illness score): as an example each symptom on a scale of <NUM> to <NUM>, except vomiting (either <NUM> or <NUM>, depending on having vomited in the previous <NUM> minutes). The Illness score is a sum over all symptoms (theoretical ranges from <NUM> to <NUM>).

It should be emphasized that the herein described embodiments relating to anti-ADM antibody or an anti-ADM antibody fragment are intended to be applied for sake of therapy or prevention of symptoms associated with illnesses, and thus are not necessarily intended for any methods of primary treatment or first line treatment to the acute disease or acute condition itself that has to be considered as underlying disease(s). This means the present invention do not provide for a therapy of healing/curing e.g. cancer, sepsis, septic shock, COPD, congestive heart disease (acute heart failure).

Furthermore, the anti-ADM antibody or an anti-ADM antibody fragment is monospecific. Monospecific anti-ADM antibody or monospecific anti-ADM antibody fragment means in particular that said antibody or antibody fragment binds to one specific region encompassing at least <NUM> amino acids within the target ADM. Monospecific anti-ADM antibody or monospecific anti-ADM antibody fragment are anti-ADM antibodies or anti-ADM antibody fragments s that all have affinity for the same antigen.

The monospecific anti-ADM antibody or monospecific anti-ADM antibody fragment in particular binds to one specific region encompassing at least <NUM> amino acids within the target ADM. Monospecific means in particular that said antibody or antibody fragment binds to one specific region encompassing preferably at least <NUM>, or at least <NUM> amino acids within the target ADM. Monospecific antibodies or fragments are antibodies or fragments that all have affinity for the same antigen. Monoclonal antibodies are monospecific, but monospecific antibodies may also be produced by other means than producing them from a common germ cell.

An antibody according to the present invention is a protein including one or more polypeptides substantially encoded by immunoglobulin genes that specifically binds an antigen. The recognized immunoglobulin genes include the kappa, lambda, alpha (IgA), gamma (IgG1, IgG2, IgG3, IgG4), delta (IgD), epsilon (IgE) and mu (IgM) constant region genes, as well as the myriad immunoglobulin variable region genes. Full-length immunoglobulin light chains are generally about <NUM> kDa or <NUM> amino acids in length. Full- length immunoglobulin heavy chains are generally about <NUM> kDa or <NUM> amino acid in length. Light chains are encoded by a variable region gene at the NH<NUM>-terminus (about <NUM> amino acids in length) and a kappa or lambda constant region gene at the COOH--terminus. Heavy chains are similarly encoded by a variable region gene (about <NUM> amino acids in length) and one of the other constant region genes.

The basic structural unit of an antibody is generally a tetramer that consists of two identical pairs of immunoglobulin chains, each pair having one light and one heavy chain. In each pair, the light and heavy chain variable regions bind to an antigen, and the constant regions mediate effector functions. Immunoglobulins also exist in a variety of other forms including, for example, Fv, Fab, and (Fab')<NUM>, as well as bifunctional hybrid antibodies and single chains (e.g., <NPL>: <NPL>; <NPL>; <NPL>. An immunoglobulin light or heavy chain variable region includes a framework region interrupted by three hypervariable regions, also called complementarity determining regions (CDR's) (see, <NPL>). As noted above, the CDRs are primarily responsible for binding to an epitope of an antigen. An immune complex is an antibody, such as a monoclonal antibody, chimeric antibody, humanized antibody or human antibody, or functional antibody fragment, specifically bound to the antigen.

Chimeric antibodies are antibodies whose light and heavy chain genes have been constructed, typically by genetic engineering, from immunoglobulin variable and constant region genes belonging to different species. For example, the variable segments of the genes from a mouse monoclonal antibody can be joined to human constant segments, such as kappa and gamma <NUM> or gamma <NUM>. In one example, a therapeutic chimeric antibody is thus a hybrid protein composed of the variable or antigen-binding domain from a mouse antibody and the constant or effector domain from a human antibody, although other mammalian species can be used, or the variable region can be produced by molecular techniques. Methods of making chimeric antibodies are well known in the art, e.g., see <CIT>. A "humanized" immunoglobulin is an immunoglobulin including a human framework region and one or more CDRs from a non-human (such as a mouse, rat, or synthetic) immunoglobulin. The non- human immunoglobulin providing the CDRs is termed a "donor" and the human immunoglobulin providing the framework is termed an "acceptor. " In one embodiment, all the CDRs are from the donor immunoglobulin in a humanized immunoglobulin. Constant regions need not be present, but if they are, they must be substantially identical to human immunoglobulin constant regions, i.e., at least about <NUM>-<NUM>%, such as about <NUM>% or more identical. Hence, all parts of a humanized immunoglobulin, except possibly the CDRs, are substantially identical to corresponding parts of natural human immunoglobulin sequences. A "humanized antibody" is an antibody comprising a humanized light chain and a humanized heavy chain immunoglobulin. A humanized antibody binds to the same antigen as the donor antibody that provides the CDRs. The acceptor framework of a humanized immunoglobulin or antibody may have a limited number of substitutions by amino acids taken from the donor framework. Humanized or other monoclonal antibodies can have additional conservative amino acid substitutions, which have substantially no effect on antigen binding or other immunoglobulin functions. Exemplary conservative substitutions are those such as gly, ala; val, ile, leu; asp, glu; asn, gin; ser, thr; lys, arg; and phe, tyr. Humanized immunoglobulins can be constructed by means of genetic engineering (e.g., see <CIT>). A human antibody is an antibody wherein the light and heavy chain genes are of human origin. Human antibodies can be generated using methods known in the art. Human antibodies can be produced by immortalizing a human B cell secreting the antibody of interest. Immortalization can be accomplished, for example, by EBV infection or by fusing a human B cell with a myeloma or hybridoma cell to produce a trioma cell. Human antibodies can also be produced by phage display methods (see, e.g., <CIT>; <CIT>; and Winter, <CIT>), or selected from a human combinatorial monoclonal antibody library (see the Morphosys website). Human antibodies can also be prepared by using transgenic animals carrying a human immunoglobulin gene (for example, see <CIT>; and <CIT>).

