Methods for treating breast cancer
The present disclosure is directed to methods of treating and preventing breast cancer or recurrence of breast cancer with compositions comprising anti-progastrin antibodies.
1. A method for treating breast cancer, comprising the step of administering to a patient in need of treatment for breast cancer a therapeutically effective amount of a composition comprising a neutralizing anti-hPG monoclonal antibody that specifically binds to hPG and competes for binding to hPG with a reference antibody selected from:
(a) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:12 and a light chain variable domain sequence of SEQ ID NO:13;
(b) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:59 and a light chain variable domain sequence of SEQ ID NO:63;
(c) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:60 and a light chain variable domain sequence of SEQ ID NO:64;
(d) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:61 and a light chain variable domain sequence of SEQ ID NO:65; and
(e) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:62 and a light chain variable domain sequence of SEQ ID NO:66.
2. The method of claim 1 , wherein said breast cancer is a primary breast cancer.
3. The method of claim 1 , wherein said breast cancer is metastatic breast cancer.
4. The method of claim 1 , wherein the step of administering said composition is effected before surgical resection of a breast tumor.
5. The method of claim 1 , wherein the step of administering said composition is effected after surgical resection of a breast tumor.
6. The method of claim 1 , wherein said composition is administered adjunctive to chemotherapy or radiation therapy.
7. The method of claim 6 , wherein said chemotherapy includes treatment with a chemotherapeutic agent selected from the group consisting of folate antagonists, purine antagonists, pyrimidine antagonists, DNA alkylating agents, DNA cross-linking drugs, antibiotics, platinum complexes, proteosome inhibitors, mitotic spindle poisons, topoisomerase inhibitors, and tyrosine kinase inhibitors.
8. The method of claim 1 , wherein the composition is administered adjunctive to a hormone therapy agent.
9. The method of claim 8 , in which the hormone therapy agent is bicalutamide, flutamide, fulvestrant, leuprolide acetate, megestrol acetate, tamoxifen, raloxifene, anastrozole, exemestane or letrozole.
10. The method of claim 1 , wherein the composition is administered adjunctive to a second therapeutic antibody that specifically binds EGFR, VEGF, or HER2.
11. The method of claim 10 , in which the second therapeutic antibody is panitumumab, bevacizumab, cetuximab or trastuzumab.
12. The method of claim 1 , wherein said composition is administered by a mode of administration selected from among the group consisting of: parenteral administration, intrathecal administration, intravenous administration, subcutaneous administration, intramuscular administration, intraperitoneal administration, infusion administration, and bolus administration.
13. The method of claim 1 , wherein said composition is administered at a neutralizing anti-hPG monoclonal antibody dose ranging from about 0.001 mg/kg to about 250 mg/kg.
14. The method of claim 13 , wherein said neutralizing anti-hPG monoclonal antibody dose is administered over a plurality of temporally spaced administrations.
15. The method of claim 1 , wherein said reference antibody comprises a heavy variable domain sequence of SEQ ID NO:12 and a light chain variable domain sequence of SEQ ID NO:13.
16. The method of claim 15 , wherein said neutralizing anti-hPG monoclonal antibody inhibits binding of the reference antibody to hPG with a K i ranging from 10 pM to 100 nM; wherein said K i is determined using an assay comprising the following steps:
(a) coating the wells of a 96-well plate with a capture anti-hPG antibody using a concentration of capture anti-hPG antibody from 1 to 10 μg/mL overnight at 4° C., wherein said capture anti-hPG antibody binds an epitope different than the neutralizing anti-hPG antibody;
(b) blocking the coated wells with a blocking buffer;
(c) incubating the coated wells with hPG at a concentration between 10 pM and 1 nM for 2 hours at 22° C.;
(d) incubating the coated wells with biotinylated reference antibody and increasing concentrations of unlabeled neutralizing anti-hPG antibodies for 1 hour at 22° C.;
(e) removing unbound antibodies;
(f) detecting bound labeled biotinylated reference antibody using streptavidin-HRP and a fluorogenic substrate.
17. The method of claim 1 , wherein said reference antibody comprises a heavy variable domain sequence of SEQ ID NO:59 and a light chain variable domain sequence of SEQ ID NO:63.
18. The method of claim 17 , wherein said neutralizing anti-hPG monoclonal antibody inhibits binding of the reference antibody to hPG with a K i ranging from 10 pM to 100 nM; wherein said K i is determined using an assay comprising the following steps:
(a) coating the wells of a 96-well plate with a capture anti-hPG antibody using a concentration of capture anti-hPG antibody from 1 to 10 μg/mL overnight at 4° C., wherein said capture anti-hPG antibody binds an epitope different than the neutralizing anti-hPG antibody;
(b) blocking the coated wells with a blocking buffer;
(c) incubating the coated wells with hPG at a concentration between 10 pM and 1 nM for 2 hours at 22° C.;
(d) incubating the coated wells with biotinylated reference antibody and increasing concentrations of unlabeled neutralizing anti-hPG antibodies for 1 hour at 22° C.;
(e) removing unbound antibodies;
(f) detecting bound labeled biotinylated reference antibody using streptavidin-HRP and a fluorogenic substrate.
