CHEMOTHERAPEUTIC

CROSS REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Application No.

62/308,639 filed March 15, 2016, U.S. Provisional Application No. 62/317,792 filed April 4, 2016, and U.S. Provisional Application No. 62/367,510 filed July 27, 2016, the disclosures of which are incorporated by reference herein in their entirety for all purposes.

REFERENCE TO A SEQUENCE LISTING

[0002] A sequence listing designated 0710-00313PC Sequence Listing ST25.txt of 16 KB created March 1, 2017, is incorporated herein by reference.

FIELD OF THE INVENTION

[0003] This invention relates to methods for the treatment of cancer comprising administering a LIVl-ADC and a chemotherapeutic.

BACKGROUND

[0004] LIV-1 is a member of the LZT (LIV-1 -ZIP Zinc Transporters) subfamily of zinc transporter proteins. Taylor et al., Biochim. Biophys. Acta 1611: 16-30 (2003). Computer analysis of the LIV- 1 protein reveals a potential metalloprotease motif, fitting the consensus sequence for the catalytic zinc -binding site motif of the zinc metalloprotease. LIV-1 mRNA is primarily expressed in breast, prostate, pituitary gland and brain tissue.

[0005] The LIV-1 protein has also been implicated in certain cancerous conditions, e.g. breast cancer and prostate cancer. The detection of LIV-1 is associated with estrogen receptor-positive breast cancer, McClelland et al., Br. J. Cancer 77: 1653-1656 (1998), and the metastatic spread of these cancers to the regional lymph nodes. Manninget al., Eur. J. Cancer 30A:675-678 (1994). [0006] SGN-LIV1A is a LIV-1 -directed antibody-drug conjugate (ADC) consisting of three components: 1) the humanized antibody hLIV22, specific for human LIV-1, 2) the microtubule disrupting agent monomethyl auristatin E (MMAE), and 3) a stable linker, valine-citrulline (vc), that covalently attaches MMAE to hLIV22. The proposed mechanism of action (MO A) is initated by SGN-LIV1A binding to LIV-1 on the cell surface followed by internalization of the ADC. Upon trafficking to lysosomes, the delivered drug (MMAE) is released through proteolytic degradation of the vc linker. Binding of the released drug to tubulin disrupts the microtubule network, leading to cell cycle arrest and apoptosis.

[0007] SGN-LIV1A has been shown to reduce tumor volumes in vivo, and is currently being evaluated in a phase 1 clinical trial for patients with LIV-1 -positive metastatic breast cancer. However, improvements are needed in cancer therapy. The present invention solves this and other problems.

SUMMARY OF THE CLAIMED INVENTION

[0008] The invention provides a method for treating a subject having or at risk of cancer. The method includes administering to the subject a LIV-1 antibody drug conjugate (LIV- l-ADC) and a chemo therapeutic. The LIV-l-ADC includes a humanized hLIV22 antibody conjugated to a vcMMAE (valine-citrulline-monomethyl auristatin E), and the chemotherapeutic is one of carboplatin, doxorubicin, paclitaxel, trastuzumab, and an mTOR inhibitor. Optionally, the subject has prostate cancer, ovarian cancer,

endrometrial cancer, pancreatic cancer, lung cancer, a cervical cancer, a melanoma, squamous cell carcinoma, or a breast cancer, such as triple negative breast cancer, triple positive breast cancer, HER2-positive breast cancer, or hormone receptor positive breast cancer.

[0009] In some embodiments, the LIV-l-ADC is administered at a dosage between 1.5 mg/kg and 4 mg/kg of the subject's body weight. In an embodiment, the LIV-l-ADC is administered at a dosage of 2.5 mg/kg of the subject's body weight. In an embodiment, the LIV-l-ADC is administered once every 3 weeks. In an embodiment, the LIV-l-ADC is administered by intravenous injection. [0010] In an embodiment, the chemotherapeutic is carboplatin, and is administered at a dosage between 200 mg/m 2 and 750 mg/m 2. In an embodiment, the carboplatin is administered by intravenous injection. In another embodiment, the chemotherapeutic is doxorubicin, and is administered at a dosage between 40 mg/m 2 and 80 mg/m 2. In an embodiment, the doxorubicin is administered by intravenous injection. In another embodiment, the chemotherapeutic is paclitaxel, and is administered at a dosage between

100 mg/m 2 to 260 mg/m 2. In an embodiment, the paclitaxel is administered by intravenous injection.

