NZ711210B2 - Combination therapy for neoplasia treatment - Google Patents

Abstract

The present invention relates to an insulin-like growth factor (IGF) receptor antagonist for use in the treatment of prostate neoplasia, including benign prostatic hyperplasia (BPH), prostate cancer, and particularly CRPC, wherein the antagonist is used in combination with an androgen receptor antagonist. An embodiment of the invention is where the androgen receptor antagonist is enzalutamide. onist. An embodiment of the invention is where the androgen receptor antagonist is enzalutamide.

Description

PCT /EP2014/054300 Combination therapy for neoplasia treatment The present invention relates to the pharmaceutical treatment of neoplasia, including benign and malignant tumors.

Background of the invention Prostate cancer is the most common malignancy diagnosed in males and a leading cause of mortality in western countries can Cancer y, 2010 (http://www.cancer.org/acs/groups/content/@epidemiologysurveilance/documents/ document/acspc-026238.pdf)). Androgens and stimulation of their receptor, androgen receptor (AR), are essential for the development and function of the normal prostate gland, and the development and progression of prostate cancer (reviewed in Basu S et al., Horm Cancer. 2010 Oct;1 (5):223-8.; Yadav N et al., Minerva Urol Nefrol. 2012 Mar;64(1):35-49). For metastatic te cancer, androgen deprivation therapy remains the standard treatment. e the fact that initially more than 90% of patients respond to androgen deprivation y, the clinical ts are temporary with tumors becoming refractory and progressing to androgen-independent/castration-resistant prostate cancer (CRPC) (Rini Bl et al., Curr Treat Options Oncol. 2002 Oct;3(5):437-46.; Carles J et al., Clin Transl Oncol. 2012 Mar;14(3):169-76). CRPC is associated with continued androgen or (AR) activation despite hormonal tion and/or treatment with currently available anti-androgens. The molecular ism of androgen stimulation of prostate cancer growth and the switch to androgen independence is not fully clear. The progression to androgen independence may be explained by s with the androgen receptor, such as amplification, mutations, or d activity of splice variants. Other possible mechanisms include tumor cell autonomous production of androgens, ligand-independent activation of AR by kinases like ERK or AKT (reviewed in Dutt SS et al., Future Oncol. 2009 Nov;5(9):1403-13. and Attar RM et al., Clin Cancer Res. 2009 May ;15(10):3251-5) or that androgens may regulate te cancer proliferation by up-regulating autocrine loops involving peptide growth factors and their cognate receptors (De Bellis A et al., J Clin Endocrinol Metab 1:4148—54.). All these mechanisms could result in independence to endocrine ens.

Benign prostatic hyperplasia (BPH) can be detected in the vast majority of men as they age (Parsons JK., Curr Bladder Dysfunct Rep. 2010 Dec;5(4):212—218). BPH can be defined as a non-cancerous enlargement of the prostate resulting from a proliferation of both benign stromal, and to a lesser extent, epithelial cells (Foster CS. Prostate 2000;9:4—14.). In both of these cell types, dihydrotestosterone (DHT), a metabolite of testosterone that is 10 times more potent e it dissociates from the androgen receptor more slowly than testosterone, binds to nuclear androgen receptors resulting in the transcription of growth factors that are mitogenic to the epithelial and stromal cells. In the prostate, testosterone is converted to DHT by the enzyme 5d-reductase, type 2. In the condition of BPH, local testosterone levels can be elevated more than 100-fold above serum levels leading to an increased availability of DHT (Gat Y et al., Andrologia 2008 (5):273-81). Therapy with 5d-reductase inhibitors, such as finasteride, markedly reduces the DHT t of the prostate and, in turn, s prostate volume and, in many cases, BPH symptoms. Androgens are t to be essential for BPH to occur, but do not seem to be the only cause for the condition.

Insulin-like growth factors(lGFs) and their binding proteins may play an important role in tanding the etiology of prostate disease, including BPH. Several lines of ce support involvementof the IGF axis in BPH. IGF ligands have mitogenic effects on the prostate, while IGF binding proteins (lGFBPs) are growth inhibitory due to their ability to regulate availability of the lGFs, other growth factors, and steroid hormones (Pollak MN et al, Nat Rev 2004;4:505—518.).

IGFBP3 is secreted at particularly low levels in stromal cells in BPH tissue (Boudon C et al., J Clin Endocrinol Metab 1996;81:612—617.) which may favor hyperplastic growth and play a role in the development of BPH. Moreover, acromegalic patients, who have very high levels of IGF1 and itantly low levels of testosterone and DHT, present with enlarged prostates and high rates of BPH (Colao A et al J Clin Endocrinol Metab 1999;84:1986—1991; Colao A et al, Eur J Endocrinol 2000;143:61—69.).

The n-like growth factor (IGF) system plays a key role in stimulating proliferation and survival of both normal tissues and cancers (reviewed in LeRoith D, Roberts CT Jr., Cancer Lett 95:127—37). High circulating lGF-1 concentrations have been associated with increased risk for te cancer in several clinical and epidemiologic studies (Price AJ et al., Cancer Epidemiol Biomarkers Prev. 2012 Sep;21(9):1531-41; Roddam AW et al., Ann Intern Med 2008;149(7):461-71). In prostate epithelial cells, increased lGF-1 expression was shown to lead to higher rates of proliferation and/or lower rates of apoptosis (Takahara K et al., Prostate. 2011 Apr;71(5):525-37). Loss of imprinting of the lGF-2 locus and increased expression of lGF-2 are observed in many cancers including prostate cancer (Jarrard DF et al., al Cancer ch 1995;1, 1471—1478.; Fu VX et al., Cancer Research 2008;68, 6797—6802) and may be related to the risk to develop prostate cancer (Belharazem D et al, Endocrine Connections 2012;1, 87—94). Furthermore, not only expression of lGF-1 and lGF-2 ligands but also their receptor, lGF-1 R, has been shown to be elevated in ed prostate tumors (Cardillo, MR et al., Anticancer Res. 2003 23, 3825— 3835; Liao, Y et al., Hum. Pathol. 2005;36 (11), 1186— 1196; Hellawell GO et al., Cancer Res. 2002 May 10):2942-50; Turney BW et al., BJU Int. 2011 May;107(9):1488—99; Krueckl SL et al., Cancer Res. 2004 Dec 1;64(23):8620-9; Figueroa, JA et al., Cancer Invest. 2001 ;19 (1), 28—34; Ryan, CJ et al., . Urol.

Oncol. 2007;25, 134—140). In recurrent and androgen-independent cancer, an increase also in AKT phosphorylation was demonstrated (Graff JR et al.,. J. Biol.

Chem 2000;275: 24500—5; Murillo H et al., Endocrinology 2001;142: 05.) Castration-resistant prostate cancer has been shown to be sensitive, but not resistant, to sustained manipulation of the androgen/AR axis. The androgen axis can be manipulated using anti-androgens (nilutamide, tamide), androgen sis tors (ketonazole, abiraterone acetate), corticosteroids (dexamethasone, prednisone) or estrogen treatment. ing the emergence of castration-refractory disease, taxane-based chemotherapy has been shown to be therapeutically efficacious and prolong survival. Patients progressing on docetaxel have been shown to benefit from abiraterone acetate, a selective cytochrome P450 17A1 inhibitor which requires inistration with glucocorticoids to l side effects. Enzalutamide (MDV-3100) is a novel AR antagonist that blocks AR ing more effectively than currently available AR antagonists (Tran et al., Science 2009;324(5928): 787-790.) and has shown impressive antitumor activity and a similar impact on overall al as abiraterone.