In a preferred embodiment antibodies according to the present invention are recombinantly produced antibodies as e.g. IgG, a typical full-length immunoglobulin, or antibody fragments containing at least the F-variable domain of heavy and/or light chain as e.g. chemically coupled antibodies (fragment antigen binding) including but not limited to Fab-fragments including Fab minibodies, single chain Fab antibody, monovalent Fab antibody with epitope tags, e.g. Fab-V5Sx2; bivalent Fab (mini-antibody) dimerized with the CH3 domain; bivalent Fab or multivalent Fab, e.g. formed via multimerization with the aid of a heterologous domain, e.g. via dimerization of dHLX domains,e.g. Fab-dHLX-FSx2; F(ab')<NUM>-fragments, scFv-fragments, multimerized multivalent or/and multispecific scFv- fragments, bivalent and/or bispecific diabodies, BITE® (bispecific T-cell engager), trifunctional antibodies, polyvalent antibodies, e.g. from a different class than G; single-domain antibodies, e.g. nanobodies derived from camelid or fish immunoglobulins and numerous others.

An antibody fragment according to the present invention is an antigen binding fragment of an antibody according to the present invention.

In a preferred embodiment the ADM antibody format is selected from the group comprising Fv fragment, scFv fragment, Fab fragment, scFab fragment, (Fab)<NUM> fragment and scFv-Fc Fusion protein. In another preferred embodiment the antibody format is selected from the group comprising scFab fragment, Fab fragment, scFv fragment and bioavailability optimized conjugates thereof, such as PEGylated fragments. One of the most preferred formats is the scFab format.

Antibodies may be produced as follows: A Balb/c mouse was immunized with 100µg ADM-Peptide-BSA-Conjugate at day <NUM> and <NUM> (emulsified in 100µI complete Freund's adjuvant) and 50µg at day <NUM> and <NUM> (in <NUM>µl incomplete Freund's adjuvant). Three days before the fusion experiment was performed, the animal received 50µg of the conjugate dissolved in <NUM>µI saline, given as one intraperitoneal and one intravenous injection. Splenocytes from the immunized mouse and cells of the myeloma cell line SP2/<NUM> were fused with <NUM> <NUM>% polyethylene glycol for <NUM> at <NUM>. After washing, the cells were seeded in <NUM>-well cell culture plates. Hybrid clones were selected by growing in HAT medium (RPMI <NUM> culture medium supplemented with <NUM>% fetal calf serum and HAT-Supplement). After two weeks the HAT medium is replaced with HT Medium for three passages followed by returning to the normal cell culture medium. The cell culture supernatants were primary screened for antigen specific IgG antibodies three weeks after fusion. The positive tested microcultures were transferred into <NUM>-well plates for propagation. After retesting, the selected cultures were cloned and recloned using the limiting-dilution technique and the isotypes were determined (see also<NPL>; <NPL>).

Antibodies may also be produced by means of phage display according to the following procedure: The human naive antibody gene libraries HAL7/<NUM> were used for the isolation of recombinant single chain F- Variable domains (scFv) against ADM peptide. The antibody gene libraries were screened with a panning strategy comprising the use of peptides containing a biotin tag linked via two different spacers to the ADM peptide sequence. A mix of panning rounds using non-specifically bound antigen and streptavidin bound antigen were used to minimize background of non-specific binders. The eluted phages from the third round of panning have been used for the generation of monoclonal scFv expressing E. coli strains. Supernatants from the cultivation of these clonal strains have been directly used for an antigen ELISA testing (see references cited in <CIT>, incorporated herein in their entirety).

