HK1109574B - Mucinous glycoprotein (muc-1) vaccine - Google Patents

Description

Mucinous glycoprotein (MUC-1) vaccine

Priority declaration

This application claims priority from U.S. provisional patent application No. 60/558,139 filed on day 4 of 2004 and U.S. provisional patent application No. 60/576,804 filed on day 4 of 2004, the entire teachings of both applications being expressly incorporated herein by reference.

Technical Field

The present invention relates to the treatment of individuals suffering from cancer, particularly non-small cell lung cancer and prostate cancer, using mucoprotein-based (MUC-1) preparations. In some cases, the MUC-1 based formulation is a BLP25 liposome vaccine.

Background

Lung cancer is the first cause of cancer-associated mortality in both genders of north america. In 2004, approximately 174,000 new lung cancer cases (54% in men, 46% in women) were diagnosed in the united states. In 2004, approximately 160,000 people died of the disease in the united states alone.

Unfortunately, at the time of diagnosis, only 25% of lung cancer patients are potentially curable by surgery. Furthermore, chemotherapy is only appropriate to improve the chances of survival in individuals with this cancer.

Non-small cell lung cancer (NSCLC) is the most prevalent lung cancer, and is represented by squamous cell carcinoma, adenocarcinoma, and large cell carcinoma. NSCLC accounts for approximately 75 to 80% of all primary lung cancers. It has been observed that the mucinous glycoprotein, MUC-1, is highly expressed in these cancers, beyond its normal expression level in epithelial cells of healthy individuals. It has also been observed that many of the sugar moieties that modify the MUC-1 protein by attachment to the MUC-1 polypeptide backbone are shorter than the sugar moieties of the MUC-1 protein that attach to normal cells. Thus, the MUC-1 polypeptide backbone in cancer cells is more exposed than the polypeptide backbone in normal cells.

Prostate cancer is the second most commonly diagnosed cancer that occurs in american men after lung cancer. Approximately 190,000 men are diagnosed with prostate cancer and nearly 30,000 men die from the disease each year in the united states.

Biochemical failure following Prostatectomy (PR) for the treatment of prostate cancer is often a predictor of clinical failure, which may shorten a patient's life expectancy. While there is a need for additional non-invasive methods of treating prostate cancer, there is a particular need for treatment for men with post-prostatectomy biochemical failure.

The present invention provides methods for treating a population of individuals with non-small cell lung cancer and prostate cancer by using MUC-1 based formulations. In many embodiments, the formulation is a lipidated (lipidated) MUC-1 liposome vaccine.

Summary of The Invention

The present invention relates to the treatment of individuals suffering from cancer, such as NSCLC or prostate cancer, using MUC-1 based formulations. The invention also encompasses the use of MUC-1 based formulations described herein to treat cancers other than NSCLC and prostate cancer.

In one embodiment of the invention, the MUC-1 based formulation may be a MUC-1 based liposome vaccine. For example, the liposomal vaccine may comprise the MUC-1 peptide in its lipid bilayer or encapsulated in its vesicle structure. The MUC-1 peptide may also be lipidated to facilitate its association with the liposomal lipid bilayer or membrane. The MUC-1 peptide may comprise the amino acid sequence depicted in SEQ ID No.1 or a variant thereof, or SEQ ID No.2 or a variant thereof. Specific properties of MUC-1 core repeat variants are described below.

In another aspect of the invention, a method ("method 1") for treating an individual having NSCLC or prostate cancer is provided. The method comprises the following steps: (A) selecting an individual having NSCLC or prostate cancer for treatment, and (B) administering to the individual a MUC-1-based formulation for a time period. In one embodiment of method 1, the MUC-1 based formulation comprises a liposome comprising at least one polypeptide having the amino acid sequence set forth in SEQ ID No.1 or a variant thereof, or SEQ ID No.2 or a variant thereof.

In particular embodiments, method 1 may further comprise step (C) comprising evaluating the treated individual. In a particular embodiment, the treated individual can be evaluated by measuring an immune response in the treated individual. In certain embodiments, measuring the immune response in the treated individual may comprise measuring T cell proliferation. In another embodiment, evaluating the treated individual may comprise determining at least one or more of: (a) tumor size, (b) tumor location, (c) nodal stage (nodal stage), (d) growth rate of NSCLC or prostate cancer, (e) survival of the individual, (f) a change in lung cancer or prostate cancer symptoms in the individual, (g) a change in PSA concentration in the individual, (h) a change in PSA concentration doubling rate (doubling rate) in the individual, (i) a change in quality of life (life) in the individual, or (j) a combination thereof.

In these embodiments, the individual may be evaluated before, during, or after a period of time. Individuals may also be evaluated before and after a period of time.

In other embodiments, the formulation is a BLP25 liposome vaccine. "BLP 25" is a specific lipidated MUC-1 core repeat identified below. The BLP25 vaccine may comprise preformed liposomes comprising MUC-1 core repeats, such as those of SEQ id nos: 1 and 2. Preformed liposomes containing MUC-1 core repeats can be freeze dried.

In one embodiment of the method, the BLP25 liposome vaccine is present in a kit and instructions for making and using the vaccine are included in the kit. Thus, the kit may comprise another liquid, such as a sodium chloride solution (0.9%, USP) that may be used to reconstitute the lyophilized material. Alternatively, the BLP25 liposome vaccine may be provided as a liquid. The kit may also contain additional adjuvants that may be added to the vaccine formulation, such as lipid a, muramyl dipeptide, alum or cytokines. Examples of these components and other adjuvants are disclosed in detail below. Thus, the kit may comprise a number of vials or containers that enable one to prepare the BLP25 vaccine for administration.

The step of administering the formulation to the individual may be performed by any suitable method, for example by injection, wherein the injection is intramuscular, subcutaneous, intranodal (intradal), intratumoral, intraperitoneal or intradermal. Alternatively, vaccine or liposome-bound MUC-1 core repeat peptides may be administered by aerosol, nasal delivery or oral delivery. Vaccine or liposome-bound MUC-1 core repeat peptides can also be administered by a formulation suitable for transdermal delivery, for example, by transdermal patch (patch).

In other embodiments, the period of administration of any of the methods described herein is at least about 2 weeks, at least about 4 weeks, at least about 8 weeks, at least about 16 weeks, at least about 17 weeks, at least about 18 weeks, at least about 19 weeks, at least about 20 weeks, at least about 24 weeks, at least about 28 weeks, at least about 32 weeks, at least about 36 weeks, at least about 40 weeks, at least about 44 weeks, at least about 48 weeks, at least about 52 weeks, at least about 60 weeks, at least about 68 weeks, at least about 72 weeks, at least about 80 weeks, at least about 88 weeks, at least about 96 weeks, or at least about 104 weeks.

In another aspect of the invention, a method for improving or maintaining the quality of life of an individual having NSCLC or prostate cancer is described ("method 2"). The method may comprise routinely administering one dose of a liposome vaccine of BLP25 to an individual diagnosed with NSCLC or prostate cancer over a period of time. In other aspects of method 2, a combined score of the individual's physical well-being, functional well-being (functional well-being) and lung cancer symptoms or prostate cancer symptoms before, during, and after a period of time can be calculated.

In one embodiment, conventional administration includes administering one dose of the BLP25 liposome vaccine once a week over a period of time. Of course, the dosing regimen may include other permutations of MUC-1 peptide delivery. That is, the vaccine may be administered 1,2, 3, 4, 5, 6 or more times per week. In another embodiment, the period of time for administering the vaccine under this regimen is at least about 6 months, at least about 12 months, at least about 18 months, or at least about 24 months.

In one embodiment, the dose of the BLP25 liposome vaccine provides approximately 1,000 μ g of BLP25MUC-1 lipopeptide, although other doses described below may be administered. See, for example, BLP25 dose below to segment the predicted dose.

The foregoing general description and the following brief description of the drawings are exemplary and explanatory and are intended to provide further explanation of the invention as claimed. Other objects, advantages and novel features will become apparent to those skilled in the art from the following detailed description of the invention.

Brief Description of Drawings

Fig. 1 is a graph depicting results from the study detailed herein, showing overall survival obtained by means between studies of patients receiving treatment with the BLP25 liposome vaccine or patients receiving only the Best Supportive Care (BSC). See example 1 below.

FIG. 2 is a graph demonstrating survival analysis of patients with stage IIIB regional (logeginal) NSCLC. Survival analysis of both groups of patients (treatment and BSC groups) included the survival distribution function of patients treated with the BLP25 liposome vaccine versus patients treated with the best supportive therapy alone. See example 1 below.

Figure 3 is a graph depicting the percent change in doubling time of prostate specific antigen ("PSA") from patient to patient who had received one dose of BLP25 liposome vaccine. See example 3 below.

Detailed Description

The present invention provides MUC-1 based formulations and methods of using MUC-1 based formulations to treat individuals having cancer, such as NSCLC or prostate cancer. The invention also encompasses the treatment of cancers other than NSCLC and prostate cancer with MUC-1-based formulations described herein.

According to the invention, the formulation may comprise MUC-1 core repeats. The MUC-1 core repeat may be an amino acid sequence that occurs any number of times in the MUC-1 protein. Preferably, the MUC-1 core repeat peptides of the present invention mimic the exposure properties of MUC-1 proteins expressed in cancer cells, which MUC-1 proteins have shorter sugar moieties attached to the backbone of the MUC-1 protein.

In one embodiment, the MUC-1 core repeat of the invention has the amino acid sequence STAPPAHGVTSAPDTRPAPGSTAPP (SEQ ID NO. 1).

The MUC-1 core repeat may also have an amino acid sequence represented by any one of the following sequences:

STAPPAHGVTSAPDTRPAPGSTAPPK (palmitoyl) G (SEQ ID NO: 2)

STAPPAHGVTSAPDTRPAPG(SEQ ID NO：3)

GSTAPPAHGVTSAPDTRPAP(SEQ ID NO：4)

GVTSAPDTRPAPGSTAPPAH(SEQ ID NO：5)

PDTRPAPGSTAPPAHGVTSA(SEQ ID NO：6)

HGVTSAPDTRPAPGSTAPPA(SEQ ID NO：7)

VTSAPDTRPAPGSTAPPAHG(SEQ ID NO：8)

The MUC-1 peptide of the present invention may also be an amino acid of any length. For example, the peptide may comprise 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100 or more amino acids in length.