Antagonists to IGF action and their use in cancer y have been described in the art. For disclosure of IGF receptor tyrosine kinase inhibitors, see WO2009/009016 and WO2010/099139. For disclosure of dies against IGF receptor, see WO2002/53596, WO2003/093317, WO2003/106621, WO2006/013472, WO2006/069202. For disclosure of dies against IGF ligand, see WO2003/093317, WO2005/028515, WO2007/022172, WO2007/070432, WO2008/155387, WO2009/137758, WO2010/066868. IGF-1 receptor antibodies, WO2008/098917, WO2009/137378) and IGF ligand antibodies (WO2007/1 18214, WO2008/155387, WO2009/137758, WO2010/066868) have been proposed for use, inter alia, in the treatment of prostate .

The state of the art is also discussed in further publications (Pollak MN et al., Cancer Metastasis Rev 1998;17:383—90; Djavan B et al., World J Urol 2001;19:225—33; Wolk A et al., J Natl Cancer Inst 1998;90:911—5; Jiang YG et al., Int. J. Urol. 2007;14:1034—9; Lin HK et al., Proc. Natl Acad. Sci. USA 2001;98: ; Wen Y et al., Cancer Res. 2000;60: 6841—5; Plymate SR et al., Prostate 2004;61:276—90; AA Lubik et al., Endocr Relat Cancer ERC0250 2013, first published on 14 January ; Nickerson T et al. Cancer Res. 2001 ;61 (16), 6276— 6280; Pandini G et al., Cancer Res., 2005 March 1;65; 1849; Bedolla R et al. Clin Cancer Res. 2007 Jul 1;13(13):3860-7; Carver BS et al., Cancer Cell 2011 May 17;19, 575—586; Mulholland DJ et al., Cancer Cell, 2011 June 14;19, 792—804) Despite advances made in the early detection and treatment of prostate neoplasia, including benign prostatic hyperplasia (BPH), prostate cancer, and ularly CRPC, there is a significant need for improvements in therapy.

Brief ption of the Figures Figure 1. Inhibitory effect of IGF and AR signaling blockade on VCaP, MDA PCa 2b and DUCaP cell proliferation VCaP, MDA PCa 2b and DUCaP cells were treated with MDV-3100 and IGF ligand-neutralizing antibodies as single agents and in combination. Figure 1 shows the inhibitory effect of the IGF mAb_1 (Fig. 1A + 1C + 1E) and IGF mAb_2 (Fig. 18 + 1D + 1F) antibodies and MDV-3100, alone and in combination, on the 2D proliferation of prostate cancer-derived VCaP cells (Fig. 1A + 18), MDA PCa 2b cells (Fig. 1C + 1D) and DUCaP cells (Fig. 1E + 1F) in 10% FCS—containing growth medium. In all three cell lines, single agent ent with both IGF antibodies and MDV-3100 resulted in inhibition of cell proliferation which could be enhanced by the combination of both agents leading to a complete inhibition of proliferation.

Figure 2. Inhibitory effect of IGF signaling and androgen synthesis blockade on VCaP, MDA PCa 2b and DUCaP cell proliferation Figure 2 demonstrates the effects of the IGF mAb_1 and IGF mAb_2 dies and abiraterone acetate (AA), as single agents and in ation, on the 2D and 3D proliferation of prostate cancer-derived VCaP, MDA PCa 2b, and DUCaP cells in 10% FCS—containing growth medium. Panel (A) displays the s of the treatment of VCaP cells with IGF mAb_1 in 2D cell proliferation assays. IGF mAb_2 was used for the treatment of VCaP cells in (B). Panel C (IGF mAb_1) and D (IGF mAb_2) show the results of MDA PCa 2b cells. The treatment of DUCaP cells with IGF mAb_1 is displayed in panel E and with IGF mAb_2 in panel F.

Single agent treatment with IGF mAb_1 and mAb_2 resulted in inhibition of cell proliferation of 70% to 90%. Abiraterone acetate treatment caused inhibition of cell eration at higher concentrations which could be ed by the combination with either of the antibodies, lowering the doses of AA needed for complete inhibition. In a 3D soft agar cell proliferation assay (G), VCaP cells were treated with abiraterone acetate and IGF mAb_2. Similar to the results observed in 2D, single agent treatment with IGF mAb_2 results in 96% inhibition of cell proliferation. Abiraterone acetate treatment caused inhibition of cell proliferation at higher trations which could be enhanced by the combination with IGF mAb_2.

Figure 3. Protein analysis in VCaP, MDA PCa 2b and DUCaP cells following IGF and AR ing inhibition Figure 3 shows the effects of IGF mAb_1 and MDV-3100, alone and in combination, on IGF-1 R, AR and PTEN levels, as well as AKT phosphorylation, in VCaP, MDA PCa 2b and DUCaP cells as assessed by Western blot es.

Cells were seeded in 6-well plates and treated for 24 hours. (A) Lysates prepared from treated VCaP cells were compared to untreated controls and insensitive PC-3 cells for protein expression of IGF-1 R, AR, PTEN and AKT and for phosphorylation of AKT-Ser473. Protein expression and AKT phosphorylation of untreated and treated MDA PCa 2b (B) and DUCaP (C) cells were ted and compared to that of VCaP cells. Untreated PC-3 cells served as a control. lmportantly, VCaP, MDA PCa 2b, and DUCaP cells were shown to express the lGF1-R, AR and PTEN, unlike the insensitive cell line PC-3. MDV-3100 treatment slightly increased AR protein levels which may be due to stabilization of the n. Concomitantly, IGF-1 R levels were slightly decreased upon MDV-3100 treatment. The combination of both agents resulted in a more complete inhibition of AKT phosphorylation than the antibody or MDV-3100 treatment alone.

Figure 4. IGF signaling pathway inhibition following single agent and combination treatment of IGF mAb_1 and MDV-3100.

Figure 4 trates the effects of IGF mAb_1 and MDV-3100 used as single agents and in combination on IGF-1 R levels and AKT phosphorylation in VCaP cells over 120 hours of ent. VCaP cells were seeded in 6-well plates and treated with MDV-3100 and IGF mAb_1 as single agents or in combination for 24, 48, 72, 96, and 120 hours. Lysates prepared from treated cells were ed to untreated controls for phosphorylation of AKT-Ser473. Combination of both agents resulted in a longer lasting inhibition of AKT phosphorylation than the antibody or 00 treatments alone.

Figure 5. Reduced proliferative activity of VCaP cells following single agent and combination treatment of IGF mAb_1 and MDV-3100 eration of VCAP cells was monitored using a H3-thymidine incorporation assay. Treated with 10 pM of MDV-3100 or 1 pM of IGF mAb_1 as single agents for 96 hours reduced erative activity by approximately 50%. Combination of IGF mAb_1 and MDV-3100 reduced thymidine incorporation by more than 95% compared to untreated controls.