Humanization of murine antibodies may be conducted according to the following procedure: For humanization of an antibody of murine origin the antibody sequence is analyzed for the structural interaction of framework regions (FR) with the complementary determining regions (CDR) and the antigen. Based on structural modelling an appropriate FR of human origin is selected and the murine CDR sequences are transplanted into the human FR. Variations in the amino acid sequence of the CDRs or FRs may be introduced to regain structural interactions, which were abolished by the species switch for the FR sequences. This recovery of structural interactions may be achieved by random approach using phage display libraries or via directed approach guided by molecular modeling (<NPL>) in a preferred embodiment the ADM antibody format is selected from the group comprising Fv fragment, scFv fragment, Fab fragment, scFab fragment, F(ab)<NUM> fragment and scFv-Fc Fusion protein. In another preferred embodiment the antibody format is selected from the group comprising scFab fragment, Fab fragment, scFv fragment and bioavailability optimized conjugates thereof, such as PEGylated fragments. One of the most preferred formats is scFab format. In another preferred embodiment, the anti-ADM antibody or anti-ADM antibody fragment, is a full length antibody or antibody fragment.

According to an example not forming part of the literal wording of the claims as granted the anti-ADM antibody or an anti-ADM antibody fragment or anti-ADM non-Ig scaffold is directed to and can bind to an epitope of at least <NUM> amino acids in length contained in ADM.

According to a further example not forming part of the literal wording of the claims as granted the anti-ADM antibody or an anti-ADM antibody fragment or anti-ADM non-Ig scaffold is directed to and can bind to an epitope of at least <NUM> amino acids in length contained in ADM.

According to an example not forming part of the literal wording of the claims as granted the anti-ADM antibody or an anti- ADM antibody fragment binds to a region of ADM that is located in the N-terminal part (amino acids <NUM>-<NUM>) of ADM.

According to a further example not forming part of the literal wording of the claims as granted said anti-ADM antibody or an anti-ADM antibody fragment recognizes and binds to the N-terminal end (aa1) of ADM. N-terminal end means that the amino acid <NUM>, that is "Y" of SEQ ID No. <NUM> or <NUM> is mandatory for antibody binding. Said antibody or fragment would neither bind N-terminal extended nor N-terminally modified ADM nor N-terminally degraded ADM.

According to an example not forming part of the literal wording of the claims as granted the anti-ADM antibody or an anti-ADM antibody fragment is directed to and can bind to an epitope of at least <NUM> amino acids in length contained in ADM, preferably in human ADM.

In one specific embodiment it is preferred to use an anti-ADM antibody or an anti- ADM antibody fragment according to the present invention, wherein said anti-ADM antibody or said anti-ADM antibody fragment is an ADM stabilizing antibody or an ADM stabilizing antibody fragment that enhances the half-life (t<NUM>/<NUM>; half retention time) of ADM in serum, blood, plasma at least <NUM> %, preferably at least <NUM> %, more preferably ><NUM> %, most preferably ><NUM>%. The half-life (half retention time) of ADM may be determined in human plasma in absence and presence of an ADM stabilizing antibody or an ADM stabilizing antibody fragment, respectively, using an immunoassay for the quantification of ADM.

A two-fold increase of the half-life of ADM is an enhancement of half-life of <NUM>%. Half Life (half retention time) is defined as the period over which the concentration of a specified chemical or drug takes to fall to half baseline concentration in the specified fluid or blood.

According to an example not forming part of the literal wording of the claims as granted the anti-ADM antibody or anti-ADM antibody fragment is a non-neutralizing antibody or fragment. A neutralizing anti-ADM antibody or anti-ADM antibody fragment would block the bioactivity of ADM to nearly <NUM>%, to at least more than <NUM>%, preferably to at least more than <NUM>%.

In contrast, a non-neutralizing anti-ADM antibody, or anti-ADM antibody fragment blocks the bioactivity of ADM less than <NUM>%, preferably to less than <NUM>%, preferably to less than <NUM>%, more preferred to less than <NUM> % and even more preferred to less than <NUM> %. This means that the residual bioactivity of ADM bound to the non-neutralizing anti-ADM antibody, or anti-ADM antibody fragment would be more than <NUM>%, preferably more than <NUM> %, preferably more than <NUM> %, more preferred more than <NUM> %, more preferred more than <NUM> %. In this context (a) molecule(s), being it an antibody, or an antibody fragment with "non-neutralizing anti-ADM activity", collectively termed here for simplicity as "non-neutralizing" anti-ADM antibody or antibody fragment, , that e.g. blocks the bioactivity of ADM to less than <NUM> %, is defined as - a molecule or molecules binding to ADM, which upon addition to a culture of an eukaryotic cell line, which expresses functional human recombinant ADM receptor composed of CRLR (calcitonin receptor like receptor) and RAMP3 (receptor-activity modifying protein <NUM>), reduces the amount of cAMP produced by the cell line through the action of parallel added human synthetic ADM peptide, wherein said added human synthetic ADM is added in an amount that in the absence of the non-neutralizing antibody to be analyzed, leads to half- maximal stimulation of cAMP synthesis, wherein the reduction of cAMP by said molecule(s) binding to ADM takes place to an extent, which is not more than <NUM>%, even when the non-neutralizing molecule(s) binding to ADM to be analyzed is added in an amount, which is <NUM>-fold more than the amount, which is needed to obtain the maximal reduction of cAMP synthesis obtainable with the non-neutralizing antibody to be analyzed. The same definition applies to the other ranges; <NUM>%, <NUM>%, <NUM>% etc..

