Improving the delivery of drugs and other agents to target cells, tissues and tumors to achieve maximal efficacy and minimal toxicity has been the focus of considerable research for many years. Though many attempts have been made to develop effective methods for importing biologically active molecules into cells, both in vivo and in vitro, none has proved to be entirely satisfactory. Optimizing the association of the drug with its intracellular target, while minimizing intercellular redistribution of the drug, e.g. to neighboring cells, is often difficult or inefficient.
Monoclonal antibody therapy has been established for the targeted treatment of patients with cancer, immunological and angiogenic disorders. One example, HERCEPTIN® (trastuzumab; Genentech, Inc.; South San Francisco, Calif.) is a recombinant DNA-derived humanized monoclonal antibody that selectively binds with high affinity in a cell-based assay (Kd=5 nM) to the extracellular domain of the human epidermal growth factor receptor2 protein, HER2 (ErbB2) (U.S. Pat. No. 5,821,337; U.S. Pat. No. 6,054,297; U.S. Pat. No. 6,407,213; U.S. Pat. No. 6,639,055; Coussens L, et al (1985) Science 230:1132-9; Slamon D J, et al (1989) Science 244:707-12). Trastuzumab is an IgG1 kappa antibody that contains human framework regions with the complementarity-determining regions (cdr) of a murine antibody (4D5) that binds to HER2. Trastuzumab binds to the HER2 antigen and thus inhibits the growth of cancerous cells. Because Trastuzumab is a humanized antibody, it minimizes any HAMA (Human Anti-Mouse Antibody) response in patients. Trastuzumab has been shown, in both in vitro assays and in animals, to inhibit the proliferation of human tumor cells that overexpress HER2 (Hudziak R M, et al (1989) Mol Cell Biol 9:1165-72; Lewis G D, et al (1993) Cancer Immunol Immunother; 37:255-63; Baselga J, et al (1998) Cancer Res. 58:2825-2831). Trastuzumab is a mediator of antibody-dependent cellular cytotoxicity, ADCC (Hotaling T E, et al (1996) [abstract]. Proc. Annual Meeting Am Assoc Cancer Res; 37:471; Pegram M D, et al (1997) [abstract]. Proc Am Assoc Cancer Res; 38:602). In vitro, Trastuzumab-mediated ADCC has been shown to be preferentially exerted on HER2 overexpressing cancer cells compared with cancer cells that do not overexpress HER2. HERCEPTIN® as a single agent is indicated for the treatment of patients with metastatic breast cancer whose tumors overexpress the HER2 protein and who have received one or more chemotherapy regimens for their metastatic disease. HERCEPTIN® in combination with paclitaxel is indicated for treatment of patients with metastatic breast cancer whose tumors overexpress the HER2 protein and who have not received chemotherapy for their metastatic disease. HERCEPTIN® is clinically active in patients with ErbB2-overexpressing metastatic breast cancers that have received extensive prior anti-cancer therapy (Baselga et al, (1996) J. Clin. Oncol. 14:737-744).
