WO2008127298A2 - Staphylococcal enterotoxin b peptide compositions and methods of use - Google Patents

Abstract

The present invention relates to bacterial peptides, specifically Staphylococcus Enterotoxin B (SEB) peptides that have therapeutic use. The invention further relates to the use of SEB peptides in the diagnosis and therapy of diseases associated with cell proliferation.

Description

STAPHYLOCOCCAL ENTEROTOXIN B PEPTIDE COMPOSITIONS

AND METHODS OF USE RELATED APPLICATIONS This application claims the benefit of US Provisional Application No

60/853,906, filed October 24, 2006. The entire contents of the aforementioned application are hereby incorporated herein by reference.

FIELD OF THE INVENTION The present invention relates to bacterial peptides, specifically Staphylococcus

Enterotoxin B (SEB) peptides that have therapeutic use. The invention further relates to the use of SEB peptides in the diagnosis and therapy of diseases associated with cell proliferation.

BACKGROUND OF THE INVENTION

There are many known hyperproliferative disorders, in which cells of various tissues and organs exhibit aberrant patterns of growth, proliferation, migration, signaling, senescence, and death. While a number of treatments have been developed to address some of these diseases, many still remain largely untreatable with existing technologies, while in other cases, while treatments are available, they are frequently less than optimal and are seldom provide a cure. For example, therapy for cancer has largely involved the use of radiation, surgery and chemotherapeutic agents. However, results with these treatment modalities, while beneficial in some tumors, has had marginal or no effect in most. Furthermore, current therapeutic agents usually involve significant drawbacks for the patient in the form of toxicity and severe side effects. Of interest is a treatment for cancer or other proliferative diseases or disorders which have antiproliferative or apoptotic activity in doses that did not induce intolerable or lasting side effects.

Therefore, the need exists for new and more effective treatment modalities in diseases and conditions, particularly in proliferative diseases, which will reduce toxicity and benefit the patient. SUMMARY

The present invention is based on the discovery that Staphylococcus Enterotoxin B (SEB) peptides and other biologically active derivatives of SEB, exhibit antiproliferative and apoptotic effects. The invention further provides SEB peptides that have proliferative effects. The present invention provides bacterial peptides, specifically Staphylococcus Enterotoxin B (SEB) peptides that have therapeutic use.

In a first aspect, the invention features a method of increasing cell proliferation comprising administering a Staphylococcus enterotoxin B (SEB) peptide capable of increasing cell proliferation in a cell that expresses digalactosylceramide receptor.

In a related aspect, the invention features a method of increasing cell proliferation in a subject in need thereof comprising administering to the subject a SEB peptide capable of increasing cell proliferation in a cell that expresses digalactosylceramide receptor.

In one embodiment, the cell that expresses digalactosylceramide receptor is selected from the group consisting of brain cells, neuronal cells, lymphocytes, leukocytes, liver cells, inner ear cells, spleen cells, pancreatic cells, urinary tract cells, bone marrow spinal cord cells, spinal root cells, skin cells, and conjunctival vessels. In another aspect, the invention features a method of increasing kidney cell proliferation in a subject comprising administering to the subject a SEB peptide capable of increasing cell proliferation, thereby stimulating kidney cell proliferation.

In one embodiment, the subject has undergone kidney transplantation or kidney resection. In another embodiment, the method stabilizes, reduces the symptoms of, or ameliorates a disease or disorder characterized by kidney dysfunction.

In a further aspect, the invention features a method of increasing immune cell proliferation comprising administering a SEB peptide capable of increasing cell proliferation, thereby stimulating immune cell proliferation. In a related aspect, the invention features a method of increasing immune cell proliferation in a subject comprising administering to the subject a SEB peptide capable of increasing cell proliferation, thereby stimulating immune cell proliferation.

In one aspect, the immune cells are T-lymphocytes. In another aspect, the method stabilizes, reduces the symptoms of, or ameliorates a disease or disorder characterized by abnormal immune cell proliferation.

In still another embodiment, the disease or disorder is anaplastic anemia or Fabry's Disease.

In one embodiment of any one of the above aspects, the SEB peptide comprises the amino acid sequence CVFSKKTNDINSHQTDKRKT (SEQ ID NO: 3) or fragments thereof. In a related embodiment, the SEB peptide consists of the amino acid set forth as SEQ ID NO: 3. In another embodiment, the SEB peptide further comprises one or more therapeutic agents. In a further embodiment, the one or more therapeutic agents is selected from the group consisting of: protein synthesis modulators, apoptosis modulators, nitric oxide modulators, bactericides, a fungicides, anti neoplastic drugs, anti thrombotic drugs, hypochoesterolemic drugs, hypotriglyceridemic drugs, hypoglycemic drugs, anti-allergic drugs, antibiotics, tissue, anti-inflammatory drugs, nerve regenerating drugs, and anti-arthritis drugs.

In one aspect, the invention features a method of decreasing cell proliferation comprising administering a SEB peptide capable of decreasing cell proliferation in a cell that expresses digalactosylceramide receptor. In a related embodiment, the invention features a method of decreasing cell proliferation in a subject in need thereof comprising administering to the subject a SEB peptide capable of decreasing cell proliferation in a cell that expresses digalactosylceramide receptor.

In one embodiment, the method further comprises increasing apoptosis. In another aspect, the invention features a method of promoting apoptosis comprising administering a SEB peptide capable of promoting apoptosis in a cell that expresses digalactosylceramide receptor.

In a related aspect, the invention features a method of promoting apoptosis in a subject in need thereof comprising administering to the subject a SEB peptide capable of promoting apoptosis in a cell that expresses digalactosylceramide receptor.

In one embodiment, the cell expresses elevated levels of digalactosylceramide compared to a control cell.

In a further embodiment of any one of the above aspects, the cell that expresses digalactosylceramide receptor is a mammalian cell. In a related embodiment, the mammalian cell is selected from the group consisting of: a selected from the group consisting of: tumor cells, kidney cells, neuronal cells, lymphocytes, inner ear cells, spleen cells and pancreatic cells.

In another aspect, the invention features a method of treating a tumor in a subject comprising administering to the subject a SEB peptide capable of promoting apoptosis or decreasing cell proliferation in a tumor that expresses digalactosylceramide receptor, thereby treating the tumor in a subject.

In one embodiment, the tumor expresses elevated levels of digalactosylceramide compared to a normal tissue. In another aspect, the invention features a method of treating a lipid metabolic disorder in a subject comprising administering to the subject a SEB peptide capable of promoting apoptosis or decreasing cell proliferation in cells that express high levels of digalactosylceramide, thereby treating the lipid metabolic disorder the subject.

In a particular embodiment, the lipid metabolic disorder is selected from the group consisting of: Fabry's disease, Metachromatic Leukodystrophy, GM2 Gangliosidosis, Tay-Sachs disease and chronic myelogenous leukemia.

In another particular aspect, the invention features a method of treating a neural disease or disorder or a kidney disease or disorder in a subject comprising administering to the subject a SEB peptide capable of promoting apoptosis or decreasing cell proliferation in neural tissue or kidney tissue that expresses digalactosylceramide receptor, thereby treating a neural or kidney disease or disorder in the subject.

In one embodiment, the neural tissue or kidney tissue expresses high levels of digalactosylceramide compared to a control cell. In another embodiment, the neural or kidney disease or disorder is selected from autism or glomerular nephritis.

In one embodiment of any of the above aspects, the SEB peptide comprises the amino acid sequence RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID NO: 5) or fragments thereof, or KKKVTAQEL (SEQ ID NO: 7) or fragments thereof. In another embodiment of any of the above aspects, the SEB peptide consists of the amino acid set forth as SEQ ID NO: 5 or SEQ ID NO: 7.

In one particular embodiment, the SEB peptide further comprises one or more therapeutic agents. In a further embodiment, the one or more therapeutic agents is selected from the group consisting of: protein synthesis modulators, apoptosis modulators, nitric oxide modulators, bactericides, a fungicides, anti neoplastic drugs, anti thrombotic drugs,hypochoesterolemic drugs, hypotriglyceridemic drugs, hypoglycemic drugs, anti-allergic drugs, antibiotics,tissue, anti-inflammatory drug, nerve regenerating drugs, and anti-arthritis drugs. In still a further embodiment, the apoptosis modulator is clodronate.

In another related embodiment, the one or more therapeutic agents is an anticancer agent. In a further embodiment, the anticancer agent is selected from the group consisting of: a chemotherapeutic agent, a peptide toxin, and a protein toxin. In a particular embodiment, the anticancer agent is rapamycin. In another embodiment of any of the above aspects, the peptide further comprises an imaging agent. In one embodiment, the imaging agent is selected from a radiolabel or a fluorescent label.

In another aspect, the invention features a method of targeting an agent to a cell that expresses digalactosylceramide receptor comprising administering an SEB peptide comprising one or more agents to a cell that expresses digalactosylceramide receptor.

In a further embodiment, the invention features a method of targeting an agent to a tumor or an atherosclerotic plaque that expresses digalactosylceramide receptor in a subject comprising administering an SEB peptide comprising one or more agents to a tumor or an atherosclerotic plaque that expresses digalactosylceramide receptor.

In another embodiment, the SEB peptide comprises the amino acid sequence ENENSFWYAMMPAPGDKFDQSKYLMMYNDN (SEQ ID NO: 6) or fragments thereof. In a further embodiment, the SEB peptide consists of the amino acid set forth as SEQ ID NO: 6. In another embodiment of the method, the one or more agents is a therapeutic agent selected from the group consisting of: protein synthesis modulators, apoptosis modulators, nitric oxide modulators, bactericides, a fungicides, anti neoplastic drugs, anti thrombotic drugs,hypochoesterolemic drugs, hypotriglyceridemic drugs, hypoglycemic drugs, anti-allergic drugs, antibiotics,tissue, anti-inflammatory drugs, nerve regenerating drugs, and anti-arthritis drugs.

In a particular embodiment, the agent is an anticancer agent. In a particular embodiment, the anticancer agent is selected from the group consisting of: a chemotherapeutic agent, a peptide toxin, and a protein toxin. In a more particular embodiment, the anticancer agent is rapamycin. In a further embodiment, the agent is an imaging agent. In one embodiment, the imaging agent is selected from a radiolabel or a fluorescent label.

In another aspect, the invention features a method of detecting a tumor or an atherosclerotic plaque that expresses digalactosylceramide receptor in a subject comprising administering an SEB peptide comprising an imaging agent to a tumor or an atherosclerotic plaque that expresses digalactosylceramide receptor in a subject.

In one embodiment, the method is used to determine course of treatment.

In another embodiment, the subject is treated with an anticancer or antiproliferative agent. In a further embodiment, the method is used to determine prognosis.

In another embodiment, the method is used to determine regression in tumor size or atherosclerotic plaque size after treatment with an anticancer or antiproliferative agent.

In another aspect, the invention features a method of treating an infection caused by a bacterial toxin in a subject, the method comprising administering to the subject a SEB peptide, wherein the peptide is capable of binding to a digalactosylceramide (DAG) receptor, thereby treating infection caused by a bacterial toxin in a subject.

In one embodiment of the method, the SEB peptide comprises the amino acid sequence ENENSFWYAMMPAPGDKFDQSKYLMMYNDN (SEQ ID NO: 6).

In another embodiment, the SEB peptide consists of the amino acid set forth as SEQ ID NO: 6.

In another aspect, the invention features a method of inhibiting activity of a digalactosylceramide (DAG) receptor in a subject comprising administering a composition selected from: an antibody, a nucleic acid or an oligomer that prevents activation of the DAG receptor.

In one embodiment, the antibody has specificity for the amino acid sequence selected from the group comprising

RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID NO: 5) and KKKVTAQEL (SEQ ID NO: 7).

In another embodiment, the nucleic acid is an inhibitory nucleic acid selected from: antisense, siRNA, shRNA, aptamers, PNA oligomers, and ribozymes. In a further embodiment, the oligomer is at least a 5-mer of the amino acid sequence comprising RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID NO: 5).

In another embodiment, the subject is suffering from an infection caused by a bacterial toxin.

In another aspect, the invention features an infection caused by a bacterial toxin in a subject, the method comprising administering a composition selected from: an antibody, a nucleic acid or an oligomer that prevents activation of the DAG receptor, wherein the peptide is capable of binding to a digalactosylceramide (DAG) receptor, thereby treating infection caused by a bacterial toxin in a subject.

In one embodiment, the bacterial toxin is from a Gram-negative bacteria. In a further embodiment, the bacterial toxin is selected from Escherichia coli or Psuedomonas aeruginosa. In a particular embodiment, the E.coli bacterial toxin is verotoxin or Shiga toxin. In another embodiment, the Escherichia coli or Pseudomonas aeruginosa binds a glycolipid receptor selected from a digalactosylceramide receptor or a Globotriosylceramide Receptor (GbOse3Cer).

In another particular embodiment of the method, the infection caused by a bacterial toxin is localized to the lung, kidney, spleen, pancreas or gastroinstestinal tract. In an embodiment of any of the above-described aspects, the subject is a human.

In another aspect, the invention features a peptide comprising the amino acid sequence CVFSKKTNDINSHQTDKRKT (SEQ ID NO: 3) or fragments thereof, wherein the peptide is capable of increasing cell proliferation in a cell that expresses digalactosylceramide receptor.

In one embodiment, the peptide comprises the amino acid set forth as SEQ ID NO: 3.

In another aspect, the invention features a peptide comprising the amino acid sequence RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID NO: 5) or fragments thereof, or KKKVTAQEL (SEQ ID NO: 7) or fragments thereof, wherein the peptide is capable of decreasing cell proliferation in a cell that expresses digalactosylceramide receptor.

In one embodiment, the peptide promotes apoptosis. In another embodiment, the peptide comprises the amino acid set forth as SEQ ID NO: 5 or 7. In another aspect, the invention features a peptide comprising the amino acid sequence ENENSFWYAMMPAPGDKFDQSKYLMMYNDN (SEQ ID NO: 6) or fragments thereof, wherein the peptide is capable of binding to a digalactosylceramide (DAG) receptor. In one embodiment, the peptide comprises the amino acid set forth as SEQ ID

NO: 6.

In another embodiment of any of the above aspects, the peptide is derived from Staphylococcus enterotoxin B (SEB). In another embodiment of any of the above-mentioned aspects, the peptide is at least a 5-mer oligomeric fragment. In another aspect, the invention features a method of making one or more antibodies that bind at least one staphylococcal enterotoxin receptor, the method comprising administering to a mammal an amount of a peptide of any one of the aspects as described herein, the amount being sufficient to elicit production of one or more antibodies. In one embodiment, the peptide further comprises one or more therapeutic agents.

In a further embodiment, the therapeutic agent is covalently linked.

In another embodiment, the one or more therapeutic agents is selected from the group consisting of: protein synthesis modulators, apoptosis modulators, nitric oxide modulators, bactericides, a fungicides, anti neoplastic drugs, anti thrombotic drugs,hypochoesterolemic drugs, hypotriglyceridemic drugs, hypoglycemic drugs, anti-allergic drugs, antibiotics,tissue, anti-inflammatory drug, nerve regenerating drugs, and anti-arthritis drugs.

In one embodiment, the one of more therapeutic agents is an anticancer agent. In another embodiment, the anticancer agent is selected from the group consisting of: a chemotherapeutic agent, a peptide toxin, and a protein toxin.

In another particular embodiment, the chemotherapeutic agent is rapamycin.

In a further embodiment of the above described aspects, the peptide further comprises an imaging agent. In a further embodiment, the imaging agent is covalently linked.

In a particular embodiment, the imaging agent is selected from a radiolabel or a fluorescent label. In another aspect, the invention features a pharmaceutical composition comprising the peptide of any one of the aspects as described herein, and a pharmaceutically acceptable carrier.

In other another aspect, the invention features kits for use according to any one of the methods of the aspects as described herein, and instructions for use.

Other aspects of the invention are described infra. *

BRIEF DESCRIPTION OF THE FIGURES

Figure 1 illustrates the location of SEB Peptides. The highlighted amino acid residues highlight the location of experimental peptides used in this study. The sequence of amino acids can also be found in Table 1. All structures were obtained from the Protein Data Bank of the Research Collaborator for Structural Bioinformatics. Downloaded files were subsequently manipulated using the RasMol program. The coordinates are based upon the original publication by Papageorgiou, et al (13).

Figure 2 are five graphs that illustrate the binding of ¹²⁵I-SEB peptides by cultured human proximal tubular cells. PT cells grown as described in the Materials and Methods section. On the sixth day of culture, medium was replaced with that containing lipoprotein deficient serum (LPDS) and incubated for 24 h. Subsequently, fresh medium containing LPDS and 0-5 μg/ml of ¹²⁵I-SEB peptide was added. To another set of dishes, prior to the addition of ¹²⁵I-toxin, 20 fold excess of corresponding unlabeled toxin was added and incubation was continued for 2 h at 37°C. Next, medium was removed and the cells were washed ten times with ice-cold PBS. Samples were then solubilized overnight with 1 N NaOH and cell associated radioactivity and protein content was measured. All assays were performed in duplicate dishes from two batches of PT cells and analyzed in duplicate. Specific binding (binding in the absence of unlabeled toxin-binding in the presence of unlabeled toxin) was calculated and plotted.

Figure 3 are four graphs that illustrate the effect of antibody against SEB peptides, human kidney glycosphingolipids and endoglycoeramidase on the binding of ¹²⁵I-SEB in PT cells. To cultured PT cells increasing concentrations of SEB peptide antibodies (closed circle), glycosphingolipids (open circle) and endoglycoceramidase (open triangle) were added prior to the addition of 1 ~g/ml of ¹²⁵I-SEB peptide. Incubation was carried out for 4 h at 37°C and the specific binding of ¹²⁵I-SEB peptide to PT cells was measured. The data represents average values obtained from duplicate dishes from two batches of PT cells analyzed in duplicate.

Figure 4 (A - C) are three graphs that illustrate the effect of SEB peptide concentration on PT cell proliferation. Confluent culture of PT cells grown in 96 well trays were incubated in serum-free medium for 24 h. Next, medium containing 1 mgl/ ml lipoprotein-deficient serum and increasing concentrations of SEB peptides (0.15- 2.5 μg/ml) were added. Following incubation for 22 h (3H)thymidine (5 μCilml) was added and incubation continued for another 2 h. Next, the cells were washed 5 times with PBS and incorporation of (3H)thymidine into DNA was measured. A parallel set of dishes was trypsinized, stained with trypan blue and subject to viable cell counting employing hemocytometer and a light microscope. The data obtained from three separate experiments and 6 micro titer wells each were analyzed. Open bars-

(3H)thymidine incorporation; solid bars-viable cell count. A - SEB 130-160; B - SEB 93-112; C - SEB 191-220. Proliferation data from SEB alone (2 separate sources): Source #1 SEB = 6894 +/- 416; Source^;#2 SEB = 8987 +/- 2698.

Figure 5 (A - D) are four panels that illustrate the effect of SEB and SEB peptides on PT cell morphology. The protocol of this experiment was identical to the legend described in Figure 4 above except cells were incubated with Media Alone (Figure 5 A), SEB (Figure 5 B), SEB 93-112 (Figure 5 C) and 130-160 (Figure 5 D) at a concentration of 1 μg/ml for 24 h and then photographed.