Figure 6. Diminished growth rate of VCaP cells following single agent and ation ent of IGF mAb_1 and MDV-3100 (A) Effect of 1 pM of IGF mAb_1 and 10 pM of MDV-3100 used as single agents and in ation on cell morphology and cell growth as observed in microscopic analyses. (B) Effect of 10 pM of MDV-3100 on the generation time of VCaP cells compared to untreated controls.

Figure 7. Combination treatment of IGF mAb_1 and MDV-3100 ses induction of apoptosis in VCaP cells Caspase 3 activity was used as a measure of the induction of apoptosis in VCaP cells upon treatment with 10 pM of MDV-3100 and 1 pM of mAb as single agents and in combination for up to 96 hours. Wheras MDV-3100 treatment did not induce caspase 3 activity within 96 hours of treatment, an increase in apoptotic events were ed upon treatmen with IGF mAb_1. The combination of both agents showed a synergistic effect on the induction of caspase 3 activity, which was appoximately 9-fold increased ed to controls and approximately 25-fold higher compared to IGF mAb_1 treatment.

Figure 8. Cell cycle profiles of VCaP cells d with MDV-3100 and IGF mAb_1 The cell cycle profiles of VCaP cells after 24 h, 48 h and 72 h of treatment with 10 pM of MDV-3100 and 1 pM of mAb as single agents and in combination as determined by propidium iodide staining detected by flow cytometry. The first population to the left is the sub-G1 population representing apoptotic cells, the second population shows the G1/G0 peak, the light grey population shows cells in the S-phase, and the population to the right represents cells in the G2/M-phase of the cell cycle. In VCaP cells treated with IGF mAb_1, and to a greater extent in cells treated with the combination of IGF mAb_1 and MDV-3100 the tic cell population increases with time, wheras in VCaP cells treated with MDV-3100 this population does not change. Instead, MDV-3100 treatment increased the G1/G0 population compared to untreated ls.

Figure 9. Protein analysis of apoptosis and cell cycle regulators following IGF signaling inhibition The effects of the treatment with 10 µM of 00 and 1 µM of IGF mAb_1 as single agents and in combination on AKT phosphorylation, PARP cleavage, p21, CDK2, Cyclin E, and PCNA levels after 8h, 24h, 48h and 72h of treatment where ed by Western blot analysis. IGF mAb_1 treatment led to blockade of AKT phosphorylation, and combined IGF mAb_1 and MDV-3100 treatment increased PARP cleavage and Cyclin E levels while it reduced CDK2 and PCNA levels. MDV- 3100 treatment increased p21 levels at 8 and 24 hours.

Brief description of the invention In a first aspect of the invention, there is provided use of an insulin-like growth factor (IGF) receptor nist in the manufacture of a medicament for the treatment of prostate neoplasia, wherein the ment is ated for stration in combination with an androgen receptor antagonist selected from enzalutamide and abiraterone acetate, wherein the IGF receptor antagonist is an antibody having heavy chain complementary determining s of SEQ ID NO: 1 (HCDR1), SEQ ID NO: 2 (HCDR2), and SEQ ID NO: 3 (HCDR3) and light chain determining regions of SEQ ID NO: 4 (LCDR1), SEQ ID NO: 5 (LCDR2), and SEQ ID NO: 6 (LCDR3), or an antibody having heavy chain complementary determining s of SEQ ID NO: 11 (HCDR1), SEQ ID NO: 12 (HCDR2), and SEQ ID NO: 13 (HCDR3) and light chain determining regions of SEQ ID NO: 14 (LCDR1), SEQ ID NO: 15 (LCDR2), and SEQ ID NO: 16 (LCDR3), or an antibody having heavy chain mentary determining regions of SEQ ID NO: 21 (HCDR1), SEQ ID NO: 22 (HCDR2), and SEQ ID NO: 23 (HCDR3) and light chain determining s of SEQ ID NO: 24 (LCDR1), SEQ ID NO: 25 (LCDR2), and SEQ ID NO: 26 (LCDR3), ), or an antibody having heavy chain complementary determining regions of SEQ ID NO: 31 ), SEQ ID NO: 32 (HCDR2), and SEQ ID NO: 33 (HCDR3) and light chain determining regions of SEQ ID NO: 34 (LCDR1), SEQ ID NO: 35 (LCDR2), and SEQ ID NO: 36 (LCDR3), or an antibody having a heavy chain variable region of SEQ ID NO: 7 and a light chain variable region of SEQ ID NO: 8, or an antibody having a heavy chain variable region of SEQ ID NO: 17 and a light chain le region of SEQ ID NO: 18, ), or an antibody having a heavy chain variable region of SEQ ID NO: 27 and a light chain variable region of SEQ ID NO: 28, ), or an antibody having a heavy chain variable region of SEQ ID NO: 37 and a light chain variable region of SEQ ID NO: 38, ), or an dy having a heavy chain variable region of SEQ ID NO: 41 and a light chain variable region of SEQ ID NO: 42, ), or an antibody having a heavy chain variable region of SEQ ID NO: 43 and a light chain variable region of SEQ ID NO: 44, or an antibody having a heavy chain of SEQ ID NO: 9, and a light chain of SEQ ID NO: 10, or an antibody having a heavy chain of SEQ ID NO: 19, and a light chain of SEQ ID NO: 20, or an antibody having a heavy chain of SEQ ID NO: 29, and a light chain of SEQ ID NO: 30, or an antibody having a heavy chain of SEQ ID NO: 39, and a light chain of SEQ ID NO: 40.

In a second aspect of the ion, there is provided use of an androgen or antagonist selected from enzalutamide and abiraterone acetate in the manufacture of a medicament for the treatment of prostate neoplasia, wherein the medicament is formulated for administration in ation with an IGF receptor antagonist, wherein the IGF receptor antagonist is an antibody having heavy chain complementary determining regions of SEQ ID NO: 1 (HCDR1), SEQ ID NO: 2 (HCDR2), and SEQ ID NO: 3 (HCDR3) and light chain determining regions of SEQ ID NO: 4 (LCDR1), SEQ ID NO: 5 (LCDR2), and SEQ ID NO: 6 (LCDR3), or an antibody having heavy chain complementary determining regions of SEQ ID NO: 11 (HCDR1), SEQ ID NO: 12 (HCDR2), and SEQ ID NO: 13 (HCDR3) and light chain determining regions of SEQ ID NO: 14 (LCDR1), SEQ ID NO: 15 (LCDR2), and SEQ ID NO: 16 (LCDR3), or an antibody having heavy chain complementary ining s of SEQ ID NO: 21 (HCDR1), SEQ ID NO: 22 (HCDR2), and SEQ ID NO: 23 (HCDR3) and light chain determining regions of SEQ ID NO: 24 (LCDR1), SEQ ID NO: 25 (LCDR2), and SEQ ID NO: 26 (LCDR3), ), or an antibody having heavy chain complementary determining regions of SEQ ID NO: 31 (HCDR1), SEQ ID NO: 32 (HCDR2), and SEQ ID NO: 33 (HCDR3) and light chain determining regions of SEQ ID NO: 34 (LCDR1), SEQ ID NO: 35 (LCDR2), and SEQ ID NO: 36 (LCDR3), or an antibody having a heavy chain variable region of SEQ ID NO: 7 and a light chain variable region of SEQ ID NO: 8, or an antibody having a heavy chain variable region of SEQ ID NO: 17 and a light chain variable region of SEQ ID NO: 18, ), or an antibody having a heavy chain variable region of SEQ ID NO: 27 and a light chain variable region of SEQ ID NO: 28, ), or an antibody having a heavy chain variable region of SEQ ID NO: 37 and a light chain variable region of SEQ ID NO: 38, ), or an antibody having a heavy chain variable region of SEQ ID NO: 41 and a light chain variable region of SEQ ID NO: 42, ), or an antibody having a heavy chain variable region of SEQ ID NO: 43 and a light chain variable region of SEQ ID NO: 44, or an antibody having a heavy chain of SEQ ID NO: 9, and a light chain of SEQ ID NO: 10, or an antibody having a heavy chain of SEQ ID NO: 19, and a light chain of SEQ ID NO: 20, or an antibody having a heavy chain of SEQ ID NO: 29, and a light chain of SEQ ID NO: 30, or an antibody having a heavy chain of SEQ ID NO: 39, and a light chain of SEQ ID NO: 40.