In certain embodiments, the core repeat may be lipidated. One such MUC-1 core repeat lipopeptide is referred to herein as BLP 25. The preparation can also be combined with liposome. The conjugation may include, but is not limited to, incorporating the peptide into a liposome or the liposome encapsulating the peptide.

A liposomal vaccine comprising BLP25 lipopeptides is referred to herein as "L-BLP 25".

The formulations of the invention may also comprise an adjuvant, such as lipid A or interleukin-2 (IL-2). Other exemplary adjuvants for use in the present invention are described below. In many cases, MUC-1 based formulations can be formulated as vaccines. In certain embodiments, the vaccine is a liposome-bound MUC-1 core repeat vaccine. In several embodiments, the vaccine formulation comprises a liposome-bound MUC-1 core repeat and an adjuvant. In many embodiments, the MUC-1 core repeat is lipidated.

In certain embodiments, the vaccine of the invention may comprise (a) a polypeptide comprising SEQ ID No.: 1 and an exogenous lipid; or (b) comprises SEQ ID No.: 1, and liposomes; or (c) comprises SEQ ID No.: 1 and liposomes and an adjuvant, or (d) a MUC-1 core repeat comprising the sequence of SEQ id No.: 1 and a liposome and an adjuvant, wherein the adjuvant is lipid a.

In certain other embodiments, the vaccine of the invention may comprise (a) a polypeptide comprising SEQ id No.: 2 and an exogenous lipid; or (b) comprises SEQ id No.: 2 and liposomes; or (c) comprises SEQ ID No.: 2 and a liposome and an adjuvant, or (d) a MUC-1 core repeat comprising the sequence of SEQ id No.: 2 and a liposome and an adjuvant, wherein the adjuvant is lipid a.

The concept of treating an individual suffering from NSCLC or prostate cancer with components of a MUC-1 based formulation and a MUC-1 based vaccine formulation (formula) is described in more detail below.

BLP25 liposome vaccine

In one embodiment, the MUC-1 based formulation comprises an amount of MUC-1 lipopeptide BLP25 and an amount of an adjuvant. Such formulations are referred to herein as BLP25 liposome vaccines ("L-BLP 25") and may be presented as liquid or lyophilized formulations. For example, the formulation, or vaccine, may comprise about 1000 μ g of the MUC-1 lipopeptide BLP25 and about 500 μ g of lipid a in a single dose.

However, other microgram amounts of MUC-1 lipopeptide and lipid A are also envisioned by the present invention. For example, the amount of BLP25 lipopeptide may be sufficient to supply multiple doses of vaccine. Thus, a MUC-1 core repeat formulation may comprise 50 μ g, about 100 μ g, about 200 μ g, about 300 μ g, about 400 μ g, about 500 μ g, about 600 μ g, about 700 μ g, about 800 μ g, about 900 μ g, about 1,000 μ g, about 1,010 μ g, about 1,020 μ g, about 1,030 μ g, about 1,040 μ g, about 1,050 μ g, about 1,060 μ g, about 1,070 μ g, about 1,080 μ g, about 1,090 μ g, about 1,100. mu.g, about 1,200. mu.g, about 1,300. mu.g, about 1,400. mu.g, about 1,500. mu.g, about 1,600. mu.g, about 1,700. mu.g, about 1,800. mu.g, about 1,900. mu.g, about 2,000. mu.g, about 3000. mu.g, about 4000. mu.g, about 5000. mu.g, about 6000. mu.g, about 7000. mu.g, about 8000. mu.g, about 9000. mu.g, about 10000. mu.g, about 15000. mu.g, about 25000. mu.g or more of MUC-1 core repeats. One particular dose of MUC-1 core repeats is in the range of about 500 μ g to about 1500 μ g, more preferably between about 500 μ g and about 1500 μ g, and more preferably about 1000 μ g.

Similarly, the amount of lipid a may be varied to match the amount of MUC-1 peptide formulated into the vaccine. Thus, the amount of lipid A can be 50 μ g, about 100 μ g, about 200 μ g, about 300 μ g, about 400 μ g, about 500 μ g, about 600 μ g, about 700 μ g, about 800 μ g, about 900 μ g, about 1,000 μ g, about 1,010 μ g, about 1,020 μ g, about 1,030 μ g, about 1,040 μ g, about 1,050 μ g, about 1,060 μ g, about 1,070 μ g, about 1,080 μ g, about 1,090 μ g, about 1,100 μ g, 1,200 μ g, 1,300 μ g, 1,400 μ g, 1,500 μ g, 1,600 μ g, 1,700 μ g, 1,800 μ g, 1,900 μ g, or about 2,000 μ g or more. In particular about 500. mu.g lipid A.

The BLP25 lipopeptide and lipid a may be associated with a lipid bilayer of liposomes that may be formed after rehydration of their dry powder.

The liposome formulation or L-BLP25 vaccine may be maintained in one or more vials, for example in a 5ml type I borosilicate glass vial. The vial containing the MUC-1 formulation may also contain other vaccine components. For example, the vial may contain additional liposomal lipids, such as dipalmitoylphosphatidylcholine, cholesterol, and dimyristoylphosphatidylglycerol. The respective amounts of these specific lipids may vary. Thus, the amount of any one of dipalmitoyl phosphatidylcholine, cholesterol and dimyristoyl phosphatidylglycerol in a vial can be about 1mg, about 2mg, about 3mg, about 4mg, about 5mg, about 6mg, about 7mg, about 8mg, about 9mg, about 10mg, about 11mg, about 12mg, about 13mg, about 14mg, about 15mg, about 16mg, about 17mg, about 18mg, about 19mg, about 20mg, about 25mg, about 30mg, about 35mg, about 40mg, about 45mg, about 50mg, about 55mg, about 60mg, about 65mg, about 70mg, about 75mg, about 80mg, about 85mg, about 90 mg, about 95mg or about 100mg or more than about 100 mg. The liposomal lipids may be contained in a vial distinct from the vial containing the MUC-1 formulation.

Of course, other embodiments exist for the purposes of the present invention. Thus, the above amounts of MUC-1 lipopeptide BLP25, adjuvant and liposomal lipid in L-BLP25 are provided by way of example only. Determining the appropriate amount of each component, including the amount of MUC-1 lipopeptide, can be readily accomplished and is routine. In some embodiments, the dosage of MUC-1 lipopeptide is greater than or less than about 1000 μ g. The vaccine need not be provided in the form present in a 5ml type I borosilicate glass vial, but may be provided in any form known in the art.

In one embodiment, the BLP25 lipopeptide is a linear 27 residue peptide comprising a lipidated amino acid derivative near its C-terminus. Specifically, BLP25 comprises palmitoyl lipid at the lysine residue at position 26 of the polypeptide. The sequence of BLP25 lipopeptide is described in SEQ ID No.: 2:

SEQ ID NO. 2: STAPPAHGVTSAPDTRPAPGSTAPPK (palmitoyl) G

In other embodiments having a MUC-1 core sequence, amino acids that may be present in the native sequence of the peptide, such as threonine, serine, lysine, arginine, or cysteine, may be convenient sites to which lipids may be attached. In some embodiments, the lipid may be linked to synthetic amino acids or amino acids not naturally found in the MUC-1 core sequence. In certain embodiments, one or more natural or synthetic amino acids may be added at either end of or within the MUC-1 core sequence to facilitate lipid attachment.

The number of amino acids that can be added to the core sequence of MUC-1 is not limited, and any number of amino acids can be added, so long as the peptide functions in the methods of the invention. As demonstrated above, two additional amino acids have been added to the BLP25 polypeptide. That is, the C-terminus of the MUC-1 core sequence ends with proline and thus it is 25 residues in length. However, in the case of the BLP25 polypeptide, lysine and glycine have been added to the C-terminal proline to facilitate palmitoyl ligation. Thus, the BLP25 polypeptide is 27 amino acids in length. One or more of these additional amino acids can be added to the peptide sequence using conventional peptide synthesis methods. Alternatively, the MUC-1 core sequence peptide or BLP25 may be recombinantly produced.

In a particular embodiment, the BLP25 liposome vaccine ("L-BLP 25") may comprise BLP25 lipopeptide, lipid a, cholesterol, DMPG, and DPPC. The BLP25 lipopeptide may comprise SEQ ID NO: 2 or a variant thereof. One dose of the BLP25 liposome vaccine may comprise about 1000. mu.g BLP25 lipopeptide, about 500. mu.g lipid A, about 17.3mg cholesterol, about 3.6mg DMPG, and about 29.1mg DPPC.

The particular vaccine composition and dosage may also be described in "per vial" amounts. Thus, a vial may contain about 300 μ g BLP25 lipopeptide, about 150 μ g lipid A, about 5.2mg cholesterol, about 1.1mg DMPG, and about 8.7mg DPPC.

The vaccine may be freeze-dried and then reconstituted, for example in a sodium chloride solution prior to administration. The above-described amount of BLP25 liposome vaccine can be reconstituted in, for example, about 0.6ml of liquid, although any volume of liquid may be used, depending on the desired dose, e.g., about 0.1ml, 0.2ml, 0.3ml, 0.4ml, 0.5ml, 0.6ml, 0.7ml, 0.8ml, 0.9ml, 10ml, 11ml, 12ml, 13ml, 14ml, 15ml, 16ml, 17ml, 18ml, 19ml, or 20ml or more than 20 ml.

The volume of liquid in which the MUC-1 vaccine for freeze-drying is reconstituted need not be the volume administered to an individual. Certain desired doses may be administered in one or more volumes of reconstituted vaccine.

A. MUC-1 core repeat variants

As SEQ ID NOs: 1-8, the formulations of the invention may incorporate homologues or variants of these MUC-1 core repeats. Accordingly, the invention includes the use of a MUC-1 core repeat peptide having the amino acid sequence shown in SEQ ID NOs: 1-8, wherein the amino acid sequence is similar but not identical. Thus, the present invention contemplates the use of a MUC-1 core repeat that is identical to the core repeat of SEQ ID NOs: 1-8, has 99%, 98%, 97%, 96%, 95%, 94%, 93%, 92%, 91%, 90%, 89%, 88%, 87%, 86%, 85%, 84%, 83%, 82%, 81%, or 80% sequence identity compared to the sequence in any of claims 1-8.