[0011] The invention also provides methods for treating a subject having or at risk of cancer, the method comprising administering to the subject a LIV-1 antibody drug conjugate (LIV-l-ADC) and either trastuzumab emtansine or pertuzumab. The LIV-l- ADC includes a humanized hLIV22 antibody conjugated to a vcMMAE (valine- citrulline-monomethyl auristatin E).

BRIEF DESCRIPTION OF THE FIGURES

[0012] Figure 1 shows the viability of multiple agents on MCF-7 cells, according to an embodiment of the invention.

[0013] Figure 2 shows the effect of SGN-LIV1A and doxorubicin on MCF-7 cells, according to an embodiment of the invention.

[0014] Figure 3 shows the effect of SGN-LIV1A and paclitaxel on MCF-7 cells, according to an embodiment of the invention.

[0015] Figure 4 shows the effect of SGN-LIV1A and gemcitabine on MCF-7 cells, according to an embodiment of the invention.

[0016] Figure 5 shows the effect of SGN-LIV1A and everolimus on MCF-7 cells, according to an embodiment of the invention.

[0017] Figure 6 shows the effects of SGN-LIV1A and either doxorubicin (left) or Abraxane (nab-paclitaxel) (right) on tumor growth in NOD Scid Gamma (NSG) mice, according to an embodiment of the invention. [0018] Figure 7 shows the effects of SGN-LIV1A and either carboplatin (left) or cyclophosphamide (right) on tumor growth in NOD Scid Gamma (NSG) mice, according to an embodiment of the invention.

[0019] Figure 8 shows the effects of SGN-LIV1A and either gemcitabine (left) or paclitaxel (right) on tumor growth in NOD Scid Gamma (NSG) mice, according to an embodiment of the invention.

DEFINITIONS

[0020] The term "antibody" includes intact antibodies and binding fragments thereof. Typically, antibody fragments compete with the intact antibody from which they were derived for specific binding to the target including separate heavy chains, light chains Fab, Fab', F(ab') ₂ , F(ab)c, diabodies, Dabs, nanobodies, and Fv. Fragments can be produced by recombinant DNA techniques, or by enzymatic or chemical separation of intact immunoglobulins. The term "antibody" also includes a diabody (homodimeric Fv fragment) or a minibody (VL-VH-CH3), a bispecific antibody or the like. A bispecific or bifunctional antibody is an artificial hybrid antibody having two different heavy/light chain pairs and two different binding sites (see, e.g., Songsivilai and Lachmann, Clin. Exp. Immunol., 79:315-321 (1990); Kostelny et al., J. Immunol., 148: 1547-53 (1992)). The term "antibody" includes an antibody by itself (naked antibody) or an antibody conjugated to a cytotoxic or cytostatic drug.

[0021] A "cytotoxic effect" refers to the depletion, elimination and/or the killing of a target cell. A "cytotoxic agent" refers to an agent that has a cytotoxic effect on a cell. Cytotoxic agents can be conjugated to an antibody or administered in combination with an antibody.

[0022] A "cytostatic effect" refers to the inhibition of cell proliferation. A "cytostatic agent" refers to an agent that has a cytostatic effect on a cell, thereby inhibiting the growth and/or expansion of a specific subset of cells. Cytostatic agents can be conjugated to an antibody or administered in combination with an antibody. [0023] The term "pharmaceutically acceptable" as used herein refers to those compounds, materials, compositions, and/or dosage forms that are, within the scope of sound medical judgment, suitable for contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problems or complications commensurate with a reasonable benefit/risk ratio. The term

"pharmaceutically compatible ingredient" refers to a pharmaceutically acceptable diluent, adjuvant, excipient, or vehicle with which an anti-LIV-1 antibody is administered.

[0024] The phrase "pharmaceutically acceptable salt," refers to pharmaceutically acceptable organic or inorganic salts of an anti-LIV-1 antibody or conjugate thereof or agent administered with an anti-LIV-1 antibody. Exemplary salts include sulfate, citrate, acetate, oxalate, chloride, bromide, iodide, nitrate, bisulfate, phosphate, acid phosphate, isonicotinate, lactate, salicylate, acid citrate, tartrate, oleate, tannate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisinate, fumarate, gluconate, glucuronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate,

benzenesulfonate, p toluenesulfonate, and pamoate (i.e., 1,1' methylene bis -(2 hydroxy 3 naphthoate)) salts. A pharmaceutically acceptable salt may involve the inclusion of another molecule such as an acetate ion, a succinate ion or other counterion. The counterion may be any organic or inorganic moiety that stabilizes the charge on the parent compound. Furthermore, a pharmaceutically acceptable salt may have more than one charged atom in its structure. Multiple counter ions may occur in instances where multiple charged atoms are part of the pharmaceutically acceptable salt. Hence, a pharmaceutically acceptable salt can have one or more charged atoms and/or one or more counterion.