19. The method of claim 1 , wherein said reference antibody comprises a heavy variable domain sequence of SEQ ID NO:60 and a light chain variable domain sequence of SEQ ID NO:64.
20. The method of claim 19 , wherein said neutralizing anti-hPG monoclonal antibody inhibits binding of the reference antibody to hPG with a K i ranging from 10 pM to 100 nM; wherein said K i is determined using an assay comprising the following steps:
(a) coating the wells of a 96-well plate with a capture anti-hPG antibody using a concentration of capture anti-hPG antibody from 1 to 10 μg/mL overnight at 4° C., wherein said capture anti-hPG antibody binds an epitope different than the neutralizing anti-hPG antibody;
(b) blocking the coated wells with a blocking buffer;
(c) incubating the coated wells with hPG at a concentration between 10 pM and 1 nM for 2 hours at 22° C.;
(d) incubating the coated wells with biotinylated reference antibody and increasing concentrations of unlabeled neutralizing anti-hPG antibodies for 1 hour at 22° C.;
(e) removing unbound antibodies;
(f) detecting bound labeled biotinylated reference antibody using streptavidin-HRP and a fluorogenic substrate.
21. The method of claim 1 , wherein said reference antibody comprises a heavy variable domain sequence of SEQ ID NO:61 and a light chain variable domain sequence of SEQ ID NO:65.
22. The method of claim 21 , wherein said neutralizing anti-hPG monoclonal antibody inhibits binding of the reference antibody to hPG with a K i ranging from 10 pM to 100 nM; wherein said K i is determined using an assay comprising the following steps:
(a) coating the wells of a 96-well plate with a capture anti-hPG antibody using a concentration of capture anti-hPG antibody from 1 to 10 μg/mL overnight at 4° C., wherein said capture anti-hPG antibody binds an epitope different than the neutralizing anti-hPG antibody;
(b) blocking the coated wells with a blocking buffer;
(c) incubating the coated wells with hPG at a concentration between 10 pM and 1 nM for 2 hours at 22° C.;
(d) incubating the coated wells with biotinylated reference antibody and increasing concentrations of unlabeled neutralizing anti-hPG antibodies for 1 hour at 22° C.;
(e) removing unbound antibodies;
(f) detecting bound labeled biotinylated reference antibody using streptavidin-HRP and a fluorogenic substrate.
23. The method of claim 1 , wherein said reference antibody comprises a heavy variable domain sequence of SEQ ID NO:62 and a light chain variable domain sequence of SEQ ID NO:66.
24. The method of claim 23 , wherein said neutralizing anti-hPG monoclonal antibody inhibits binding of the reference antibody to hPG with a K i ranging from 10 pM to 100 nM; wherein said K i is determined using an assay comprising the following steps:
(a) coating the wells of a 96-well plate with a capture anti-hPG antibody using a concentration of capture anti-hPG antibody from 1 to 10 μg/mL overnight at 4° C., wherein said capture anti-hPG antibody binds an epitope different than the neutralizing anti-hPG antibody;
(b) blocking the coated wells with a blocking buffer;
(c) incubating the coated wells with hPG at a concentration between 10 pM and 1 nM for 2 hours at 22° C.;
(d) incubating the coated wells with biotinylated reference antibody and increasing concentrations of unlabeled neutralizing anti-hPG antibodies for 1 hour at 22° C.;
(e) removing unbound antibodies;
(f) detecting bound labeled biotinylated reference antibody using streptavidin-HRP and a fluorogenic substrate.
25. The method of claim 1 , wherein said neutralizing anti-hPG monoclonal antibody comprises a heavy chain variable region in which complementarity determining region (CDR) 1 comprises the amino acid sequence of V H CDR 1.3 (SEQ ID NO:1), CDR2 comprises the amino acid sequence of V H CDR 2.3 (SEQ ID NO:2), and CDR3 comprises the amino acid sequence of V H CDR 3.3 (SEQ ID NO:3), and a light chain variable region in which CDR1 comprises the amino acid sequence of V L CDR 1.3 (SEQ ID NO:4), CDR2 comprises the amino acid sequence of V L CDR 2.3 (SEQ ID NO:5), and CDR3 comprises the amino acid sequence of V L CDR 3.3 (SEQ ID NO:6).