The MUC-1 core repeat proteins of the invention may also be modified to include conservative variations, or may be modified to alter non-critical residues or residues in non-critical regions. Non-critical amino acids are identified by methods known in the art, such as site-directed mutagenesis, crystallization, nuclear magnetic resonance, photoaffinity labeling or alanine scanning mutagenesis (Cunningham et al, Science, 244: 1081-. Modified proteins can be readily tested for their ability to activate or induce an immune response by methods such as protease binding to a substrate, cleavage, in vitro activity or in vivo activity.

Specifically, the MUC-1 core repeat variant may incorporate 1 to 5 amino acid substitutions that increase the stability of the MUC-1 core repeat, or may have different hydrophobic amino acids that increase the stability of the MUC-1 core repeat against oxidation, or may have different amino acids that increase the stability of the MUC-1 core repeat against proteases.

Thus, a "variant" MUC-1 core repeat polypeptide of the invention may have an amino acid sequence that is identical to that of SEQ id nos: 1 or 2 are different. Any one of the variants may be prepared so as to contain an amino acid substitution that replaces a given amino acid with another amino acid having the same characteristics. Conservative substitutions include exchanges between aliphatic amino acids of alanine, valine, leucine, and isoleucine; the exchange of the hydroxyl residues serine and threonine, the exchange of the acidic residues aspartic acid and glutamic acid, the substitution between the amide residues asparagine and glutamine, the exchange of the basic residues lysine and arginine and the substitution between the aromatic residues phenylalanine and tyrosine. See, Bowie et al, Science, 247: 1306-1310(1990).

B. MUC-1 core repeat fusion proteins

Also, a polypeptide having the sequence of SEQ ID NOs: 1 or 2 or a variant thereof to another polypeptide which is not normally linked. Thus, the MUC-1 core repeat peptide may be operably linked at its N-terminus or C-terminus to a heterologous polypeptide having an amino acid sequence substantially heterologous to the MUC-1 core repeat. By "operably linked" is meant that both the MUC-1 core repeat peptide and the heterologous polypeptide are in frame.

The fusion protein may or may not affect the ability of the MUC-1 core repeat or functional variant thereof to induce an immunological response from the host system. For example, the fusion protein may be a glutathione S-transferase (GST) -fusion protein in which the MUC-1 core repeat is fused to a GST sequence or C-terminus of an influenza HA tag. Other types of fusion proteins include, but are not limited to, enzymatic fusion proteins, such as beta galactosidase fusion proteins, yeast two-hybrid GAL fusion proteins, polyhistidine fusions, and Ig fusions. These fusion proteins, particularly polyhistidine fusion proteins, facilitate the purification of recombinantly produced MUC-1 core repeats useful in the present invention. In certain host cells, expression and/or secretion of the protein may be increased by using a heterologous signal sequence fused to a protease, which signal peptide transports the MUC-1 core repeat peptide to the extracellular matrix or localizes the MUC-1 core repeat protein to the cell membrane.

Other fusion proteins may affect the ability of the MUC-1 core repeat to induce an immunological response. For example, a subregion of the MUC-1 core repeat may be replaced with a corresponding domain or subregion, e.g., from another region of the MUC-1 protein. Thus, chimeric MUC-1 core repeats can be generated. Also, the affinity for the substrate may be altered or even proteolysis of the substrate may be prevented. Thus, one can use a polypeptide having, for example, SEQ ID NO: 1 or 2 or a variant thereof as a competitive inhibitor of another MUC-1 core repeat peptide.

C. Modification of MUC-1 core repeats

MUC-1 core repeat variants also include derivatives or analogs in which (i) an amino acid is replaced with an amino acid residue not encoded by the genetic codon, (ii) a mature polypeptide is fused to another compound such as polyethylene glycol, or (iii) additional amino acids are fused to the MUC-1 polypeptide, such as a leader or secretory sequence or a sequence used for purification of the polypeptide.

Typical modifications include, but are not limited to, acetylation, acylation, ADP-ribosylation, amidation, covalent attachment of flavin, covalent attachment of a heme moiety, covalent attachment of a nucleotide or nucleotide derivative, covalent attachment of a lipid or lipid derivative, covalent attachment of phosphatidylinositol, cross-linking, cyclization, disulfide bond formation, demethylation, formation of covalent cross-links, formation of cystine, formation of pyroglutamate, formylation, gamma carboxylation, glycosylation, GPI anchor formation, hydroxylation, iodination, methylation, myristoylation, oxidation, proteolytic processing, phosphorylation, prenylation, racemization, selenoylation, sulfation, transport RNA mediated addition of amino acids to proteins such as arginylation and ubiquitination.

Particularly common peptide modifications that may be used for the MUC-1 core repeat include glycosylation, lipid attachment, sulfation, gamma carboxylation of glutamic acid residues, hydroxylation and ADP-ribosylation. See, t.e. creatton, Proteins-structures and Molecular Properties, 2 nd edition (w.h. freeman and Company, New York (1993)); world, F., Posttranslation volatile Modification of Proteins, B.C. Johnson, Ed. (Academic Press, New York 1-12 (1983)); seifter et al, meth. enzymol, 182: 626 + 646 (1990); and Rattan et al, ann.n.y.acad.sci., 663: 48-62(1992).

Modifications may be made at any position of the MUC-1 core repeat polypeptide, including the peptide backbone, the amino acid side chains and the amino or carboxyl termini. Blocking of amino groups or carboxyl groups or both in polypeptides produced by covalent modification is common in naturally occurring and synthetic polypeptides.

II. Dose of BLP25

When MUC-1 based formulations, including MUC-1 core peptides, BLP25 polypeptides, or BLP25 liposomal vaccines are administered to an individual, one skilled in the art will appreciate that the dosage may depend on several factors, including, but not limited to, the weight of the individual, the size of the tumor, or the progression of the tumor. Generally, as used herein, the individual receiving the MUC-1 based formulation is a single organism. In certain embodiments, the subject is a mammal. In particular, the subject may be a human, including a male or female. In many embodiments, the individual is a patient, or an individual awaiting or receiving medical treatment and therapy.

In single or cumulative use, a dose of polypeptide of about 50 μ g, about 100 μ g, about 200 μ g, about 300 μ g, about 400 μ g, about 500 μ g, about 600 μ g, about 700 μ g, about 800 μ g, about 900 μ g, about 1,000 μ g, about 1,010 μ g, about 1,020 μ g, about 1,030 μ g, about 1,040 μ g, about 1,050 μ g, about 1,060 μ g, about 1,070 μ g, about 1,080 μ g, about 1,090 μ g, about 1,100 μ g, about 1,200 μ g, about 1,300 μ g, about 1,400 μ g, about 1,500 μ g, about 1,600 μ g, about 1,700 μ g, about 1,800 μ g, about 1,900 μ g, about 2,000 μ g, about 3000 μ g, about 5000 μ g, about 6000 μ g, about 3500 μ g, about 10000 μ g, about 15000 μ g, about 200 μ g, about 1,200, the polypeptide is present in BLP25 liposome vaccine. In a specific embodiment, the dose administered to an individual is about 1,000 μ g per week of a MUC-1 based formulation.

However, an individual may receive a dose of such a MUC-1-based formulation, e.g., multiple times per day, once every other day, once per week, or any other suitable dosing regimen. In some embodiments, at least 5 doses are administered to the individual over a period of time. In another embodiment, more or less than 5 doses are administered to the individual. Thus, an individual may receive a dose of about 1,000 μ g of MUC-1 lipidated polypeptide per week. Alternatively, the subject may receive a dose of 500 μ g in two doses, 2 times a week, or a dose of 100 μ g per day over a period of 5 days.

These dosage examples are not limiting and serve only to illustrate a particular dosing regimen for administering approximately 1,000 μ g of a MUC-1 lipidated polypeptide. For example, if a suitable dose is 1,000 μ g per week for a given situation, the dose may be broken down into any number of permutations. This is also true if the appropriate dose is more or less than 1,000 μ g for a particular condition.

The period of time for administering the MUC-1 based formulation to an individual may be any suitable period of time. Examples of such suitable periods of time include, but are not limited to, at least about 6 months, at least about 12 months, at least about 18 months, or at least about 24 months or longer. Thus, administration may continue for the entire treatment period, which is at least about 3 months, at least about 4 months, at least about 5 months, at least about 6 months, at least about 7 months, at least about 8 months, at least about 9 months, at least about 10 months, at least about 11 months, at least about 12 months, at least about 13 months, at least about 14 months, at least about 15 months, at least about 16 months, at least about 17 months, at least about 18 months, at least about 19 months, at least about 20 months, at least about 21 months, at least about 22 months, at least about 23 months, or at least about 24 months. The treatment period may also last longer than 24 months, if desired, for example, 30 months, 31 months, 32 months, 33 months, 34 months, 35 months, 36 months, or longer than 36 months. The physician can determine the amount of time that the individual should remain on the MUC-1 based formulation. In some cases, it may be advantageous to administer a MUC-1 based formulation for the remainder of the patient's life.

MUC-1 based formulations can be administered at different stages of treatment. For example, MUC-1 based formulations can be administered during the treatment phase and the maintenance phase. In some embodiments, the treatment phase may include administering the MUC-1 based formulation at a once weekly dose, while the maintenance phase may be for a longer period of time, e.g., every 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, or longer. In some cases, more doses will be given during the treatment phase than during the maintenance phase. However, the treatment and maintenance phases can be designed for specific individuals, and thus the time and dosage between treatment and maintenance phases can be significantly different from the above examples. In general, the maintenance phase may begin at any time deemed appropriate. For example, in some embodiments, the treatment phase may be eight weeks, while the maintenance phase will last for the entire lifetime of the individual. In another embodiment, only the treatment phase or maintenance phase is performed.