[0025] The term "effective amount" refers to the amount of a LIV-1-ADC, e.g., SGN- LIV1A, that is sufficient to inhibit the occurrence or ameliorate one or more clinical or diagnostic symptoms of a LIV-1 -associated disorder in a subject. An effective amount of an agent is administered according to the methods described herein in an "effective regimen." The term "effective regimen" refers to a combination of amount of the agent and dosage frequency adequate to maintain high LIV-1 occupancy, which may accomplish treatment or prevention of a LIV-1 -associated disorder. In a preferred embodiment, an effective regimen maintains near complete, e.g., greater than 90%, LIV- 1 occupancy on LIV-1 -expressing cells during dosing intervals.

[0026] The terms "treatment" and "therapy," and the like, as used herein, are meant to include therapeutic or suppressive measures for a disease or disorder leading to any clinically desirable or beneficial effect, including, but not limited to, alleviation or relief of one or more symptoms, regression, slowing or cessation of progression of the disease or disorder. For example, treatment can include a decrease or elimination of a clinical or diagnostic symptom of a LIV-1 -expressing disorder after the onset of the clinical or diagnostic symptom by administration of an anti- LIV-1 antibody or other LIV-1 binding agent to a subject. Treatment can be evidenced as a decrease in the severity of a symptom, the number of symptoms, or frequency of relapse.

[0027] Except when noted, the terms "subject" or "patient" are used interchangeably and refer to mammals such as human patients and non-human primates, as well as experimental animals such as rabbits, dogs, cats, rats, mice, and other animals.

Accordingly, the term "subject" or "patient" as used herein means any mammalian patient or subject to which the LIV-1 binding agents of the invention can be administered. In preferred embodiments, the terms subject or patient are used to refer to human patients. Subjects of the present invention include those that have been diagnosed with a LIV-1 expressing cancer, including, for example, breast cancer, prostate cancer, ovarian cancer, endrometrial cancer, pancreatic cancer, lung cancer, cervical cancer, a melanoma, or squamous cell carcinoma. In certain embodiments, the subject will have a refractory or relapsed LIV-1 expressing cancer.

[0028] Compositions or methods "comprising" one or more recited elements may include other elements not specifically recited. For example, a composition that comprises antibody may contain the antibody alone or in combination with other ingredients.

[0029] Designation of a range of values includes all integers within or defining the range.

[0030] As used herein, the term "about" denotes an approximate range of plus or minus 10% from a specified value. For instance, the language "about 20%" encompasses a range of 18-22%. As used herein, about also includes the exact amount. Hence, "about 20%" means "about 20%" and also "20%.

DETAILED DESCRIPTION

I. General

[0031] The invention provides methods for treating cancer, in particular breast cancer. The present inventors have discovered that combination therapy with two different classes of anticancer compounds: antibody-drug conjugate compounds and

chemotherapeutic agents, can improve a therapeutic benefit for subjects suffering from cancer. In particular, the present inventors have found that combination therapy with (1) an anti-LIV-1 antibody conjugated to an auristatin compound and (2) a chemotherapeutic agent provides synergistic therapeutic effects in the treatment of cancer.

II. LIV-1-ADC

[0032] Unless otherwise indicated, a LIV-1 -antibody drug conjugate (LIV-1-ADC) includes an antibody specific for the human LIV-1 protein conjugated to a cytotoxic agent. An exemplary human LIV-1 sequence is assigned Swiss Prot accession number Q13433. Q13433 is included herein as SEQ ID NO: l. Three variant isoforms and one polymorphism are known. A second version of the human LIV-1 protein, accession number AAA96258.2, is included herein as SEQ ID NO:2. Four extracellular domains are bounded by residues 29-325, 377-423, 679-686 and 746-755 of Q13433 respectively.