In a further aspect, the present ion pertains to an insulin-like growth factor (IGF) receptor nist for use in the treatment of prostate neoplasia, ing benign tic lasia (BPH), prostate cancer, and particularly CRPC in combination with an androgen receptor antagonist.

In another embodiment, the invention relates to a method of treatment of prostate neoplasia, including benign prostatic hyperplasia (BPH), prostate , and particularly CRPC comprising administering a therapeutically effective amount of an IGF receptor antagonist to a patient in need thereof, and additionally administering a therapeutically ive amount of an androgen receptor PCT /EP2014/054300 antagonist to the same patient on the same day, or one, two, three, four, five, six or seven days before or after admistration of the IGF receptor antagonist.

Detailed description of the ion The present invention relates to the treatment of prostate neoplasia.

By "prostate neoplasia", the aspects of the invention include where the prostate neoplasia is prostate cancer, including benign and malignant tumours, and particularly castration resistant prostate cancer; and also benign prostatic hyperplasia.

In one aspect, the t invention pertains to an insulin-like growth factor (IGF) receptor antagonist for use in the treatment of prostate cancer. In r embodiment, the prostate cancer is hormone-sensitive prostate cancer. In another embodiment, the te cancer is prostate cancer after combined androgen blockade. In another embodiment, the prostate cancer is prostate cancer treated with antiangiogenic therapy. In another embodiment the prostate cancer has been, or will be, treated with a chemotherapeutic agent. In another embodiment, the prostate cancer is prostate cancer treated, or will be treated, with ion therapy. In another embodiment, the prostate cancer is prostate cancer treated, or will be treated, with bone loss therapy, for example mab, and e ablation.

In another embodiment, the prostate cancer is castration ant prostate cancer (CRPC). In another embodiment the castration ant prostate cancer has been, or will be, treated with a chemotherapeutic agent. In r embodiment, the castration resistant prostate cancer has been, or will be, treated with radiation therapy. In another embodiment, the prostate cancer is castration resistant PCT /EP2014/054300 prostate cancer in a pre- or post-docetaxel setting. In another embodiment, the prostate cancer is castration resistant prostate cancer after cabazitaxel treatment.

In another embodiment, the prostate cancer is castration resistant prostate cancer after ent with androgen sis inhibitors, for example erone acetate. In another embodiment, the prostate cancer is tion resistant prostate cancer after treatment with en receptor antagonists, for example enzalutamide. In another embodiment, the prostate cancer is castration resistant prostate cancer after treatment with immune-modulating agents, for example Sipuleucel-T.

In another aspect, the present invention pertains to an insulin-like growth factor (IGF) receptor antagonist for use in the treatment of prostate cancer incombination with an androgen or antagonist. In another embodiment, the prostate cancer is hormone-sensitive prostate cancer. In r embodiment, the prostate cancer is te cancer after combined androgen blockade. In another ment, the prostate cancer is prostate cancer treated with antiangiogenic y. In another embodiment the prostate cancer has been, or will be, d with a chemotherapeutic agent. In another embodiment, the prostate cancer is prostate cancer treated, or will be treated, with radiation therapy. In another embodiment, the prostate cancer is prostate cancer treated, or will be treated, with bone loss therapy, for example denosumab, and hormone ablation.

In another embodiment, the prostate cancer is castration ant prostate cancer.

In r embodiment the castration resistant prostate cancer has been, or will be, treated with a chemotherapeutic agent. In another embodiment, the castration resistant prostate cancer has been, or will be, treated with radiation y. In another embodiment, the te cancer is castration resistant prostate cancer in a pre- or post-docetaxel setting. In another embodiment, the prostate cancer is castration resistant prostate cancer after cabazitaxel treatment. In another embodiment, the prostate cancer is tion resistant prostate cancer after treatment with androgen synthesis inhibitors, for example abiraterone acetate. In another embodiment, the prostate cancer is castration resistant prostate cancer PCT /EP2014/054300 after treatment with en or antagonists, for e enzalutamide.. In another embodiment, the prostate cancer is castration ant prostate cancer after treatment with immune-modulating agents, for example Sipuleucel-T.

In another aspect, the present invention pertains to an insulin-like growth factor (IGF) receptor antagonist for use in the treatment of benign tic hyperplasia.

In another aspect, the present invention pertains to an insulin-like growth factor (IGF) receptor antagonist for use in the treatment of benign prostatic hyperplasia in combination with an androgen receptor antagonist.

An IGF receptor nist within the context of the invention is a compound that interferes with, either directly or indirectly, and reduces or blocks IGF receptor signaling. Preferably, an IGF receptor antagonist is a compound that reduces or blocks binding of IGF ligand to its receptor, or ts the tyrosine kinase activity of the IGF receptor.

In a further embodiment, the IGF receptor antagonist of the present invention is an antibody that binds to IGF ligand and thus reduces or prevents binding of the ligand to the receptor. In another embodiment, the IGF receptor antagonist is an antibody that binds to the IGF-1 receptor and thus s or prevents binding of the ligand to the or. By blocking receptor-ligand binding, ligand-induced or signaling through the tyrosine kinase activity of the receptor is reduced or prevented. Such antibodies are generally referred to as neutralizing antibodies. In another aspect, the present invention pertains to an IGF or nist that neutralizes the growth promoting properties of the insulin-like growth s, IGF- 1 and IGF-2.

The term "antibody" encompasses antibodies, antibody fragments, antibody-like molecules and conjugates with any of the above. Antibodies include, but are not limited to, poly- or monoclonal, chimeric, humanized, human, mono-, bi- or multispecific antibodies. The term "antibody" shall encompass complete PCT /EP2014/054300 immunoglobulins as they are produced by lymphocytes and for example present in blood sera, monoclonal antibodies secreted by hybridoma cell lines, polypeptides produced by inant expression in host cells, which have the binding specificity of immunoglobulins or monoclonal antibodies, and molecules which have been derived from such immunoglobulins, monoclonal antibodies, or polypeptides by further processing while retaining their g specificity. In particular, the ntibody" es te immunoglobulins comprising two heavy chains and two light chains. In another embodiment, the term asses a fragment of an immunoglobulin, like Fab fragments. In r embodiment, the term "antibody" encompasses a polypeptide having one or more variable domains derived from an immunobulin, like single chain antibodies (scFv), single domain dies, and the like.