In other embodiments, MUC-1 based formulations can be administered prophylactically. In these embodiments, administration of a MUC-1 based formulation can prevent the individual from developing cancer, e.g., NSCLC or prostate cancer. When MUC-1 based formulations are used for prophylactic purposes, dosages and schedules can be readily determined.

III. Liposomes

As indicated above, in many embodiments, MUC-1 based formulations can be used with liposomes. Liposomes are microscopic vesicles consisting of one or more lipid bilayers surrounding an aqueous compartment. See, e.g., Bakker-Woudenberg et al, eur.j.clin.microbiol.infect.dis.12 (supply.1): s61(1993), and Kim, Drugs, 46: 618(1993). Because liposomes can be formulated with a large number of lipid molecules that are also found in natural cell membranes, liposomes are generally safe to administer and are biodegradable. Thus, liposomes are commonly used for drug delivery.

Depending on the method of preparation, liposomes can be unilamellar liposomes or multilamellar liposomes, and can range in size from about 0.02 μm to diameters in excess of about 10 μm. A variety of agents may be encapsulated or inserted into liposomes. The hydrophobic agent is distributed in the bilayer and the hydrophilic agent is distributed in the internal aqueous space. See, e.g., Machy et al, lipomes incellobiology AND PHARMACOLOGY (John Libbey, 1987), AND Ostro et al, American j.hosp.pharm.46: 1576(1989).

Indeed, liposomes can be adsorbed to any type of cell and then release the incorporated agent. In some cases, the liposome can be fused to a target cell, thereby emptying the contents of the liposome into the target cell. Alternatively, the liposomes may be endocytosed by phagocytes. Following endocytosis, intralysosomal (intracytosomal) degradation of the liposome lipids occurs and the encapsulated agent is released. Scherphof et al, ann.n.y.acad.sci., 446: 368(1985).

In addition, liposomes can be used to present active agents, e.g., polypeptides, on their surface and, thus, induce various events, e.g., signal cascades that amplify or initiate biochemical pathways, without fusion to the target cell or surfaces as mentioned in the preceding paragraph. Thus, for example, a polypeptide can be incorporated into a lipid bilayer, e.g., a lipid bilayer of a liposome, by a lipid attached to the polypeptide.

Liposomes are used as delivery vehicles when used with MUC-1 based formulations of the invention. Exemplary suitable liposomes that can be used in the methods of the invention include Multilamellar Liposomes (MLV), Oligolamellar Liposomes (OLV), unilamellar liposomes (UV), small unilamellar liposomes (SUV), medium unilamellar liposomes (MUV), large unilamellar Liposomes (LUV), giant unilamellar liposomes (GUV), multivesicular vesicles (MVV), unilamellar or oligolamellar liposomes prepared by reverse phase evaporation (REV), multilamellar liposomes prepared by reverse phase evaporation (MLV-REV), stable multilamellar liposomes (SPLV), frozen and thawed MLV (fatmlv), vesicles prepared by extrusion methods (VET), vesicles prepared by French Press (FPV), vesicles prepared by Fusion (FUV), dehydration-rehydration vesicles (DRV) and bubbelsomes (bsv). However, as understood by those skilled in the art, the type of liposome is not limited and may include any liposome prepared in any manner that is compatible with the methods of the present invention. Techniques for preparing liposomes are well known in the art. See, COLLOIDAL DRUG DELIVERY SYSTEMS, volume 66 (j. kreuter, ed., Marcel Dekker, inc., 1994).

IV. Lipid

As previously mentioned, in many embodiments, MUC-1 based formulations may be lipidated, for example, as set forth in SEQ ID No.: 2. As used herein, a "lipid" may be a myristyl, palmitoyl, or lauryl molecule that may be attached to an amino acid with a functional oxygen, nitrogen, or sulfur group. As mentioned above, these amino acids include, but are not limited to, threonine, serine, lysine, arginine, and cysteine.

"Monolipopeptide (monolipopeptide)" is a peptide to which only one lipid chain is attached. Similarly, a "lipopeptide (dilipopeptide)" is a peptide having two lipid chains attached to one or two amino acids. If two lipid chains are attached to two amino acid residues, these residues may be separated by any number of amino acids. In the case of more than one lipid attachment, the lipids may be the same lipid or may be different lipids. Similarly, if more than two lipids are attached, the two or more lipids may be the same or all the lipids may be different.

It is believed that lipopeptides such as BLP25 can be incorporated into liposomes because the lipid portion of the peptide spontaneously incorporates into the lipid bilayer of the liposome. Thus, in this case, the lipopeptide may be present on the "surface" of the liposome. Alternatively, the peptide may be encapsulated within a liposome. Techniques for preparing and formulating liposomes using molecules such as peptides are well known.

V. Exemplary adjuvants

The MUC-1 based formulations of the present invention may also comprise an adjuvant. Alternatively, the adjuvant may be administered prior to, together with, or after administration of the MUC-1-based formulation of the invention.

As is recognized, an adjuvant is a substance that works with a specific antigenic stimulus to enhance a specific response to an antigen. For example, monophosphoryl lipid a (mpla) is an effective adjuvant that results in increased presentation of liposomal antigens to specific T lymphocytes. Lving, c.r., immunobiol, 187: 430-446(1993). MPLA binds to toll-like receptors that lead to activation of defense signaling pathways that control expression of various immune response genes.

Lipid-based adjuvants, such as lipid A and its derivatives, are suitable for use with MUC-1 based formulations. Muramyl Dipeptide (MDP) or alum has also been shown to increase adjuvanticity when incorporated into liposomes (Gupta RK et al, Adjuvants-A balancebeen toxicity and adjuvancity, "Vaccine, 11, 293-" 306 (1993)).

Another class of adjuvants useful in the present invention includes stimulating cytokines, such as interleukin-2 (IL-2). Thus, the liposome vaccine of the present invention can be formulated with IL-2, or IL-2 can be administered separately for optimal antigenic response. In many embodiments, it is beneficial to formulate IL-2 and liposomes together.

MUC-1 based formulations can also be co-formulated with synthetic adjuvant mimics. In this aspect, the lipid a mimetic can be used with a liposomal vaccine. One particular type of lipid a mimetic is one in which one or both of the sugar units of the lipid a disaccharide is replaced with at least one carbon backbone of pentaerythritol. See, for example, WO 03/094850, which is incorporated herein by reference.

VI. Exemplary vaccine formulations

When the MUC-1 based formulation is a vaccine, the vaccine may also be formulated with pharmaceutically acceptable excipients. The nature of such excipients is well known in the art, but generally includes excipients that are physiologically tolerated and inert or that promote the vaccine properties of the compositions of the present invention. Non-limiting examples of pharmaceutically acceptable excipients include liquid carriers such as sterile, physiological saline. The excipients may be added at any time during the formulation of the liposome vaccine or they may be mixed with the complete vaccine composition. One can readily determine when to add excipients and suitable excipients for use with the vaccines of the present invention.

One specific vaccine formulation may comprise about 300 μ g of SEQ id no: 2, about 150 μ g lipid a, and about 15mg of one or more additional liposomal lipids, such as dipalmitoylphosphatidylcholine, cholesterol (DPMC), and Dimyristoylphosphatidylglycerol (DPMG).

One specific vaccine formulation may comprise about 1000 μ g of SEQ ID NO: 2, about 500 μ g lipid a and about 29.1mg dipalmitoylphosphatidylcholine, about 17.3mg cholesterol and about 3.6mg Dimyristoylphosphatidylglycerol (DPMG).

VII. Cyclophosphamide

Prior to treatment with MUC-1 based formulations, subjects may be "pre-treated" with cyclophosphamide. In many embodiments, the dose of cyclophosphamide is about 50mg/m²、100mg/m²，200mg/m²、300mg/m²About 400mg/m²About 500mg/m²Or about 600mg/m². At about 300mg/m²Doses of cyclophosphamide within the range are considered low doses. In certain embodiments, the cyclic squamous amide is administered as a single dose. In other embodiments, more than one dose of cyclophosphamide may be administered over a period of time.

Cyclophosphamide dosage is e.g. 300mg/m²Can partially overcome the immunosuppression seen in some cancer patients. Cyclophosphamide has been shown to increase delayed-type hypersensitivity, increase antibody production, eliminate tolerance, and enhance immunity against tumors in certain individuals in various animal models. Other drugs that affect the immune system in a similar manner to cyclophosphamide may also be used in a pretreatment regimen using the formulations of the present invention.

VIII. Routes of administration and targeting of L-BLP25 vaccine

MUC-1 based formulations of the invention, including vaccines, can be formulated for a variety of routes of administration. Specific routes include any suitable method of administration, for example by intramuscular, subcutaneous or intradermal injection, aerosol, transdermal, pulmonary, nasal or oral administration, or by a combination of these routes, either simultaneously or in multiple unit doses.

The administration of vaccines is well known and ultimately depends on the particular formulation and the judgment of the attending physician. MUC-1 based formulations, such as L-BLP25, may be maintained as a suspension, or may be lyophilized and then hydrated to produce a useful formulation.

In some embodiments, such as the embodiment of example 1, one dose of a MUC-1 based formulation can be injected into several different sites. For example, in the embodiment of example 1,1,000 μ g of MUC-1 based polypeptide may be administered in 4 sub-doses of about 250 μ g each. In the case of injection, the amount injected is immaterial, so long as an appropriate dose or sub-dose of the composition of the invention is administered. For example, one injection may be 1cc (ml) and another injection with the exact same dose may be 5cc (ml). Furthermore, the amounts in the sub-doses are merely non-limiting examples, and embodiments are contemplated in which the sub-doses are more or less than the full dose of 1/4.

The sub-dose or doses may be administered in the deltoid or triceps region of the upper arm and in the left and right anterior lateral directions of the abdomen. However, these injection sites are merely examples. In some embodiments, only two sub-doses are administered and these sub-doses may be administered in any of the regions described above. In other embodiments, sub-doses or full doses may be administered in distinct regions. If MUC-1 based formulations are injected, the appropriate injection site can be readily determined.