[0033] SGN-LIV1 A is a LIV- 1 -directed antibody-drug conjugate (ADC) produced by the conjugation of the drug linker vcMMAE (monomethyl auristatin E with a valine- citrulline linker) to the humanized antibody hLIV22. hLIV22 is a humanized form of the mouse BR2-22a antibody, described in US Patent No. 9,228,026. The hLIV22 antibody is essentially the same as BR2-22a within experimental error and contains seven back mutations. Methods of making the hLIV22 antibody are also disclosed in US Patent No. 9,228,026. The amino acid sequence of the light chain variable region of hLIV22 is provided herein as SEQ ID NO: 3. The amino acid sequence of the heavy chain variable region of hLIV22 is provided herein as SEQ ID NO: 4. Synthesis and conjugation of the drug linker vcMMAE (shown below; also referred to as 1006) are further described in US Patent No. 9,228,026 and US Patent Pub. No. 20050238649.

vcMMAE

III. Combination therapy of chemotherapeutic agents and LIV-1-ADC

[0034] Cancer can be treated using a combination of SGN-LIV1A and a

chemotherapeutic agent. The chemotherapeutic agent is either carboplatin, doxorubicin or paclitaxel, trastuzumab, a checkpoint inhibitor, or an mTOR inhibitor (such as

Everolimus). Carboplatin (PARAPLATIN®; Bristol Myers Squibb, New York, NY) is an alkylating agent. Doxorubicin (ADRIAMYCIN®, RUB EX®, DOXIL®, MYOCEL®, or

CAELYX®) is an anthracycline antibiotic with antineoplastic activity. Paclitaxel

(ABRAXANE®; Celgene, Summit, NJ) is a taxane that inhibits microtubule breakdown.

Trastuzumab (HERCEPTIN®; Genentech, South San Francisco, CA) is a monoclonal antibody that binds the HER2 receptor. Examples of checkpoint inhibitors (inhibitors that block immune checkpoints) include antibodies such as anti-PD-1 antibodies (e.g.,

MEDI0680, AMP-224, nivolumab, pembrolizumab, and pidilizumab), anti-PD-Ll antibodies (e.g., MEDI4736 and MPDL3280A), and anti-CTLA4 antibodies (e.g., ipilimumab and tremelimumab). Other checkpoint inhibitors/activators include B7-DC-

Fc, LAG3 and TIM3.

[0035] The combination of SGN-LIV1A and a chemotherapeutic agent can be given to subjects at levels that inhibit cancer cell growth, while at the same time are tolerated by the subject. In an embodiment, administration of the combination decreases the toxic effects caused by administration of a chemotherapeutic agent alone. In some

embodiments, the combination of SGN-LIV1A and a chemotherapeutic agent is synergistic or additive. For some combinations, each agent in the combination can be effectively administered at lower levels than when administered alone.

[0036] As discussed above, the combination therapies of the invention can be used to treat cancer. Some such cancers show detectable levels of LIV-1 measured at either the protein (e.g., by immunoassay using one of the exemplified antibodies) or mRNA level. Some such cancers show elevated levels of LIV-1 relative to noncancerous tissue of the same type, preferably from the same patient. An exemplary level of LIV-1 on cancer cells amenable to treatment is 5000-150000 LIV-1 molecules per cell, although higher or lower levels can be treated. Optionally, a level of LIV-1 in a cancer is measured before performing treatment.

[0037] Examples of cancers associated with LIV-1 expression and amenable to treatment with the combination therapies of the invention include breast cancer, prostate cancer, ovarian cancer, endometrial cancer, pancreatic cancer, cervical, liver, gastric, kidney, and squamous cell carcinomas (e.g., bladder, head, neck and lung), skin cancers, e.g., melanoma, small lung cell carcinoma or lung carcinoid. The treatment can be applied to patients having primary or metastatic tumors of these kinds. The treatment can also be applied to patients who are refractory to conventional treatments (e.g., for breast cancer: hormones, tamoxifen, HERCEPTIN®), or who have relapsed following a response to such treatments. The methods can also be used on triple negative breast cancers. A triple negative breast cancer is a term of art for a cancer lacking detectable estrogen and progesterone receptors and lacking overexpression of HER2/neu when stained with an antibody to any of these receptors, such as described in the examples. The methods can also be used on triple positive breast cancers, hormone receptor positive breast cancers, and HER2 positive breast cancers. Staining can be performed relative to an irrelevant control antibody and lack of expression shown from a background level of straining the same or similar to that of the control within experimental error. Likewise, lack of overexpression is shown by staining at the same or similar level within experimental error of noncancerous breast tissue, preferably obtained from the same patient. Alternatively or additionally, triple negative breast cancers are characterized by lack of responsiveness to hormones interacting with these receptors, aggressive behavior and a distinct pattern of metastasis.