Figure 6 is two panels (upper, A - F, and lower) that illustrate the effect of SEB and SEB peptides on apoptosis. Confluent culture of human kidney proximal tubular cells grown on glass cover slips were switched to 1% serum containing medium. SEB, SEB peptide 130-160 (1 and 2 μg/ml medium) and SEB peptide 93- 112 (2 μg/ml) were added. To other dishes SEB and SEB peptide 130- 160 antibodies (2 μl/ml) were added first followed by the addition of SEB and SEB peptide 130-160. After incubation for 24 h the number of cells shed in the medium were counted following staining with trypan blue. Cells on the glass cover slips were fixed with ethanol-acetic acid (3:2 v/v) for 10 min, washed with PBS and stained with DAPI reagent. Representative areas were photographed (Figure 6 A). A - Control (no apoptotic cells were observed); B -SEB(2 μg /ml); C-SEB 130-160 (1 μg /ml); D - SEB 93-112; E - SEB 30-160(1 μg /ml)+SEB Ab; F - SEB130-160(2 μg /ml)+ SEB 130- 160 Ab. Five different areas on the microscope field were counted for apoptotic and nonapoptotic cells and percentage of apoptotic cells plotted (Figure 6 B). 1- Control; 2- SEB (1 μg/ ml); 3- SEB (2 μg/ ml); 4- SEB 130 - 160 (1 μg/ ml); 5- SEB 130 - 160 (2 μg/ ml); 6- SEB 93 - 112 (2 μg/ ml); 7- SEB Ab (lμl/ml); 8- SEB (2 μg/ ml) + SEB Ab (lμl/ml); 9- SEB 130 - 160 (1 μg/ ml) + SEB Ab (lμl/ml); 10- SEB 130 - 160 Ab (lμl/ml); 11- SEB (2 μg/ ml) + SEB 130 - 160 Ab (2μl/ml); 12- SEB 130 - 160 (2 μl/ ml) + SEB 130 - 160Ab (lμl/ml). SEB and SEB peptide 130- 160 dose-dependently induced apoptosis in PT cells. Antibody against SEB completely reversed SEB and/or SEB peptide 130-160-induced apoptosis, whereas antibody against SEB peptide 130-160 mitigates SEB and SEB peptide 130-160 induced apoptosis.

Figure 7 (A and B) illustrates the effect of SEB and SEB peptide on the activity of neutral sphingomyelinase (N-SMase) (Figure 7 A) and the level of ceramide and sphingomyelin (Figure 7 B). Cells grown in 6 well trays were incubated with SEB and SEB peptide 130-160 (1 μg/ml) for 15 min at 37°C. Cells were harvested and the activity of N-SMase was measured using (14C) sphingomyelin as substrate (A). Cell were also metabolically labeled for 24 h with (14C)serine (5 μCi/ml), washed and then incubated with fresh medium containing SEB (1 μg/ml) for 15 min at 37°C. Next, medium was removed, cells were extracted with hexane- isopropanol (3;2 v/v) for 10 min. The total lipid extract was dried in nitrogen and fractionated by HPTLC. Gel area corresponding to ceramide and sphingomyelin were scraped and radioactivity was measured.

Figure 8 shows SEQ ID NOs: 1 - 7.

DEFINITIONS

Unless defined otherwise, all technical and scientific terms used herein have the meaning commonly understood by a person skilled in the art to which this invention belongs. The following references provide one of skill with a general definition of many of the terms used in this invention: Singleton et al, Dictionary of Microbiology and Molecular Biology (2nd ed. 1994); The Cambridge Dictionary of Science and Technology (Walker ed., 1988); The Glossary of Genetics, 5th Ed., R. Rieger et al. (eds.), Springer Verlag (1991); and Hale & Marham, The Harper Collins Dictionary of Biology (1991). As used herein, the following terms have the meanings ascribed to them unless specified otherwise.

In this disclosure, "comprises," "comprising," "containing" and "having" and the like can have the meaning ascribed to them in U.S. Patent law and can mean " includes," "including," and the like; "consisting essentially of or "consists essentially" likewise has the meaning ascribed in U.S. Patent law and the term is open-ended, allowing for the presence of more than that which is recited so long as basic or novel characteristics of that which is recited is not changed by the presence of more than that which is recited, but excludes prior art embodiments.

By the term "agent" is meant a polypeptide, polynucleotide, or fragment, or analog thereof, small molecule, or other biologically active molecule. By the term "amino acid" is meant to refer to either natural and/or unnatural or synthetic amino acids, including glycine and both D or L optical isomers, and amino acid analogs and peptidomimetics. A peptide of three or more amino acids is commonly called an oligopeptide if the peptide chain is short. If the peptide chain is long (e.g., greater than about 10 amino acids), the peptide is commonly called a polypeptide or a protein. While the term "protein" encompasses the term "polypeptide", a "polypeptide" may be a less than full-length protein. By the term "apoptosis" is meant programmed cell death. By the term "antibody" is meant to refer to any immunoglobulin, including antibodies and fragments thereof, that binds a specific epitope. The term encompasses polyclonal, monoclonal, and chimeric antibodies (e.g., bispecific antibodies). An "antibody combining site" is that structural portion of an antibody molecule comprised of heavy and light chain variable and hypervariable regions that specifically binds antigen. Exemplary antibody molecules are intact immunoglobulin molecules, substantially intact immunoglobulin molecules, and those portions of an immunoglobulin molecule that contains the paratope, including Fab, Fab¹, F(ab')₂ and F(v) portions, which portions are preferred for use in the therapeutic methods described herein.

By the terms "cancer," "neoplasm," and "tumor," are used interchangeably and in either the singular or plural form, are meant to refer to cells that have undergone a malignant transformation that makes them pathological to the host organism. Primary cancer cells (that is, cells obtained from near the site of malignant transformation) can be readily distinguished from non-cancerous cells by well-established techniques, particularly histological examination. The definition of a cancer cell, as used herein, includes not only a primary cancer cell, but any cell derived from a cancer cell ancestor. This includes metastasized cancer cells, and in vitro cultures and cell lines derived from cancer cells. When referring to a type of cancer that normally manifests as a solid tumor, a "clinically detectable" tumor is one that is detectable on the basis of tumor mass; e.g., by procedures such as CAT scan, MR imaging, X-ray, ultrasound or palpation, and/or which is detectable because of the expression of one or more cancer-specific antigens in a sample obtainable from a patient.

By the term "chemotherapy" is meant the treatment of disease with chemical substances.

By the phrase "an effective amount" is meant the amount of an agent required to ameliorate the symptoms of a disease relative to an untreated patient. The effective amount of active agent(s) used to practice the present invention for therapeutic treatment of a neoplasia varies depending upon the manner of administration, the age, body weight, and general health of the subject. Ultimately, the attending physician or veterinarian will decide the appropriate amount and dosage regimen. Such amount is referred to as an "effective" amount.

By the terms a "therapeutically effective amount" is meant to refer to an amount sufficient to prevent, correct and/or normalize an abnormal physiological response. In one aspect, a "therapeutically effective amount" is an amount sufficient to reduce by at least about 30 percent, more preferably by at least 50 percent, most preferably by at least 90 percent, a clinically significant feature of pathology, such as for example, size of an ischemic region, size of a tumor mass, elevated blood pressure, fever or white cell count, etc.

By the term "inhibitory nucleic acid" is meant a single or double-stranded RNA, siRNA (short interfering RNA), shRNA (short hairpin RNA), or antisense RNA, or a portion thereof, or a mimetic thereof, that when administered to a mammalian cell results in a decrease (e.g., by 10%, 25%, 50%, 75%, or even 90- 100%) in the expression of a target gene. Typically, a nucleic acid inhibitor comprises or corresponds to at least a portion of a target nucleic acid molecule, or an ortholog thereof, or comprises at least a portion of the complementary strand of a target nucleic acid molecule.

By the term "antisense nucleic acid", it is meant a non-enzymatic nucleic acid molecule that binds to target RNA by means of RNA--RNA or RNA-DNA interactions and alters the activity of the target RNA (for a review, see Stein et al. 1993; Woolf et al, U.S. Pat. No.5, 849, 902).

By "Staphylococcal enterotoxin B (SEB)" is meant an exotoxin produced by the Staphylococcus aureus bacterium.

By the term "siRNA" refers to small interfering RNA; a siRNA is a double stranded RNA that "corresponds" to or matches a reference or target gene sequence. This matching need not be perfect so long as each strand of the siRNA is capable of binding to at least a portion of the target sequence. SiRNA can be used to inhibit gene expression, see for example Bass, 2001, Nature, 411, 428 429; Elbashir et al., 2001, Nature, 411, 494 498; and Zamore et al., Cell 101 :25-33 (2000). By the term "Gram-negative bacterial cell" is intended to include the art recognized definition. Typically, Gram-negative bacteria include Gluconobacter, Rhizobium, Bradyrhizobium, Alcaligenes, Rhodobacter, Rhodococcus. Azospirillum, Rhodospirillum, Sphingomonas, Burkholderia, Desulfomonas, Geospirillum, Succinomonas, Aeromonas, Shewanella, Halochromatium, Citrobacter, Escherichia, Klebsiella, Zymomonas (e.g., Zymomonas mobilis), Zymobacter (e.g., Zymobacter palmae), and Acetobacter (e.g., Acetobacter pasteurianus).

By "fragment" is meant a portion (e.g., at least 2, 3, 4, 5,10, 12, 14, 16, 20, 22, 24, 25, 28, 30, or more amino acids) of the peptides of the invention that retains the biological activity of the reference peptide, e.g. antiproliferative or apoptotic activity. By the term "inhibit" or "inhibiting" is meant decrease, suppress, attenuate, diminish, arrest, or stabilize the development or progression of a disease.

By the term "nucleic acid" is intended to include nucleic acid molecules, e.g., polynucleotides which include an open reading frame encoding a polypeptide, and can further include non-coding regulatory sequences, and introns. In addition, the terms are intended to include one or more genes that map to a functional locus. In addition, the terms are intended to include a specific gene for a selected purpose. The gene can be endogenous to the host cell or can be recombinantly introduced into the host cell, e.g., as a plasmid maintained episomally or a plasmid (or fragment thereof) that is stably integrated into the genome. By the term "subject" is meant vertebrates, preferably a mammal. Mammals include, but are not limited to, humans.

By the term "target site" is meant to refer to regions, aggregates, or populations of cells or tissues. A target site can be accessed in vitro or in vivo. By the term "rapamycin" is meant an immunosuppressive macrolide antibiotic.

DETAILED DESCRIPTION OF THE INVENTION

The present invention provides bacterial peptides, specifically Staphylococcus Enterotoxin B (SEB) peptides that have therapeutic use. The invention further relates to the use of SEB peptides in the diagnosis and therapy of diseases associated with cell proliferation and apoptosis.

I. Staphylococcal enterotoxin B (SEB) Staphylococcal enterotoxin B (SEB), a 28kDa exoprotein produced by gram- positive S. aureus has been well studied for its potent role as a T lymphocyte mitogen (7). SEB, along with the other enterotoxins, Toxic Shock Syndrome Toxin- 1, and a host of other viral and bacterial proteins have been termed superantigens (sAg). These molecules are able to bypass conventional antigen presentation and stimulate up to 20% of the host T-cell repertoire (7). SEB achieves this through extracellularly binding to the major histocompatability complex II (MHC-II) on antigen presenting cells while subsequently binding specific variable regions on the T-cell receptor (TcR) (6). This interaction initiates T cell proliferation with concomitant cytokine production. Mutational analysis of SEB suggests that the region of SEB implicated in MHC-II binding are residues 13-17 and 44-52 (9). Much of the pathophysiology after SEB exposure has been thought to occur via the massive production of inflammatory cytokines, tumor necrosis factor alpha (TNF-a) and interleukin 2 (IL-2) (7). Interestingly the emetic actions of the SE have been found to occur independent of mitogenic events. In fact a histidine substitution in SEA at position 61 showed that superantigenic and emetic activity can be separated (5). Further, the active region for emesis has been speculated to correspond to amino acid residues 113-126 (16) an area remote from the immunologically active regions. In addition to the discrepancy in explanation of emetic events, there is an increasing body of evidence suggesting that SEB has the ability to interact and induce lesions in several non- immunological mammalian tissues (1, 7, 15, 17). SEB has been studied as a potential biological agent of war, since it easily can be aerosolized, is very stable, and can cause widespread systemic damage, multiorgan system failure, and even shock and death when inhaled at very high dosages. Localization and metabolic turnover studies in experimental animals reveal that the kidney is a major site of toxin sequestration (2, 10-12). In vivo studies show that the clearance of 121-SEB is markedly altered by perturbations in renal blood flow (10, 14). Other studies indicate that SEB is predominantly confined (more than 75% of the injected dose) to proximal tubular (PT) cells (11). Later it was determined that SEB bound digalactosylceramide, a neutral glycosphingolipid found on human renal proximal tubular cells (3, 4).

H. PEPTIDES

In a preferred embodiment, one or more bacterially derived peptides from SEB are present in a therapeutic composition. These peptides can either be produced recombinantly, by chemical synthesis or purified from native sources, using methods known in the art. See, for example, Ranelli, D. M. et al., Proc. Natl. Acad. Sci. USA 82:850-854 (1985); Iandolo, J. J. Annu. Rev. Microbiol. 43:375 (1989); Kappler, J. W. et al., J Exp. Med 175:387 (1992); Rahim, A. et al., J Exp. Med. 180:615 (1994; Lando, P. A. et al., Cane. Immunol. Immunother. 33:231 (1991) Dohlsten, M. et al., 88:9287 (1991); Dohlsten, M. et al., Immunology 79:520 (1993); Dohlsten, M. Proc. Natl. Acad. Sci. USA (1994); Marrack, P. et al., Science 248:750 (1990); and Terman, D. S. et al., PCT Publication W091/10680 (1991).

Preferred peptides according to the invention are SEB peptides. However, peptides from other enterotoxins are envisioned by the instant invention, for example from Staphylococcus aureus enterotoxins A, C 1, C2, D or E (SEA, SEC 1, SEC2, SED, SEE). Other examples of other enterotoxins include, but are not limited to Streptococcus pyogenes toxins A and C (SPE-A and SPE-C; Staphylococcus aureus toxic shock syndrome-associated toxin (TSST-I); Staphylococcus aureus exfoliating toxins A and B (ETA and ETB) and Staphylococcus aureus alpha toxin. Also included are toxins from Mycoplasma arthritides and Yersinia enterocolitica. Various enterotoxins share differing degrees of immunological relatedness (Bergdoll, M. S. et al., Infect. Immun. 4: 593 (1971); Bergdoll, M. S., Enterotoxins. In: STAPHYLOCOCCIAND STAPHYLOCOCCI INFECTIONS, C. S. F. Easmon et al., eds, pp. 559-598, 1983, London, Academic Press; Freer, J. H. et, J Pharmacol. Pharm. Ther. 19:55 (1983). Immunologic cross-reactivity between SPE-A, SEB and SEC 1 suggests the presence of a conserved domain. SEA, SEB, SEC, SED, TSST-I and the pyrogenic exotoxins share considerable DNA and amino acid sequence homology. The enterotoxins, the pyrogenic exotoxins and TSST-I therefore appear to be evolutionarily related and all belong to a common generic group of proteins. SPE-A and SPE-C are about as similar to each of the Staphylococcal toxins as they are to each other. Exfoliative toxins have sizes similar to SEB and SEA and similar modes of action. They share several regions of sequence similarity to the Staphylococcal enterotoxins. Overall there are several stretches of protein having similarities throughout the total group of Staphylococcal enterotoxins, Streptococcal pyrogenic exotoxins and Staphylococcal exfoliative toxins. The structural homologies between the enterotoxins and the S. pyogenes, toxins, above, apparently are responsible for the identity of clinical responses to them. These toxins induce hypotension, fever, chills and septic shock in humans, apparently by inducing cytokines such as interleukin- 1 , interleukin-2, tumor necrosis factors, interferons and procoagulant activity which are the prime mediators of the clinical symptoms.

Previously, a series of thirteen synthetic SEB peptides (spanning the entire toxin sequence), have been used to evaluate their effects on T-cell proliferation in a culture system containing human peripheral blood lymphocytes incubated with a specific ratio of mononuclear cells (8). In this previous study four peptide regions were identified that inhibit SEB-induced peripheral blood mononuclear cell (PBMC) proliferation. The regions included sequences 1-30, 51-92 (such sequences have been related to the T-cell receptor site), 93-112 (a linear sequence corresponding to the 16 loop), and 130-160 (containing highly conserved sequence, KKKVT AOEL) (Figure 1). Moreover, neutralizing antibodies against the latter domain were capable of abrogating SEB-induced proliferation. The peptide 130-160 also inhibited the binding of 1251 -SEB to lymphocytes.

In another study investigating the functionality of the SEB 130-160 region, peptide KKKVTAOELD and anti-peptide sera to this region was observed to impressively block SEB movement across cultured gut monolayers. Further peptide and sera were also shown to block the movement of SEA, SEE, and to a lesser extent TSST-I (15). These studies reveal that the residues containing and surrounding the sequence, KKKVTAOEL(D), may not only be critical in SEB-induced T-cell proliferation but also in lesions of non-immunological tissue. This region of the molecule may of therapeutic interest.

Neutral sphingomyelinase is a cell membrane associated phospholipase that cleaves sphingomyelin to ceramide and phosphocholine. N-SMase has been implicated in a variety of cell systems to mediate the effects of cytokines such as TNF-a, IL-I, and interferon-gamma. The basic mechanism may involve the binding of these cytokines to receptors. This, in turn, activates the N-SMase. N-SMase then cleaves the sphingomyelin to ceramide (22). Ceramide, in turn, stimulates programmed cell death (23) presumably by activating nuclear factors, such as NFkB, pBcI and the ICE family (24).

Glycosphingolipids (GSLs) are integral components of cell membranes and serve as receptors for bacterial toxins (25). GSL are composed of sphingosine, fatty acids and sugars (26). Ceramide comprised of sphingosine and a fatty acid is the backbone of all GSL, to which monosaccharide units are attached. GSL are synthesized in the Golgi apparatus via sequential addition of monosaccharide units from nucleotide sugars to ceramide via specific glycosyltransferases (26). The GSL are then transported to various subcellular organelles. In mammalian cells lactosylceramide has been shown to be located within cytoplasmic membranes (21). In PT cells, GSL have been shown to be 14.

Glycosphingolipids (GSLs) are integral components of cell membranes and serve as receptors for bacterial toxins (25). GSL are composed of sphingosine, fatty acids and sugars (26). Ceramide comprised of sphingosine and a fatty acid is the backbone of all GSL, to which monosaccharide units are attached. GSL are synthesized in the Golgi apparatus via sequential addition of monosaccharide units from nucleotide sugars to ceramide via specific glycosyltransferases (26). The GSL are then transported to various subcellular organelles. In mammalian cells lactosylceramide has been shown to be located within cytoplasmic membranes (21). In PT cells, GSL have been shown to be localized with the apical and basolateral membranes (27). Whether the topology of a GSL is a determinant of its functional role as a receptor for bacterial or viral proteins is not known.