To provide greater specificity, and thus theoretically reduce the risk of toxicity or other unwanted effects during in vivo administration, in some embodiments, the inventive compositions are targeted to cells through which the compositions are designed to act, i.e., antigen presenting cells. This is conveniently achieved using conventional targeting techniques to direct liposomes containing immunogenic peptides to specific locations in the body. To target antigen presenting cells, for example, the mannose and Fc portions of an antibody may be chemically conjugated to an antigenic peptide, or a targeting peptide may be recombinantly fused to an immunogenic lipopeptide. In addition, similar strategies are familiar to those skilled in the art. Nonetheless, in some embodiments, the compositions of the present invention are not targeted to a particular cell type or organ.

IX. The treated individual

Any individual diagnosed with NSCLC or prostate cancer may be treated with the MUC-1-based formulation described herein. Alternatively, any individual exhibiting symptoms of any stage of NSCLC or any stage of prostate cancer, but not yet formally diagnosed as having NSCLC or prostate cancer, may also be treated with a MUC-1-based formulation. Furthermore, as noted above, MUC-1 based formulations can be administered prophylactically to prevent an individual from developing NSCLC or prostate cancer.

In selecting an individual suffering from NSCLC and/or prostate cancer for treatment with a MUC-1 based formulation, it may be beneficial to determine the level of MUC-1 in the serum of the individual prior to or during treatment. In some cancer patients, high serum MUC-1 levels have been associated with poor prognosis. See, e.g., Pihl et al, Pathology, 12: 439-447(1980). Because abnormal amounts of circulating MUC-1 may inhibit or reduce the efficacy of the interaction of an exogenous MUC-1 based formulation, knowledge of the amount of endogenous MUC-1 may help determine the appropriate dosage of MUC-1 based formulation to be administered to an individual.

X. Individuals with NSCLC

When an individual suffering from NSCLC is to be treated with a MUC-1-based formulation of the invention, an individual diagnosed with stage IIIB regional Locality (LR), stage IIIB or stage IV NSCLC with malignant pleural effusion can be specifically treated. However, the invention also includes treating NSCLC individuals other than individuals with stage III locoregional, stage III pleural effusion, and stage IV disease. Thus, the present invention contemplates treating patients diagnosed with stage IA, stage IB, stage IIA, stage IIB, stage IIIA, stage IIIB locoregional, stage IIIB pleural effusion, and stage IV NSCLC. See mount c.f., chest; 111(6): 1710-7(1997), which are incorporated herein by reference.

Indeed, the present invention contemplates treating patients at varying severity of NSCLC, not just patients exhibiting advanced NSCLC cancer. For example, according to the present invention, an individual diagnosed with stage IIIA NSCLC can be treated.

A. Staging of Lung cancer

Generally, when a MUC-1 based formulation is used in an individual with NSCLC, the stage of NSCLC in the individual may be determined before, after, or during treatment. The staging of lung cancer is summarized below:

normally in lung cancer, an increased number of "stages" and worse prognosis are associated. To diagnose individuals at a particular stage, the size and location of the primary tumor (the "T" value) as well as the degree of nodal involvement (the "N" value) and the increased probability of metastasis are considered. Also of note when diagnosing individuals is the absence of metastasis ("M0") or the presence of metastasis ("M1").

1. Class T

The T class consists of subclasses T1 to T4, where numbers increasing from 1 to 4 represent increasing size and local invasion of the primary tumor. T1 and T2 differ mainly in size, e.g., T1 is less than 3cm and T2 is greater than 3 cm. T3 tumors typically involve the chest wall and include, but are not limited to, the superior sulcus (lung), diaphragm (diaphragm), mediastinum pleura, pericardium, or proximal main stem bronchus (progiamal brain bronchus), but are resectable. The T4 tumor cannot be surgically excised because it may have invaded the mediastinum and may involve the heart, large blood vessels, trachea, keel valve or esophagus, or in the case of malignant pleural effusion, the pleura.

2. Class N

The nodular phase is divided into N1, N2 and N3. The N1 nodules generally include bronchial or ipsilateral portal nodules (ipsilateral tubular nodules). These nodules are localized within the pleural membrane. The N2 nodules generally include ipsilateral mediastinal or subclavian (subarachnal) nodules. The N3 nodules generally include the contralateral portal or mediastinal nodules, any oblique muscle nodules, or supraclavicular nodules.

3. Stage NSCLC

Thus, the "stage" of NSCLC represents a distinct classification of NSCLC based on various T, N and M-value permutations. The recognized NSCLC stage is as follows:

latent cancer (Occult Carcinoma): in this category, the patient is classified as TX N0M0, meaning that it has malignant cells detected in secretions of its broncho-pulmonary, but no tumor confirmed by bronchoscopy or radiography.

Stage IA and stage IB: stage IA was graded as T1N 0M0 based on 5-year survival outcomes significantly better than patients with stage IB disease (T2N 0M 0). Surgery is preferably performed on these patients. In 1997, the 5-year survival rate for patients undergoing surgery graded as stage IA was 67%, while that for stage IB was 57%.

Stage IIA and stage IIB: stage IIA disease is defined as T1N 1M 0 and has a 5-year survival rate of 55% based on surgical staging. Stage IIB disease consists of T2N 1M 0 and T3N 0M 0. The designation T3N 0M0 indicates extrapulmonary spread of the tumor but no lymph node involvement. The classification of categories T3N 0M0 and T2N 1M 0 into one group is due to the 5-year survival rate of their respective surgically staged diseases, with no significant difference between 38% and 39%. Surgery is also the primary treatment for these individuals.

Stage IIIA: stage IIIA patients were considered resectable, although IIIB patients failed. Stage IIIA patients were defined by lesions with extrapulmonary spread (T3) and limited lymph node involvement (N1 or N2). Nodal involvement may spread to the ipsilateral mediastinal and/or subclavian lymph nodes. These patients were classified as T3N 1M 0 or TI-3N 2M 0. In 1997, the 5-year survival rate for stage IIIA disease was 23%.

Stage IIIB: stage IIIB classification refers to patients with extrapulmonary involvement including, but not limited to, contralateral mediastinal or portal lymph nodes, ipsilateral or contralateral supraclavicular lymph nodes or scalene muscle nodules, extensive mediastinal nodules without distant metastasis, or cytologically positive malignant pleural effusion. These patients can be classified as T1-3N 3M 0 or T4N 0-3M 0. In 1997, the 5-year survival rate of clinically staged disease with multidisciplinary therapy was 5%.

Stage IV: stage IV is defined by any metastatic involvement. These patients were classified as M1 with any T and any N. In 1997, more than a quarter of patients with NSCLC had clinical stage IV.

XI. Individuals with prostate cancer

Similar to the low survival rate of individuals with advanced lung cancer undergoing multidisciplinary therapy, men with prostate cancer who have undergone biochemical failure following prostatectomy have few treatment options. One treatment option that it does have is androgen ablation therapy (ADT). Unfortunately, this treatment has significant morbidity, particularly when used over long periods of time.

In individuals with prostate cancer, it is known that prostate-specific antigen ("PSA") levels in the blood rise, for example, when the prostate gland is enlarged. Thus, PSA is a good biological or tumor marker for prostate cancer. In individuals with more advanced disease, treatment-induced decreases in PSA and an increased survival association (Scher, et al, J.Natl.cancer Inst.; 91 (3): 244-51 (1999)).

XII. Treatment of individuals with NSCLC or prostate cancer

The present invention includes treating NSCLC individuals with MUC-1-based formulations of the invention at all stages of NSCLC and treating individuals with prostate cancer, including individuals with prostate cancer that has failed PSA after radical prostatectomy. The phrase "treating" or "treatment" as used herein refers to the use of a formulation or vaccine to prevent, cure, reverse, attenuate, lessen, minimize, inhibit or halt the deleterious effects of a disease state, disease process, disease causative agent or other abnormal condition.

In some embodiments, prior to treatment with the compositions of the present invention, an individual having NSCLC or prostate cancer may have previously received radiation therapy or surgery. The subject may also be treated with chemotherapy, radiation therapy or surgery before, during or after the subject has been treated with the MUC-1-based formulation of the invention. In the case of these individuals, any acceptable cancer treatment may be administered before, during or after treatment with the MUC-1-based formulation.

Inclusion and exclusion criteria (inclusion and exclusion criteria) may be used when selecting individuals for treatment using the formulations and vaccines of the present invention. For example, in one embodiment, when treating an individual with NSCLC with a MUC-1 based formulation, the individual to be treated may be a male or female with an age of over 18 years who is stable in their disease, or who responds to treatment after completion of their first line (first line) standard of chemotherapy. Individuals other than those described above may also be treated with MUC-1 based formulations. Indeed, some individuals treated with the compositions of the present invention may not be treated with chemotherapy prior to treatment with a MUC-1-based formulation.

XIII. Possible inclusion and exclusion criteria for individuals with NSCLC

In another embodiment, the individual with NSCLC selected for treatment has an Eastern Cooperative Oncology Group (ECOG) performance status of ≦ 2 with a neutrophil count ≧ 1.5X 10⁹The platelet count is more than or equal to 100 multiplied by 10⁹/L、WBC2.5×10⁹The ratio of the concentration of the hemoglobin to the concentration of the hemoglobin is 90 g/L. However, while ECOG numbers can be used to evaluate an individual undergoing treatment, no particular ECOG number is required before, during, or after treatment.

Other inclusion criteria may include expected survival for 4 months and when the individual has understood and signed written consent. Of course, these are not constant inclusion criteria, and treatment of individuals with shorter life expectancy is envisioned. Furthermore, when MUC-1 based formulations become the mainstream cancer treatment, it will be possible for an individual to prescribe the composition of the invention without the need for signed written consent.

For individuals with NSCLC who may be excluded from treatment, the exclusion criteria are only instructive. In many cases, individuals exhibiting one or more exclusion criteria (including all exclusion criteria) may still be treated with a MUC-1-based formulation. Examples of exclusion criteria for NSCLC individuals include: (a) surgery or immunotherapy within 4 weeks prior to treatment, (b) immunosuppressive drugs including systemic corticosteroids (c) having past or present tumor history in addition to lung cancer, (d) autoimmune diseases or recognized immunodeficiency disorders, (e) clinically significant liver or kidney dysfunction, (f) major heart disease or active infection, or (g) individuals who have undergone a splenectomy, are administered within 3 weeks prior to treatment.