[0038] In some embodiments, SGN-LIV1A and a chemotherapeutic agent are administered in such a way that the combination provides a synergistic or additive effect in the treatment of LIV-1 -associated cancer in a patient. Administration can be by any suitable means provided that the administration provides the desired therapeutic effect. In preferred embodiments, SGN-LIVIA and a chemotherapeutic agent are administered during the same cycle of therapy, e.g., during one cycle of therapy, e.g., a three or four week time period.

[0039] SGN-LIVIA and a chemotherapeutic agent are administered in an effective regime meaning a dosage, route of administration and frequency of administration that delays the onset, reduces the severity, inhibits further deterioration, and/or ameliorates at least one sign or symptom of cancer. If a patient is already suffering from cancer, the regime can be referred to as a therapeutically effective regime. If the patient is at elevated risk of the cancer relative to the general population but is not yet experiencing symptoms, the regime can be referred to as a prophylactically effective regime. In some instances, therapeutic or prophylactic efficacy can be observed in an individual patient relative to historical controls or past experience in the same patient. In other instances, therapeutic or prophylactic efficacy can be demonstrated in a preclinical or clinical trial in a population of treated patients relative to a control population of untreated patients.

[0040] Exemplary dosages for SGN-LIVIA are 0.1 mg/kg to 50 mg/kg of the patient's body weight, more typically 1 mg/kg to 30 mg/kg, 1 mg/kg to 20 mg/kg, 1 mg/kg to 15 mg/kg, 1 mg/kg to 12 mg/kg, 1 mg/kg to 10 mg/kgl, 2 mg/kg to 30 mg/kg, 2 mg/kg to 20 mg/kg, 2 mg/kg to 15 mg/kg, 2 mg/kg to 12 mg/kg, 2 mg/kg to 10 mg/kg, 3 mg/kg to 30 mg/kg, 3 mg/kg to 20 mg/kg, 3 mg/kg to 15 mg/kg, 3 mg/kg to 12 mg/kg, or 3 mg/kg to 10 mg/kg . In some methods, the patient is administered a dose of at least 1.5 mg/kg, at least 2 mg/kg or at least 3 mg/kg, administered once every three weeks or greater. In an embodiment, the patient is administered a dose of 2.5 mg/kg. In a further embodiment, the patient is administered a dose of 2.5 mg/kg, administered once every three weeks. The dosage depends on the frequency of administration, condition of the patient and response to prior treatment, if any, whether the treatment is prophylactic or therapeutic and whether the disorder is acute or chronic, among other factors.

[0041] In combination with carboplatin, SGN-LIVIA is administered at a dose between 0.5 mg/kg and 6 mg/kg. Other appropriate dose ranges of SGN-LIVIA in the

combination are 1 mg/kg to 5 mg/kg, and 2 mg/kg to 3 mg/kg. In an embodiment, SGN- LIVIA is administered at a dose of 2.5 mg/kg in combination with carboplatin. In combination with SGN-LIVIA, carboplatin is administered at a dose between 100 mg/m 2

and 950 mg/m . Other appropriate dose ranges of carboplatin in the combination are 200 mg/m 2 to 750 mg/m 2 , and 300 mg/m 2 to 600 mg/m 2. In an embodiment, carboplatin is administered at a dose of 300 mg/m in combination with SGN-LIVIA. In another embodiment, carboplatin is administered at a dose of AUC 6 IV in combination with SGN-LIVIA.

[0042] In combination with doxorubicin, SGN-LIVIA is administered at a dose between 0.5 mg/kg and 6 mg/kg. Other appropriate dose ranges of SGN-LIVIA in the

combination are 1 mg/kg to 5 mg/kg, and 2 mg/kg to 3 mg/kg. In an embodiment, SGN- LIVIA is administered at a dose of 2.5 mg/kg in combination with doxorubicin. In combination with SGN-LIVIA, doxorubicin is administered at a dose between 30 mg/m and 90 mg/m . Other appropriate dose ranges of doxorubicin in the combination are 40 mg/m 2 to 80 mg/m 2 , and 60 mg/m 2 to 75 mg/m 2. In an embodiment, doxorubicin is administered at a dose of 60 mg/m in combination with SGN-LIVIA.

[0043] In combination with paclitaxel, SGN-LIVIA is administered at a dose between 0.5 mg/kg and 6 mg/kg. Other appropriate dose ranges of SGN-LIVIA in the

combination are 1 mg/kg to 5 mg/kg, and 2 mg/kg to 3 mg/kg. In an embodiment, SGN- LIVIA is administered at a dose of 2.5 mg/kg in combination with paclitaxel. In combination with SGN-LIVIA, paclitaxel is administered at a dose between 50 mg/m and 300 mg/m . Other appropriate dose ranges of paclitaxel in the combination are 100 mg/m 2 to 260 mg/m 2 , and 135 mg/m 2 to 175 mg/m 2. In an embodiment, paclitaxel is administered at a dose of 175 mg/m in combination with SGN-LIVIA. In another embodiment, paclitaxel is administered at a dose of 80 mg/m in combination with SGN- LIVIA.