Kidney PT cells have previously been reported to contain high affinity saturable binding receptors for 121 SEB (3). Pretreatment of cells with endoglycoceramidase (that specifically cleaves the oligosaccharide backbone of GSL) or human kidney neutral GSLs, completely compromised the binding of 121 SEB to PT cells (3). Furthermore, direct binding of 1251 SEB to GSL separated on thin layer plates and solid phase binding assays on microtiter plates, has identified digalactosylceramide (diGalCer) as a receptor for SEB in human kidney and PT cells. This GSL was not found in rat kidney cells; however, feeding these cells diGalCer resulted in saturable binding of 121 -SEB (4). This finding may suggest a possible biochemical basis for the discrepancy seen in lower mammals challenged with SEB.

The invention provides in one aspect a peptide comprising the amino acid sequence CVFSKKTNDINSHQTDKRKT (SEQ ID NO: 3), where the peptide is capable of increasing cell proliferation in a cell that expresses digalactosylceramide receptor. The peptide, in certain examples, comprises the amino acid set forth as SEQ ID NO: 3.

The invention provides in other aspects, a peptide comprising the amino acid sequence RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID NO: 5) or KKKVTAQEL (SEQ ID NO: 7), where the peptide is capable of decreasing cell proliferation in a cell that expresses digalactosylceramide receptor. The peptide, in certain examples, comprises the amino acid set forth as SEQ ID NO: 5 or 7. The peptide of SEQ ID NO: 5 or SEQ ID NO: 7 can also, in certain embodiments, promote apoptosis. In certain cases, the peptide decreases cell proliferation and promotes apoptosis.

Methods of assaying cell growth and proliferation are known in the art. See, for example, Kittler et al. (Nature. 432 (7020): 1036-40, 2004) and by Miyamoto et al. (Nature 416(6883):865-9, 2002). Assays for cell proliferation generally involve the measurement of DNA synthesis during cell replication. In one example, proliferation is measured using a tetrazolium compound in the CELL TITER 96^® Assay. Alternatively, [³H] -Thymidine or 5-bromo-2'-deoxyuridine [BrdU], can be added to cells (or animals) and then the incorporation of these precursors into genomic DNA during the S phase of the cell cycle (replication) can be detected (Ruefli-Brasse et al., Science 302(5650): 1581-4, 2003; Gu et al., Science 302 (5644):445-9, 2003).

Assays for measuring cell viability are known in the art, and are described, for example, by Crouch et al. (J. Immunol. Meth. 160, 81-8); Kangas et al. (Med. Biol.62, 338^3, 1984); Lundin et al., (Meth. Enzymol.133, 27^2, 1986); Petty et al. (Comparison of J. Biolum. Chemilum.lO, 29-34, 1995); and Cree et al. (Anticancer Drugs 6: 398-404, 1995). Cell viability can be assayed using a variety of methods, including MTT (3-(4,5-dimethylthiazolyl)-2,5-diphenyltetrazolium bromide) (Barltrop, Bioorg. & Med. Chem. Lett.l : 611, 1991; Cory et al., Cancer Comm. 3, 207-12, 1991; Paull J. Heterocyclic Chem. 25, 911, 1988). Assays for cell viability are also available commercially. These assays include CELLTITER-GLO

Luminescent Cell Viability Assay (Promega), which uses luciferase technology to detect ATP and quantify the health or number of cells in culture, and the CellTiter- Glo^® Luminescent Cell Viability Assay, which is a lactate dehyrodgenase (LDH) cytotoxicity assay. Assays for measuring cell apoptosis are known to the skilled artisan.

Apoptotic cells are characterized by characteristic morphological changes, including chromatin condensation, cell shrinkage and membrane blebbing, which can be clearly observed using light microscopy. The biochemical features of apoptosis include DNA fragmentation, protein cleavage at specific locations, increased mitochondrial membrane permeability, and the appearance of phosphatidylserine on the cell membrane surface. Assays for apoptosis are known in the art. Exemplary assays include TUNEL (Terminal deoxynucleotidyl Transferase Biotin-dUTP Nick End Labeling) assays, caspase activity (specifically caspase-3) assays, and assays for fas- ligand and annexin V. Commercially available products for detecting apoptosis include, for example, Apo-ONE^® Homogeneous Caspase-3/7 Assay, FragEL TUNEL kit (ONCOGENE RESEARCH PRODUCTS, San Diego, CA), the ApoBrdU DNA Fragmentation Assay (BIOVISION, Mountain View, CA), and the Quick Apoptotic DNA Ladder Detection Kit (BIOVISION, Mountain View, CA).

The invention provides in other aspects, a peptide comprising the amino acid sequence ENENSFWYAMMPAPGDKFDQSKYLMMYNDN (SEQ ID NO: 6), wherein the peptide is capable of binding to a digalactosylceramide (DAG) receptor. The peptide, in certain examples, comprises the amino acid set forth as SEQ ID NO: 6.

In certain examples, the peptide is derived from Staphylococcus enterotoxin B (SEB). As discussed herein, the peptide can be produced recombinantly, by chemical synthesis or purified from native sources, using methods known in the art. The peptide is not limited to a SEB peptide, as discussed above.

In certain examples, the peptides may comprise one or more other agents. The number of agents that the peptide comprises are limited only by the physician or skilled practitioner administering the peptides and the need of the subject. The agents are linked to the peptides by covalent linkage. Various moieties may be used to attach the agents to the peptides, including but not limited to linkage to an amino, carboxyl, hydroxyl, sulfide, carbon, or oxygen group on the peptides; however essentially any location in the peptide is appropriate for linkage of the agent. In certain cases, a linker sequence may be used. Suitable linker sequences are known in the art and generally include chemically reactive groups on each end of a suitable polymeric sequence such as an amino acid sequence.

In certain examples, any of the peptides of the invention may further comprise one or more therapeutic agents. The therapeutic agent is limited only by the physician or skilled practicioner administering the peptide. In certain examples, the therapeutic agents is selected from the group consisting of, but not limited to, protein synthesis modulators, apoptosis modulators, nitric oxide modulators, bactericides, a fungicides, anti neoplastic drugs, anti thrombotic drugs, hypochoesterolemic drugs, hypotriglyceridemic drugs, hypoglycemic drugs, anti-allergic drugs, antibiotics, tissue, anti-inflammatory drug, nerve regenerating drugs, and anti-arthritis drugs.

Without wishing to be bound by theory, it is believed the compositions of the peptides and agents of the (i.e. at least one identified SEB peptide in combination with an agent, such as a therapeutic agent, for example an anticancer agent) can significantly enhance efficacy of the drug, e.g., of an anti-cancer agent.

Moreover, by virtue of the combination of the peptide and agent, the invention provides for the presentation to the subject cell essentially simultaneously, an effect that may not be readily achieved by administering the same compounds in a drug "cocktail" formulation without linking the compounds. In preferred example, the one or more therapeutic agents is an anticancer agent. The anticancer agents may be selected from the group consisting of, but not limited to, a chemotherapeutic agent, a peptide toxin, and a protein toxin.

In other certain cases, the one or more therapeutic agents is an anticancer agent. The anticancer agent can be selected from, but not limited to, a chemotherapeutic agent, a peptide toxin, and a protein toxin. The anticancer agent can be any anticancer agent that is known to a clinician for use in anticancer therapy. Other examples of anti-cancer drugs that may be used in the various embodiments of the invention, including pharmaceutical compositions and dosage forms and kits of the invention, include, but are not limited to: acivicin; aclarubicin; acodazole hydrochloride; acronine; adozelesin; aldesleukin; altretamine; ambomycin; ametantrone acetate; aminoglutethimide; amsacrine; anastrozole; anthramycin; asparaginase; asperlin; azacitidine; azetepa; azotomycin; batimastat; benzodepa; bicalutamide; bisantrene hydrochloride; bisnafide dimesylate; bizelesin; bleomycin sulfate; brequinar sodium; bropirimine; busulfan; cactinomycin; calusterone; caracemide; carbetimer; carboplatin; carmustine; carubicin hydrochloride; carzelesin; cedefingol; chlorambucil; cirolemycin; cisplatin; cladribine; crisnatol mesylate; cyclophosphamide; cytarabine; dacarbazine; dactinomycin; daunorubicin hydrochloride; decitabine; dexormaplatin; dezaguanine; dezaguanine mesylate; diaziquone; docetaxel; doxorubicin; doxorubicin hydrochloride; droloxifene; droloxifene citrate; dromostanolone propionate; duazomycin; edatrexate; eflornithine hydrochloride; elsamitrucin; enloplatin; enpromate; epipropidine; epirubicin hydrochloride; erbulozole; esorubicin hydrochloride; estramustine; estramustine phosphate sodium; etanidazole; etoposide; etoposide phosphate; etoprine; fadrozole hydrochloride; fazarabine; fenretinide; floxuridine; fludarabine phosphate; fluorouracil; flurocitabine; fosquidone; fostriecin sodium; gemcitabine; gemcitabine hydrochloride; hydroxyurea; idarubicin hydrochloride; ifosfamide; ilmofosine; interleukin II (including recombinant interleukin II, or rIL2), interferon alfa-2a; interferon alfa-2b; interferon alfa-nl; interferon alfa-n3; interferon beta-I a; interferon gamma-I b; iproplatin; irinotecan hydrochloride; lanreotide acetate; letrozole; leuprolide acetate; liarozole hydrochloride; lometrexol sodium; lomustine; losoxantrone hydrochloride; masoprocol; maytansine; mechlorethamine, mechlorethamine oxide hydrochloride rethamine hydrochloride; megestrol acetate; melengestrol acetate; melphalan; menogaril; mercaptopurine; methotrexate; methotrexate sodium; metoprine; meturedepa; mitindomide; mitocarcin; mitocromin; mitogillin; mitomalcin; mitomycin; mitosper; mitotane; mitoxantrone hydrochloride; mycophenolic acid; nocodazole; nogalamycin; ormaplatin; oxisuran; paclitaxel; pegaspargase; peliomycin; pentamustine; peplomycin sulfate; perfosfamide; pipobroman; piposulfan; piroxantrone hydrochloride; plicamycin; plomestane; porfimer sodium; porfiromycin; prednimustine; procarbazine hydrochloride; puromycin; puromycin hydrochloride; pyrazofurin; riboprine; rogletimide; safingol; safingol hydrochloride; semustine; simtrazene; sparfosate sodium; sparsomycin; spirogermanium hydrochloride; spiromustine; spiroplatin; streptonigrin; streptozocin; sulofenur; talisomycin; tecogalan sodium; tegafur; teloxantrone hydrochloride; temoporfin; teniposide; teroxirone; testolactone; thiamiprine; thioguanine; thiotepa; tiazofurin; tirapazamine; toremifene citrate; trestolone acetate; triciribine phosphate; trimetrexate; trimetrexate glucuronate; triptorelin; tubulozole hydrochloride; uracil mustard; uredepa; vapreotide; verteporfin; vinblastine sulfate; vincristine sulfate; vindesine; vindesine sulfate; vinepidine sulfate; vinglycinate sulfate; vinleurosine sulfate; vinorelbine tartrate; vinrosidine sulfate; vinzolidine sulfate; vorozole; zeniplatin; zinostatin; zorubicin hydrochloride, improsulfan, benzodepa, carboquone, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide, trimethylolomelamine, chlornaphazine, novembichin, phenesterine, trofosfamide, estermustine, chlorozotocin, gemzar, nimustine, ranimustine, dacarbazine, mannomustine, mitobronitol,aclacinomycins, actinomycin F(I), azaserine, bleomycin, carubicin, carzinophilin, chromomycin, daunorubicin, daunomycin, 6-diazo-5-oxo-l- norleucine, doxorubicin, olivomycin, plicamycin, porfiromycin, puromycin, tubercidin, zorubicin, denopterin, pteropterin, 6-mercaptopurine, ancitabine, 6- azauridine, carmofur, cytarabine, dideoxyuridine, enocitabine, pulmozyme, aceglatone, aldophosphamide glycoside, bestrabucil, defofamide, demecolcine, elfornithine, elliptinium acetate, etoglucid, flutamide, hydroxyurea, lentinan, phenamet, podophyllinic acid, 2-ethylhydrazide, razoxane, spirogermanium, tamoxifen, taxotere, tenuazonic acid, triaziquone, 2,2',2"-trichlorotriethylamine, urethan, vinblastine, vincristine, vindesine and related agents. 20-epi-l ,25 dihydroxyvitamin D3; 5-ethynyluracil; abiraterone; aclarubicin; acylfulvene; adecypenol; adozelesin; aldesleukin; ALL-TK antagonists; altretamine; ambamustine; amidox; amifostine; aminolevulinic acid; amrubicin; amsacrine; anagrelide; anastrozole; andrographolide; angiogenesis inhibitors; antagonist D; antagonist G; antarelix; anti-dorsalizing morphogenetic protein- 1; antiandrogen, prostatic carcinoma; antiestrogen; antineoplaston; antisense oligonucleotides; aphidicolin glycinate; apoptosis gene modulators; apoptosis regulators; apurinic acid; ara-CDP- DL-PTBA; arginine deaminase; asulacrine; atamestane; atrimustine; axinastatin 1 ; axinastatin 2; axinastatin 3; azasetron; azatoxin; azatyrosine; baccatin III derivatives; balanol; batimastat; BCR/ ABL antagonists; benzochlorins; benzoylstaurosporine; beta lactam derivatives; beta-alethine; betaclamycin B; betulinic acid; bFGF inhibitor; bicalutamide; bisantrene; bisaziridinylspermine; bisnafide; bistratene A; bizelesin; breflate; bropirimine; budotitane; buthionine sulfoximine; calcipotriol; calphostin C; camptothecin derivatives; canarypox IL-2; capecitabine; carboxamide-amino-triazole; carboxyamidotriazole; CaRest M3; CARN 700; cartilage derived inhibitor; carzelesin; casein kinase inhibitors (ICOS); castanospermine; cecropin B; cetrorelix; chlorlns; chloroquinoxaline sulfonamide; cicaprost; cisporphyrin; cladribine; clomifene analogues; clotrimazole; collismycin A; collismycin B; combretastatin A4; combretastatin analogue; conagenin; crambescidin 816; crisnatol; cryptophycin 8; cryptophycin A derivatives; curacin A; cyclopentanthraquinones; cycloplatam; cypemycin; cytarabine ocfosfate; cytolytic factor; cytostatin; dacliximab; decitabine; dehydrodidemnin B; deslorelin; dexamethasone; dexifosfamide; dexrazoxane; dexverapamil; diaziquone; didemnin B; didox; diethylnorspermine; dihydro-5- azacytidine; dihydrotaxol, 9-; dioxamycin; diphenyl spiromustine; docetaxel; docosanol; dolasetron; doxifluridine; droloxifene; dronabinol; duocarmycin SA; ebselen; ecomustine; edelfosine; edrecolomab; eflornithine; elemene; emitefur; epirubicin; epristeride; estramustine analogue; estrogen agonists; estrogen antagonists; etanidazole; etoposide phosphate; exemestane; fadrozole; fazarabine; fenretinide; filgrastim; finasteride; flavopiridol; flezelastine; fluasterone; fludarabine; fluorodaunorunicin hydrochloride; forfenimex; formestane; fostriecin; fotemustine; gadolinium texaphyrin; gallium nitrate; galocitabine; ganirelix; gelatinase inhibitors; gemcitabine; glutathione inhibitors; hepsulfam; heregulin; hexamethylene bisacetamide; hypericin; ibandronic acid; idarubicin; idoxifene; idramantone; ilmofosine; ilomastat; imidazoacridones; imiquimod; immunostimulant peptides; insulin-like growth factor- 1 receptor inhibitor; interferon agonists; interferons; interleukins; iobenguane; iododoxorubicin; ipomeanol, 4-; iroplact; irsogladine; isobengazole; isohomohalicondrin B; itasetron; jasplakinolide; kahalalide F; lamellarin-N triacetate; lanreotide; leinamycin; lenograstim; lentinan sulfate; leptolstatin; letrozole; leukemia inhibiting factor; leukocyte alpha interferon; leuprolide+estrogen+progesterone; leuprorelin; levamisole; liarozole; linear polyamine analogue; lipophilic disaccharide peptide; lipophilic platinum compounds; lissoclinamide 7; lobaplatin; lombricine; lometrexol; lonidamine; losoxantrone; lovastatin; loxoribine; lurtotecan; lutetium texaphyrin; lysofylline; lytic peptides; maitansine; mannostatin A; marimastat; masoprocol; maspin; matrilysin inhibitors; matrix metalloproteinase inhibitors; menogaril; merbarone; meterelin; methioninase; metoclopramide; MIF inhibitor; mifepristone; miltefosine; mirimostim; mismatched double stranded RNA; mitoguazone; mitolactol; mitomycin analogues; mitonafide; mitotoxin fibroblast growth factor-saporin; mitoxantrone; mofarotene; molgramostim; monoclonal antibody, human chorionic gonadotrophin; monophosphoryl lipid A+myobacterium cell wall sk; mopidamol; multiple drug resistance gene inhibitor; multiple tumor suppressor 1 -based therapy; mustard anticancer agent; mycaperoxide B; mycobacterial cell wall extract; myriaporone; N-acetyldinaline; N-substituted benzamides; nafarelin; nagrestip; naloxone+pentazocine; napavin; naphterpin; nartograstim; nedaplatin; nemorubicin; neridronic acid; neutral endopeptidase; nilutamide; nisamycin; nitric oxide modulators; nitroxide antioxidant; nitrullyn; 06- benzylguanine; octreotide; okicenone; oligonucleotides; onapristone; ondansetron; ondansetron; oracin; oral cytokine inducer; ormaplatin; osaterone; oxaliplatin; oxaunomycin; taxel; taxel analogues; taxel derivatives; palauamine; palmitoylrhizoxin; pamidronic acid; panaxytriol; panomifene; parabactin; pazelliptine; pegaspargase; peldesine; pentosan polysulfate sodium; pentostatin; pentrozole; perflubron; perfosfamide; perillyl alcohol; phenazinomycin; phenylacetate; phosphatase inhibitors; picibanil; pilocarpine hydrochloride; pirarubicin; piritrexim; placetin A; placetin B; plasminogen activator inhibitor; platinum complex; platinum compounds; platinum-triamine complex; porfimer sodium; porfiromycin; prednisone; propyl bis-acridone; prostaglandin J2; proteasome inhibitors; protein A-based immune modulator; protein kinase C inhibitor; protein kinase C inhibitors, microalgal; protein tyrosine phosphatase inhibitors; purine nucleoside phosphorylase inhibitors; purpurins; pyrazoloacridine; pyridoxylated hemoglobin polyoxyethylene conjugate; raf antagonists; raltitrexed; ramosetron; ras farnesyl protein transferase inhibitors; ras inhibitors; ras-GAP inhibitor; retelliptine demethylated; rhenium Re 186 etidronate; rhizoxin; ribozymes; RII retinamide; rogletimide; rohitukine; romurtide; roquinimex; rubiginone Bl; ruboxyl; safingol; saintopin; SarCNU; sarcophytol A; sargramostim; Sdi 1 mimetics; semustine; senescence derived inhibitor 1 ; sense oligonucleotides; signal transduction inhibitors; signal transduction modulators; single chain antigen binding protein; sizofiran; sobuzoxane; sodium borocaptate; sodium phenylacetate; solverol; somatomedin binding protein; sonermin; sparfosic acid; spicamycin D; spiromustine; splenopentin; spongistatin 1; squalamine; stem cell inhibitor; stem-cell division inhibitors; stipiamide; stromelysin inhibitors; sulfinosine; superactive vasoactive intestinal peptide antagonist; suradista; suramin; swainsonine; synthetic glycosaminoglycans; tallimustine; tamoxifen methiodide; tauromustine; tazarotene; tecogalan sodium; tegafur; tellurapyrylium; telomerase inhibitors; temoporfin; temozolomide; teniposide; tetrachlorodecaoxide; tetrazomine; thaliblastine; thiocoraline; thrombopoietin; thrombopoietin mimetic; thymalfasin; thymopoietin receptor agonist; thymotrinan; thyroid stimulating hormone; tin ethyl etiopurpurin; tirapazamine; titanocene bichloride; topsentin; toremifene; totipotent stem cell factor; translation inhibitors; tretinoin; triacetyluridine; triciribine; trimetrexate; triptorelin; tropisetron; turosteride; tyrosine kinase inhibitors; tyrphostins; UBC inhibitors; ubenimex; urogenital sinus-derived growth inhibitory factor; urokinase receptor antagonists; vapreotide; variolin B; vector system, erythrocyte gene therapy; velaresol; veramine; verdins; verteporfin; vinorelbine; vinxaltine; vitaxin; vorozole; zanoterone; zeniplatin; zilascorb; and zinostatin stimalamer. Preferred additional anti-cancer drugs are 5- fluorouracil and leucovorin. Additional cancer therapeutics include monoclonal antibodies such as rituximab, trastuzumab and cetuximab.