The use of antibody-drug conjugates (ADC), i.e. immunoconjugates, for the local delivery of cytotoxic or cytostatic agents to kill or inhibit tumor cells in the treatment of cancer (Syrigos and Epenetos (1999) Anticancer Research 19:605-614; Niculescu-Duvaz and Springer (1997) Adv. Drg Del. Rev. 26:151-172; U.S. Pat. No. 4,975,278) theoretically allows targeted delivery of the drug moiety to tumors, and intracellular accumulation therein, where systemic administration of these unconjugated drug agents may result in unacceptable levels of toxicity to normal cells as well as the tumor cells sought to be eliminated (Baldwin et al., (1986) Lancet pp. (Mar. 15, 1986):603-05; Thorpe, (1985) “Antibody Carriers Of Cytotoxic Agents In Cancer Therapy: A Review,” in Monoclonal Antibodies '84: Biological And Clinical Applications, A. Pinchera et al. (ed.s), pp. 475-506). Maximal efficacy with minimal toxicity is sought thereby. Efforts to design and refine ADC have focused on the selectivity of monoclonal antibodies (mAbs) as well as drug-linking and drug-releasing properties. Both polyclonal antibodies and monoclonal antibodies have been reported as useful in these strategies (Rowland et al., (1986) Cancer Immunol. Immunother., 21:183-87). Drugs used in these methods include daunomycin, doxorubicin, methotrexate, and vindesine (Rowland et al., (1986) supra). Toxins used in antibody-toxin conjugates include bacterial toxins such as diphtheria toxin, plant toxins such as ricin, small molecule toxins such as geldanamycin (Mandler et al (2000) Jour. of the Nat. Cancer Inst. 92 (19):1573-1581; Mandler et al (2000) Bioorganic & Med. Chem. Letters 10:1025-1028; Mandler et al (2002) Bioconjugate Chem. 13:786-791), maytansinoids (US 20050169933 A1; EP 1391213; Liu et al., (1996) Proc. Natl. Acad. Sci. USA 93:8618-8623), and calicheamicin (Lode et al (1998) Cancer Res. 58:2928; Hinman et al (1993) Cancer Res. 53:3336-3342). The toxins may effect their cytotoxic and cytostatic effects by mechanisms including tubulin binding, DNA binding, or topoisomerase inhibition. Some cytotoxic drugs tend to be inactive or less active when conjugated to large antibodies or protein receptor ligands.
ZEVALIN® (ibritumomab tiuxetan, Biogen/Idec) is an antibody-radioisotope conjugate composed of a murine IgG1 kappa monoclonal antibody directed against the CD20 antigen found on the surface of normal and malignant B lymphocytes and 111In or 90Y radioisotope bound by a thiourea linker-chelator (Wiseman et al (2000) Eur. Jour. Nucl. Med. 27 (7):766-77; Wiseman et al (2002) Blood 99 (12):4336-42; Witzig et al (2002) J. Clin. Oncol. 20 (10):2453-63; Witzig et al (2002) J. Clin. Oncol. 20 (15):3262-69). Although ZEVALIN has activity against B-cell non-Hodgkin's Lymphoma (NHL), administration results in severe and prolonged cytopenias in most patients. MYLOTARG™ (gemtuzumab ozogamicin, Wyeth Pharmaceuticals), an antibody drug conjugate composed of a hu CD33 antibody linked to calicheamicin, was approved in 2000 for the treatment of acute myeloid leukemia by injection (Drugs of the Future (2000) 25 (7):686; U.S. Pat. No. 4,970,198; U.S. Pat. No. 5,079,233; U.S. Pat. No. 5,585,089; U.S. Pat. No. 5,606,040; U.S. Pat. No. 5,693,762; U.S. Pat. No. 5,739,116; U.S. Pat. No. 5,767,285; U.S. Pat. No. 5,773,001). Cantuzumab mertansine (Immunogen, Inc.), an antibody drug conjugate composed of the huC242 antibody linked via the disulfide linker SPP to the maytansinoid drug moiety, DM1, is advancing into Phase II trials for the treatment of cancers that express CanAg, such as colon, pancreatic, gastric, and others. MLN-2704 (Millennium Pharm., BZL Biologics, Immunogen Inc.), an antibody drug conjugate composed of the anti-prostate specific membrane antigen (PSMA) monoclonal antibody linked to the maytansinoid drug moiety, DM1, is under development for the potential treatment of prostate tumors. The auristatin peptides, auristatin E (AE) and monomethylauristatin (MMAE) synthetic analogs of dolastatin, were conjugated to chimeric monoclonal antibodies including: cBR96 (specific to Lewis Y on carcinomas); cAC10 (specific to CD30 on hematological malignancies); and other antibodies (US 20050238649 A1) and are under therapeutic development (Doronina et al (2003) Nature Biotechnology 21 (7):778-784).