[0044] In combination with trastuzumab, SGN-LIVIA is administered at a dose between 0.5 mg/kg and 2.8 mg/kg. In an embodiment, SGN-LIVIA is administered at a dose between 1 mg/kg and 2.8 mg/kg in combination with trastuzumab. In another embodiment, SGN-LIVIA is administered at a dose of 2.5 mg/kg in combination with trastuzumab. In an embodiment, SGN-LIVIA is administered once a week. In another embodiment, SGN-LIVIA is administered once every three weeks. In an embodiment, in combination with SGN-LIVIA, trastuzumab is administered at an initial dose of 8 mg/kg over a 90 minute IV infusion, and then 6 mg/kg over 30-90 minutes of an IV infusion. In a further embodiment, the 6 mg/kg over 30-90 minutes of an IV infusion is administered every 3 weeks. In a further embodiment, the 6 mg/kg over 30-90 minutes of an IV infusion is administered every 3 weeks for 52 weeks. In another embodiment, in combination with SGN-LIVIA, trastuzumab is administered at an initial dose 4 mg/kg over a 90 minute IV infusion, and then 2 mg/kg over a 30 minute IV infusion. In a further embodiment, the 2 mg/kg over a 30 minute IV infusion is administered once a week. In a further embodiment, the 2 mg/kg over a 30 minute IV infusion is administered once a week for 52 weeks.

[0045] In combination with a checkpoint inhibitor, SGN-LIVIA is administered at a dose between 0.5 mg/kg and 2.8 mg/kg. . In an embodiment, SGN-LIVIA is administered at a dose between 1 mg/kg and 2.8 mg/kg in combination with trastuzumab. In another embodiment, SGN-LIVIA is administered at a dose of 2.5 mg/kg in combination with trastuzumab. In an embodiment, SGN-LIVIA is administered once a week. In another embodiment, SGN-LIVIA is administered once every three weeks.

[0046] Administration of SGN-LIVIA, carboplatin, doxorubicin, paclitaxel, trastuzumab, or an mTOR inhibitor can be parenteral, intravenous, oral, subcutaneous, intra-arterial, intracranial, intrathecal, intraperitoneal, topical, intranasal or intramuscular. In an embodiment, SGN-LIVIA is administered by intraperitoneal injection. In another embodiment, SGN-LIVIA is administered by intravenous injection. In another embodiment, carboplatin is administered by intravenous injection. In another

embodiment, paclitaxel is administered by intravenous injection. In another embodiment, doxorubicin is administered by intravenous injection. Administration can also be localized directly into a tumor. Administration into the systemic circulation by intravenous or subcutaneous administration is preferred. Intravenous administration can be, for example, by infusion over a period such as 30-90 min or by a single bolus injection.

[0047] The frequency of administration of each agent of the combination (SGN- LIVIA, carboplatin, doxorubicin, paclitaxel, trastuzumab, or an mTOR inhibitor) depends on its half-life in the circulation, the condition of the patient and the route of administration among other factors. The frequency can be daily, weekly, monthly, quarterly, or at irregular intervals in response to changes in the patient's condition or progression of the cancer being treated. An exemplary frequency for intravenous administration is between twice a week and quarterly over a continuous course of treatment, although more or less frequent dosing is also possible. Other exemplary frequencies for intravenous administration are between weekly or three out of every four weeks over a continuous course of treatment, although more or less frequent dosing is also possible. In an embodiment, one or both agents of the combination is administered once every three weeks. In another embodiment, one or both agents of the combination is administered once every four weeks. For subcutaneous administration, an exemplary dosing frequency is daily to monthly, although more or less frequent dosing is also possible.

[0048] The number of dosages of SGN-LIV1A administered depends on the nature of the cancer (e.g., whether presenting acute or chronic symptoms) and the response of the disorder to the treatment. For acute disorders or acute exacerbations of a chronic disorder between 1 and 10 doses are often sufficient. Sometimes a single bolus dose, optionally in divided form, is sufficient for an acute disorder or acute exacerbation of a chronic disorder. Treatment can be repeated for recurrence of an acute disorder or acute exacerbation. For chronic disorders, an antibody can be administered at regular intervals, e.g., weekly, fortnightly, monthly, quarterly, every six months for at least 1, 5 or 10 years, or the life of the patient.