In other uses, the peptide can comprise an imaging agent. The imaging agent can be selected from a radiolabel or a fluorescent label. A peptide of the invention includes any substituted analog or chemical derivative of a peptide as described herein, so long as the peptide is capable promoting cell proliferation, or decreasing cell proliferation and promoting apoptosis as described herein . Therefore, a peptide can be subject to various changes that provide for certain advantages in its use. Peptide oligomers

In certain examples, peptide oligomers are used in the methods of the invention as described herein. The oligomers can be inhibitory oligomers, e.g. oligomers that inhibit DAG receptor activity. The oligomers can in preferred examples inhibit cell proliferation or apoptosis. The oligomers can be stimulatory oligomers, e.g. oligomers that stimulate DAG receptor activity. The oligomers can in preferred examples stimulate cell proliferation.

In certain preferred examples, the oligomer is at least a 3-mer, 4-mer, 5-mer,

6-mer, 7-mer, 8-mer, 9-mer or 10-mer of the amino acid sequence comprising

CVFSKKTNDINSHQTDKRKT (SEQ ID NO: 3), RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID NO: 5),

ENENSFWYAMMPAPGDKFDQSKYLMMYNDN (SEQ ID NO: 6) or

KKKVTAQEL (SEQ ID NO: 7). The oligomers can be of any length so long as they retain their inhibitory or stimulatory activity. The oligomers can be modified, for example with enzymatic or chemical modification. The modification can be an agent that is used for targeting.

III. METHODS OF THE INVENTION The invention features methods that use the peptides as described herein.

Methods of increasing cell proliferation

In one aspect of the invention, the invention features a method of increasing cell proliferation comprising administering a peptide that is capable of increasing cell proliferation in a cell that expresses digalactosylceramide receptor. In preferred embodiments, the peptide is a Staphylococcus enterotoxin B (SEB) peptide, however the invention is not limited to SEB peptides.

The invention features methods of increasing cell proliferation in a subject. The methods comprise administering to the subject a peptide, for example, but not limited to an SEB peptide, that is capable of increasing cell proliferation in a cell that expresses digalactosylceramide receptor.

Any cell that expresses digalactosylceramide (DAG) receptor is suitable for use in the invention, however in certain examples, the cell that expressed DAG receptor is selected from the group consisting of: brain cells, neuronal cells, lymphocytes, leukocytes, liver, inner ear cells, spleen, pancreas, urinary tract, bone marrow spinal cord, spinal roots, skin cells, and conjunctival vessels.

Also encompassed by the invention are methods for increasing kidney cell proliferation in a subject comprising administering to the subject a peptide capable of increasing cell proliferation, thereby stimulating kidney cell proliferation. In preferred embodiments, the peptide is a Staphylococcus enterotoxin B (SEB) peptide, however the invention is not limited to SEB peptides.

In certain cases, the method is useful where the subject has undergone kidney transplantation or kidney resection. The method can be of use to stabilze, reduces the symptoms of, or ameliorates a disease or disorder characterized by kidney dysfunction. The method can be used, in other examples, to increase immune cell proliferation. The method is used, in certain cases, to increase immune cell proliferation in a subject. The method comprises administering a peptide capable of increasing cell proliferation, thereby stimulating immune cell proliferation. In preferred embodiments, the peptide is a Staphylococcus enterotoxin B (SEB) peptide, however the invention is not limited to SEB peptides.

When the method is used to increase immune cell proliferation, the immune cells are T-lymphocytes. In certain examples, the method stabilizes, reduces the symptoms of, or ameliorates a disease or disorder characterized by abnormal immune cell proliferation.

Exemplary diseases that can be treated by the invention are anaplastic anemia or Fabry's Disease. Anaplastic Anemia (AA) is a rare disease in which the bone marrow is unable to produce adequate blood cells; leading to pancytopenia (deficiency of all types of blood cells). AA may occur at any age, but there is a peak in adolescence / early adulthood, and again in old age. Slightly more males than females are diagnosed with AA, also the disease is more common in the Far East. Patients successfully treated for aplastic anemia have a higher risk of developing other diseases later in life, including cancer.

Fabry's disease is caused by the lack of or faulty enzyme needed to metabolize lipids, fat-like substances that include oils, waxes, and fatty acids. The enzyme is known as ceramide trihexosidase, also called alpha-galactosidase-A. A mutation in the gene that controls this enzyme causes insufficient breakdown of lipids, which build up to harmful levels in the eyes, kidneys, autonomic nervous system, and cardiovascular system. Since the gene that is altered is carried on a mother's X chromosome, her sons have a 50 percent chance of inheriting the disorder and her daughters have a 50 percent chance of being a carrier. Some women who carry the genetic mutation may have symptoms of the disease. Symptoms usually begin during childhood or adolescence and include burning sensations in the hands that gets worse with exercise and hot weather and small, raised reddish-purple blemishes on the skin. Some boys will also have eye manifestations, especially cloudiness of the cornea. Lipid storage may lead to impaired arterial circulation and increased risk of heart attack or stroke. The heart may also become enlarged and the kidneys may become progressively involved. Other symptoms include decreased sweating, fever, and gastrointestinal difficulties, particularly after eating. Fabry disease is one of several lipid storage disorders. Information on Fabry's Disease is publicly available on the world wide web at http://www.ninds.nih.gov/disorders/fabrys/fabrys.htm. As described herein in preferred embodiments, the peptide is a Staphylococcus enterotoxin B (SEB) peptide. The peptide can comprise the amino acid sequence CVFSKKTNDINSHQTDKRKT (SEQ ID NO: 3), including variants as described herein. In certain examples, the SEB peptide further comprises one or more therapeutic agents. The therapeutic agent may be any one of, but not limited to protein synthesis modulators, apoptosis modulators, nitric oxide modulators, bactericides, a fungicides, anti neoplastic drugs, anti thrombotic drugs,hypochoesterolemic drugs, hypotriglyceridemic drugs, hypoglycemic drugs, anti-allergic drugs, antibiotics,tissue, anti-inflammatory drug, nerve regenerating drugs, and anti-arthritis drugs.

Methods of decreasing cell proliferation or promoting apoptosis

Another method of the invention uses peptides, in certain preferred embodiments, SEB peptides, in methods to decrease cell proliferation in a cell that expresses digalactosylceramide receptor. The method can also be used to decrease cell proliferation in a subject in need thereof, where the method comprises administering to the subject a peptide, for example an SEB peptide, that is capable of decreasing cell proliferation in a cell that expresses digalactosylceramide receptor. In certain preferred examples, the method further comprises increasing apoptosis. Accordingly, a method of the invention includes a method of promoting apoptosis comprising administering a SEB peptide capable of promoting apoptosis in a cell that expresses digalactosylceramide receptor. The method can also be used to promote apoptosis in a subject in need thereof comprising administering to the subject a SEB peptide capable of promoting apoptosis in a cell that expresses digalactosylceramide receptor.

The method of the invention is particularly useful in cases where the cell expresses elevated levels of digalactosylceramide compared to a control cell. In certain examples, the cell that expresses digalactosylceramide receptor is a mammalian cell. The mammalian cell can be selected from the group consisting of a selected from the group consisting of: tumor cells, kidney cells, neuronal cells, lymphocytes, inner ear cells, spleen cells and pancreatic cells.

The invention as described herein can be used to in cancer treatment. By "cancer" is meant any disease that is caused by or results in inappropriately high levels of cell division, inappropriately low levels of apoptosis, or both. For example, cancer is an example of a neoplasia. Examples of cancers include, without limitation, leukemias (e.g., acute leukemia, acute lymphocytic leukemia, acute myelocytic leukemia, acute myeloblastic leukemia, acute promyelocyte leukemia, acute myelomonocytic leukemia, acute monocytic leukemia, acute erythroleukemia, chronic leukemia, chronic myelocytic leukemia, chronic lymphocytic leukemia), polycythemia vera, lymphoma (Hodgkin's disease, non-Hodgkin's disease), Waldenstrom's macroglobulinemia, heavy chain disease, and solid tumors such as sarcomas and carcinomas (e.g., fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's tumor, leiomyosarcoma, rhabdomyosarcoma, colon carcinoma, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinomas, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma, nile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilm's tumor, cervical cancer, uterine cancer, testicular cancer, lung carcinoma, small cell lung carcinoma, bladder carcinoma, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodenroglioma, schwannoma, meningioma, melanoma, neuroblastoma, and retinoblastoma). Lymphoproliferative disorders are also considered to be proliferative diseases.

The methods of the invention are particularly useful in certain cases for treating tumors. Accordingly, in one aspect the invention features a method of treating a tumor in a subject comprising administering to the subject a SEB peptide capable of promoting apoptosis or decreasing cell proliferation in a tumor that expresses digalactosylceramide receptor, thereby treating a tumor in a subject.

Many tumors express elevated levels of glycosaminoglycans (Altered growth behavior of malignant cells associated with changes in externally labeled glycoprotein and glycolipid. Proc Natl Acad Sci U S A. 1973 Dec;70(12):3329-33). Accordingly, in certain embodiments of the method, the tumor expresses elevated levels of digalactosylceramide compared to a normal tissue. The invention also features methods of treating a lipid metabolic disorder in a subject comprising administering to the subject a SEB peptide capable of promoting apoptosis or decreasing cell proliferation in cells that express high levels of digalactosylceramide, thereby treating the lipid metabolic disorder the subject. In certain examples the lipid metabolic disorder is selected from the group consisting of Fabry's disease,Metachromatic Leukodystrophy, GM2 Gangliosidosis, Tay-Sachs disease and chronic myelogenous leukemia.

Metachromatic leukodystrophy (MLD) is one of a group of genetic disorders called the leukodystrophies. These diseases impair the growth or development of the myelin sheath, the fatty covering that acts as an insulator around nerve fibers. Myelin, which lends its color to the white matter of the brain, is a complex substance made up of at least 10 different enzymes. The leukodystrophies are caused by genetic defects in how myelin produces or metabolizes these enzymes. Each of the leukodystrophies is the result of a defect in the gene that controls one (and only one) of the enzymes. MLD is caused by a deficiency of the enzyme arylsulfatase A. MLD is one of several lipid storage diseases, which result in the toxic buildup of fatty materials (lipids) in cells in the nervous system, liver, and kidneys. There are three forms of MLD: late infantile, juvenile, and adult. In the late infantile form, which is the most common MLD, affected children have difficulty walking after the first year of life. Symptoms include muscle wasting and weakness, muscle rigidity, developmental delays, progressive loss of vision leading to blindness, convulsions, impaired swallowing, paralysis, and dementia. Children may become comatose. Most children with this form of MLD die by age 5. Children with the juvenile form of MLD (between 3-10 years of age) usually begin with impaired school performance, mental deterioration, and dementia and then develop symptoms similar to the infantile form but with slower progression. The adult form commonly begins after age 16 as a psychiatric disorder or progressive dementia. Adult-onset MLD progresses more slowly than the infantile form. More information can be found publicly on the world wide web at http://www.ninds.nih.gov/disorders/metachromatic_leukodystrophy/metachromatic_le ukodystrophy.htm.

The GM2 gangliosidoses are a group of lysosomal lipid storage disorders caused by mutations in at least 1 of 3 recessive genes: HEXA, HEXB, and GM2A. Normal products of all 3 genes are required for normal catabolism of the GM2 ganglioside substrate. Deficient activity of these enzymes leads to accumulation of the substrate inside neuronal lysosomes, leading to cell death. The products of the 3 genes are, respectively, the alpha subunits of b-hexosaminidase A (Hex A; EC 3.2.1.52), the beta subunits of Hex A (EC 3.2.1.52), and the GM2 activator protein. Hydrolysis of GM2 ganglioside requires a normal GM2 ganglioside-GM2 activator-Hex A complex. Hex A is a dimer and has the structure alpha-beta. The alpha subunit is encoded by the HEXA gene at band 15q23-q24; the beta subunit is encoded by the HEXB gene at band 5ql3. A site on the alpha subunit acts against negatively charged sulfated substrates, while a site on the beta subunit acts against neutral water-soluble substrates. b-Hexosaminidase B (Hex B) is a dimer of beta chains. It hydrolyzes GM2 and its neutral asialo derivative GA2. Both subunit precursors acquire the mannose 6- phosphate marker for recognition by lysosomes. Hexosaminidase S (Hex S) is a dimer of alpha chains; it is a normal constituent of plasma and degrades a wide range of glycoconjugates containing b-linked N-acetylhexosaminyl residues.

Type I GM2 gangliosidosis is also known as classic infantile acute TSD, B variant, pseudo-AB variant, or Hex A deficiency. Type III GM2 gangliosidosis is also known as juvenile subacute TSD. The Bl variant of GM2 gangliosidosis is also known as late infantile subacute-to-chronic TSD; it is characterized by a defect in formation and stabilization of the alpha subunit active site. GM2 gangliosidosis, also known as adult chronic-type TSD is characterized by a pseudodeficiency mutation in one or both HEXA alleles.

Tay-Sachs disease is a fatal genetic lipid storage disorder in which harmful quantities of a fatty substance called ganglioside GM2 build up in tissues and nerve cells in the brain. The condition is caused by insufficient activity of an enzyme called beta-hexosaminidase A that catalyzes the biodegradation of acidic fatty materials known as gangliosides. Gangliosides are made and biodegraded rapidly in early life as the brain develops. Infants with Tay-Sachs disease appear to develop normally for the first few months of life. Then, as nerve cells become distended with fatty material, a relentless deterioration of mental and physical abilities occurs. The child becomes blind, deaf, and unable to swallow. Muscles begin to atrophy and paralysis sets in. Other neurological symptoms include dementia, seizures, and an increased startle reflex to noise. A much rarer form of the disorder occurs in patients in their twenties and early thirties and is characterized by an unsteady gait and progressive neurological deterioration. Persons with Tay-Sachs also have "cherry-red" spots in their eyes. The incidence of Tay-Sachs is particularly high among people of Eastern European and Askhenazi Jewish descent. Patients and carriers of Tay-Sachs disease can be identified by a simple blood test that measures beta-hexosaminidase A activity. Both parents must carry the mutated gene in order to have an affected child. In these instances, there is a 25 percent chance with each pregnancy that the child will be affected with Tay-Sachs disease. Prenatal diagnosis is available if desired. More information is available publicly on the world wide web at http://www.ninds.nih.gov/disorders/taysachs/taysachs.htm.

Chronic myelogenous leukemia is cancer of the bone marrow. CML can occur in adults (usually middle-aged) and children. The disease affects 1 to 2 people per 100,000 and accounts for 7 - 20% cases of leukemia. It is usually associated with a chromosome abnormality called the Philadelphia chromosome. CML causes rapid growth of the blood-forming cells (myeloid precursors) in the bone marrow, peripheral blood, and body tissues. A wealth of information is available on the internet, for example at http://www.nlm.nih.gov/medlineplus/ency/article/000570.htm.

Other diseases may include GM2 gangliosidosis type I, TSD, amaurotic idiocy, GM2 gangliosidosis B variant, classic infantile acute TSD, hexosaminidase A deficiency, HEXA deficiency, GM2 gangliosidosis type III, juvenile subacute TSD, GM2 gangliosidosis adult chronic type, adult TSD, adult hexosaminidase A deficiency, TSD variant Bl, TSD pseudo-AB variant, GM2 gangliosidosis type II, Sandhoff disease, GM2 gangliosidosis O variant, hexosaminidases A and B deficiency, Sandhoff disease infantile, Sandhoff disease juvenile type, Sandhoff disease adult type, GM2 gangliosidosis type AB, GM2 activator deficiency, hexosaminidase activator deficiency, hexosaminidase B deficiency, HEXB deficiency.

The invention described herein includes methods of treating a neural disease or disorder or a kidney disease or disorder in a subject comprising administering to the subject a SEB peptide capable of promoting apoptosis or decreasing cell proliferation in neural tissue or kidney tissue that expresses digalactosylceramide receptor, thereby treating a neural disease or disorder or a kidney disease in the subject.

The neural tissue or kidney tissue, in certain examples, expresses high levels of digalactosylceramide compared to a control cell. A number of different kidney diseases or disorders can be treated by the methods of the invention. Certain exemplary diseases or disorders include glomerular diseases, such as, but not limited to, glomerular nephritis. Glomerulonephritis is a type of kidney disease caused by inflammation of the internal kidney structures (glomeruli), which help filter waste and fluids from the blood. Glomerulonephritis may be caused by specific problems with the body's immune system, but the precise cause of some cases is unknown. Damage to the glomeruli causes blood and protein to be lost in the urine. The condition may develop after survival of the acute phase of rapidly progressive glomerulonephritis. In about a quarter of people with chronic glomerulonephritis there is no prior history of kidney disease and the disorder first appears as chronic renal failure. Specific disorders that are associated with glomerulonephritis include focal segmental glomerulosclerosis, Goodpasture syndrome, IgA nephropathy, Lupus nephritis, Membranoproliferative GN I, Membranoproliferative GN II, Post-streptococcal GN, Rapidly progressive glomerulonephritis.

A number of different neural diseases can be treated by the methods of the invention. One such disorder is autism. Autism (sometimes called "classical autism") is the most common condition in a group of developmental disorders known as the autism spectrum disorders (ASDs). Autism is characterized by impaired social interaction, problems with verbal and nonverbal communication, and unusual, repetitive, or severely limited activities and interests. Other ASDs include Asperger syndrome, Rett syndrome, childhood disintegrative disorder, and pervasive developmental disorder not otherwise specified (usually referred to as PDD-NOS). Experts estimate that three to six children out of every 1 ,000 will have autism. Males are four times more likely to have autism than females. More information on autism is available publicly on the worlds wide web at http://www.mnds. nih.gov/disorders/autism/detail_autism.htm.

In the methods of decreasing cell proliferation or promoting apoptosis, and the diseases and disorders treated as described herein, in certain examples the SEB peptide comprises the amino acid sequence

RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID NO: 5) or KKKVTAQEL (SEQ ID NO: 7).