XIV. Possible inclusion and exclusion criteria for individuals with prostate cancer

Similar to individuals with NSCLC, individuals with prostate cancer may also receive inclusion and exclusion criteria. Again, these criteria are only instructive, and individuals with prostate cancer who do not meet any one of the inclusion criteria or meet any one or all of the exclusion criteria may still be treated under the methods of the invention. For individuals with prostate cancer, inclusion criteria may include: (a) a radical prostatectomy performed at least 6 months prior to treatment, (b) three consecutive increases in serum PSA values after radical prostatectomy with an increase of at least 50% above the nadir after prostatectomy, (c) no evidence of malignancy in a pretreatment evaluation as evidenced by negative pelvic CT and bone scans, (d) ECOG performance status of 0, 1, (e) normal hematologic, hepatic and renal function tests, (f) understanding and signing written informed consent; and (g) individuals who have been treated with hormone therapy for prostate cancer (i.e., neoadodjuvanttreatment (pro-RP) prior to radical prostatectomy) must have serum testosterone in the normal range. As noted above, these inclusion criteria are only instructive, and many individuals with different criteria may be treated using the methods of the invention. For example, individuals who have had prostate cancer who have not undergone a complete prostatectomy may be treated. In addition, individuals who do not have increased serum PSA, or whose serum PSA is not continuously increased or does not increase by more than 50% compared to nadir after prostatectomy, may also be treated.

For individuals with prostate cancer, exclusion criteria that may be used (although are not required) include: (a) hormone therapy within 6 months prior to treatment, (b) immunotherapy within 4 weeks prior to treatment, (c) radiation therapy to the prostate bed (prostate bed) within one year prior to treatment, (d) treatment with immunosuppressive drugs, such as cyclosporin or adrenocorticotropic hormone (ACTH) or long-term therapy requiring the use of corticosteroids, (e) known autoimmune or immunodeficiency disorders, or (f) clinically significant heart disease or active infection. Again, these exclusion criteria are merely exemplary. For example, in individual cases, individuals with prostate cancer and clinically significant heart disease may be treated with the methods of the present invention.

XV. Effect of the treatment

Treatment with the MUC-1 based formulations described herein can result in a variety of effects. One effect of treating an individual diagnosed with NSCLC, particularly an individual diagnosed with stage IIIB NSCLC, with a MUC-1-based formulation is an increase in the length of survival. Similarly, administration of the MUC-1 based formulation to an individual may affect the "quality of life" or "health related quality of life" of the individual. An increase in survival and an impact on quality of life can also be seen in treated individuals with prostate cancer. Furthermore, in certain individuals with prostate cancer, treatment with MUC-1 based formulations will result in lower PSA, stabilized PSA, or reduced PSA doubling rates.

A comparison of the effect of treatment with a MUC-1 based formulation can be made between treated individuals and individuals who are not undergoing treatment or who undergo best support therapy (BSC). BSCs include many alternative types of treatment that do not include treatment with MUC-1 based formulations. For example, a BSC, although generally arbitrarily dependent on the environment, may include psychosocial support, analgesia, and nutritional support. In some embodiments, the comparison of therapeutic efficacy is performed between individuals receiving different amounts of a MUC-1-based formulation. In other embodiments, the individual receives the BSC in combination with treatment with a MUC-1-based formulation.

Prior to treating an individual with a MUC-1 based formulation of the invention, the individual may receive an assessment of pre-treatment. Non-limiting examples of pre-treatment assessments include complete medical history and physical examination. The physical examination may include such items as a CT scan or X-ray. The subject may also be evaluated for treatment during the course of treatment. Treatment assessment may include monitoring vital signs of an individual, examining injection sites, and analyzing blood samples.

The treated individual may also be evaluated by determining the factors: (a) tumor size, (b) tumor location, (c) nodal stage, (d) growth rate of NSCLC or prostate cancer, (e) survival rate of the individual, (f) change in lung or prostate cancer symptoms in the individual, (g) change in PSA concentration in the individual, (h) change in the rate of doubling of PSA concentration in the individual, or (i) change in quality of life of the individual.

XVI. Increased survival time in NSCLC individuals by administration of MUC-1 based formulations or BLP25 liposome vaccines

One advantage of treating an individual suffering from NSCLC or prostate cancer with a MUC-1-based formulation of the invention is that the individual may have a longer survival time than an individual not receiving treatment with a composition of the invention. Survival can be determined by comparing the current number of survivors to the number of individuals starting treatment with a MUC-1 based formulation. In other embodiments, survival rates can be compared to the survival rates disclosed for particular types of cancer. Generally, survival can be measured at any time after treatment is initiated.

For example, survival may be measured at less than 6 months, greater than 6 months but less than 1 year, 1 year or greater than 1 year but less than 2 years, 2 years or greater than 2 years but less than 5 years, or 5 years or greater than 5 years after initiation of treatment. In some embodiments, increased survival rates will demonstrate that MUC-1-based formulations of the invention affect a particular individual.

XVII. Maintenance of quality of life and lung cancer symptoms by administration of MUC-1 based formulations

As mentioned above, another advantageous aspect of treating an individual suffering from NSCLC or prostate cancer with a MUC-1-based formulation of the invention is to maintain or improve the quality of life of the individual. Clinicians and regulatory agencies recognize that an individual's "quality of life" ("QoL") is an important endpoint in cancer clinical trials. See, e.g., Plunkett et al, clin. lung Cancer, 5 (1): 28-32(2003), and Cella et al, j.clin.epidemol., 55 (3): 285-95(2002), each of which is incorporated herein by reference.

The 4 most important quality of life indicators are physical and occupational function, psychological state, social interaction and somatic sensation. In this regard, two lung Cancer questionnaires, namely the European Organization for research and Treatment of Cancer ("EORTC") and the evaluation of Cancer Treatment function ("FACT-L"), in individuals with NSCLC can be used to evaluate the individual's, particularly the individual's, health-related quality of life before, during and after Treatment with the MUC-1-based formulation described herein.

It is contemplated that the methods of the invention may be used in conjunction with the evaluation of various sub-markers (subscales) for monitoring the Physical Well-being (PWB), Social/Family Well-being (SWB), Emotional Well-being (EWB), functional Well-being (FWB) and Lung Cancer symptom sub-markers (Lung Cancer symptomSubscale) (LCS) of an individual. Although the secondary markers of lung cancer symptoms are clearly tailored to the individual with lung cancer, different secondary markers may be used for different cancers. Thus, different side-markers may be used for individuals with prostate cancer. Depending on which "health" scores are combined, one can obtain either a "FACT-L Score" (sum of all subtotals) or a "trial result Score (TOI)" (sum of PWB, FWB and LCS subtotals). TOI is a reliable indicator of meaningful changes in quality of life. See, Cella et al, supra.

Individuals may be evaluated for their FACT-L and TOI scores before, during and after treatment with MUC-1 based formulations of the invention. For example, TOI scores may be obtained at baseline, i.e., prior to treatment, and then at various time intervals after treatment initiation, i.e., over 4 weeks, 8 weeks, 19 weeks, 31 weeks, or 43 weeks or more. These different time intervals are merely exemplary, and the quality of life indicator may be obtained at any suitable time. For example, a first TOI score may be obtained after a first treatment, rather than at baseline. Then, the change in score between time points can then be calculated to determine trends relating to improvement, worsening or maintenance of quality of life.

It has been calculated that a 3 or more point decrease in LCS from baseline is a clinically significant worsening in lung cancer symptoms, while an increase of 3 or more points is a clinically significant improvement in lung cancer symptoms. Also, for the TOI score, a decrease of 7 points or more indicates a deterioration in quality of life, and an increase of 7 points or more indicates an improvement in quality of life.

In some embodiments, a clinical improvement in lung cancer symptoms or quality of life demonstrates that a MUC-1 based formulation affects a particular individual.

Thus, administration of a MUC-1 based formulation of the invention may be used to improve or maintain the quality of life of a treated individual having NSCLC or prostate cancer. In measuring the effect on quality of life, the magnitude of the effect can be determined from a baseline or from any treatment point. In some embodiments, an impact magnitude between 0.2 and < 0.49 indicates a small effect, 0.5 to 0.79 indicates a medium effect, and 0.8 or greater indicates a large effect. These numbers are exemplary only, and the magnitude of the effect may vary for treatment of certain individuals.

Administration of MUC-1 based formulations can also be used to prevent the deterioration in quality of life seen in many cancer patients over time. For example, in some embodiments, administration of a MUC-1 based formulation, such as the BLP25 liposome vaccine, may result in a quality of life index that remains substantially unchanged or does not reach a level that worsens or improves quality of life.

In one embodiment, the invention comprises improving or maintaining the quality of life or improving or stabilizing lung cancer symptoms in an individual diagnosed with NSCLC by determining the TOI or LCS score of the individual before, during and after treatment with a BLP25MUC-1 based formulation described herein.

XVIII. Reduction of PSA doubling time

In some embodiments, treatment of an individual with prostate cancer with a MUC-1-based formulation of the invention will result in a decrease in PSA concentration, stabilization of PSA concentration, or a decrease in PSA doubling time. In general, the effect of MUC-1 based formulations on PSA concentration or PSA doubling time can be measured at any time. For example, while the concentration of PSA after treatment can be compared to a baseline value, the concentration of PSA can also be compared between treatment points or between a particular treatment point and a treatment endpoint. In certain embodiments, the PSA response is determined during treatment.

XIX. Evaluation of treatment with immune function

In some embodiments, an assay of immune function, such as a T cell proliferation response assay, can be used to measure an individual's response to a MUC-1-based formulation. In some embodiments, the results from the T-cell proliferation response assay can be used to determine whether treatment with a MUC-1-based formulation affects an individual. The results from these assays can also be used to determine the individual's response to the agent at various time points during treatment.

The assay to measure proliferating T-cells is not particularly limited and can be performed by any method known in the art. A comparison of T cell proliferative responses can be made to compare pre-treatment versus post-treatment responses and to compare immune responses during treatment.

XX. Other cancers

The invention also encompasses the treatment of cancers other than NSCLC and prostate cancer with MUC-1-based formulations described herein.