DNA intercalation is a proposed mechanism for inhibiting the progression of tumorigenesis. Bis-1,8 naphthalimide compounds strongly bind DNA and may disrupt the DNA-topoisomerase II complex (Bailly et al (2003) Biochemistry 42:4136-4150) by stacking with purine nucleobases on opposite strands (Gallego et al (1999) Biochemistry 38 (46):15104-15115). Bis-1,8 naphthalimide compounds have been investigated for their anti-cancer properties (Brana et al (2004) Jour. Med. Chem. 47 (6):1391-1399; Brana et al (2004) J. Med. Chem. 47:2236-2242; Bailly et al (2003) Biochemistry 42:4136-4150; Carrasco et al (2003) 42:11751-11761; Brana, M. F. and Ramos, A. (2001) Current Med. Chem.-Anti-Cancer Agents 1:237-255; Mekapati et al (2001) Bioorganic & Med. Chem. 9:2757-2762; U.S. Pat. No. 5,641,782; U.S. Pat. No. 5,376,664). The investigational antitumor drug bis-1,8 naphthalimide mesylate (LU79553, also known as elinafide dimesylate, N,N-bis[1,8-naphthalimido)ethyl]-1,3-diaminopropane bismethane sulfonate; or N,N′-Bis[2-(1,3-dioxo-2,3-dihydro-1H-benz[de]isoquinolin-2-yl)ethyl]-1,3-diaminopropane dimethanesulfonate; 2,2′-Propane-1,3-diylbis(iminoethylene)bis(2,3-dihydro-1H-benz[de]isoquinoline-1,3-dione) dimethanesulfonate, (Abbott Laboratories, Knoll A G, Ludwigshafen, D E)), is composed of two tricyclic 1,8-naphthalimide chromophores separated by an aminoalkyl linker chain and designed to permit bisintercalation of the drug into DNA (Villalona-Calero et al (2001) Jour. Clinical Oncology 19 (3):857-869; Bousquet et al (1995) Cancer Res. 55:1176-1180; U.S. Pat. No. 4,874,863; U.S. Pat. No. 5,416,089; U.S. Pat. No. 5,616,589; U.S. Pat. No. 5,789,418; WO 95/05365). Clinical trials with Elinafide were conducted in Germany (Awada et al (2003) Euro. J. of Cancer 39 (6):742-747). Unlike most other known topoisomerase II inhibitors, elinafide does not cause significant DNA cleavage suggesting Elinafide inhibits topoisomerase II via a different mechanism. This could mean that cancer cells resistant to coventional topoisomerase II inhibitors may not be cross-resistant to elinafide. Elinafide also inhibits topoisomerase I isolated from calf thymus with and IC50 value of 5 μMolar assessed by a supercoiled DNA relaxation assay. In mouse xenograft models, repeated dosing regimens with elinafide demonstrated antitumor activity and did not demonstrate a strong schedule dependency (Bousquet et al (1995) Cancer Res. 55:1176-1180). In human xenograft models, complete regression of MX-1 (mammary carcinoma) xenografts was observed when LU-79553 was administered iv for five daily doses at 20 mg/kg (2 cycles, beginning on days 6 and 20), or every 3 days for two doses at 55 mg/kg (2 cycles, beginning on days 6 and 13) or weekly for four doses (Bousquet et al (1995) Cancer Res. 55:1176-1180). Elinafide has been shown to be curative also in human melanoma (LOX) models and give partial and complete tumor regression, as well as some cures, in human lung (LX-1) and human colon (CX-1) carcinoma xenograft models.
It is desirable to test further analogs of bis-1,8 naphthalimide compounds for their anticancer properties. It is desirable to discover such analogs with optimized biological in vivo properties such as pharmacokinetics, pharmacodynamics, metabolism, potency, safety, and bioavailability. It is also desirable to discover such analogs with optimized physical properties such as increased aqueous solubility and stability.
It is further desirable to the known anticancer properties of bis-1,8 naphthalimide compounds by conjugation to antibodies in order to improve their delivery to target cells, and achieve maximal efficacy and minimal toxicity.