In other examples, the SEB peptide further comprises one or more therapeutic agents, which can be selected from, but not limited to, protein synthesis modulators, apoptosis modulators, nitric oxide modulators, bactericides, a fungicides, anti neoplastic drugs, anti thrombotic drugs,hypochoesterolemic drugs, hypotriglyceridemic drugs, hypoglycemic drugs, anti-allergic drugs, antibiotics,tissue, anti-inflammatory drug, nerve regenerating drugs, and anti-arthritis drugs. It is a general feature of the invention that because the peptides are targeting the therapeutic agent to the site of therapy (e.g. the tumor), that the dosage required (e.g. the dose of anticancer agent) will be less than the amount typically used in traditional therapy. In this way, the toxicity of many drugs can be reduces d or eliminated. In certain cases, the apoptosis modulator is clodronate. Clodronate is the disodium salt of a nitrogen-free bisphosphonate analog of naturally occurring pyrophosphate. Clodronate binds to calcium and inhibits osteoclastic bone resorption and hydroxyapatite crystal formation and dissolution, resulting in a reduction of bone turnover. Clodronate may control malignancy-associated hypercalcemia, inhibit osteolytic bone metastasis and decrease pain.

In other certain cases, the one or more therapeutic agents is an anticancer agent. The anticancer agent can be selected from, but not limited to, a chemotherapeutic agent, a peptide toxin, and a protein toxin. The anticancer agent can be any anticancer agent that is known to a clinician for use in anticancer therapy. Other examples of anti-cancer drugs that may be used in the various embodiments of the invention, including pharmaceutical compositions and dosage forms and kits of the invention, include, but are not limited to: acivicin; aclarubicin; acodazole hydrochloride; acronine; adozelesin; aldesleukin; altretamine; ambomycin; ametantrone acetate; aminoglutethimide; amsacrine; anastrozole; anthramycin; asparaginase; asperlin; azacitidine; azetepa; azotomycin; batimastat; benzodepa; bicalutamide; bisantrene hydrochloride; bisnafide dimesylate; bizelesin; bleomycin sulfate; brequinar sodium; bropirimine; busulfan; cactinomycin; calusterone; caracemide; carbetimer; carboplatin; carmustine; carubicin hydrochloride; carzelesin; cedefingol; chlorambucil; cirolemycin; cisplatin; cladribine; crisnatol mesylate; cyclophosphamide; cytarabine; dacarbazine; dactinomycin; daunorubicin hydrochloride; decitabine; dexormaplatin; dezaguanine; dezaguanine mesylate; diaziquone; docetaxel; doxorubicin; doxorubicin hydrochloride; droloxifene; droloxifene citrate; dromostanolone propionate; duazomycin; edatrexate; eflornithine hydrochloride; elsamitrucin; enloplatin; enpromate; epipropidine; epirubicin hydrochloride; erbulozole; esorubicin hydrochloride; estramustine; estramustine phosphate sodium; etanidazole; etoposide; etoposide phosphate; etoprine; fadrozole hydrochloride; fazarabine; fenretinide; floxuridine; fludarabine phosphate; fluorouracil; flurocitabine; fosquidone; fostriecin sodium; gemcitabine; gemcitabine hydrochloride; hydroxyurea; idarubicin hydrochloride; ifosfamide; ilmofosine; interleukin II (including recombinant interleukin II, or rIL2), interferon alfa-2a; interferon alfa-2b; interferon alfa-nl; interferon alfa-n3; interferon beta-I a; interferon gamma-I b; iproplatin; irinotecan hydrochloride; lanreotide acetate; letrozole; leuprolide acetate; liarozole hydrochloride; lometrexol sodium; lomustine; losoxantrone hydrochloride; masoprocol; maytansine; mechlorethamine, mechlorethamine oxide hydrochloride rethamine hydrochloride; megestrol acetate; melengestrol acetate; melphalan; menogaril; mercaptopurine; methotrexate; methotrexate sodium; metoprine; meturedepa; mitindomide; mitocarcin; mitocromin; mitogillin; mitomalcin; mitomycin; mitosper; mitotane; mitoxantrone hydrochloride; mycophenolic acid; nocodazole; nogalamycin; ormaplatin; oxisuran; paclitaxel; pegaspargase; peliomycin; pentamustine; peplomycin sulfate; perfosfamide; pipobroman; piposulfan; piroxantrone hydrochloride; plicamycin; plomestane; porfimer sodium; porfiromycin; prednimustine; procarbazine hydrochloride; puromycin; puromycin hydrochloride; pyrazofurin; riboprine; rogletimide; safingol; safingol hydrochloride; semustine; simtrazene; sparfosate sodium; sparsomycin; spirogermanium hydrochloride; spiromustine; spiroplatin; streptonigrin; streptozocin; sulofenur; talisomycin; tecogalan sodium; tegafur; teloxantrone hydrochloride; temoporfin; teniposide; teroxirone; testolactone; thiamiprine; thioguanine; thiotepa; tiazofurin; tirapazamine; toremifene citrate; trestolone acetate; triciribine phosphate; trimetrexate; trimetrexate glucuronate; triptorelin; tubulozole hydrochloride; uracil mustard; uredepa; vapreotide; verteporfin; vinblastine sulfate; vincristine sulfate; vindesine; vindesine sulfate; vinepidine sulfate; vinglycinate sulfate; vinleurosine sulfate; vinorelbine tartrate; vinrosidine sulfate; vinzolidine sulfate; vorozole; zeniplatin; zinostatin; zorubicin hydrochloride, improsulfan, benzodepa, carboquone, triethylenemelamine, triethylenephosphoramide, tri ethyl enethiophosphoramide, trimethylolomelamine, chlornaphazine, novembichin, phenesterine, trofosfamide, estermustine, chlorozotocin, gemzar, nimustine, ranimustine, dacarbazine, mannomustine, mitobronitol,aclacinomycins, actinomycin F(I), azaserine, bleomycin, carubicin, carzinophilin, chromomycin, daunorubicin, daunomycin, 6-diazo-5-oxo-l- norleucine, doxorubicin, olivomycin, plicamycin, porfiromycin, puromycin, tubercidin, zorubicin, denopterin, pteropterin, 6-mercaptopurine, ancitabine, 6- azauridine, carmofur, cytarabine, dideoxyuridine, enocitabine, pulmozyme, aceglatone, aldophosphamide glycoside, bestrabucil, defofamide, demecolcine, elfornithine, elliptinium acetate, etoglucid, flutamide, hydroxyurea, lentinan, phenamet, podophyllinic acid, 2-ethylhydrazide, razoxane, spirogermanium, tamoxifen, taxotere, tenuazonic acid, triaziquone, 2,2',2"-trichlorotriethylamine, urethan, vinblastine, vincristine, vindesine and related agents. 20-epi-l,25 dihydroxyvitamin D3; 5-ethynyluracil; abiraterone; aclarubicin; acylfulvene; adecypenol; adozelesin; aldesleukin; ALL-TK antagonists; altretamine; ambamustine; amidox; amifostine; aminolevulinic acid; amrubicin; amsacrine; anagrelide; anastrozole; andrographolide; angiogenesis inhibitors; antagonist D; antagonist G; antarelix; anti-dorsalizing moφhogenetic protein- 1; antiandrogen, prostatic carcinoma; antiestrogen; antineoplaston; antisense oligonucleotides; aphidicolin glycinate; apoptosis gene modulators; apoptosis regulators; apurinic acid; ara-CDP-

DL-PTBA; arginine deaminase; asulacrine; atamestane; atrimustine; axinastatin 1 ; axinastatin 2; axinastatin 3; azasetron; azatoxin; azatyrosine; baccatin III derivatives; balanol; batimastat; BCR/ ABL antagonists; benzochlorins; benzoyl staurosporine; beta lactam derivatives; beta-alethine; betaclamycin B; betulinic acid; bFGF inhibitor; bicalutamide; bisantrene; bisaziridinylspermine; bisnafide; bistratene A; bizelesin; breflate; bropirimine; budotitane; buthionine sulfoximine; calcipotriol; calphostin C; camptothecin derivatives; canarypox IL-2; capecitabine; carboxamide-amino-triazole; carboxyamidotriazole; CaRest M3; CARN 700; cartilage derived inhibitor; carzelesin; casein kinase inhibitors (ICOS); castanospermine; cecropin B; cetrorelix; chlorlns; chloroquinoxaline sulfonamide; cicaprost; cisporphyrin; cladribine; clomifene analogues; clotrimazole; collismycin A; collismycin B; combretastatin A4; combretastatin analogue; conagenin; crambescidin 816; crisnatol; cryptophycin 8; cryptophycin A derivatives; curacin A; cyclopentanthraquinones; cycloplatam; cypemycin; cytarabine ocfosfate; cytolytic factor; cytostatin; dacliximab; decitabine; dehydrodidemnin B; deslorelin; dexamethasone; dexifosfamide; dexrazoxane; dexverapamil; diaziquone; didemnin B; didox; diethylnorspermine; dihydro-5- azacytidine; dihydrotaxol, 9-; dioxamycin; diphenyl spiromustine; docetaxel; docosanol; dolasetron; doxifluridine; droloxifene; dronabinol; duocarmycin SA; ebselen; ecomustine; edelfosine; edrecolomab; eflornithine; elemene; emitefur; epirubicin; epristeride; estramustine analogue; estrogen agonists; estrogen antagonists; etanidazole; etoposide phosphate; exemestane; fadrozole; fazarabine; fenretinide; filgrastim; finasteride; flavopiridol; flezelastine; fluasterone; fludarabine; fluorodaunorunicin hydrochloride; forfenimex; formestane; fostriecin; fotemustine; gadolinium texaphyrin; gallium nitrate; galocitabine; ganirelix; gelatinase inhibitors; gemcitabine; glutathione inhibitors; hepsulfam; heregulin; hexamethylene bisacetamide; hypericin; ibandronic acid; idarubicin; idoxifene; idramantone; ilmofosine; ilomastat; imidazoacridones; imiquimod; immunostimulant peptides; insulin-like growth factor- 1 receptor inhibitor; interferon agonists; interferons; interleukins; iobenguane; iododoxorubicin; ipomeanol, 4-; iroplact; irsogladine; isobengazole; isohomohalicondrin B; itasetron; jasplakinolide; kahalalide F; lamellarin-N triacetate; lanreotide; leinamycin; lenograstim; lentinan sulfate; leptolstatin; letrozole; leukemia inhibiting factor; leukocyte alpha interferon; leuprolide+estrogen+progesterone; leuprorelin; levamisole; liarozole; linear polyamine analogue; lipophilic disaccharide peptide; lipophilic platinum compounds; lissoclinamide 7; lobaplatin; lombricine; lometrexol; lonidamine; losoxantrone; lovastatin; loxoribine; lurtotecan; lutetium texaphyrin; lysofylline; lytic peptides; maitansine; mannostatin A; marimastat; masoprocol; maspin; matrilysin inhibitors; matrix metalloproteinase inhibitors; menogaril; merbarone; meterelin; methioninase; metoclopramide; MIF inhibitor; mifepristone; miltefosine; mirimostim; mismatched double stranded RNA; mitoguazone; mitolactol; mitomycin analogues; mitonafide; mitotoxin fibroblast growth factor-saporin; mitoxantrone; mofarotene; molgramostim; monoclonal antibody, human chorionic gonadotrophin; monophosphoryl lipid

A+myobacterium cell wall sk; mopidamol; multiple drug resistance gene inhibitor; multiple tumor suppressor 1 -based therapy; mustard anticancer agent; mycaperoxide

B; mycobacterial cell wall extract; myriaporone; N-acetyldinaline; N-substituted benzamides; nafarelin; nagrestip; naloxone+pentazocine; napavin; naphterpin; nartograstim; nedaplatin; nemorubicin; neridronic acid; neutral endopeptidase; nilutamide; nisamycin; nitric oxide modulators; nitroxide antioxidant; nitrullyn; 06- benzylguanine; octreotide; okicenone; oligonucleotides; onapristone; ondansetron; ondansetron; oracin; oral cytokine inducer; ormaplatin; osaterone; oxaliplatin; oxaunomycin; taxel; taxel analogues; taxel derivatives; palauamine; palmitoylrhizoxin; pamidronic acid; panaxytriol; panomifene; parabactin; pazelliptine; pegaspargase; peldesine; pentosan polysulfate sodium; pentostatin; pentrozole; perflubron; perfosfamide; perillyl alcohol; phenazinomycin; phenylacetate; phosphatase inhibitors; picibanil; pilocarpine hydrochloride; pirarubicin; piritrexim; placetin A; placetin B; plasminogen activator inhibitor; platinum complex; platinum compounds; platinum-triamine complex; porfimer sodium; porfiromycin; prednisone; propyl bis-acridone; prostaglandin J2; proteasome inhibitors; protein A-based immune modulator; protein kinase C inhibitor; protein kinase C inhibitors, microalgal; protein tyrosine phosphatase inhibitors; purine nucleoside phosphorylase inhibitors; purpurins; pyrazoloacridine; pyridoxylated hemoglobin polyoxyethylene conjugate; raf antagonists; raltitrexed; ramosetron; ras farnesyl protein transferase inhibitors; ras inhibitors; ras-GAP inhibitor; retelliptine demethylated; rhenium Re 186 etidronate; rhizoxin; ribozymes; RII retinamide; rogletimide; rohitukine; romurtide; roquinimex; rubiginone Bl; ruboxyl; safingol; saintopin; SarCNU; sarcophytol A; sargramostim; Sdi 1 mimetics; semustine; senescence derived inhibitor 1 ; sense oligonucleotides; signal transduction inhibitors; signal transduction modulators; single chain antigen binding protein; sizofiran; sobuzoxane; sodium borocaptate; sodium phenylacetate; solverol; somatomedin binding protein; sonermin; sparfosic acid; spicamycin D; spiromustine; splenopentin; spongistatin 1 ; squalamine; stem cell inhibitor; stem-cell division inhibitors; stipiamide; stromelysin inhibitors; sulfinosine; superactive vasoactive intestinal peptide antagonist; suradista; suramin; swainsonine; synthetic glycosaminoglycans; tallimustine; tamoxifen methiodide; tauromustine; tazarotene; tecogalan sodium; tegafur; tellurapyrylium; telomerase inhibitors; temoporfin; temozolomide; teniposide; tetrachlorodecaoxide; tetrazomine; thaliblastine; thiocoraline; thrombopoietin; thrombopoietin mimetic; thymalfasin; thymopoietin receptor agonist; thymotrinan; thyroid stimulating hormone; tin ethyl etiopurpurin; tirapazamine; titanocene bichloride; topsentin; toremifene; totipotent stem cell factor; translation inhibitors; tretinoin; triacetyluridine; triciribine; trimetrexate; triptorelin; tropisetron; turosteride; tyrosine kinase inhibitors; tyrphostins; UBC inhibitors; ubenimex; urogenital sinus-derived growth inhibitory factor; urokinase receptor antagonists; vapreotide; variolin B; vector system, erythrocyte gene therapy; velaresol; veramine; verdins; verteporfϊn; vinorelbine; vinxaltine; vitaxin; vorozole; zanoterone; zeniplatin; zilascorb; and zinostatin stimalamer. Preferred additional anti-cancer drugs are 5- fluorouracil and leucovorin. Additional cancer therapeutics include monoclonal antibodies such as rituximab, trastuzumab and cetuximab. In preferred examples, the anticancer agent is rapamycin. Rapamycin is an immunosuppressive macrolide antibiotic. Rapamycin inhibits T and B-cell proliferation. Rapamycin inhibits TOR (target of rapamycin) in the Ras/MAP kinase signalling pathway. The peptides of the invention may also comprise an imaging agent. The imaging agent may be a radiolabel or a fluorescent label. For cancer therapy, inclusion of an imaging agent will allow for the practitioner to monitor the size of the tumor in response to therapy.

Methods of Targeting

The invention also features methods of targeting an agent to a cell that expresses digalactosylceramide receptor comprising administering an SEB peptide comprising one or more agents to a cell that expresses digalactosylceramide receptor. For instance, an exemplary method is a method of targeting an agent to a tumor or an atherosclerotic plaque that expresses digalactosylceramide receptor in a subject comprising administering an SEB peptide comprising one or more agents to a tumor or an atherosclerotic plaque that expresses digalactosylceramide receptor.

For methods of targeting, in certain examples, the SEB peptide comprises the amino acid sequence ENENSFWYAMMPAPGDKFDQSKYLMMYNDN (SEQ ID NO: 6).

It is advantageous that the peptide, for example the SEB peptide, in certain examples, comprises a therapeutic or imaging agent. For example, the SEB peptide comprising the amino acid sequence of SEQ ID NO: 6 and an agent, for example a therapeutic or an imaging agent. The therapeutic agents may be selected from the group consisting of, but not limited to, protein synthesis modulators, apoptosis modulators, nitric oxide modulators, bactericides, a fungicides, anti neoplastic drugs, anti thrombotic drugs,hypochoesterolemic drugs, hypotriglyceridemic drugs, hypoglycemic drugs, anti-allergic drugs, antibiotics,tissue, anti-inflammatory drug, nerve regenerating drugs, and anti-arthritis drugs.

In other certain cases, the one or more therapeutic agents is an anticancer agent. The anticancer agent can be selected from, but not limited to, a chemotherapeutic agent, a peptide toxin, and a protein toxin. The anticancer agent can be any anticancer agent that is known to a clinician for use in anticancer therapy. The peptides of the invention may also comprise an imaging agent. The term "imaging agent" is meant to refer to compounds which can be detected.

Suitable imaging agents include commercially available agents used in positron emission tomography (PET), computer assisted tomography (CAT), single photon emission computerized tomography, x-ray, fluoroscopy, and magnetic resonance imaging (MRI).

Imaging agents include metals, radioactive isotopes and radioopaque agents (e.g., gallium, technetium, indium, strontium, iodine, barium, bromine and phosphorus-containing compounds), radiolucent agents, contrast agents, dyes (e.g., fluorescent dyes and chromophores) and enzymes that catalyze a colorimetric or fluorometric reaction. In general, such agents may be attached or entrapped using a variety of techniques as described above, and may be present in any orientation. See, e.g., U.S. Pat. Nos. 6,159,443 and 6,391,280, both of which are expressly incorporated by reference herein. Contrast agents according to the present invention are useful in the imaging modalities, such as X-ray contrast agents, light imaging probes, spin labels or radioactive units.

Examples of suitable materials for use as contrast agents in MRI include gadolinium chelates currently available, such as diethylene triamine pentaacetic acid (DTPA) and gadopentotate dimeglumine, as well as iron, magnesium, manganese, copper, and chromium.

Examples of materials useful for CAT and x-rays include iodine based materials, such as ionic monomers typified by diatrizoate and iothalamate, non-ionic monomers such as iopamidol, isohexol, and ioversol, non-ionic dimers, such as iotrol and iodixanol, and ionic dimers, for example, ioxagalte.

Both luminescent cyclen-based lanthamide chelates and those primarily yielding magnetic resonance signatures have been shown to be sensitive to changes in pH. Luminescent probes used for sensing pH changes typically detect changes in the fluorescence lifetime of the lanthamide ion as a function of pH. Analogously, magnetic resonance contrast agents which modulate the water proton relaxivity via changes in pH are useful in the instant invention. In both cases, by changing the pH in a given system, one can envision agents with enhanced contrast.