According to an example not forming part of the literal wording of the claims as granted an anti-ADM antibody or an anti- ADM antibody fragment is used, wherein said antibody or antibody fragment blocks the bioactivity of ADM to less than <NUM> %, preferably less than <NUM>% (of baseline values). This is in the sense of blocking the circulating ADM of not more than <NUM>% or not more than <NUM>%, respectively. It has been understood that limited blocking of the bioactivity of ADM occurs even at excess concentration of the antibody or antibody fragment, meaning an excess of the antibodyor antibody fragment in relation to ADM. Said limited blocking is an intrinsic property of the ADM binder itself. This means that said antibody or antibody fragment have a maximal inhibition of <NUM>% or <NUM>%, respectively. By implication, this means that <NUM>% or <NUM>% residual ADM bioactivity remains present although appropriate amounts or excess amounts of antibody or antibody fragment are administered, respectively.

According to an example not forming part of the literal wording of the claims as granted said anti-ADM antibody, anti-ADM antibody fragment would block the bioactivity of ADM at least <NUM> %. By implication, this means residual <NUM>% circulating ADM bioactivity remains present. This is the lower threshold of bioactivity remaining after administration of said anti-ADM antibody, anti-ADM antibody fragment. The bioactivity is defined as the effect that a substance takes on a living organism or tissue or organ or functional unit in vivo or in vitro (e.g. in an assay) after its interaction. In case of ADM bioactivity this may be the effect of ADM in a human recombinant ADM receptor cAMP functional assay. According to an example not forming part of the literal wording of the claims as granted bioactivity is defined via an ADM receptor cAMP functional assay. The following steps may be performed in order to determine the bioactivity of ADM in such an assay:.

A maximal (at maximal dose) inhibition by said ADM stabilizing antibody of <NUM>% means that said ADM antibody or said ADM antibody fragment, respectively, blocks the bioactivity to <NUM>% of baseline values. A maximal inhibition in said ADM bioassay of <NUM>% means that said anti-ADM antibody or said anti- ADM antibody fragment, respectively, blocks the bioactivity of ADM to <NUM>%. This is in the sense of blocking the ADM bioactivity to not more than <NUM>%.

According to an example not forming part of the literal wording of the claims as granted a modulating anti-ADM antibody or a modulating anti-ADM antibody fragment is used. A "modulating" anti-ADM antibody or a modulating anti-ADM antibody fragment is an antibody or an ADM antibody fragment that enhances the half-life (t half retention time) of ADM in serum, blood, plasma at least <NUM> %, preferably at least, <NUM> %, more preferably ><NUM> %, most preferably ><NUM> % and blocks the bioactivity of ADM to less than <NUM> %, preferably less than <NUM> % and wherein said anti-ADM antibody or anti-ADM antibody fragment would block the bioactivity of ADM at least <NUM> %. These values related to half-life and blocking of bioactivity have to be understood in relation to the before-mentioned assays in order to determine these values. This is in the sense of blocking the circulating ADM of not more than <NUM>% or not more than <NUM>%, respectively. This means <NUM>% residual ADM bioactivity remains present, or <NUM>% residual ADM bioactivity remains present, respectively. Such a modulating anti-ADM antibody or a modulating anti-ADM antibody fragment offers the advantage that the dosing of the administration is facilitated. The combination of partially blocking or partially reducing ADM bioactivity and increase of the in vivo half-life (increasing the ADM bioactivity) leads to beneficial simplicity of anti- ADM antibody or an anti-ADM antibody fragment dosing. In a situation of excess endogenous ADM the activity lowering effect is the major impact of the antibody or fragment, limiting the (negative) effect of ADM. In case of low or normal endogenous ADM concentrations, the biological effect of anti- ADM antibody or an anti-ADM antibody fragment is a combination of lowering (by partially blocking) and increase by increasing the ADM half-life. Thus, the non-neutralizing and modulating ADM antibody or ADM antibody fragment acts like an ADM bioactivity buffer in order to keep the bioactivity of ADM within a certain physiological range.

The anti-ADM antibody or anti-ADM antibody fragment according to the present invention exhibits an affinity towards human ADM that the affinity constant is greater than <NUM>-<NUM> M, preferred <NUM>-<NUM> M, preferred affinity is greater than <NUM>-<NUM> M, most preferred higher than <NUM>-<NUM> M. A person skilled in the art knows that it may be considered to compensate lower affinity by applying a higher dose of compounds and this measure would not lead out-of-the-scope of the invention. The affinity constants may be determined according to the method as described in Example <NUM> of <CIT>.

Subject of the present invention is further a pharmaceutical formulation comprising an anti-ADM antibody or anti-ADM antibody fragment as defined herein for the use according to the present invention. Subject of the present invention is further a pharmaceutical formulation according to the present invention wherein said pharmaceutical formulation is a solution, preferably a ready-to-use solution. In another embodiment subject of the present invention is further a pharmaceutical formulation according to the present invention wherein said pharmaceutical formulation is in a dried state to be reconstituted before use. Said pharmaceutical formulation may be administered intra-muscular. Said pharmaceutical formulation may be administered intra-vascular. Said pharmaceutical formulation may be administered via infusion. In another embodiment subject of the present invention is further a pharmaceutical formulation according to the present invention wherein said pharmaceutical formulation is in a freeze-dried state.