In a further embodiment, the IGF receptor antagonist of the invention is an antibody t IGF-1, an antibody against IGF-2, an antibody binding both IGF-1 and IGF-2, an antibody against IGF-1 receptor (IGF-1R), or an inhibitor of IGF-1R tyrosine kinase activity.

In another embodiment, the IGF receptor nist is an IGF ligand antibody having heavy chain complementary ining regions of SEQ ID NO: 1 (HCDR1), SEQ ID NO: 2 (HCDR2), and SEQ ID NO: 3 (HCDR3) and light chain determining regions of SEQ ID NO: 4 (LCDR1 ), SEQ ID NO: 5 (LCDR2), and SEQ ID NO: 6 (LCDR3).

In another embodiment, the IGF receptor antagonist is an IGF ligand antibody having heavy chain complementary determining regions of SEQ ID NO: 11 (HCDR1), SEQ ID NO: 12 (HCDR2), and SEQ ID NO: 13 (HCDR3) and light chain determining regions of SEQ ID NO: 14 (LCDR1), SEQ ID NO: 15 (LCDR2), and SEQ ID NO: 16 (LCDR3).

In another embodiment, the IGF receptor antagonist is an IGF ligand antibody having heavy chain complementary determining regions of SEQ ID NO: 21 PCT /EP2014/054300 (HCDR1), SEQ ID NO: 22 (HCDR2), and SEQ ID NO: 23 (HCDR3) and light chain determining regions of SEQ ID NO: 24 (LCDR1), SEQ ID NO: 25 (LCDR2), and SEQ ID NO: 26 (LCDR3).

In another preferred embodiment, the IGF receptor antagonist is an IGF ligand antibody having heavy chain complementary determining regions of SEQ ID NO: 31 (HCDR1), SEQ ID NO: 32 (HCDR2), and SEQ ID NO: 33 (HCDR3) and light chain determining regions of SEQ ID NO: 34 (LCDR1), SEQ ID NO: 35 ), and SEQ ID NO: 36 (LCDR3). An example of an antibody containing these complementary determining regions is designated herein as IGF mAb_1.

In another ment, the IGF receptor antagonist is an IGF ligand dy having a heavy chain le region of SEQ ID NO: 7 and a light chain variable region of SEQ ID NO: 8.

In another ment, the IGF receptor antagonist is an IGF ligand antibody having a heavy chain variable region of SEQ ID NO: 17 and a light chain variable region of SEQ ID NO: 18.

In another embodiment, the IGF receptor antagonist is an IGF ligand antibody having a heavy chain variable region of SEQ ID NO: 27 and a light chain variable region of SEQ ID NO: 28.

In another preferred embodiment, the IGF receptor antagonist is an IGF ligand antibody having a heavy chain variable region of SEQ ID NO: 37 and a light chain variable region of SEQ ID NO: 38. An example of an antibody containing these variable regions is ated herein as IGF mAb_ 1.

In another embodiment, the IGF receptor antagonist is an IGF ligand antibody having a heavy chain variable region of SEQ ID NO: 41 and a light chain variable region of SEQ ID NO: 42.

PCT /EP2014/054300 In another embodiment, the IGF or antagonist is an IGF ligand antibody having a heavy chain variable region of SEQ ID NO: 43 and a light chain variable region of SEQ ID NO: 44.

In another embodiment, the IGF receptor antagonist is an IGF ligand antibody having a heavy chain of SEQ ID NO: 9, and a light chain of SEQ ID NO: 10.

In another embodiment, the IGF receptor antagonist is an IGF ligand antibody having a heavy chain of SEQ ID NO: 19, and a light chain of SEQ ID NO: 20.

In another embodiment, the IGF receptor antagonist is an IGF ligand antibody having a heavy chain of SEQ ID NO: 29, and a light chain of SEQ ID NO: 30.

In another preferred embodiment, the IGF receptor antagonist is an IGF ligand dy having a heavy chain of SEQ ID NO: 39, and a light chain of SEQ ID NO: 40. An example of an antibody containing these heavy and light chains is designated herein as IGF mAb_1.

In another embodiment, the IGF receptor antagonist is an IGF receptor antibody having a heavy chain of SEQ ID NO: 45, and a light chain of SEQ ID NO: 46.

In another ment, the IGF or antagonist is mumab, dalotuzumab, cixutumumab, robatumumab, or ganitumab.

In another embodiment, the IGF receptor antagonist is linsitinib.

Preferably the IGF receptor antagonist is IGF mAb_1, as defined above.

Manufacture and therapeutic use of the aforementioned antibodies is disclosed in /53596, WO2007/070432, WO2008/152422, WO2008/155387, and WO2010/066868.

PCT /EP2014/054300 In one embodiment, the antibody is produced by inant expression in a mammalian host cell, purified by a series of chromatographic and nonchromatographic steps, and formulated in an aqueous buffer composition for parenteral (intravenous) infusion or injection at an dy concentration of 10 mg/ml, said buffer comprisingfor example 25 mM Na citrate pH 6, 115 mM NaCl, and 0.02 % polysorbate 20. For intravenous infusion, the pharmaceutical composition may be diluted with a physiological solution, e.g. with 0.9 % sodium chloride or G5 solution.

The antibody may be administered to the patient at a dose between 1 mg/kg to 20 mg/kg, by one or more separate administrations, or by continuous infusion, e.g. infusion over 1 hour. A typical treatment schedule y involves administration of the antibody once every week to once every three weeks. For example, a weekly dose could be 5, 10, or 15 mg/kg. Preferably, the antibody is prepared at a concentration of 10 mg/ml of IGF mAb_1. The antibody may preferably be stered to a patient as a 750 mg (up to 1000 mg) total dose by one hour i.v. infusion, to be repeated once a week until disease progression The IGF receptor antagonist is administered to the patient in combination with administration of an androgen receptor antagonist. "In ation" means that both drugs are administered to the same patient within a certain time frame to achieve a therapeutic effect caused by the combined effects of both modes of action. In one aspect, the androgen receptor antagonist is administered on the same day as the IGF receptor antagonist. In another aspect of the invention, the androgen receptor antagonist is administered one, two, three, four, five, six or seven days before or after admistration of the IGF or antagonist.

In another embodiment, both active compounds are present within the same pharmaceutical composition. Hence, in another ment, the invention ns to a pharmaceutical ition, comprising an IGF receptor antagonist and an androgen receptor antagonist, together with a ceutically acceptable carrier.

PCT /EP2014/054300 An androgen receptor antagonist (AR nist) is a compound that blocks androgen or (AR) signaling. Androgen receptor antagonists prevent androgens from expressing their biological effects on responsive tissues. Such compounds may alter the androgen pathway by blocking the respective receptors, competing for binding sites on the receptor, affecting r translocation, DNA binding of the receptor, or affecting en production. In the context of the present invention the androgen receptor antagonist can be an anti-androgen, an androgen synthesis inhibitor, a 17 a-hydroxylase/C17,20 lyase (CYP17A1) inhibitor, a 5-alpha-reductase inhibitor, a corticosteroid, a luteinizing hormonereleasing hormone (LH-RH) agonist, or an estrogen agonist.