Accordingly, a pH sensitive contrast agent can be utilized at or near a cancer cell with the peptides of the invention. In this way, a change in pH causes the nuclear magnetic resonance relaxation properties of water protons or other nuclei in the aqueous medium to be changed in a manner that is reflective of pH. Examples of pH sensitive contrast agents that may be utilized include those agents that contain a lanthamide metal, such as Ce, Pr, Nd, Sm, Eu, Gd, Db, Dy, Ho, Er, Tm, Yb, and the like, or another paramagnetic element, such as Fe, Mn, 170, or the like. Specific contrast agents that may be utilized include H (2)(17)0, GdDOTA-4AmP(5-) which is described in Magn Reson Med. 2003 February;49(2):249 57, and Fe(III)meso-tetra(4- sulfonatophenyl)porphine (Fe-TPPS4) as described in Helpern et al. (1987) Magnetic Resonance in Medicine 5:302 305 and U.S. Pat. No. 6,307,372, which is incorporated herein by reference. In addition, Gd based with polyion, as described in Mikawa et al. Acad. Radiol (2002) 9(suppl 1): S 109 Sl 111, may be used in the invention.

As another alternative, a shift reagent may be provided in the aqueous medium surrounding the cancer cell. The shift reagent is configured such that a change in pH affects the chemical shift properties of the water protons or other nuclei in a manner that is reflective of pH. The change in chemical shift properties may then be measured using nuclear magnetic resonance to determine whether the active agent is biologically active. Exemplary shift reagents that may be used include those containing a lanthamide metal, such as Ce, Pr, Nd, Sm, Eu, Gd, Db, Dy, Ho, Er, Tm, or Yb, or another paramagnetic element. Examples of specific shift reagents that may be utilized include Tm(DOTP) (5-), the thulium (III) complex of 1 ,4,7, 10- tetraazacylododecane-N, N^N^W'-tetratmethylenephospate). Dy(PPP) (2)(7)- dysprosium tripolyphosphate, and the like.

A dual-contrast-agent strategy using two gadolinium agents, such as the pH- insensitive GdDOTP(5-) and the pH-sensitive GdDOTA-4AmP(5-), may be utilized to generate pH maps by MRI, as described in Magn Reson Med (2003) February;49(2):249 57.

Preferred agents for use with PET scan include 13N and fluorodeoxyglucose (FDG).

In preferred embodiments, the imaging agent may be a radiolabel or a fluorescent label. An imaging agent is especially useful for determining that an agent of interest, e.g. a therapeutic agent, has been targeted to the site of interest.

The invention also features methods of detecting a tumor or an atherosclerotic plaque that expresses digalactosylceramide receptor in a subject comprising administering an SEB peptide comprising an imaging agent to a tumor or an atherosclerotic plaque that expresses digalactosylceramide receptor in a subject.

The tumor or atherosclerotic plaque can be detected at different time points, using an imaging agent. For example, the tumor or atherosclerotic plaque can be determined prior to treatment, after a first round of therapy or treatment, after a second round of therapy or treatment and after any number of subsequent rounds of therapy or treatment. In this way, the clinician can monitor the size of the tumor or atherosclerotic plaque and determine the course of treatment. By "course of treatment" is meant the aggressiveness of treatment, including the dose of drug, the frequency of dosing or therapy.

The prognosis of the patient can be determined using this method. The method is also used to determine regression in tumor size or atherosclerotic plaque size after treatment with an anticancer or antiproliferative agent. Accordingly, The subject can be treated with at the same time as the administration of the peptides of the invention with an anticancer or antiproliferative agent.

The invention features methods of treating infection caused by a bacterial toxin in a subject, the method comprising administering to the subject a peptide, for example a SEB peptide, wherein the peptide is capable of binding to a digalactosylceramide (DAG) receptor, thereby treating infection caused by a bacterial toxin in a subject.

In certain preferred examples, the SEB peptide comprises the amino acid sequence ENENSFWYAMMPAPGDKFDQSKYLMMYNDN (SEQ ID NO: 6). Also featured are methods of inhibiting activity of a digalactosylceramide

(DAG) receptor in a subject comprising administering a composition selected from: an antibody, a nucleic acid or an oligomer that prevents activation of the DAG receptor. In certain cases, the subject is suffering from an infection caused by a bacterial toxin. The antibody, in preferred examples, has specificity for the amino acid sequence selected from RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID

NO: 5) and KKKVTAQEL (SEQ ID NO: 7) The nucleic acid can be any inhibitory nucleic acid. In certain preferred examples, the nucleic acid is selected from, but not limited to antisense, siRNA, shRNA, aptamers, PNA oligomers, and ribozymes.

Inhibitory nucleic acid molecules are essentially nucleobase oligomers that may be employed as single-stranded or double-stranded nucleic acid molecule to inhibit DAG activity. In one approach, the DAG inhibitory nucleic acid molecule is a double-stranded RNA used for RNA interference (RNAi)-mediated knock-down of Nrf2 gene expression. In one embodiment, a double-stranded RNA (dsRNA) molecule is made that includes between eight and twenty-five (e.g., 8, 10, 12, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25) consecutive nucleobases of a nucleobase oligomer of the invention. The dsRNA can be two complementary strands of RNA that have duplexed, or a single RNA strand that has self-duplexed (small hairpin (sh)RNA). Typically, dsRNAs are about 21 or 22 base pairs, but may be shorter or longer (up to about 29 nucleobases) if desired. Double stranded RNA can be made using standard techniques (e.g., chemical synthesis or in vitro transcription). Kits are available, for example, from Ambion (Austin, Tex.) and Epicentre (Madison, Wis.). Methods for expressing dsRNA in mammalian cells are described in Brummelkamp et al. Science 296:550-553, 2002; Paddison et al. Genes & Devel. 16:948-958, 2002. Paul et al. Nature Biotechnol. 20:505-508, 2002; Sui et al. Proc. Natl. Acad. Sci. USA 99:5515- 5520, 2002; Yu et al. Proc. Natl. Acad. Sci. USA 99:6047-6052, 2002; Miyagishi et al. Nature Biotechnol. 20:497-500, 2002; and Lee et al. Nature Biotechnol. 20:500- 505 2002, each of which is hereby incorporated by reference. An inhibitory nucleic acid molecule that "corresponds" to an DAG gene comprises at least a fragment of the double-stranded gene, such that each strand of the double-stranded inhibitory nucleic acid molecule is capable of binding to the complementary strand of the target DAG gene. The inhibitory nucleic acid molecule need not have perfect correspondence to the reference DAG sequence. In one embodiment, an siRNA has at least about 85%, 90%, 95%, 96%, 97%, 98%, or even 99% sequence identity with the target nucleic acid. For example, a 19 base pair duplex having 1-2 base pair mismatch is considered useful in the methods of the invention. In other embodiments, the nucleobase sequence of the inhibitory nucleic acid molecule exhibits 1, 2, 3, 4, 5 or more mismatches.

By "antisense nucleic acid", it is meant a non-enzymatic nucleic acid molecule that binds to target RNA by means of RNA- RNA or RNA-DNA interactions and alters the activity of the target RNA (for a review, see Stein et al. 1993; Woolf et al., U.S. Pat. No.5, 849, 902). Typically, antisense molecules are complementary to a target sequence along a single contiguous sequence of the antisense molecule. An antisense molecule can bind to substrate such that the substrate molecule forms a loop, and/or an antisense molecule can bind such that the antisense molecule forms a loop. Thus, the antisense molecule can be complementary to two (or even more) noncontiguous substrate sequences or two (or even more) non-contiguous sequence portions of an antisense molecule can be complementary to a target sequence or both. For a review of current antisense strategies, see Schmajuk NA et al, 1999; Delihas N et al, 1997; Aboul-Fadl T, 2005.).

RNA interference (RNAi) is a method for decreasing the cellular expression of specific proteins of interest (reviewed in Tuschl, Chembiochem 2:239-245, 2001; Sharp, Genes & Devel. 15:485-490, 2000; Hutvagner and Zamore, Curr. Opin. Genet. Devel. 12:225-232, 2002; and Harmon, Nature 418:244-251, 2002). In RNAi, gene silencing is typically triggered post-transcriptionally by the presence of double- stranded RNA (dsRNA) in a cell. This dsRNA is processed intracellular^ into shorter pieces called small interfering RNAs (siRNAs). The introduction of siRNAs into cells either by transfection of dsRNAs or through expression of shRNAs using a plasmid-based expression system is currently being used to create loss-of-function phenotypes in mammalian cells. The inhibitory nucleic acid can bind to and inhibit the activity of the DAG receptor, in certain examples. siRNAs that target DAG Receptor decrease DAG activity in vivo.

In certain preferred examples, oligomers are used. The oligomers can be inhibitory oligomers. In certain preferred examples, the oligomer is at least a 3-mer, 4-mer, 5-mer, 6-mer, 7-mer, 8-mer, 9-mer or 10-mer of the amino acid sequence comprising RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID NO: 5). The oligomers can be of any length so long as they retain their inhibitory activity. Modifications to the oligomers are possible, as described herein.

The invention also features methods of treating infection caused by a bacterial toxin in a subject, the method comprising administering a composition selected from: an antibody, a nucleic acid or an oligomer that prevents activation of the DAG receptor, wherein the peptide is capable of binding to a digalactosylceramide (DAG) receptor, thereby treating infection caused by a bacterial toxin in a subject. The bacterial toxin can be from a Gram negative bacteria. In certain preferred examples, the bacterial toxin is selected from Escherichia coli or Psudomonas aeruginosa. The E. coli bacterial toxin is, in preferred examples, verotoxin or Shiga toxin. The Escherichia coli or Pseudomonas aeruginosa binds a glycolipid receptor selected from a digalactosylceramide receptor or a Globotriosylceramide Receptor (GbOse3Cer).

In any of the cases, the infection caused by a bacterial toxin can be localized anywhere, however preferred locations are the lung, kidney, spleen, pancreas or gastroinstestinal tract.

In any of the above-mentioned methods, the subject to be treated is a mammal. In preferred examples, the subject is a human.

IV. ANTIBODIES The term "antibodies" is used herein to refer to immunoglobulin molecules and immunologically active portions of immunoglobulin molecules. Exemplary antibody molecules are intact immunoglobulin molecules, substantially intact immunoglobulin molecules and portions of an immunoglobulin molecule, including those portions known in the art as Fab, Fab¹, F(ab')-sub.2 and F(v) as well as chimeric antibody molecules.

Included in the invention are methods of making one or more antibodies that bind at least one staphylococcal enterotoxin receptor, where the method comprise administering to a mammal an amount of any of the SEB peptides as described herein. The amount of peptide that is administered to the mammal is given in an amount that is sufficient to elicit production of one or more antibodies.

Antibodies according to the present invention can be inhibitory antibodies, for example antibodies that bind to the DAG receptor to inhibit receptor activity.

An antibody of the present invention is typically produced by immunizing a mammal with an immunogen or vaccine containing one or more peptides of the invention, or a structurally and/or antigenically related molecule, to induce, in the mammal, antibody molecules having immunospecificity for the immunizing peptide or peptides. The peptide(s) or related molecule(s) may be monomeric, polymeric, conjugated to a carrier, and/or administered in the presence of an adjuvant. The antibody molecules may then be collected from the mammal if they are to be used in immunoassays or for providing passive immunity.

The antibody molecules of the present invention may be polyclonal or monoclonal. Monoclonal antibodies may be produced by methods known in the art. Portions of immunoglobulin molecules may also be produced by methods known in the art.

The antibody of the present invention may be contained in various carriers or media, including blood, plasma, serum (e.g., fractionated or unfractionated serum), hybridoma supernatants and the like. Alternatively, the antibody of the present invention is isolated to the extent desired by well known techniques such as, for example, by using DEAE Sephadex, or affinity chromatography. The antibodies may be purified so as to obtain specific classes or subclasses of antibody such as IgM, IgG, IgA, IgG.sub.l, IgG.sub.2, IgG.sub.3, IgG.sub.4 and the like. Antibody of the IgG class are preferred for purposes of passive protection. The presence of the antibodies of the present invention, either polyclonal or monoclonal, can be determined by various assays. Assay techniques include, but are not limited to, immunobinding, immunofluorescence (IF), indirect immunofluorescence, immunoprecipitation, ELISA, agglutination and Western blot techniques. The antibodies of the present invention have a number of diagnostic and therapeutic uses. The antibodies can be used as an in vitro diagnostic agent to test for the presence of various staphylococcal and streptococcal pyrogenic exotoxins in biological samples in standard immunoassay protocols and to aid in the diagnosis of various diseases related to the presence of bacterial pyrogenic exotoxins. Preferably, the assays which use the antibodies to detect the presence of bacterial pyrogenic exotoxins in a sample involve contacting the sample with at least one of the antibodies under conditions which will allow the formation of an immunological complex between the antibody and the toxin that may be present in the sample. The formation of an immunological complex if any, indicating the presence of the toxin in the sample, is then detected and measured by suitable means. Such assays include, but are not limited to, radioimmunoassays, (RIA), ELISA, indirect immunofluorescence assay, Western blot and the like. The antibodies may be labeled or unlabeled depending on the type of assay used. Labels which may be coupled to the antibodies include those known in the art and include, but are not limited to, enzymes, radionucleotides, fluorogenic and chromogenic substrates, cofactors, biotin/avidin, colloidal gold and magnetic particles. Modification of the antibodies allows for coupling by any known means to carrier proteins or peptides or to known supports, for example, polystyrene or polyvinyl microtiter plates, glass tubes or glass beads and chromatographic supports, such as paper, cellulose and cellulose derivatives, and silica.

Such assays may be, for example, of direct format (where the labeled first antibody reacts with the antigen), an indirect format (where a labeled second antibody reacts with the first antibody), a competitive format (such as the addition of a labeled antigen), or a sandwich format (where both labeled and unlabelled antibody are utilized), as well as other formats described in the art. In one such assay, the biological sample is contacted to antibodies of the present invention and a labeled second antibody is used to detect the presence of staphylococcal and streptococcal pyrogenic exotoxins, to which the antibodies are bound. The antibodies of the present invention are also useful as therapeutic agents in the prevention and treatment of diseases caused by the deleterious effects of staphylococcal and streptococcal pyrogenic exotoxins.

In certain embodiments, the peptide antibodies are useful to mitigate multidrug resistant bacterial infections or Methicillin resistant bacterial infection. Methicillin resistant Staphylococcus aureus (MRSA) is the term used for bacteria of the Staphylococcus aureus group that are resistant to the usual antibiotics used in the treatment of infections with such organisms. Traditionally MRSA stood for Methicillin resistance, but the term has become increasingly known in the art to refer to a multi-drug resistant group of bacteria. Such bacteria often have resistance to many antibiotics traditionally used against S. aureus.

Drug-resistant bacteria can complicate treatment after many surgical procedures. In particular, Methicillin-resistant Staphylococcus aureus (MRSA), which has been found in many healthcare settings, can be a serious post-operative complication. In a study published in the April issue of the American Journal of Ophthalmology, researchers found MRSA infections in the eyes of 12 patients after refractive surgery. Moreover, Colonization of MRSA has been found in 1.5% of the general population, but as many as 9.4% of those were exposed to a healthcare facility. However, strains of MRSA are emerging in the community. These so-called community strains tend to be resistant only to beta-lactam antibiotics, unlike the hospital strains, which usually demonstrate multiple drug resistance. Community-acquired MRSA is becoming a significant problem, with the prevalence of MRSA among community isolates expected to reach as high as 25% in the next decade, as reported in Science News Daily on the world wide web at http://www.sciencedaily.com/releases/2007/04/070410091413.htm.

The antibodies are generally administered with a physiologically acceptable carrier or vehicle therefor. A physiologically acceptable carrier is one that does not cause an adverse physical reaction upon administration and one in which the antibodies are sufficiently soluble and retain their activity to deliver a therapeutically effective amount of the compound. The therapeutically effective amount and method of administration of the antibodies may vary based on the individual patient, the indication being treated and other criteria evident to one of ordinary skill in the art. A therapeutically effective amount of the antibodies is one sufficient to attenuate the dysfunction without causing significant side effects such as non-specific T cell lysis or organ damage. The route(s) of administration useful in a particular application are apparent to one or ordinary skill in the art.

Routes of administration of the antibodies include, but are not limited to, parenteral, and direct injection into an affected site. Parenteral routes of administration include but are not limited to intravenous, intramuscular, intraperitoneal and subcutaneous.

The present invention includes compositions of the antibodies described above, suitable for parenteral administration including, but not limited to, pharmaceutically acceptable sterile isotonic solutions. Such solutions include, but are not limited to, saline and phosphate buffered saline for intravenous, intramuscular, intraperitoneal, subcutaneous or direct injection into a joint or other area.

Antibodies for use to elicit passive immunity in humans are preferably obtained from other humans previously inoculated with compositions comprising one or more of the SEB peptide sequences of the invention. Alternatively, antibodies derived from other species may also be used. Such antibodies used in therapeutics suffer from several drawbacks such as a limited half-life and propensity to elicit an immune response. Several methods have been proposed to overcome these drawbacks. Antibodies made by these methods are encompassed by the present invention and are included herein. One such method is the "humanizing" of non- human antibodies by cloning the gene segment encoding the antigen binding region of the antibody to the human gene segments encoding the remainder of the antibody. Only the binding region of the antibody is thus recognized as foreign and is much less likely to cause an immune response. An article describing such antibodies is Reichmann et al., "Reshaping Human Antibodies for Therapy", Nature 332:323-327 (1988), which is incorporated herein by reference.

In providing the antibodies of the present invention to a recipient mammal, preferably a human, the dosage of administered antibodies will vary depending upon such factors as the mammal's age, weight, height, sex, general medical condition, previous medical history and the like. An appropriate dosage can be determined by a skilled practitioner.

In general, it is desirable to provide the recipient with a dosage of antibodies which is in the range of from about 5 mg/kg to about 20 mg/kg body weight of the mammal, although a lower or higher dose may be administered. In general, the antibodies will be administered intravenously (IV) or intramuscularly (IM). The antibodies of the present invention are intended to be provided to the recipient subject in an amount sufficient to prevent, or attenuate the severity, extent or duration of the deleterious effects of staphylococcal and streptococcal pyrogenic exotoxins.

V. PHARMACEUTICAL COMPOSITIONS

While one or more compounds of the invention may be administered alone, the invention features in other aspects pharmaceutical compositions comprising any of the peptides as described herein and a pharmaceutically acceptable carrier, i.e., pharmaceutically acceptable organic or inorganic carrier substances suitable for parenteral, oral or other desired administration and which do not deleteriously react with the active compounds and are not deleterious to the recipient thereof. Suitable pharmaceutically acceptable carriers include but are not limited to water, salt solutions, alcohol, vegetable oils, polyethylene glycols, gelatin, lactose, amylose, magnesium stearate, talc, silicic acid, viscous paraffin, perfume oil, fatty acid monoglycerides and diglycerides, petroethral fatty acid esters, hydroxymethyl- cellulose, polyvinylpyrrolidone, etc. The pharmaceutical preparations can be sterilized and if desired mixed with auxiliary agents, e.g., lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, colorings, flavorings and/or aromatic substances and the like which do not deleteriously react with the active compounds.