In another more preferred embodiment the present invention provides for a pharmaceutical formulation comprising an anti-ADM antibody or an anti-ADM antibody fragment binding to ADM as defined herein for use in therapy or prevention of symptoms of illness in a patient as defined herein, wherein said pharmaceutical formulation is to be administered to a patient in need thereof.

In another embodiment of the present invention the pharmaceutical formulation according to the present invention is to be administered to a patient for therapy and/or prevention of symptoms associated with illnesses as defined above with the proviso that said patient is in need of such treatment.

According to an example not forming part of the literal wording of the claims as granted the ADM antibody or an ADM antibody fragment according to the invention is a non-neutralizing ADM antibody or a non-neutralizing ADM antibody fragment.

As used herein, the antibody format is selected from the group comprising Fv fragment, scFv fragment, Fab fragment, scFab fragment, (Fab)<NUM> fragment and scFv-Fc Fusion protein.

In embodiments of the present invention, the ADM antibody or an ADM antibody fragment according to any of the preceding embodiments for use in a treatment of a patient in need thereof wherein said antibody or fragment may be administered in a dose of at least <NUM> / Kg body weight, particularly at least <NUM>/kg body weight, more particularly, from <NUM> to <NUM>/kg body weight, e.g., from <NUM> to <NUM>/kg body weight, from <NUM> to <NUM>/kg body weight, or from <NUM> to <NUM>/kg body weight.

In embodiments of the present invention, the symptoms of illness referred to herein are not associated with either disease selected from the group comprising sepsis, diabetes, cancer, heart failure (acute heart failure), and septic shock.

In preferred embodiments of the present invention, the symptoms of illness that are treated or prevented using any of the ADM antibody or an ADM antibody fragment according to any of preceding embodiments are associated with migraine.

In preferred embodiments of the present invention, the symptoms of illness that are treated or prevented using any of the ADM antibody or an ADM antibody fragment according to any of preceding embodiments are associated with virus infections, wherein said viruses are selected from the group comprising hepadnaviridae, adenoviridae, herpesviridae, influenza viruses, arenaviridae, filoviridae, togaviridae, noroviruses, flaviviridae, retroviridae, measles virus, reoviridae, enteroviridae, picomaviridae, caliciviridae, etc..

In preferred embodiments of the present invention, the symptoms of illness that are treated or prevented using any of the ADM antibody or an ADM antibody fragment according to any of preceding embodiments are associated with drug-treatment of primary diseases, such as chemotherapy, therapy with biologics (e.g., antibodies, or fragments thereof), antibiotics, or any medicaments causing any of the above mentioned symptoms of illness.

Further preferred embodiments of the present invention relate to methods of therapy (e.g., treatment, curing, alleviating, improving, amelioration, etc.) or prevention of symptoms as defined in any of the foregoing embodiments comprising administering to a subject in need thereof the ADM antibody or an ADM antibody fragment according to any of preceding embodiments. The subject is preferably a human.

Administration of the ADM antibody or an ADM antibody fragment can be by any means known in the art, including: orally, intravenously, subcutaneously, intraarterially, intramuscularly, intracardially, intraspinally, intrathoracically, intraperitoneal, intraventricular, sublingually, transdermal, and/or via inhalation. Administration may be systemic, e.g. intravenously, or localized.

In one aspect, the present invention provides a method for treating or preventing headache, preferably primary headache, more preferred migraine in an individual comprising administering to the individual an effective amount of an ADM antibody or an ADM antibody fragment as defined herein.

In a further embodiment, the invention provides methods for ameliorating, controlling, reducing incidence of, or delaying the development or progression of headache preferably primary headache, more preferred migraine in an individual comprising administering to the individual an effective amount of the ADM antibody or an ADM antibody fragment.

The ADM antibody or an ADM antibody fragment may be administered prior to, during and/or after headache. In some embodiments, the ADM antibody or an ADM antibody fragment is administered prior to the attack of headache.

In some embodiments ADM antibody or an ADM antibody fragment as defined herein may be administered as defined herein in conjunction with another agent, such as another agent for treating headache.

"Development" or "progression" of headache means initial manifestations and/or ensuing progression of the disorder. Development of headache can be detectable and assessed using standard clinical techniques as well known in the art. However, development also refers to progression that may be undetectable. For purpose of this invention, development or progression refers to the biological course of the symptoms. "Development" includes occurrence, recurrence, and onset. As used herein "onset" or "occurrence" of headache includes initial onset and/or recurrence.