Any individual with a MUC-1 expressing cancer can be targeted for treatment with MUC-1 based formulations. For example, individuals with adenocarcinoma expressing MUC-1 protein may be targeted for treatment with BLP25 liposome vaccine. Examples of adenocarcinomas include, but are not limited to, ovarian cancer, liver cancer, e.g., invasive cholangiocarcinoma of the liver, colon cancer, breast cancer, pancreatic cancer, e.g., invasive ductal carcinoma of the pancreas, and renal cancer. Another MUC-1 expressing cancer is a cancer of the head and neck.

The following examples are intended to illustrate but not limit the invention. Although it is representative of how the methods of the present invention are used, other methods that are consistent with the spirit of the present invention are also contemplated and used. Throughout this specification, any and all references to publicly available documents, including U.S. patents, are specifically incorporated herein by reference.

Example 1

Phase II study of liposomal MUC1 vaccine for treatment of NSCLC

This example demonstrates the efficacy of the L-BLP25 vaccine in the treatment of individuals with stage IIIB locoregional or stage IV NSCLC.

Patients treated with the L-BLP25 vaccine exhibited increased survival rates. Furthermore, the maintenance of stable physical health and the maintenance of the quality of life of the individual throughout the treatment and maintenance phases of the treatment, as measured by the FACT-L total score and the test result Index (Trial Outcome Index), demonstrates the clear advantage of adding BLP25 liposome vaccine to the optimal supportive care regimen compared to the optimal supportive care regimen alone.

Method of producing a composite material: the controlled, open label phase IIb trial recruited 171 patients. Of the 171 patients enrolled, 65 had regional IIIB disease. Of these patients, 35 were randomly selected to receive treatment and 30 were randomly selected to receive the best standard of care. The groups were adequately balanced by age and race. More women and ECOG0 patients were randomized for treatment versus best standard therapy (BSC) (51.4% and 36.7%, and 40.0% and 26.7%), and more patients with treatment regimens received radiation therapy for cancer treatment in addition to chemotherapy prior to trial enrollment (91.4 versus 76.7%).

The L-BLP25 vaccine used in this particular experiment was a lyophilized formulation consisting of (1)1000 μ g of a BLP25 lipopeptide, e.g., comprising SEQ ID NO: 2, (2)500 μ g of immunoadjuvant monophosphoryl lipid a, and (3) three lipids that form a liposome product: (i)17.3mg cholesterol, (ii)3.6mg dimyristoyl phosphatidyl glycerol, and (iii)29.1mg dipalmitoyl phosphatidyl choline.

All patients receiving at least 5 vaccinations with the L-BLP25 regimen, 96.6% of these patients completed the primary phase and 69.3% continued with the maintenance phase of the treatment plan. Second-line therapy under investigation consists mainly of chemotherapy (two or three lines), radiotherapy and surgery. During the primary treatment period of the study, 5 patients on the L-BLP25 approach and 10 patients on the BSC approach received second line therapy. Of the patients who continued the maintenance phase of the study, 43 patients underwent the L-BLP25 approach and 45 patients underwent the BSC approach received second line therapy.

To enhance antigenic stimulation of more draining lymph nodes (draining lymph nodes), 4 anatomical sites were administered with the vaccine. A1000 μ g dose of L-BLP25 was administered by 4 subcutaneous injections of 0.5mL, each injection containing one quarter of the total dose. Sub-doses were administered in the deltoid or triceps muscle of the upper arm and in the left and right anterior directions of the abdomen.

Generally, as used in this experiment, survival time is defined as the time from the date when random selection was performed to the date of death. For patients who were alive or no longer followed up at the time of analysis, the interval between the date when the random selection was made and the date when the patient was last known to be alive was calculated and used as a checked observation in the analysis (censorediboservation). In this example, survival was monitored at 3 month intervals over 12 months after patient natural growth (patient actual) was completed.

The FACT-L QoL questionnaire was administered to all patients at specific time points. QoL analysis includes evaluation of mean FACT-L individual change scores from baseline to fourth and eighth weeks, graphical representation of QoL scores over time, and analysis of the area under the curve for total and subtest scores. The magnitude of the impact of the quality of life change between treatments is determined from baseline. An influence magnitude between 0.2 and < 0.49 indicates a small effect, 0.5 to 0.79 indicates a medium effect, and 0.8 or more indicates a large effect.

Results: as shown in fig. 2, the observed 2-year survival of stage IIIB regional patients was 36.7% of the vaccine approach versus the control approach, demonstrating a significant increase in life expectancy of 23.3%. In the total patient population, the two-year survival was 43.2% of the vaccine approach versus 28.9% of the control approach, demonstrating a 14.3% increase in life expectancy. See fig. 1. At 13.3 months, median survival for the phase IIIB regional patients on best standard therapy alone was similar to the overall median survival for the entire group on best standard therapy. In contrast, the median overall median survival for phase IIIB regional patients treated with L-BLP25 was still unsatisfied at a minimum median of 24 months, demonstrating an increase in life expectancy of at least 10.7 months. This is surprising and unexpected because prior to the introduction of the MUC-1 compositions of the invention, there were no viable treatment options for this class of patients that could produce such results.

With respect to quality of life, a significant advantageous aspect of the L-BLP25 approach was demonstrated compared to the BSC approach. More patients treated with the L-BLP25 approach showed clinically meaningful improvement or no change compared to patients treated with the BSC approach. In the BSC-only approach, more patients showed clinically meaningful exacerbations in the test result index (TOI).

Method of producing a composite material: a T-test was used for comparative analysis of regional area of IIIB (LR) disease and malignant Pleural Effusion (PE)/IV subgroup IIIB between patients receiving treatment and BSC only patients, with regard to the FACT-L total score, various side-markers and TOI. A negative total/TOI change score indicates a deterioration in QoL, while a positive total/TOI change score indicates an improvement. Subgroup analysis showed that stage IIIB LR patients treated with L-BLP25 had better QoL. This is consistent with the previous data demonstrating a clinically meaningful improvement in survival in stage IIIB LR patients treated with BLP25 (p ═ 0.0692).

The results of the quality of life comparison are shown in table 1 below:

design of research

Week-2: the FACT-L QoL questionnaire is administered.

Week-2: patients were randomly selected for either L-BLP25+ optimal standard therapy or only optimal standard therapy (optimal standard therapy includes palliative radiation therapy and/or second-line chemotherapy according to current clinical practice, and may also include psychosocial support, analgesia, and nutritional support).

Week-2: pretreatment evaluation (complete history, physical examination and clinical laboratory studies). If clinically approved, evaluation of other potential disease sites is performed to exclude progressive disease in other areas. Women with childbearing potential require a negative pregnancy (HCG) test prior to treatment.

Day-3: the treatment means patients receive 300mg/m²A single intravenous dose of cyclophosphamide.

Week 0 to 7: L-BLP25 was inoculated with #1 to #8 (primary treatment period). Patients undergoing the L-BLP25 regimen were subjected to vital sign assessment and previous injection sites were examined prior to each L-BLP25 treatment. Vital signs were also monitored one hour after each L-BLP25 treatment. Patients were diary-checked after each vaccination to record any adverse events, and previous injection sites were assessed at each follow-up visit (sequent visit). Toxicity was graded according to CALGB ExpandedCriteria.

Week 4: the evaluation of the treatment of patients taking therapeutic measures and the safety and immunological blood work are performed. The FACT-L QoL questionnaire was performed on all patients.

Week 8: treatment assessments (physical examination, ECOG status, vital signs, examination of the treatment site with the means of L-BLP25 and assessment of adverse events) were performed. Blood samples were also drawn and analyzed for standard safety (hematology and chemistry) and immune responses. The FACT-L QoL questionnaire was performed on all patients.

Week 19+ d: maintenance vaccination (6 week intervals) and treatment evaluation (12 week intervals). Patients who underwent the L-BLP25 procedure underwent treatment evaluation and safe blood work at each maintenance vaccination and an immunological profile check one week after the first maintenance vaccination. EACT-L QoL questionnaires were performed on all patients.

Patient population

Inclusion criteria

1. Men and women over the age of 18 with NSCLC who are stable or who respond to treatment following completion of one line of standard chemotherapy.

2. Easter Cooperative Oncology Group (ECOG) performance status less than or equal to 2, neutrophil count more than or equal to 1.5 × 10⁹The platelet count is more than or equal to 100 multiplied by 10⁹/L、WBC≥2.5×10⁹L and hemoglobinIs 90 g/L.

3. Expected survival of 4 months.

4. Written consent for understanding and signing.

Exclusion criteria

1. Surgery or immunotherapy was performed within 4 weeks prior to study entry.

2. Immunosuppressive drugs, including systemic corticosteroids, were administered within 3 weeks prior to study entry.

3. Past or present tumor history, except for lung cancer.

4. Autoimmune diseases or recognized immunodeficiency diseases.

6. Clinically significant liver or kidney dysfunction.

7. Obvious heart disease or active infection, or patients who have undergone splenectomy.

Example 2

T cell proliferation response assay

This example demonstrates that the MUC-1 formulation of the invention is directly responsible for the median increase in survival shown in example 1.

Using the patients enrolled in the study of example 1, lymphoproliferation assays were performed before and after vaccination to monitor the MUC1 antigen-specific TH response (proliferation of helper T cells) to measure the kinetics of the patient's immune response against MUC1 cells. T cell proliferation assays were performed on patients using the L-BLP25 approach before immunization and at several time points after immunization.

In patients using the L-BLP25 protocol, 78 patients were evaluated for T cell proliferative response. 16 patients were determined to have a positive MUC1 specific T cell proliferation response (which was absent prior to immunization) induced by the L-BLP25 vaccine. Of the 16 patients who developed an immune response, 2 had stage IIIB regional disease, and the remaining patients had stage IV disease. Patients with positive proliferative responses had a median survival of 27.6 months using the L-BLP approach, while patients with negative proliferative responses had a median survival of 16.7 months. These results demonstrate that the MUC-1 formulation of the invention is directly responsible for an increase in median survival expected for 10.9 months of life.

Example 3

Phase II study of liposomal MUC1 vaccine in PSA failure following Radical Prostatectomy (RP)

This example shows the immunotherapeutic effect of the L-BLP25 vaccine on PSA levels in men with elevated PSA following radical prostatectomy.