In another embodiment, the androgen or antagonist is ide, mide, enzalutamide, bicalutamide, ketonazole, abiraterone, abiraterone acetate, orteronel, finasteride, dutasteride, bexlosteride, eride, turosteride, ride, dexamethasone, prednisone, leuprolide, goserelin, triptorelin, histrelin, or estrogen.

In another embodiment, the androgen receptor antagonist is enzalutamide (Tran et al., Science 2009, 324(5928): 787-790.) Enzalutamide can be obtained from, for e, Medivation or Astellas under the name Xtandi ®. Enzalutamide is preferably administered as a dosage of 160 mg once daily during each cycle of treatment In another embodiment, the androgen receptor antagonist is abiraterone, for example in the form of abiraterone acetate (Agarwal et al., Future Oncology 2010, 6(5): 665-679). Abieraterone can be obtained from, for example, Janssen Biotech, Inc.

Manufacture, ation, and use of the androgen receptor antagonist depends on the actual compound chosen and can be found in the state of the art.

PCT /EP2014/054300 Another ment of the invention is an en receptor antagonist for use in the treatment of prostate cancer in combination with an IGF receptor antagonist. In another embodiment the use of an androgen receptor antagonist in combination with an IGF or nist is for the treatment of benign prostatic hyperplasia. In a further embodiment, said en or antagonist is flutamide, nilutamide, enzalutamide, bicalutamide, ketonazole, abiraterone acetate, orteronel, finasteride, dutasteride, bexlosteride, izonsteride, turosteride, episteride, dexamethasone, prednisone, leuprolide, goserelin, triptorelin, histrelin, or estrogen.

Another embodiment of the invention pertains to a method of treatment of prostate neoplasia comprising administering a therapeutically effective amount of an IGF receptor antagonist to a t in need thereof, and additionally administering a therapeutically effective amount of an androgen receptor antagonist to the same patient on the same day, or one, two, three, four, five, six or seven days before or after admistration of the IGF receptor antagonist.

By "prostate neoplasia", this aspect of the ion include where the prostate neoplasia is prostate cancer, including benign and malignant tumours, and particularly castration resistant prostate ; and also benign prostatic lasia.

A "therapeutically effective amount" of the IGF or androgen receptor antagonist to be administered is the minimum amount necessary to prevent, ameliorate, or treat a prostate neoplasia, in particular castration-resistant prostate cancer, or benign prostatic hyperplasia.

In another embodiment, the invention pertains to the use of an IGF receptor antagonist for the manufacture of a medicament for the ent of prostate neoplasia , wherein the IGF receptor antagonist is to be used in combination with an androgen receptor antagonist.

PCT /EP2014/054300 By "prostate neoplasia", this aspect of the invention include where the te neoplasia is prostate cancer, including benign and malignant tumours, and particularly castration resistant prostate cancer; and also benign tic hyperplasia.

In another embodiment, the invention pertains to the use of an en receptor nist for the manufacture of a medicament for the treatment of prostate cancer neoplasia, wherein the androgen receptor antagonist is to be used in combination with an IGF receptor antagonist.

By "prostate neoplasia", this aspect of the invention include where the prostate neoplasia is prostate , ing benign and malignant tumours, and particularly tion resistant prostate cancer; and also benign prostatic hyperplasia.

EXAMPLES Materials and Methods Compounds IGF mAb_1 is an antibody against IGF ligand having a heavy chain of SEQ ID NO: 39 and a light chain of SEQ ID NO: 40. Its manufacture has been disclosed in WO 2010/066868.

IGF mAb_2 is an antibody against IGF ligand having a heavy chain of SEQ ID NO: 29 and a light chain of SEQ ID NO: 30. Its manufacture has been disclosed in WO 2010/066868.

PCT /EP2014/054300 Cell culture DU-145 (ATCC, HTB-81), BM-1604 (DSMZ, ACC 298), PC-3 (ATCC, CRL-1435), 22Rv1 (ATCC, CRL-2505), LNCaP (ATCC, CRL-1740), and DUCaP cells (generated in the lab of Prof KJ. Pienta, Hallym University, College of Medicine, Seoul, Korea; Lee YG et al., In Vivo 2001 ;15(2):157-62) were cultivated in RPMI 1640 growth medium (GIBCO, #31870) supplemented with 10 % heat inactivated fetal calf serum (FCS; JRH, #12103), and 2 mM L-glutamine (GIBCO, #25030); NCI-H660 (ATCC, CRL-5813) were grown in RPMI supplemented with 5% FCS, 4 mM L-glutamine, 5 µg/ml insulin, 0.01 mg/ml transferrin, 30 nM sodium selenite, nM beta estradiol and 10 nM hydrocortisone. C4-2 and C4-2b (both licensed from MD Anderson Cancer Center; Thalmann GN et al., Cancer Res. 1994; 54:2577-2581) and VCaP (ATCC, CRL-2876) were cultivated in DMEM , #12-604F) supplemented with 10 % heat inactivated FCS, 2 mM L-glutamine and R1881 (Sigma, #R0908; VCaP with 0.1 nM and 4-2b with 1 nM). -2b (ATCC, CRL-2422) were grown in F-12K , #21127) supplemented with 20% nactivated FCS, 25 ng/ml cholera toxin, 0.005 mM ethanolamine, 100 pg/ml hydrocortisone, and 45 nM ous acid. Bob cells (ECACC, #10021102) were cultured in keratinocyte-SFM (lnvitrogen, #37010-022) supplemented with prequalified human recombinant epidermal growth factor 1-53, bovine pituitary extract and glutamine, 2 ng/ml leukemia inhibitory , 2 ng/ml stem cell factor, 100 ng/ml cholera toxin, and 1 ng/ml ocyte macrophage colony stimulating factor. Shmac 4 , #10112302), Shmac 5 (ECACC, #10112303) and P4E6 cells (ECACC, 301) were grown in Stemline Keratinocyte Medium II (Sigma, #S0196) with Stemline Keratinocyte Growth Supplement (Sigma, #S9945), 2mM L-glutamine and 2% FCS. The cells were maintained in 75 cm2 tissue culture flasks (Nunc, #178905) at 37°C in 5 % CO2 in a humidified atmosphere. 2D Cell Proliferation Assay The following method was used to determine the inhibitory effect of IGF ligandneutralizing mAbs and androgen signaling inhibitors on the growth of prostate WO 35611 PCT /EP2014/054300 cancer cell lines. Assays were performed in cell growth medium containing 10% serum.

Adherent cells were detached with trypsin/EDTA solution (GIBCO, #043-9031FU), resuspended in growth medium, centrifuged, resuspended in assay medium (supplemented with 10% heat inactivated FCS and 2 mM amine) and diluted to 5,000 - 40,000 cells per ml. 100 µL/well cell suspension was added to each well of a sterile flat-bottom white 96-well plate (PerkinElmer, #6005280) and plates were incubated overnight in a humidified incubator set at +37°C and 5% CO2. On the next day supernatants were aspirated and 35 µL/well assay medium was added to all wells.