26. The method of claim 1 , wherein said neutralizing anti-hPG monoclonal antibody comprises a heavy chain variable region in which CDR1 comprises the amino acid sequence of V H CDR 1.8 (SEQ ID NO:37), CDR2 comprises the amino acid sequence of V H CDR 2.8 (SEQ ID NO:41), and CDR3 comprises the amino acid sequence of V H CDR 3.8 (SEQ ID NO:45), and a light chain variable region in which CDR1 comprises the amino acid sequence of V L CDR 1.8 (SEQ ID NO:49), CDR2 comprises the amino acid sequence of V L CDR 2.8 (SEQ ID NO:52), and CDR3 comprises the amino acid sequence of V L CDR 3.8 (SEQ ID NO:55).
27. The method of claim 1 , wherein said neutralizing anti-hPG monoclonal antibody comprises a heavy chain variable region in which CDR1 comprises the amino acid sequence of V H CDR 1.13 (SEQ ID NO:38), CDR2 comprises the amino acid sequence of V H CDR 2.13 (SEQ ID NO:42), and CDR3 comprises the amino acid sequence of V H CDR 3.13 (SEQ ID NO:46), and a light chain variable region in which CDR1 comprises the amino acid sequence of V L CDR 1.13 (SEQ ID NO:50), CDR2 comprises the amino acid sequence of V L CDR 2.13 (SEQ ID NO:53), and CDR3 comprises the amino acid sequence of V L CDR 3.13 (SEQ ID NO:56).
28. The method of claim 1 , wherein said neutralizing anti-hPG monoclonal antibody comprises a heavy chain variable region in which CDR1 comprises the amino acid sequence of V H CDR 1.16 (SEQ ID NO:39), CDR2 comprises the amino acid sequence of V H CDR 2.16 (SEQ ID NO:43), and CDR3 comprises the amino acid sequence of V H CDR 3.16 (SEQ ID NO:47), and a light chain variable region in which CDR1 comprises the amino acid sequence of V L CDR 1.16 (SEQ ID NO:50), CDR2 comprises the amino acid sequence of V L CDR 2.16 (SEQ ID NO:53), and CDR3 comprises the amino acid sequence of V L CDR 3.16 (SEQ ID NO:57).
29. The method of claim 1 , wherein said neutralizing anti-hPG monoclonal antibody comprises a heavy chain variable region in which CDR1 comprises the amino acid sequence of V H CDR 1.19 (SEQ ID NO:40), CDR2 comprises the amino acid sequence of V H CDR 2.19 (SEQ ID NO:44), and CDR3 comprises the amino acid sequence of V H CDR 3.19 (SEQ ID NO:48), and a light chain variable region in which CDR1 comprises the amino acid sequence of V L CDR 1.19 (SEQ ID NO:51), CDR2 comprises the amino acid sequence of V L CDR 2.19 (SEQ ID NO:54), and CDR3 comprises the amino acid sequence of V L CDR 3.19 (SEQ ID NO:58).
30. The method of any one of claims 1 and 15 - 29 , wherein said neutralizing anti-hPG monoclonal antibody is humanized.
31. The method of any one of claims 1 and 15 - 24 , wherein said neutralizing anti-hPG monoclonal antibody has an hPG binding affinity in a range of about 0.001 nM to about 5000 nM.
32. A method for preventing recurrence of breast cancer, comprising the step of administering to a patient in need of prevention of recurrence of breast cancer a composition comprising a neutralizing anti-hPG monoclonal antibody that specifically binds to hPG in an amount effective to prevent recurrence of breast cancer, wherein said breast cancer is hPG sensitive, and wherein said neutralizing anti-hPG monoclonal antibody competes for binding to hPG with a reference antibody selected from:
(a) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:12 and a light chain variable domain sequence of SEQ ID NO:13;
(b) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:59 and a light chain variable domain sequence of SEQ ID NO:63;
(c) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:60 and a light chain variable domain sequence of SEQ ID NO:64;
(d) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:61 and a light chain variable domain sequence of SEQ ID NO:65; and
(e) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:62 and a light chain variable domain sequence of SEQ ID NO:66.
33. A method of inhibiting proliferation of a breast cancer stem cell, comprising exposing a breast cancer stem cell with a CD44(+)/CD24(−), CD44(+)/CD24(low) or CD44(+)/CD24(−)/ESA(+) marker phenotype to an amount of a neutralizing anti-hPG monoclonal antibody effective to inhibit its proliferation, wherein said neutralizing anti-hPG monoclonal antibody specifically binds to hPG and competes for binding to hPG with a reference antibody selected from:
(a) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:12 and a light chain variable domain sequence of SEQ ID NO:13;
(b) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:59 and a light chain variable domain sequence of SEQ ID NO:63;
(c) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:60 and a light chain variable domain sequence of SEQ ID NO:64;
(d) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:61 and a light chain variable domain sequence of SEQ ID NO:65; and
(e) a monoclonal antibody comprising a heavy variable domain sequence of SEQ ID NO:62 and a light chain variable domain sequence of SEQ ID NO:66.