A pharmaceutical composition according to this invention comprises the novel peptide or peptide conjugate described herein in a formulation that, as such, is known in the art. Thus the compositions may be in the form of a lyophilized particulate material, a sterile or aseptically produced solution, a tablet, an ampule, etc. Vehicles, such as water or other aqueous solutions preferably buffered to a physiologically acceptable pH (as in phosphate buffered saline) or other inert solid or liquid material such as normal saline or various buffers may be present. The particular vehicle is not critical, and those skilled in the art will readily know which vehicle to use for any particular utility described herein. In certain embodiments, the polymer or polymer conjugate may be maintained in liquid or lyophilized form. The peptide may be mixed with an adjuvant. The peptide also may be bound to a non-toxic non-host protein carrier to form a conjugate or it may be bound to a saccharide carrier and/or a non-toxic non-host protein carrier to form a conjugate.

In general terms, a pharmaceutical composition is prepared by mixing, dissolving, binding or otherwise combining the peptide or peptide and agent(s) conjugate of this invention with one or more water-insoluble or water-soluble aqueous or non-aqueous vehicles. If necessary, another suitable additive or adjuvant is included. It is imperative that the vehicle, carrier or excipient, as well as the conditions for formulating the composition are such that do not adversely affect the biological or pharmaceutical activity of the peptide or peptide and agent(s) conjugate.

For parenteral application, particularly suitable are solutions, preferably oily or aqueous solutions as well as suspensions, emulsions, or implants, including suppositories. Ampules are convenient unit dosages.

For enteral application, particularly suitable are tablets, dragees or capsules having talc and/or carbohydrate carrier binder or the like, the carrier preferably being lactose and/or corn starch and/or potato starch. A syrup, elixir or the like can be used wherein a sweetened vehicle is employed. Sustained release compositions can be formulated including those wherein the active component is protected with differentially degradable coatings, e.g., by microencapsulation, multiple coatings, etc.

Therapeutic compounds of the invention also may be incorporated into liposomes. The incorporation can be carried out according to known liposome preparation procedures, e.g. sonication and extrusion. Suitable conventional methods of liposome preparation are also disclosed in e.g. A. D. Bangham et al., J. MoI. Biol.,

23:238-252 (1965); F. Olson et al., Biochim. Biophys. Acta, 557:9-23 (1979); F.

Szoka et al., Proc. Nat. Acad. Sci., 75:4194-4198 (1978); S. Kim et al., Biochim.

Biophys. Acta, 728:339-348 (1983); and Mayer et al., Biochim. Biophys. Acta, 858:161-168 (1986).

The administration of the agents including peptide and antibody compositions of the invention may be for either "prophylactic" or "therapeutic" purpose. In certain cases, it may be desirable to administer the peptides for a prophylactic use. Accordingly, when provided prophylactically, the agents are provided in advance of any symptom. The prophylactic administration of the agent serves to prevent or ameliorate any subsequent deleterious effects of the disease, disorder or condition being treated.

When provided therapeutically, the agent is provided at (or shortly after) the onset of a symptom of the disease, disorder or condition. The pharmaceutical compositions of the present invention may, thus, be provided either prior or after the disease, disorder or condition to be treated.

Compositions and methods of the invention may be used in combination with any conventional therapy known in the art. In one embodiment, peptides having anti proliferative or apoptotic activity may be used in combination with any anti- neoplastic therapy known in the art. Exemplary anti-neoplastic therapies include, for example, chemotherapy, cryotherapy, hormone therapy, radiotherapy, and surgery.

Anticancer agents have been described herein.

The liposome may be made from one or more of the conjugates discussed above alone, or more preferably, in combination with any of the conventional synthetic or natural phospholipid liposome materials including phospholipids from natural sources such as egg, plant or animal sources such as phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, sphingomyelin, phosphatidylserine or phosphatidylinositol. Synthetic phospholipids also may be used e.g., dimyristoylphosphatidylcholine, dioleoylphosphatidylcholine, dioleoylphosphatidycholine and corresponding synthetic phosphatidylethanolamines and phosphatidylglycerol s. Cholesterol or other sterols, cholesterol hemisuccinate, glycolipids, l,2-bis(oleoyloxy)-3-(tr- imethyl ammonio)propane (DOTAP), N-[I -(2,3- dioleoyl)propyl]-N,N,N-trimethy- lammonium chloride (DOTMA), and other cationic lipids may be incorporated into the liposomes. The relative amounts of the one or more compounds and additives used in the liposomes may vary relatively widely. Liposomes of the invention suitably contain about 60 to 90 mole percent of natural or synthetic phospholipid; cholesterol, cholesterol hemisuccinate, fatty acids or cationic lipids may be used in amounts ranging from 0 to 50 mole percent; and the one or more therapeutic compounds of the invention may be suitably present in amounts of from about 0.01 to about 50 mole percent.

It will be appreciated that the actual preferred amounts of active compounds used in a given therapy will vary according to the specific compound being utilized, the particular compositions formulated, the mode of application, the particular site of administration, and the disease or disorder being treated. Optimal administration rates for a given protocol of administration can be readily ascertained by those skilled in the art using conventional dosage determination tests conducted with regard to the foregoing guidelines.

A variety of administration routes are available. The methods of the invention, generally speaking, may be practiced using any mode of administration that is medically acceptable, meaning any mode that produces effective levels of the active compounds without causing clinically unacceptable adverse effects. Administration of compounds of the invention may be made by a variety of suitable routes including oral, topical (including transdermal, buccal or sublingal), nasal and parenteral (including intraperitoneal, subcutaneous, intravenous, intradermal or intramuscular injection) Other modes of administration include rectal, intraocular, buccal, intravaginal, intracisternal, intracerebroventricular, intratracheal, nasal, transdermal, within/on implants, e.g., fibers such as collagen, osmotic pumps, or grafts comprising appropriately transformed cells, etc., or parenteral routes. Compounds of the invention may be used in therapy in conjunction with other medicaments such those with recognized pharmacological activity to treat any of the conditions as described herein.

VI. KITS In yet another aspect, the invention provides kits or pharmaceutical packs.

For all therapeutic, prophylactic and diagnostic uses, the peptides and antibodies of the invention, alone or linked to a therapeutic agent or carrier, as well as antibodies and other necessary reagents and appropriate devices and accessories may be provided in kit form so as to be readily available and easily used. Kits or pharmaceutical packs can be used according to any one of the methods as described herein, and accordingly provide instructions for use.

The kits or pharmaceutical packs may comprise a containers, e.g., a flexible packet, vial, ampoule, bottle and the like, filled with one or more of the ingredients of the compositions of the invention. Optionally associated with such container(s) can be a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use or sale for human administration. In a preferred embodiment, the compositions of the present invention can be presented as single- or multi-dose forms in a flexible packet. Preferably, the compositions of the present invention are packaged in the container with an appropriate dosage and instructions for use.

The following examples are offered by way of illustration, not by way of limitation. While specific examples have been provided, the above description is illustrative and not restrictive. Any one or more of the features of the previously described embodiments can be combined in any manner with one or more features of any other embodiments in the present invention. Furthermore, many variations of the invention will become apparent to those skilled in the art upon review of the specification. The scope of the invention should, therefore, be determined not with reference to the above description, but instead should be determined with reference to the appended claims along with their full scope of equivalents.

EXAMPLES Studies suggest that Staphylococcal Enterotoxin B (SEB) is initially harbored in the kidney by binding to digalactosylceramide molecules in the proximal tubular cells. However, little is known about the peptide motif within SEB that binds to these cells and imparts toxic effects. The results presented herein employ human kidney proximal tubular cells (PT) in a systematic study on the binding of various peptides and peptide analogs of SEB, and establish a structure-function relationship in regard to peptide motifs specific to induce cell proliferation and apoptosis.

Here we have investigated a series of synthetic SEB peptides to evaluate their binding to PT cells, digalactosylceramide; a putative receptor, and established structure function relationship in regard to peptide motif specific to induce cell proliferation and apoptosis.

Example 1. Effect Of ¹²⁵I-SEB peptide concentration and various inhibitory molecules on the on the binding of ¹²⁵I-SEB peptides to human PT cells.

Effect of ¹²⁵I-SEB peptide concentration on binding to cultured human proximal tubular cells.

Using cultured human proximal tubular (PT) cells a high affinity binding of ¹²⁵I-SEB peptides (Figure 2) was first described. SEB 191 -220 bound to PT cells with the highest affinity, as compared to four other SEB peptides as shown in Figure 2.

Effect of SEB peptide antibody, human kidney neutral glycosphingolipids and endoglycoceramidase on the binding of^{1 5}I-SEB pp tides to human PT cells. Peptide 191 -220 was bound with the highest affinity, as shown in Figure 1. This binding was inhibited by the presence of glycolipids and endoglycoceramidase, but this effect was less so when antibody against SEB was used, as shown in Figure 3. These findings suggest that SEB peptide 191-220 represents the major peptide domain that binds to the glycosphigolipid domain in the kidney cells. This finding is in agreement with previous studies that show that digalactosylceramide is the receptor for SEB in cultured human kidney cells (3, 4). The other peptides that were examined did not bind to cells with the same affinity as peptide 191-220; however binding was still affected by the use of antibody to SEB, glycolipids and by treatment with endoglycoceramidase. For example for peptide 93-120, binding was significantly affected by the use of SEB antibody but relatively less so with the use of glycolipids and endoglycoceramidase treatment. One explanation for these results is that the clustering of the glycosphigolipid on the lipid rafts and the dissolution/ perturbation of the lipid raft due to endoglycoceramidase treatment may have a general adverse effect on the binding of these SEB peptides.

Example 2. Effect of SEB peptide concentration on human proximal tubular cell proliferation.

Cell proliferation was measured employing multiple criteria. These were (3H)thymidine incorporation into DNA (shown in open bars in Figure 4) and employing trypan blue dye exclusion viable cell count assay (solid bars in Figure 4). As shown in Figure 4 A, incubation of PT cells with SEB 130-160 resulted in a concentration-dependent decrease in cell proliferation. In contrast, in PT cells SEB 93-112 exerted a concentration-dependent increase in cell proliferation, as shown in Figure 4B. Maximum increase in cell proliferation (-2.5 fold) occurred with

0.625 μg /ml SEB 93-112. However, further increasing the amount of SEB 93-112 did not increase cell proliferation. SEB peptide 191-220 had a modest increase - 30% in cell proliferation compared to control at a concentration of 0.15-0.3 μg /ml (Figure 4C). However, at relatively higher concentrations (0.625-2.5 g/ml) SEB 191-220 did not alter cell proliferation significantly (Figure 4C). Light microscope studies revealed that PT cells incubated with SEB 93-112 were highly densely packed (Figure 5C) and appeared small in size as compared to control, untreated cells or SEB treated cells, respectively (Figure 5A,B). In contrast, PT cells incubated with peptide 130-160 were relatively less dense and had a polygonal morphology (Figure 5D).

Example 3. Effects of amino acid composition of SEB peptide 130-160 on cell proliferation and apoptosis.

Cell Proliferation. SEB 130-160 exerted a 6-fold decrease in cell proliferation as compared to control, as shown in Table 1 below. In Table 1, SEB 130-160 and its amino acid substitutes were prepared as described in Methods, below. These peptides were added to PT cells grown in 96 well trays. (3H)-thymidine incorporation into DNA was measured as described previously and known to one of skill in the art. The data was obtained from two separate experiments and analyzed in six micro liter wells each. Amino acid substitution of this sequence, as well as deletion of some amino acids within this sequence imparted a partial or complete restoration in PT cell proliferation.

Table 1

Apoptosis,

To determine phenotypic changes that may accompany SEB peptide 130-160 in inhibition of cell proliferation we examined the effect of SEB, SEB peptide 130- 160 and SEB peptide 93-112 on apoptosis. The ability of antibody against SEB and SEB peptide 130-160 to mitigate apoptosis was also examined by staining the cells with 4',6-Diamidino-2-phenylindole (DAPI) reagent. DAPI stains intact nuclei blue; whereas fragmented DNA, for example apoptotic cells, is stained white. Figure 6A shows the DAPI-stained cells and Figure 6B in the corresponding quantitative bar chart represent the percentage of apoptotic cells.

SEB (1-2 μg/ml) exerted a concentration-dependent increase in number in apoptotic cells as compared to control (Figure 6B). SEB peptide 130-160 also induced apoptosis in these kidney cells. However, SEB peptide 93-112 did not alter apoptosis. Moreover, SEB and/or SEB peptide 130-160-induced apoptosis was completely reversed by the use of ether SEB or SEB 130-160 antibodies (Figure 6B).

Example 4. Effect of SEB and SEB peptide 130-160 on neutral sphingomyelinaselceramide pathway.

To determine biochemical mechanism by which SEB and SEB peptide 130- 160 may induce apoptosis we examined the effect of these compounds on the activity of neutral sphingomyelinase and ceramide and sphingomyelin mass. As shown in Figure 7 A, incubation of PT cells with SEB and SEB peptide 130-160 stimulated the activity of neutral sphingomyelinase"" 1.5 fold and -1.8 fold, respectively. Metabolic labeling using C4C) palmitic acid followed by quantitation revealed that SEB peptide 130-160 increased the cellular level of ceramide and decreased the cellular level of sphingomyelin as compared to control. The data presented herein employed cultured human kidney proximal tubular cells and revealed that SEB peptide 191-220 binds to the digalactosylceramide receptor in PT cells with high affinity and specificity. Moreover, this SEB peptide imparts a modest effect on PT cell proliferation. SEB 93-112 was also found to bind PT cells but with a lower affinity as compared to SEB 191-220. However, SEB 191- 220 exerted a marked increase in PT cell proliferation as compared to SEB 93-112. In contrast, SEB 130-160 exhibited less binding but exerted a concentration-dependent and profound inhibition (4-6 fold) in cell proliferation. Moreover, peptides from this 30-mer which did not contain the KKKVTAQEL (AA 152-160) sequence restored cell proliferation. Furthermore, the mechanistic studies revealed that SEB 130-160 induces apoptosis via activation of the neutral sphingomyelinase-ceramide pathway. Moreover, SEB and SEB peptide 130-160 mediated apoptosis was mitigated by preincubation of cells with either SEB antibody or antibody against SEB peptide 130- 160.

In sum, the data show that specific peptide domains within the SEB molecule (SEB 190-220 ) bind to the digalactosylceramide receptor present in PT cells. SEB 93-112 binding results in the activation of downstream signaling events that ultimately lead to cell proliferation. In contrast, SEB peptide 130-160, which is bound to PT cells, activates the N-SMase-ceramide pathway to induce apoptosis and that in turn, may contribute to a marked decrease in cell proliferation. Most important, the toxic/apoptotic effect of SEB peptide 130-160 was completely reversed by the use of corresponding antibodies.

Methods

The present invention was performed with the following methods and materials.

Isotopes and chemicals.

¹²⁵I-SEB (specific activity 644 MBq/J,g iodine) was purchased from Dupont, New England Nuclear. All other biochemicals were purchased from Sigma Chemical Co., St.Lous, MO. Rhodococcus endoglycoceramidase was purchased from Genzyme Corporation, Boston. Human kidney neutral glycosphingolipids were prepared in our laboratory (18) and characterized employing HPTLC and HPLC techniques (4). Lipoprotein deficient serum (LPDS) was prepared from lipoprotein-deficient plasma by precipitation with thrombin as described and determined to be free from glycosphingolipids and cholesterol (19)

Radiolabeling of SEB peptides with ¹²⁵I-SEB.

SEB synthetic peptides were prepared by Peninsula Labs (now a subsidiary of Bachem, Torrance, CA). Such peptides were classified based upon the SEB amino acid sequence they represented. SEB peptides were labeled with ¹²⁵I-SEB using iodogen (20), solubilized in sample buffer and subjected to polyacrylamide gel electrophoresis on phorcast gels at 12.5 miliamp/gel for 24 h at room temperature. Appropriate standard proteins of known MW were also electrophoresed, a portion of the gel including the standard molecular weight proteins was excised and stained with coomassie blue at 6O⁰C for 5-10 min. The gel area corresponding to individual SEB was excised, eluted and dialyzed. The material was freeze-dried, solubilized and assessed for purity by SDS-PAGE analysis. Such preparations were free from contaminating proteins.

Cells.

Cultured human PT cells were prepared from autopsy kidney as described previously (21). Cells were trypsinized and seeded (I x 105) in 60 x 15 mm plastic Petri dishes and grown for 6 days in minimum essential medium containing heat inactivated-dialyzed 10% fetal calf serum and no antibiotics. On the 6th day, medium was removed, cells were washed with phosphate buffered saline (PBS) and incubation continued for 24 h in medium containing LPDS (1 mg protein/ml).

⁵I-SEB peptide binding assay.

Unless otherwise described in the text, the following assay was adopted to measure the binding of ¹²⁵I-SEB peptide to PT cells. Medium was removed from cells primed with LPDS. Next, fresh medium (1 ml) and ¹²⁵I-SEB peptide (1 μg/ml) plus a twenty fold excess of unlabeled SEB/peptide was added and incubation continued for 2 h at 37⁰C. Next, the medium was discarded and the cells washed with 5 ml of PBS containing 0.2% bovine serum albumin (maintained at 4°C) and 5 times with PBS. The monolayer was solubilized in 1 N NaOH, protein and radioactivity was measured according to Lowry et.al (12) and scintillation spectrometry, respectively. Specific binding of 1251 toxin peptide was calculated by subtracting the data obtained in the absence of unlabeled toxin/peptide from the data obtained in the presence of 20 fold excess of unlabeled toxin/peptide (3, 4).

Incubation of cells with endoglycoceramidase.

Cells preincubated with medium containing LPDS were further incubated with endoglycoceramidase (0.15 milliunits-0.6 millunits/ml) for 1 h at 37°C, washed and the binding of ¹²⁵I-SEB peptide pursued as described above.

Incubation of cells with glycosphingolipids and SEB peptide antibodies.

Glycosphingolipids were taken into a sterile glass tube and dried in N2 atmosphere. Then medium containing LPDS was added, sonicated and suitable aliquots added to the assay mixture. Cells preincubated with medium containing ' LPDS were further incubated with fresh medium containing human kidney neutral glycosphingolipids and ¹²⁵I-SEB peptide. After incubation for 2 h at 37°C the assay was terminated and the binding of toxin to PT cells measured. Similarly, cells were incubated with SEB peptide antibodies for 1 h prior to the ¹²⁵I-SEB peptide binding assay.

Measurement of cell proliferation.

(3H)Thymidine incorporation into DNA and trypan blue exclusion assay were employed to measure the effect of SEB, and SEB peptides on cell proliferation. Further details are described in the legend to individual figures.

The present invention has been described in detail, including the preferred embodiments thereof. However, it will be appreciated that those skilled in the art, upon consideration of the present disclosure, may make modifications and/or improvements of this invention and still be within the scope and spirit of this invention as set forth in the following claims.

All publications and patent documents cited in this application are incorporated by reference in their entirety for all purposes to the same extent as if each individual publication or patent document were so individually denoted. By their citation of various references in this document, Applicants do not admit any particular reference is "prior art" to their invention.

REFERENCES

1. Campbell WN, Fitzpatrick M, Ding X, Jett M, Gemski P, Goldblum SE.SEB is cytotoxic and alters EC barrier function through protein tyrosine phosphorylation in vitro. Am J Physiol273 (1 Pt 1): L31-9, 1997.

2. Canonico PG, Henriksen EL, Ayala E, Bowman DG. Subcellular localization of staphylococcal enterotoxin B in rabbit kidney. Am J Physiol 226 (6): 1333-7,1974.