Serial dilutions of IGF mAb_1 and mAb_2 (1 µM highest concentration), MDV- 3100 (10 µM highest concentration), abiraterone e (100 µM highest concentration) were prepared on a separate plate in assay medium (no growth factors or hormones supplemented). All agents were tested as single agents or in combination. All samples were tested in triplicate wells (100 µL/well assay). Plates were incubated for 5 days in a humidified incubator at +37°C and 5% CO2. After this incubation period, CellTiter-Glo buffer, substrate and test plates were brated to RT. CellTiter-Glo is a bioluminescent assay ga, #G7571) designed to determine the number of viable cells in culture, in which the generation of a luminescent signal is proportional to the amount of ATP present in cells. 100 µL of freshly mixed CellTiter-Glo reagent was added to each well. After 2 min on an l shaker (MTS 2/4, IKA) and 10 min incubation at RT, luminescence was recorded (luminescence reader (Genios Pro, Tecan or Victor X4, Perkin Elmer), integration time 1 sec).

Generation of Cell Lysates and lmmunoblotting One x 106 and 4 x 106 cells were plated in 6-well plates and 10 cm dishes, tively, in medium containing 10% heat-inactivated FCS and after over night incubation the cells were treated with 1 µM of MDV-3100 and 100 nM of IGF mAb_1 or a combination of antibody and AR signaling inhibitor. After 24 hours the PCT /EP2014/054300 cells were lyzed on the plates, total protein was isolated and protein concentration was determined by Bradford assay. Cell lysates were snap frozen and stored at - 80°C.

Western blotting was performed loading 30-50 µg of total protein s on a 4- 12% Bis-Tris PAG (Bio Rad) and blotting with the Bio Rad blot® Turbo system using a PVDF ne. Membranes were incubated over night at 4°C with antibodies against the following proteins: IGF-1R beta (#3027, Cell Signaling; 1:1000), p-S473 AKT (#4060, Cell Signaling; 1:2000), AKT (#9272, Cell Signaling; 1:1000), PTEN (#9559, Cell Signaling; 1:1000), AR (N-20, # , Santa Cruz; , and GAPDH (#7298, Cell Signaling; 1 :1000)(which served as loading control). Cell cycle regulators and markers of proliferation and apoptosis were analyzed using the following antibodies: p21 Waf1/Cip1 (12D1;#2947, Cell ing; 1:1000), CDK2 (78B2; #2546, Cell Signaling; 1:1000), Cyclin E (C-19; sc-198, Santa Cruz; 1:1000), PCNA (#2586, Cell Signaling; 1:2000), and PARP (#9542, Cell Signaling; 1:1000).

Antibody ons were prepared in 5% BSA or 5% non-fat dry milk in 5% 0 (TBS-T). Following washes in TBS-T membranes were incubated with a polyclonal HRP-conjugated goat anti-rabbit secondary antibody (DAKO, ) for 1 hour and after further washes in TBS-T antibody reactivity was detected by means of EGL/Super EGL (GE Healthcare) and exposure on lmageQuant LAS4000. For the detection of total protein levels, membranes incubated with antiphospho antibodies were stripped in Restore Western Blot Stripping Buffer (Thermo, #21059) for 15-20 min, blocked, and incubated with the antibody against the total protein before the membrane was processed as described above.

Cell Cycle Analysis using Flow Cytometry 4 x 105 VCaP cells were treated with 1 µM of IGF mAb_ 1 and 10 µM of MDV- 3100, and the combination of both agents, and incubated in 6-well plates at 37°C for 24 h, 48 h and 72 h. uently, the supernatant was transferred to FAGS tubes, nt cells were detached with trypsin and collected in the respective PCT 4/054300 FAGS tubes. After centrifugation, the medium was discarded and the cell pellet was fixed in ice-cold 70% ethanol for a minimum of 2 h at 4 °C. After removing the ethanol entirely, fixed cells were stained with propidium iodide (10 µg/ml; Sigma; P4864-10ml) in a hypotonic buffer solution (0.1 % sodium citrate, 0.1 % (v/v) triton X-100, 100 µg/ml DNase-free RNase A) and incubated in the dark at room temperature for 30 minutes. Cells were analyzed using the Becton Dickinson FAGS Canto II Flow ter and data was evaluated with the FAGS Diva software.

Thymidine Incorporation Assay VCaP cells were treated with 1 µM of IGF mAb_1 and 10 µM of MDV-3100 and the combination of both agents and incubated as triplicates in ottom l plates for 96 hours at a density of 5 x 104 cells per well, in the absence of R1881.

For the last 24 hours of incubation, H-thymidine (0.4 µCi/well; PerkinElmer, NET355001 MC) was added. Afterwards, the plates were frozen and incubated at - °C for 24h. For harvesting, the plates were thawed and 40 µL Trypsin was added to each well to detach the cell fragments. The suspension was transferred to filter plates. The plates were then washed three times with distilled water and dried at 60°C for 3h. 25 µL per well Microscint were added and the proliferation rate was ined by measuring thymidine incorporation (CPM; counts per ) using a liquid scintillation counter (1450 Microbeta Wallac Trilux, Perkin­ Elmer).

Analysis of Cellular Doubling Time 3 x 105 cells/well VCaP cells were seeded in 2 ml cell culture medium per well. 24 hrs post g the cell culture medium was removed and replaced with DMEM +10% FCS without R1881. 24 hr following the medium change the pre-treatment wells were harvested and counted with the Beckman Coulter™ Vi-CELL XR 2.03, and 10 µM of MDV-3100 was added to the remaining cells. Four times every 24 hr VCaP cell number was determined in 3 wells for each time point. The mean value was calculated from these triplicates. To determine the generation time, following formula is used: PCT /EP2014/054300 . . log2 :.: rnltin1tion hours [h] [h generation time] = 1og,�·- 1og.�o . . _ log2 )( rnltimtion hours [h] [h generationtm e -] 1 1og,v- • • 1ogno �, No= cell count at TO N= cell count after cultivation Assessment of Caspase 3 Activity To acquire live cell images of cells undergoing caspase-3/7 mediated apoptosis upon treatment with different concentrations of MDV-3100, IGF mAb_1, and the combination of both agents, the CellPlayer™ 96-Well Kinetic Caspase-3/7 Reagent (Essen Bioscience; # 4440) was used. 50000 VCaP cells/ 100µ1/ well were seeded and treated on the next day with the respective concentrations of both agents in growth medium in the absence of R1881. The Caspase-3/7 reagent was ed to a final concentration of 5 µM in 100 µI per well of growth medium and added to the . The plate was placed within a microplate tray into the lncuCyte™ 2011A and 3 images per well were acquired every 4 hours for 7 days using the phase st and fluorescence channels.

Example 1 Inhibitory effect of IGF and AR ing blockade on prostate cancer cell proliferation In order to test the anti-proliferative effects of the combination of AR and IGF-1/2 inhibition, 10 different prostate cancer cell lines (Bob, C4-2, C2B, DUCaP, MDA PCa 2b, P4E6, PC-3, Shmac 4, Shmac 5, VCaP) were treated with the AR antagonist MDV-3100 and fully human monoclonal antibodies against the IGF ligands (IGF mAb_1 and IGF mAb_2), as single agents and in combination, in 2D cell proliferation assays (Table 1). Three of the tested cell lines (VCaP and DUCaP - both cell lines were derived from the same te cancer patient from different sites of asis, and MDA PCa 2b) showed single agent anti-proliferative PCT /EP2014/054300 response to both the AR and IGF signaling inhibition alone, and an enhanced effect when ed (Figure 1).