[0049] Pharmaceutical compositions for parenteral administration of SGN-LIV1A are preferably sterile and substantially isotonic and manufactured under GMP conditions. Pharmaceutical compositions can be provided in unit dosage form (i.e., the dosage for a single administration). Pharmaceutical compositions can be formulated using one or more physiologically acceptable carriers, diluents, excipients or auxiliaries. The formulation depends on the route of administration chosen. For injection, SGN-LIV1A can be formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hank's solution, Ringer's solution, or physiological saline or acetate buffer (to reduce discomfort at the site of injection). The solution can contain formulatory agents such as suspending, stabilizing and/or dispersing agents. Alternatively SGN-LIV1A can be in lyophilized form for constitution with a suitable vehicle, e.g., sterile pyrogen-free water, before use. The concentration of SGN-LIV1A in a liquid formulation can be e.g., 1-100 mg/ml, such as 10 mg/ml.

[0050] Treatment with the combination therapies of the invention can be further combined with additional chemotherapy, radiation, stem cell treatment, surgery other treatments effective against the disorder being treated. Useful classes of other agents that can be administered with the combination therapies of the invention include, for example, antibodies to other receptors expressed on cancerous cells, including other antibodies to the HER2 receptor (e.g., rastuzumab emtansine (KADCYLA®, Genentech, South San Francisco, CA), antitubulin agents (e.g., auristatins), pertuzumab (PERJETA®,

Genentech, South San Francisco, CA)), or other antibody drug conjugates such as sacituzumab govitecan, checkpoint inhibitors (e.g., anti-PD-1, anti-PD-Ll), DNA minor groove binders, DNA replication inhibitors, alkylating agents (e.g., platinum complexes such as czs-platin, mono(platinum), bis(platinum) and tri-nuclear platinum complexes and carboplatin), anthracyclines, antibiotics, antifolates, antimetabolites, chemotherapy sensitizers, duocarmycins, etoposides, fluorinated pyrimidines, ionophores, lexitropsins, nitrosoureas, platinols, pre-forming compounds, purine antimetabolites, puromycins, radiation sensitizers, steroids, taxanes, topoisomerase inhibitors, vinca alkaloids, and the like.

[0051] Treatment with the combination therapies of the invention can increase the median progression-free survival or overall survival time of patients with tumors (e.g., breast, prostate, melanoma), especially when relapsed or refractory, by at least 30% or 40% but preferably 50%, 60% to 70% or even 100% or longer, compared to the same treatment (e.g., chemotherapy) but without the combination therapies of the invention. In addition or alternatively, treatment (e.g., standard chemotherapy) including the combination therapies of the invention can increase the complete response rate, partial response rate, or objective response rate (complete + partial) of patients with tumors by at least 30% or 40% but preferably 50%, 60% to 70% or even 100% compared to the same treatment (e.g., chemotherapy) but without the combination therapies of the invention.

[0052] All patent filings, website, other publications, accession numbers and the like cited above or below are incorporated by reference in their entirety for all purposes to the same extent as if each individual item were specifically and individually indicated to be so incorporated by reference. If different versions of a sequence are associated with an accession number at different times, the version associated with the accession number at the effective filing date of this application is meant. The effective filing date means the earlier of the actual filing date or filing date of a priority application referring to the accession number if applicable. Likewise if different versions of a publication, website or the like are published at different times, the version most recently published at the effective filing date of the application is meant unless otherwise indicated. Any feature, step, element, embodiment, or aspect of the invention can be used in combination with any other unless specifically indicated otherwise. Although the present invention has been described in some detail by way of illustration and example for purposes of clarity and understanding, it will be apparent that certain changes and modifications may be practiced within the scope of the appended claims.

EXAMPLES

The following examples are offered to illustrate, but not to limit the claimed invention.

I. In vitro cytotoxicity assay and combination assays Materials

[0053] Docetaxel, paclitaxel, doxorubicin, gemcitabine, and NVP-BEZ235 were purchased from LC Laboratories (Woburn, MA) and reconstituted in DMSO for in vitro assays. Cyclophosphamide was purchased from Selleckchem (Houston, TX). SGN- LIV1A was conjugated with average of 4 MMAE per antibody. US Patent No. 9,228,026 discloses further methods for the conjugation of vcMMAE to hLIV22.