As used herein, an "effective dosage" or "effective amount" of drug, compound, or pharmaceutical composition is an amount sufficient to effect beneficial or desired results. For prophylactic use, beneficial or desired results include results such as eliminating or reducing the risk, lessening the severity, or delaying the outset of the disease, including biochemical, histological and/or behavioral symptoms of the disease, its complications and intermediate pathological phenotypes presenting during development of the disease. For therapeutic use, beneficial or desired results include clinical results such as reducing pain intensity, duration, or frequency of headache attack, and decreasing one or more symptoms resulting from headache (biochemical, histological and/or behavioral), including its complications and intermediate pathological phenotypes presenting during development of the disease, increasing the quality of life of those suffering from the disease, decreasing the dose of other medications required to treat the disease, enhancing effect of another medication, and/or delaying the progression of the disease of patients. An effective dosage can be administered in one or more administrations. For purposes of this invention, an effective dosage of drug, compound, or pharmaceutical composition is an amount sufficient to accomplish prophylactic or therapeutic treatment either directly or indirectly. As is understood in the clinical context, an effective dosage of a drug, compound, or pharmaceutical composition may or may not be achieved in conjunction with another drug, compound, or pharmaceutical composition. Thus, an "effective dosage" may be considered in the context of administering one or more therapeutic agents, and a single agent may be considered to be given in an effective amount if, in conjunction with one or more other agents, a desirable result may be or is achieved.

Several human and murine antibodies were produced and their affinity constants were determined (see Table <NUM>).

Peptides for immunization were synthesized, see Table <NUM>, (JPT Technologies, Berlin, Germany) with an additional N-terminal Cysteine (if no Cysteine is present within the selected ADM-sequence) residue for conjugation of the peptides to Bovine Serum Albumin (BSA). The peptides were covalently linked to BSA by using Sulfolink-coupling gel (Perbio Science, Bonn, Germany). The coupling procedure was performed according to the manual of Perbio.

The murine antibodies were generated according to the following method:
A Balb/c mouse was immunized with 100µg Peptide-BSA-Conjugate at day <NUM> and <NUM> (emulsified in 100µl complete Freund's adjuvant) and 50µg at day <NUM> and <NUM> (in 100µl incomplete Freund's adjuvant). Three days before the fusion experiment was performed, the animal received 50µg of the conjugate dissolved in 100µl saline, given as one intraperitoneal and one intra-venous injection.

Splenocytes from the immunized mouse and cells of the myeloma cell line SP2/<NUM> were fused with <NUM> <NUM>% polyethylene glycol for <NUM> at <NUM>. After washing, the cells were seeded in <NUM>-well cell culture plates. Hybrid clones were selected by growing in HAT medium (RPMI <NUM> culture medium supplemented with <NUM>% fetal calf serum and HAT-Supplement). After two weeks the HAT medium is replaced with HT Medium for three passages followed by returning to the normal cell culture medium.

The cell culture supernatants were primary screened for antigen specific IgG antibodies three weeks after fusion. The positive tested microcultures were transferred into <NUM>-well plates for propagation. After retesting, the selected cultures were cloned and recloned using the limiting-dilution technique and the isotypes were determined (see also <NPL>;<NPL>).

Antibodies were produced via standard antibody production methods (<NPL>) and purified via Protein A. The antibody purities were > <NUM>% based on SDS gel electrophoresis analysis.

Human Antibodies were produced by means of phage display according to the following procedure:
The human naive antibody gene libraries HAL7/<NUM> were used for the isolation of recombinant single chain F-Variable domains (scFv) against ADM peptide. The antibody gene libraries were screened with a panning strategy comprising the use of peptides containing a biotin tag linked via two different spacers to the ADM peptide sequence. A mix of panning rounds using non-specifically bound antigen and streptavidin bound antigen were used to minimize background of non-specific binders. The eluted phages from the third round of panning have been used for the generation of monoclonal scFv expressing E. coli strains. Supernatant from the cultivation of these clonal strains has been directly used for an antigen ELISA testing (see also <NPL>;<NPL>).

Positive clones have been selected based on positive ELISA signal for antigen and negative for streptavidin coated micro titer plates. For further characterizations the scFv open reading frame has been cloned into the expression plasmid pOPE107 (<NPL>), captured from the culture supernatant via immobilized metal ion affinity chromatography and purified by a size exclusion chromatography.

To determine the affinity of the antibodies to ADM, the kinetics of binding of ADM to immobilized antibody was determined by means of label-free surface plasmon resonance using a Biacore <NUM> system (GE Healthcare Europe GmbH, Freiburg, Germany). Reversible immobilization of the antibodies was performed using an anti-mouse Fc antibody covalently coupled in high density to a CM5 sensor surface according to the manufacturer's instructions (mouse antibody capture kit; GE Healthcare) (<NPL>).

The monoclonal antibodies were raised against the below depicted ADM regions of human and murine ADM, respectively. The following table represents a selection of obtained antibodies used in further experiments. Selection was based on target region:.

The generation of Fab and F(ab)<NUM> fragments was done by enzymatic digestion of the murine full length antibody NT-M. Antibody NT-M was digested using a) the pepsin-based F(ab)<NUM> Preparation Kit (Pierce <NUM>) and b) the papain-based Fab Preparation Kit (Pierce <NUM>). The fragmentation procedures were performed according to the instructions provided by the supplier. Digestion was carried out in case of F(ab)<NUM>-fragmentation for <NUM> at <NUM>. The Fab-fragmentation digestion was carried out for <NUM>, respectively.