3. Chatterjee S, Jett M. Glycosphingolipids: the putative receptor for Staphylococcus aureus enterotoxin-B in human kidney proximal tubular cells. MoI Cell Biochem 113 (1): 25-31, 1992.

4. Chatterjee S, Khullar M, Shi \N. Digalactosylceramide is the receptor for staphylococcal enterotoxin-B in human kidney proximal tubular cells. Glycobiology 5 (3): 327-33, 1995.

5. Hoffman M, Tremaine M, Mansfield J, Betley M. Biochemical and mutational analysis of the histidine residues of staphylococcal enterotoxin A. Infect Immun 64 (3): 885-90, 1996.

6. Jardetzky TS, Brown JH, Gorga JC, Stern LJ, Urban RG, Chi YI, Staufacher C, Strominger JL, Wiley DC. Three-dimensional structure of a human class II histocompatibility molecule complexed with superantigen. Nature 368 (6473): 711-8, 1994.

7. Jett M, lonin, B., Das, R., and Neill, R. The Staphylococcal Enterotoxins. Molecular Medical Microbiology, M. Sussman Ed, 2001. 18 Effects of SEB peptides on Human Renal Cells Chatterjee, et al

8. Jett M, Neill R, Welch C, Boyle T, Bernton E, Hoover D, Lowell G, Hunt RE, Chatterjee S, Gemski P. Identification of staphylococcal enterotoxin B sequences important for induction of lymphocyte proliferation by using synthetic peptide fragments of the toxin. Infect Immun 62 (8): 3408-15,

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9. Kappler JW, Herman A, Clements J, Marrack P. Mutations defining functional regions of the superantigen staphylococcal enterotoxin B. J Exp Med 175 (2): 387-96, 1992.

10. Morris EL, Hodoval LF, Beisel WR. The unusual role of the kidney during intoxication of monkeys by intravenous staphylococcal enterotoxin B. J Infect Dis 1 17 (4): 273-84, 1967.

1 1. Normann SJ, Jaeger RP, Johnsey RT. Pathology of experimental enterotoxemia. The in vivo localization of staphylococcal enterotoxin B. Lab Invest 20 (1): 17-25, 1969.

12. Normann SJ, Stone CM. Renal lysosomal catabolism of staphylococcal enterotoxin B. Lab Invest 27 (2): 236-41, 1972.

13. Papageorgiou AC, Tranter HS, Acharya KR. Crystal structure of microbial superantigen staphylococcal enterotoxin B at 1.5 A resolution: implications for superantigen recognition by MHC class II molecules and T-cell receptors. J MoI Bioi 277 (1): 61-79, 1998.

14. Rapoport MI, Hodoval LF, Beisel WR. Influence of thorotrast blockade and acute renal artery ligation on disappearance of staphylococcal enterotoxin B from blood. J Bacteriol93 (3): 779-83, 1967.

15. Shupp JW, Jett M, Pontzer CH. Identification of a transcytosis epitope on staphylococcal enterotoxins. Infect Immun 70 (4): 2178-86, 2002.

16. Spero L, Morlock BA. Biological activities of the peptides of staphylococcal enterotoxin C formed by limited tryptic hydrolysis. J Bioi Chem 253 (24): 8787 -91, 1978. 17. van Gessel VA, Mani S, Bi S, Hammamieh R, Shupp JW, Das R, Coleman GD, Jett M. Functional piglet model for the clinical syndrome and postmortem findings induced by staphylococcal enterotoxin B. Exp Bioi Med (Maywood) 229 (10): 1061-71,2004

18. Chatterjee S, Sekerke CS, Kwiterovich PO Jr. Increased urinary excretion of glycosphingolipids in familial hypercholesterolemia. J Lipid Res. 23(4):513-522, 1982

19. Chatterjee S, Kwiterovich PO. Glycosphingolipids of human plasma lipoproteins. Lipids. 11 :462-6, 1976

20. Markwell MA, Fox CF Surface-specific iodination of membrane proteins of viruses and eucaryotic cells using l^Aό-tetrachloro-Salpha^alphadiphenylglycoluril. Biochem istry. 17(22):4807-17, 1978

21. Chatterjee S, Kwiterovich PO Jr, Gupta P, Erozan VS, Alving CR, RichardsRL. Localization of urinary lactosylceramide in cytoplasmic vesicles of renaltubular cells in homozygous familial hypercholesterolemia. Proc Natl Acad Sci USA. 80(5): 1313-7, 1983

22. Chatterjee S. Neutral sphingomyelinase: past, present and future. Chem Phys Lipids.1 02(1-2): 79-96, 1999.

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24. Luberto C, Yoo DS, Suidan HS, Bartoli GM, Hannun VA. Differential effects of sphingomyelin hydrolysis and resynthesis on the activation of NF-kappa B in normal and SV40-transformed human fibroblasts. J BioilChem.275(19):14760-6,2000. 25. Lycke N, Bromander AK, Ekman L, Karlsson U, Holmgren J. Cellular basis of immunomodulation by cholera toxin in vitro with possible association to the adjuvant function in vivo. J Immunol. 142(l):20-7, 1989.

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Claims

WHAT IS CLAIMED IS:

1. A method of increasing cell proliferation comprising administering a Staphylococcus enterotoxin B (SEB) peptide capable of increasing cell proliferation in a cell that expresses digalactosylceramide receptor.

2. A method of increasing cell proliferation in a subject in need thereof comprising administering to the subject a SEB peptide capable of increasing cell proliferation in a cell that expresses digalactosylceramide receptor.

3. The method of claim 1 or claim 2, wherein the cell that expresses digalactosylceramide receptor is selected from the group consisting of: brain cells, neuronal cells, lymphocytes, leukocytes, liver cells, inner ear cells, spleen cells, pancreatic cells, urinary tract cells, bone marrow spinal cord, spinal root cells, skin cells, and conjunctival vessels.

4. A method of increasing kidney cell proliferation in a subject comprising administering to the subject a SEB peptide capable of increasing cell proliferation, thereby stimulating kidney cell proliferation.

5. The method of claim 4, wherein the subject has undergone kidney transplantation or kidney resection.

6. The method of claim 4, wherein the method stabilizes, reduces the symptoms of, or ameliorates a disease or disorder characterized by kidney dysfunction.

7. A method of increasing immune cell proliferation comprising administering a SEB peptide capable of increasing cell proliferation, thereby stimulating immune cell proliferation.

8. A method of increasing immune cell proliferation in a subject comprising administering to the subject a SEB peptide capable of increasing cell proliferation, thereby stimulating immune cell proliferation.

9. The method of claim 7 or claim 8, wherein the immune cells are T-lymphocytes.

10. The method of claim 7 or claim 8, wherein the method stabilizes, reduces the symptoms of, or ameliorates a disease or disorder characterized by abnormal immune cell proliferation.

11. The method of claim 10, wherein the disease or disorder is anaplastic anemia or Fabry's Disease.

12. The method of any one of claims 1 - 11, wherein the SEB peptide comprises the amino acid sequence CVFSKKTNDINSHQTDKRKT (SEQ ID NO: 3) or fragments thereof.

13. The method of any one of claims 1 - 11, wherein the SEB peptide consists of the amino acid set forth as SEQ ID NO: 3.

14. The method of claim 12, wherein the SEB peptide further comprises one or more therapeutic agents.

15. The method of claim 14, wherein the one or more therapeutic agents is selected from the group consisting of: protein synthesis modulators, apoptosis modulators, nitric oxide modulators, bactericides, a fungicides, anti neoplastic drugs, anti thrombotic drugs, hypochoesterolemic drugs, hypotriglyceridemic drugs, hypoglycemic drugs, anti-allergic drugs, antibiotics, tissue, anti-inflammatory drugs, nerve regenerating drugs, and anti- arthritis drugs.

16. A method of decreasing cell proliferation comprising administering a SEB peptide capable of decreasing cell proliferation in a cell that expresses digalactosylceramide receptor.

17. A method of decreasing cell proliferation in a subject in need thereof comprising administering to the subject a SEB peptide capable of decreasing cell proliferation in a cell that expresses digalactosylceramide receptor.

18. The method of claim 16 or claim 17, wherein the method further comprises increasing apoptosis.

19. A method of promoting apoptosis comprising administering a SEB peptide capable of promoting apoptosis in a cell that expresses digalactosylceramide receptor.

20. A method of promoting apoptosis in a subject in need thereof comprising administering to the subject a SEB peptide capable of promoting apoptosis in a cell that expresses digalactosylceramide receptor.

21. The method of claim 19 or claim 20, wherein the cell expresses elevated levels of digalactosylceramide compared to a control cell.

22. The method of any one of claims 16 - 21, wherein the cell that expresses digalactosylceramide receptor is a mammalian cell.

23. The method of claim 22, wherein the mammalian cell is selected from the group consisting of: a selected from the group consisting of: tumor cells, kidney cells, neuronal cells, lymphocytes, inner ear cells, spleen cells and pancreatic cells.

24. A method of treating a tumor in a subject comprising administering to the subject a SEB peptide capable of promoting apoptosis or decreasing cell proliferation in a tumor that expresses digalactosylceramide receptor, thereby treating the tumor in a subject.

25. The method of claim 24, wherein the tumor expresses elevated levels of digalactosylceramide compared to a normal tissue.

26. A method of treating a lipid metabolic disorder in a subject comprising administering to the subject a SEB peptide capable of promoting apoptosis or decreasing cell proliferation in cells that express high levels of digalactosylceramide, thereby treating the lipid metabolic disorder the subject.

27. The method of claim 26, wherein the lipid metabolic disorder is selected from the group consisting of: Fabry's disease, Metachromatic Leukodystrophy, GM2 Gangliosidosis, Tay-Sachs disease and chronic myelogenous leukemia.

28. A method of treating a neural disease or disorder or a kidney disease or disorder in a subject comprising administering to the subject a SEB peptide capable of promoting apoptosis or decreasing cell proliferation in neural tissue or kidney tissue that expresses digalactosylceramide receptor, thereby treating a neural or kidney disease in the subject.

29. The method of claim 28, wherein the neural tissue or kidney tissue expresses high levels of digalactosylceramide compared to a control cell.

30. The method of claim 28, wherein the neural or kidney disease or disorder is selected from autism or glomerular nephritis.

31. The method of any one of claims 16 - 30, wherein the SEB peptide comprises the amino acid sequence RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID NO: 5) or fragments thereof, or KKKVTAQEL (SEQ ID NO: 7) or fragments thereof.

32. The method of any one of claims 16 - 30, wherein the SEB peptide consists of the amino acid set forth as SEQ ID NO: 5 or SEQ ID NO: 7.

33. The method of claim 28, wherein the SEB peptide further comprises one or more therapeutic agents.

34. The method of claim 33, wherein the one or more therapeutic agents is selected from the group consisting of: protein synthesis modulators, apoptosis modulators, nitric oxide modulators, bactericides, a fungicides, anti neoplastic drugs, anti thrombotic drugs,hypochoesterolemic drugs, hypotriglyceridemic drugs, hypoglycemic drugs, antiallergic drugs, antibiotics,tissue, anti-inflammatory drug, nerve regenerating drugs, and anti-arthritis drugs.

35. The method of claim 34, wherein the apoptosis modulator is clodronate.

36. The method of claim 33, wherein the one or more therapeutic agents is an anticancer agent.

37. The method of claim 36, wherein the anticancer agent is selected from the group consisting of: a chemotherapeutic agent, a peptide toxin, and a protein toxin.

38. The method of claim 37, wherein the anticancer agent is rapamycin.

39. The method of any one of claims 16 - 38, wherein the peptide further comprises an imaging agent.

40. The method of claim 39, wherein the imaging agent is selected from a radiolabel or a fluorescent label.

41. A method of targeting an agent to a cell that expresses digalactosylceramide receptor comprising administering an SEB peptide comprising one or more agents to a cell that expresses digalactosylceramide receptor.

42. A method of targeting an agent to a tumor or an atherosclerotic plaque that expresses digalactosylceramide receptor in a subject comprising administering an SEB peptide comprising one or more agents to a tumor or an atherosclerotic plaque that expresses digalactosylceramide receptor.

43. The method of claim 41 or 42, wherein the SEB peptide comprises the amino acid sequence ENENSFWYAMMPAPGDKFDQSKYLMMYNDN (SEQ ID NO: 6) or fragments thereof.

44. The method of claim 43, wherein the SEB peptide consists of the amino acid set forth as SEQ ID NO: 6.

45. The method of claim 41 or 42, wherein the one or more agents is a therapeutic agent selected from the group consisting of: protein synthesis modulators, apoptosis modulators, nitric oxide modulators, bactericides, a fungicides, anti neoplastic drugs, anti thrombotic drugs,hypochoesterolemic drugs, hypotriglyceridemic drugs, hypoglycemic drugs, anti-allergic drugs, antibiotics,tissue, anti-inflammatory drugs, nerve regenerating drugs, and anti-arthritis drugs.

46. The method of claim 45, wherein the agent is an anticancer agent.

47. The method of claim 46, wherein the anticancer agent is selected from the group consisting of: a chemotherapeutic agent, a peptide toxin, and a protein toxin.

48. The method of claim 46, wherein the anticancer agent is rapamycin.

49. The method of any one of claims 40 - 42, wherein the agent is an imaging agent.

50. The method of claim 49, wherein the imaging agent is selected from a radiolabel or a fluorescent label.

51. A method of detecting a tumor or an atherosclerotic plaque that expresses digalactosylceramide receptor in a subject comprising administering an SEB peptide comprising an imaging agent to a tumor or an atherosclerotic plaque that expresses digalactosylceramide receptor in a subject.

52. The method of claim 51, wherein the method is used to determine course of treatment.

53. The method of claim 51, wherein the subject is treated with an anticancer or antiproliferative agent.

54. The method of claim 51, wherein the method is used to determine prognosis.

55. The method of claim 51, wherein the method is used to determine regression in tumor size or atherosclerotic plaque size after treatment with an anticancer or antiproliferative agent.

56. A method of treating infection caused by a bacterial toxin in a subject, the method comprising administering to the subject a SEB peptide, wherein the peptide is capable of binding to a digalactosylceramide (DAG) receptor, thereby treating infection caused by a bacterial toxin in a subject.

57. The method of claim 51 or 56, wherein the SEB peptide comprises the amino acid sequence ENENSFWYAMMPAPGDKFDQSKYLMMYNDN (SEQ ID NO: 6).

58. The method of claim 57, wherein the SEB peptide consists of the amino acid set forth as SEQ ID NO: 6.

59. A method of inhibiting activity of a digalactosylceramide (DAG) receptor in a subject comprising administering a composition selected from: an antibody, a nucleic acid or an oligomer that prevents activation of the DAG receptor.

60. The method of claim 59, wherein the antibody has specificity for the amino acid sequence selected from the group comprising:

RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID NO: 5) and KKKVTAQEL (SEQ ID NO: 7)

61. The method of claim 60, wherein the nucleic acid is an inhibitory nucleic acid selected from: antisense, siRNA, shRNA, aptamers, PNA oligomers, and ribozymes.

62. The method of claim 59, wherein the oligomer is at least a 5-mer of the amino acid sequence comprising RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID NO: 5).

63. The method of claim 59, wherein the subject is suffering from an infection caused by a bacterial toxin.

64. A method of treating infection caused by a bacterial toxin in a subject, the method comprising administering a composition selected from: an antibody, a nucleic acid or an oligomer that prevents activation of the DAG receptor, wherein the peptide is capable of binding to a digalactosylceramide (DAG) receptor, thereby treating infection caused by a bacterial toxin in a subject.

65. The method of claim 56 or 64, wherein the bacterial toxin is from a Gram- negative bacteria.

66. The method of claim 65, wherein the bacterial toxin is selected from Escherichia coli or Psuedomonas aeruginosa.

67. The method of claim 66, wherein the E. coli bacterial toxin is verotoxin or Shiga toxin.

68. The method of claim 66, wherein the Escherichia coli or Pseudomonas aeruginosa binds a glycolipid receptor selected from a digalactosylceramide receptor or a Globotriosylceramide Receptor (GbOse3Cer).

69. The method of claim 56 or 64, wherein the infection caused by a bacterial toxin is localized to the lung, kidney, spleen, pancreas or gastroinstestinal tract.

70. The method of any one of the above claims, wherein the subject is a human.

71. A peptide comprising the amino acid sequence CVFSKKTNDINSHQTDKRKT (SEQ ID NO: 3) or fragments thereof, wherein the peptide is capable of increasing cell proliferation in a cell that expresses digalactosylceramide receptor.

72. The peptide of claim 71, wherein the peptide comprises the amino acid set forth as SEQ ID NO: 3.

73. A peptide comprising the amino acid sequence

RSITVRVFEDGKNLLSFDVQTNKKKVTAQEL (SEQ ID NO: 5) or fragments thereof, or KKKVTAQEL (SEQ ID NO: 7) or fragments thereof, wherein the peptide is capable of decreasing cell proliferation in a cell that expresses digalactosylceramide receptor.

74. The peptide of claim 73, wherein the peptide promotes apoptosis.

75. The peptide of claim 73 or 74, wherein the peptide comprises the amino acid set forth as SEQ ID NO: 5 or 7.

76. A peptide comprising the amino acid sequence

ENENSFWYAMMPAPGDKFDQSKYLMMYNDN (SEQ ID NO: 6) or fragments thereof, wherein the peptide is capable of binding to a digalactosylceramide (DAG) receptor.

77. The peptide of claim 76, wherein the peptide comprises the amino acid set forth as SEQ ID NO: 6.

78. The peptide of any one of claims 71 - 77, wherein the peptide is derived from Staphylococcus enterotoxin B (SEB).

79. The peptide of any one of claims 71, 73 or 76, wherein the peptide is at least a 5- mer oligomeric fragment.

80. A method of making one or more antibodies that bind at least one staphylococcal enterotoxin receptor, said method comprising administering to a mammal an amount of a peptide of any one of claims 71 - 77 sufficient to elicit production of one or more antibodies.

81. The peptide of any one of claims 71 - 79, wherein the peptide further comprises one or more therapeutic agents.

82. The peptide of claim 81 , wherein the agent is covalently linked the peptide.

83. The peptide of claim 81 , wherein the one or more therapeutic agents is selected from the group consisting of: protein synthesis modulators, apoptosis modulators, nitric oxide modulators, bactericides, a fungicides, anti neoplastic drugs, anti thrombotic drugs,hypochoesterolemic drugs, hypotriglyceridemic drugs, hypoglycemic drugs, antiallergic drugs, antibiotics,tissue, anti-inflammatory drug, nerve regenerating drugs, and anti -arthritis drugs.

84. The peptide of claim 83, wherein the one or more therapeutic agents is an anticancer agent.

85. The peptide of claim 84, wherein the anticancer agent is selected from the group consisting of: a chemotherapeutic agent, a peptide toxin, and a protein toxin.

86. The peptide of claim 85, wherein the chemotherapeutic agent is rapamycin.

87. The peptide of any one of claims 71 - 86, wherein the peptide further comprises an imaging agent.

88. The peptide of claim 87, wherein the imaging agent is selected from a radiolabel or a fluorescent label.

89. A pharmaceutical composition comprising the peptide of any one of claims 71 - 88 and a pharmaceutically acceptable carrier.

90. A kit for use according to any one of the methods of claims 1 - 70 and instructions for use.