Assays

[0054] MCF-7 (breast carcinoma) cells were obtained from ATCC. One day prior to assay, the MCF-7 cells were plated at a density of 2000 cells per well in 96-well plates, in Earles' Minimum Essential Media supplemented with 10% fetal bovine serum and 0.01 μg/mL insulin.

[0055] The in vitro cytotoxicity assay was used to determine activity of each drug on MCF-7 cells. Briefly, cells were incubated with the indicated drugs at a titration of concentrations for 120 hours. Viability of each drug was measured using Cell-titer Glo (Promega, Wisconsin) (Figure 1, top). In another assay, viability of SGN-LIVIA (hLIV22-vcMMAE) was also measured using Cell-titer Glo (Figure 1, bottom).

[0056] Combinations of SGN-LIVIA and one of the agents docetaxel, paclitaxel, doxorubicin, gemcitabine, NVP-BEZ235, everolimus, or cyclophosphomaide were added to cells the following day and kept in a 5% C0 ₂ incubator at 37°C for 120 hours. Cells were then lysed with Cell Titer Glo (Promega; Seattle, WA) for 60 minutes before viability was determined on an Envision plate reader (PerkinElmer; Waltham, MA).

[0057] To evaluate combination indexes (CI), SGN-LIVIA, doxorubicin, gemecitabine, and paclitaxel were combined at their ED50 concentrations or 2 fold higher or lower concentrations to obtain a 10 points dataset. The CI value was determined using

CalcuSyn software (Biosoft; Cambridge, UK), which performs multiple drug dose-effect calculations using the Median Effect methods described by Chou and Talalay. See Chou TC (2010). Drag combination studies and their synergy quantification were performed using the Chou-Talaiay method. Cancer Res 70: 440-446. Table 1 illustrates the results below:

Table 1: SGN-LIVIA Combinations

[0058] Table 1 and Figure 2 show that SGN-LIVIA and doxorubicin were moderately synergistic at their respective ED50 and ED75 concentrations (CI<1), and slightly antagonistic at their ED 90 concentrations. Table 1 and Figure 3 show that SGN-LIVIA and paclitaxel were strongly synergistic at the ED50 concentration, but antagonistic at ED75 or ED90. Table 1 and Figure 4 show that SGN-LIV1A and gemcitabine were synergistic at the ED50 concentration, but not at the ED75 or ED90 concentrations. Table 1 and Figure 5 show that SGN-LIV1A and everolimus were synergistic at the ED50, ED75, and ED90 concentrations.

II. In vivo assays

[0059] NOD Scid Gamma (NSG) mice were obtained from Jackson Laboratory and implanted with 17-P-estradiol pellet (Innovative Research of America; Sarasota, FL) one day prior to cell implant. Five million MCF-7 cells were implanted subcutaneously with Matrigel (BD Biosciences; Franklin Lakes, NJ) and tumor volume was determined by a digital caliper twice a week, using the formula: volume= 1/2X length width .

[0060] Mice received treatments when the average tumor volume reached 100 mm . The treatment regimen for each agent was: doxorubicin, lmg/kg q4dx3 intravenous (IV); SGN-LIV1, 1 mg/kg q4dx4 intraperitoneal (IP); nab-paclitaxel (nanoparticle albumin- bound paclitaxel) 20 mg/kg q4dx3 IP; carboplatin 10 mg/kg q7dx3 IP;

cyclophosphamide, 40 mg/kg q7dx3 rV; gemcitabine 20 mg/kg q7dx2 IP; and paclitaxel 10 mg/kg q3dx5 IP. Combination groups received the first dose of SGN-LIV1A and combination drug on the same day. n=6 for each group. The effects of each drug and their combination on tumor growth were plotted in Figures 6-8.

[0061] To determine the combined benefit in vivo, tumor growth inhibition (TGI) was compared using the area under the curve between untreated, single agent groups, and combined groups on the date the untreated animals were euthanized (Table 2). The following formula was used to calculate the TGI: Thus, the expected additive TGI for combination is (l-TGI _d r _{ug x} )X(l-TGI _d r _{ug y} )- When the TGI for the combination group is 10% greater than the expected TGI, the combination regimen is considered synergistic in vivo. The data in Table 2 shows that doxorubicin, 10 mg/kg nab-paclitaxel, and carboplatin provide the highest combinatorial benefit. Furthremore, the 20 mg/kg gemcitabine and 20 mg/kg nab-paclitaxel were found to be nearly additive when combined with SGN-LIV1A.

Table 2: Summary of In Vivo Tumor Growth Inhibition (TGI)