The immobilized papain was equilibrated by washing the resin with <NUM> of Digestion Buffer and centrifuging the column at <NUM> × g for <NUM> minute. The buffer was discarded afterwards. The desalting column was prepared by removing the storage solution and washing it with digestion buffer, centrifuging it each time afterwards at <NUM> × g for <NUM> minutes. <NUM> of the prepared IgG sample where added to the spin column tube containing the equilibrated Immobilized Papain. Incubation time of the digestion reaction was done for <NUM> on a tabletop rocker at <NUM>. The column was centrifuged at <NUM> × g for <NUM> minute to separate digest from the Immobilized Papain. Afterwards the resin was washed with <NUM> PBS and centrifuged at <NUM> × g for <NUM> minute. The wash fraction was added to the digested antibody that the total sample volume was <NUM>. The NAb Protein A Column was equilibrated with PBS and IgG Elution Buffer at room temperature. The column was centrifuged for <NUM> minute to remove storage solution (contains <NUM>% sodium azide) and equilibrated by adding <NUM> of PBS, centrifuge again for <NUM> minute and the flow-through discarded. The sample was applied to the column and resuspended by inversion. Incubation was done at room temperature with end-over-end mixing for <NUM> minutes. The column was centrifuged for <NUM> minute, saving the flow-through with the Fab fragments. (References: <NPL>. ; <NPL>; <NPL>. ; <NPL>; <NPL>;<NPL>;<NPL>).

The immobilized Pepsin was equilibrated by washing the resin with <NUM> of Digestion Buffer and centrifuging the column at <NUM> × g for <NUM> minute. The buffer was discarded afterwards. The desalting column was prepared by removing the storage solution and washing it with digestion buffer, centrifuging it each time afterwards at <NUM> × g for <NUM> minutes. <NUM> of the prepared IgG sample where added to the spin column tube containing the equilibrated Immobilized Pepsin. Incubation time of the digestion reaction was done for <NUM> on a tabletop rocker at <NUM>. The column was centrifuged at <NUM> × g for <NUM> minute to separate digest from the Immobilized Papain. Afterwards the resin was washed with <NUM> PBS and centrifuged at <NUM> × g for <NUM> minute. The wash fraction was added to the digested antibody that the total sample volume was <NUM>. The NAb Protein A Column was equilibrated with PBS and IgG Elution Buffer at room temperature. The column was centrifuged for <NUM> minute to remove storage solution (contains <NUM>% sodium azide) and equilibrated by adding <NUM> of PBS, centrifuge again for <NUM> minute and the flow-through discarded. The sample was applied to the column and resuspended by inversion. Incubation was done at room temperature with end-over-end mixing for <NUM> minutes. The column was centrifuged for <NUM> minute, saving the flow-through with the Fab fragments. (References: <NPL>; <NPL>;<NPL>;<NPL>; <NPL>; <NPL>;<NPL>; <NPL>; <NPL>; <NPL>; <NPL>).

The antibody fragment was humanized by the CDR-grafting method (<NPL>).

The following steps where executed to achieve the humanized sequence:.

Annotation for the antibody fragment sequences (SEQ ID No.: <NUM> to <NUM>): bold and underlined are the CDR <NUM>, <NUM>, <NUM> in numeric order from the N-terminus to the C-terminus; italic are constant regions; hinge regions are highlighted with bold, underlined letters and the histidine tag at the C-terminus with bold and italic letters.

A randomized, double-blind, placebo-controlled phase I study in healthy male volunteers during experimental endotoxemia with single, escalating per group doses of Adrecizumab administered as i. infusion over a <NUM> hour period. A continuous LPS (lipopolysaccharide) model was used to induce experimental endotoxemia; administration of LPS in an initial bolus of <NUM> ng/kg followed by continuous infusion at <NUM> ng/kg/h for <NUM> hours. Subject will receive one course of treatment with study medication (Adrecizumab or placebo), <NUM> hour after start of LPS administration. The model of systemic inflammation has been recently described (<NPL>).

Adrecizumab was administrated as single infusion over a <NUM> hour period in a dose of <NUM>/kg and was increased up to <NUM> and <NUM>/kg in subsequent treatment groups (see Table <NUM>).

The study population consisted of healthy young male volunteers. To be included into the trial, subjects had to meet all inclusion criteria and none of the exclusion criteria.

To achieve a controlled inflammatory state, subjects received lipopolysaccharide (LPS, U. reference endotoxin [Escherichia coli O:<NUM>, List Biological Laboratories Inc. , Campbell, California, US]) intravenously.

The pre-dose blood samples were drawn on T=<NUM> (baseline). Vital signs were measured after <NUM> of resting in supine or semi-supine position. Schedule for blood samples and control visits (see table <NUM> for more detailed information):.

The following hematology parameters were analyzed: Hemoglobin, Hematocrit, RBC, MCV, MCH, MCHC, WBC, Platelets, Differential blood count.

Blood samples were collected in serum separation tubes and the following biochemistry parameters were analyzed: CRP, AF, ALT, AST, γ-GT, Creatine kinase, LDH, Urea-N, Creatinine, Sodium, Potassium, PT and APTT.