At the end of the primary treatment period (week 8), 8/16 patients had stable PSA. One patient maintained stable PSA until the end of the study period (week 49). There was a significant extension of PSA doubling time ("PSADT") for all enrolled patients, but except one. Doubling time is the length of time it takes for an individual to double PSA levels, and it is a factor used to predict survival after surgery in individuals with prostate cancer. This data shows that doubling time is over 50% in 6/16 patients.

Method of producing a composite material: males with biochemical failure evidenced by 3 elevations of PSA after prostatectomy were enrolled. This included 16 patients with a median age of 60 years, an ECOG score of 0 or 1, and a median Gleason score of 7. The primary endpoints are efficacy (as measured by PSA response) and safety of MUC1 liposome vaccine (L-BLP 25). Changes in PSA doubling time (PSADT) were also estimated. The patient receives 300mg/m²Cyclophosphamide (C)TX), then 8 times per week 1 time subcutaneous inoculation with L-BLP25 (treatment) containing 1,000 μ g of antigen. Subsequent inoculations were given at 6 week intervals up to week 49 (maintenance). PSA concentrations were measured during the treatment and maintenance phases and PSADT was calculated for these time intervals and compared to PSADT prior to enrollment.

All 16 patients received CTX and 15/16 completed the treatment period. 10 patients completed the maintenance period. The most common adverse events after treatment were nausea (31%) and fatigue (25%); however, none of these deleterious effects worsen beyond level 1.

Results: after induction, 8/15 evaluable patients had stabilized or reduced PSA (as defined by the PSA Working Group). In the PSA measurement in the last study, one patient maintained stable PSA. 6/15 had > 50% prolongation of PSADT compared to pre-study PSADT.

The primary endpoint of PSA stabilization or reduction in this population of individuals by use of the L-BLP25 vaccine was assessed as follows:

-8/16, the individual having PSA stability after a primary treatment period;

-1/16 individuals retain PSA stability at the end of the maintenance period; and

-elongation of PSADT in a subject of 14/15 by use of a vaccine; 6/16 individuals had > 50% PSADT prolongation.

Design of research：

Week-2: pretreatment evaluation (physical examination, PSA concentration measurement, pelvic CT, and bone scan).

Day-3: cyclophosphamide pretreatment.

Week 0 to 7: L-BLP25 was inoculated with #1 to #8 (primary treatment period).

Week 8: primary treatment period evaluation, including PSA response.

Week 13; confirmation of PSA reaction.

Weeks 13, 19, 25, 31, 37, 43 and 49: L-BLP25 inoculated with #9 to #15 (maintenance period).

Week 43: estimation of PSA reaction.

Week 49: confirmation of PSA reaction.

At week 50: maintenance treatment evaluation.

Population of individuals:

inclusion criteria

1. Radical prostatectomy was performed at least 6 months prior to study entry.

2.3 consecutive increases in serum PSA values after radical prostatectomy with at least a 50% increase above the nadir after prostatectomy.

3. There were no signs of malignant disease at the time of pretreatment evaluation as evidenced by negative pelvic CT and bone scans.

4.0, 1 ECOG performance status.

5. Normal hematology, liver and kidney function tests.

6. Understand and sign written informed consent.

7. All patients who had been treated for prostate cancer with hormone therapy (i.e., neoadjuvant treatment prior to radical prostatectomy) had a normal range of serum testosterone.

Exclusion criteria

1. Hormone therapy was received within 6 months prior to study enrollment.

2. Immunotherapy was received within 4 weeks prior to study enrollment.

3. Radiation therapy was performed on the prostate bed within 1 year prior to study enrollment.

4. Treatment with immunosuppressive drugs such as cyclosporin or adrenocorticotropic hormone (ACTH) or long-term treatment requiring the use of corticosteroids.

5. Known autoimmune or immunodeficiency diseases.

6. Clinically significant heart disease or active infection.

Table 3: patient characterization for prostate cancer study

Initial diagnosis to study group (years)

Mean value. + -. S.D. 3.8. + -. 2.5

Median value 3.2

In the range of 1.0 to 9.5

Lowest point after prostatectomy to study group (years)

Mean value. + -. S.D. 3.1. + -. 2.3

Median value of 2.8

In the range of 0.6 to 9.1

Baseline PSA μ g/L:

average value of 3.8

Median value 0.4

25％/75％ 0.1/0.8

Receive the treatment

Total number of treatments n (%)

Cyclophosphamide 16(100.0)

Primary treatment period vaccination

1 16(100.0)

2 16(100.0)

3 16(100.0)

4 16(100.0)

5 16(100.0)

6 16(100.0)

7 15(93.8)

8 15(93.8)

Vaccination during maintenance therapy

9 14(87.5)

10 14(87.5)

11 13(81.3)

12 12(75.0)

13 11(68.8)

14 10(62.5)

15 10(62.5)

Table 4: PSA value

Change in PSA from baseline to week 8

(Primary treatment period)

Each patient

Adverse event

Total number of patients with adverse events, n (%) 14(87.5)

Patients reporting adverse events with a frequency of 10% or more

Nausea 5(31.3)

Fatigue 4(25.0)

Diarrhea NOS (not otherwise specified) 3(18.8)

Influenza-like disease 3(18.8)

Nasopharyngitis 3(18.8)

Constipation 2(12.5)

Headache NOS 2(12.5)

Painful NOS 2(12.5)

Of the 16 patients 6 reported no injection site reactions of any kind. Erythema was reported in 9 patients. No ulcer occurred. No serious or event precluded further vaccination occurred.

Table 5: PSA doubling time

Patient ID	Time interval APSADT (sky)	Time interval BPSADT (sky)	Time interval A and timePSADT Difference between intervals B
				002	173	476	175％
003	133	291	119％
				004	345	393	14％
005	302	342	13％
				006	172	185	8％
007	309	347	12％
				008	173	332	92％
009	404	637	58％
				010	508	595	17％
011	165	257	56％
				012	241	97	-60％
013	84	112	33％
				014	479	844	76％
015	227	288	27％
				016	344	385	12％
Mean value of	271	372	44％

Time interval a before study (first to baseline from 3 consecutive increases in PSA before study group)

Time interval B is maintained (from week 8 to the end of the study)

It is to be understood that all ranges disclosed herein also encompass any and all possible subranges or combinations of subranges thereof. Thus, it can be readily appreciated that any listed range is sufficient to describe and break down the same range into at least the same halves, thirds, quarters, fifths, tenths, etc. As non-limiting examples, the ranges described herein may be readily broken down into a lower third (lower third), a middle third (middle third), and an upper third (upper third), among others. It is also to be understood that all terms such as "up to," "at least," "greater than," "less than," "exceeding," and the like, include the referenced number and refer to ranges that can be subsequently broken down into the above-described subranges. Likewise, all ratios disclosed herein also include all sub ratios (subratios) falling within the broader range of ratios.

Likewise, when members are grouped together in a common manner, such as in the Markush (Markush) group, the invention encompasses not only the entire group as a whole, but also the individual members of the group individually and all possible subgroups of the main group. Thus, for all purposes, the present invention includes not only the main group, but also the main group lacking one or more group members. The present invention also contemplates that one or more of any one of the group members is specifically excluded from the claimed invention.

All references, patents, and publications disclosed herein are expressly incorporated herein by reference. "A" or "an" mean "one or more" unless indicated otherwise.

It will be apparent to those skilled in the art that many modifications and variations can be made in the methods and compositions of the present invention without departing from the spirit or scope of the invention. Thus, it is intended that the present invention cover the modifications and variations of this invention provided they come within the scope of the appended claims and their equivalents.

Claims (16)

1. Use of a MUC-1 based formulation in the manufacture of a medicament for treating an individual having stage IIIB localized area non-small cell lung cancer without malignant pleural effusion, wherein the formulation comprises liposomes comprising at least one MUC-1 peptide having an amino acid sequence as set forth in the amino acid sequence of SEQ ID No.1 or the amino acid sequence of SEQ ID No. 2.

2. The use of claim 1, wherein the formulation further comprises at least one adjuvant.

3. The use of claim 2, wherein the adjuvant is selected from the group consisting of lipid a, muramyl dipeptide, alum and a cytokine.

4. The use of claim 3, wherein said lipid A is monophosphoryl lipid A or a mimic of synthetic lipid A.

5. The use of claim 3, wherein the cytokine is interleukin-2.

6. The use of any one of claims 1 to 5, wherein the formulation comprises a BLP25 liposome vaccine, wherein the BLP25 liposome vaccine comprises (i) a MUC-1 peptide having the amino acid sequence set forth in SEQ ID NO: 1 or 2, (ii) an adjuvant, and (iii) one or more additional liposomal lipids.

7. The use of claim 6, wherein the BLP25 liposome vaccine is provided in a kit.

Use of a BLP25 liposome vaccine in the manufacture of a medicament for improving or maintaining the quality of life of an individual diagnosed with localized area non-small cell lung cancer in phase IIIB without malignant pleural effusion, wherein the BLP25 liposome vaccine comprises (i) a MUC-1 peptide having an amino acid sequence as set forth in SEQ ID NO: 1 or SEQ ID NO: 2, (ii) an adjuvant, and (iii) one or more additional liposomal lipids.

9. The use of claim 2 or 8, wherein the dosage of MUC-1 is 1000 μ g and the dosage of adjuvant is 500 μ g.

10. The use of claim 1 or 8 wherein the amount of MUC-1 peptide is 300 μ g.

11. The use of any one of claims 2 or 8, wherein the adjuvant is lipid A.

12. The use of claim 11, wherein the amount of lipid a is 150 μ g.

13. Use according to claim 6 or 8, wherein the amount of additional liposomal lipid is 15 mg.

14. The use of claim 1 or 8, wherein the amino acid sequence of the MUC-1 peptide is as set forth in SEQ ID NO: 1 is shown in sequence.

15. The use of claim 1 or 8, wherein the amino acid sequence of the MUC-1 peptide is as set forth in SEQ ID NO: 2, as shown in the figure.

16. The use of claim 14, wherein the MUC-1 peptide is lipidated.