Primary endpoint variables are: Adverse Events; vital signs during the first <NUM> hours after Adrecizumab administration and at follow-up periods (T=<NUM> hours, T=<NUM> days, T=<NUM> days, T=<NUM> days, T=<NUM> days, T=<NUM> days), e.g. heart rate, blood pressure, oxygen saturation, temperature; Local tolerability at site of i. infusion; safety laboratory parameters (Hb, Ht, leukocytes, thrombocytes, leukocyte differential blood count, sodium, potassium, creatinine, urea, alkaline phosphatase, ALT, AST, GGT, CK, CRP, PT, APTT); <NUM>-lead electrocardiogram (ECG), <NUM> and <NUM> hours after Adrecizumab administration.

Secondary endpoints are: Pharmacokinetics of Adrecizumab during experimental endotoxemia (including AUC, Cmax, Terminal T1/<NUM>, Cl, V),; plasma levels of inflammatory mediators on the endotoxemia day (including but not limited to TNFα, IL-<NUM>, IL-<NUM>, IL-<NUM>); Symptom Illness score; Optional: Kidney damage markers (including but not limited to creatinine clearance, pro-Enkephalin and KIM-<NUM>).

No Serious Adverse Events (SAEs) or Suspected Unsuspected Serious Adverse Reactions (SUSARs) were observed during the complete duration of the study, including the <NUM> day follow-up period.

The Illness Score drops quicker under Adrecizumab treatment compared to placebo.

Both, <NUM> and <NUM>/kg have a significantly lower illness score than placebo between the time points at <NUM> (T210) to <NUM> (T450) (p=<NUM> and <NUM>, respectively; GLM repeated measures) (<FIG>).

<FIG> shows the distribution of the illness scores by treatment arm at T270 (time point <NUM>) and T300 (time point <NUM>), respectively (<FIG>)) supporting the surprisingly beneficial effect of the antibody of the present invention.

The contribution of single illness score components to the effect is illustrated in <FIG>) nausea, <FIG>) headache, <FIG>) muscle aches, <FIG>) back pain, and <FIG>) shivering. All symptoms shown contribute significantly to the effect. Vomiting occurred only in two patients once each and could not be analyzed.

<FIG> shows an alternative evaluation. Here another score, the Sickness score was evaluated using the single question regarding the overall sickness, on a scale from <NUM> to <NUM>. Again, the sickness score drops quicker in anti-ADM-antibody (Adrecizumab)-treated patients.

The study was conducted in healthy male subjects as a randomized, double-blind, placebo-controlled, study with single escalating doses of NT-H antibody administered as intravenous (i. ) infusion in <NUM> sequential groups of <NUM> healthy male subjects each (1st group <NUM>,<NUM>/kg, 2nd group <NUM>/kg, 3rd group <NUM>/kg) of healthy male subjects (n=<NUM> active, n = <NUM> placebo for each group).

The main inclusion criteria were written informed consent, age <NUM><NUM> - <NUM> years, agreement to use a reliable way of contraception and a BMI between <NUM> and <NUM>/m<NUM>.

Subjects received a single i. dose of NT-H antibody (<NUM>/kg; <NUM>/kg; <NUM>/kg) or placebo by slow infusion over a <NUM>-hour period in a research unit.

The baseline ADM-values in the <NUM> groups did not differ. Median ADM values were <NUM> pg/mL in the placebo group, <NUM> pg/mL in the first treatment group (<NUM>/kg), <NUM> pg/mL in second treatment group (<NUM>/kg) and <NUM> pg/mL in the third treatment group (<NUM>/mL).

The results show, that ADM-values rapidly increased within the first <NUM> hours after administration of NT-H antibody in healthy human individuals, then reached a plateau and slowly declined (<FIG>).

Claim 1:
Anti-adrenomedullin (ADM) antibody or an anti-adrenomedullin antibody fragment for use in therapy or prevention of symptoms of illness in a subject in need thereof, wherein
the subject is human, and
wherein the symptoms of illness are selected from the group of nausea, headache, muscle aches, back pain, and/or shivering,
wherein the illness is characterized by an amount of C-reactive protein (CRP) in a sample which is ≥ <NUM>/L, and/or wherein the amount of TNF in a sample is ≥ <NUM> pg/ml, and/or wherein the amount of bio-ADM in a sample is ≥ 43pg/ml, and wherein said sample is selected from the group consisting whole blood, plasma, serum, and
wherein said antibody or fragment is a humanized monoclonal antibody or fragment that binds to ADM or a fragment thereof,
wherein the heavy chain comprises the sequences:
CDR1: SEQ ID NO: <NUM>
GYTFSRYW
CDR2: SEQ ID NO: <NUM>
ILPGSGST
CDR3: SEQ ID NO: <NUM>
TEGYEYDGFDY
and wherein the light chain comprises the sequences:
CDR1: SEQ ID NO: <NUM>
QSIVYSNGNTY
CDR2:
RVS
CDR3: SEQ ID NO: <NUM>
FQGSHIPYT.