Example 2 Inhibitory effect of IGF signaling and androgen synthesis blockade on prostate cancer cell proliferation As a second approach to test the combination potential of androgen and IGF signaling inhibition, 8 different prostate cancer cell lines (22Rv1, BM 1604, DU- 145, DUCaP, LNCaP, MDA PCa 2b, PC-3, VCaP) were treated with abiraterone e, which selectively inhibits CYP17A and thus de novo synthesis of androgens, alone and in combination with IGF-ligand neutralizing monoclonal antibodies (IGF mAb_1 and IGF mAb_2). The results from these assays also identified VCaP, MDA PCa 2b, and DUCaP cells to be the only cell lines which are responsive to both single agent and combination treatments. Treatment with erone acetate, however, implies autocrine androgen production by the tumor cells for abiraterone acetate to show anti-proliferative effects. This might limit the number of cells sensitive to abiraterone acetate ent. Results of 2D and 3D eration assays for VCaP and 2D assays of MDA PCa 2b and DUCaP cells are shown in Figure 2. These data t that the single agent effects of abiraterone acetate on cell proliferation can be ed by the combination with antibodies neutralizing IGF s.

Example 3 The presence of androgen receptor and IGF-1 R as well as expression of PTEN and wt PIK3CA characterizes prostate cancer cells sensitive to the combination of androgen and IGF signaling inhibitors Figure 3 shows signaling protein expression in the VCaP, MDA PCa 2b, and DUCaP cell lines, which are sensitive to AR and IGF signaling inhibition, in comparison to the insensitive cell line PC-3. Cells were treated with MDV-3100 and IGF mAb_1 as single agents, or in combination, for 24 hours and protein lysates were ed to untreated controls for protein expression of IGF-1R, AR, PCT /EP2014/054300 PTEN and AKT, and for phosphorylation of AKT-Ser473. Responsive cell lines expressed wt AR, IGF-1 R, and PTEN. These characteristics were not present in PC-3 or the other tested cell lines which did not show an anti-proliferative responce to either one of the single agent treatments or the combination of both agents (Table 1 ).

These results indicate that in the presence of androgen receptor, IGF-1 R, and expression of PTEN (and wt PIK3CA), the combination of androgen and IGF signaling inhibitors results in an increased efficacy in blocking prostate cancer cell proliferation in vitro.

Example 4 Prolonged AKT orylation inhibition following combined treatment of MDV-3100 and IGF mAb 1 The effects of MDV-3100 and IGF ligand mAb (IGF mAb_1) as single agents, and combined treatment, on the tion of AKT phosphorylation were analyzed by Western blot from 4h until 120h of treatment. The combination of both agents resulted in a more complete and longer g inhibition of AKT orylation than the antibody treatment alone (Figure 4).

Combined treatment with IGF mAb_1 and MDV-3100 leads to a istic effect on apoptotic induction in VCaP cells In support of the data shown in Figure 1, results from tritiated thymidine incorporation assays shown in Figure 5 demonstrates that both MDV-3100 and IGF mAb_1 alone have an inhibitory effect on cell proliferation (approximately 50%), r, the combination of both agents was much more ive. ent of VCaP cells with IGF mAb 1 alone led to a modest increase in apoptosis as assessed by phase contrast microscopy (Figure 6A), caspase 3 activity (Figure 7), FAGS-based cell cycle analysis (Figure 8), and PARP cleavage (Figure 9). In contrast, the reduced cell number seen after treatment with MDV- PCT /EP2014/054300 3100 alone (Figure 6A) was due to prolonged cellular doubling time (Figure 6B). 00 did not induce caspase 3 activity (Figure 7), sub-G1 apoptosis cell population (Figure 8), or PARP cleavage e 9). However, when IGF mAb_1 and MDV-3100 were combined a synergistic effect on caspase 3 activity was observed (Figure 7), in addition to enhanced sub-G1 apoptotic cell population (Figure 8) and cleaved PARP (Figure 9).

Example 6: Proposed study of IGF mAb_1 in combination with enzalutamide Introduction The study proposed here investigates the safety and anti-tumour activity of IGF mAb_ 1 in combination with enzalutamide, compared to enzalutamide given alone, in CRPC patients This randomised, open label, study will be conducted to explore the anti-tumour activity and safety profile of the combination of IGF mAb_ 1 and enzalutamide (Arm A), compared to enzalutamide (Arm B). A tolerability and safety phase lb will be performed to determine the maximum tolerated dose (MTD), and/or recommended phase II dose, in addition to any safety issues before commencement of the ised trial..

IGF mAb_ 1 will be administered weekly in 28 day cycles of ent by a one hour intravenous on at the start of each treatment cycle. Enzalutamide will be administered daily by uous oral dosing during each treatment cycle.

Background IGF mAb_ 1 is a fully human monoclonal dy (HumAb) of the lgG1 isotype.

The Ab binds with high affinity to IGF-1 and IGF-2, and potently neutralizes the erative and prosurvival cellular signaling triggered by both proteins.

PCT 4/054300 Enzalutamide is an en receptor nist that acts on different steps in the androgen or ling pathway. The al name is 4-{3-[4-cyano (trifluoromethyl )phenyl]-5,5-dimethyloxosulfanylideneimidazolidinyl} fluoro-N-methylbenzamide. The molecular weight is 464.44 and molecular formula is C21H16F4N4O2S. Enzalutamide is indicated for the treatment of patients with metastatic castration-resistant prostate cancer (CRPC) Administration IGF mAb_ 1 will be administered weekly in 28 day cycles of treatment, by a one hour intravenous infusion at the start of each treatment cycle. Enzalutamide will be administered daily by continuous oral dosing during each treatment cycle.

Selection of trial population A total of up to approximately 140 patients may be recruited into the study.

Approximately 15-18 patients will be entered into the part I tolerability and safety phase of the study to ensure the safety of the combination therapy and determine the part II recommended dose. In part II of the study, 120 patients will be randomised onto one of the two study arms, with 60 patients randomised to each arm (Arm 8=60, Arm 8=60).

Part I of the study will be performed in 3 or more centres. Part II of the study will be performed in 10 or more centres globally.

A log of all patients included into the study (i.e. having given informed consent) will be maintained in the ISF at the investigational site irrespective of whether they have been treated with investigational drug or not.

PCT /EP2014/054300 Main sis for study entry Patients to be included in this study must have diagnosed and histologically, or cytologically, confirmed metastatic CRPC and have received and progressed after one line of docetaxol treatment. Patients may, or may not, have received and failed prior abiraterone, or taxel treatment, in any setting.

Inclusion criteria 1. The patient has ogically, or cytologically, confirmed adenocarcinoma of the prostate. 2. Male patient aged � 18 years old. 3. Patients with radiographic evidence of atic prostate cancer (stage M1 or D2). Distant metastases evaluable by radionuclide bone scan, CT scan, or MRI within 28 days of start of study treatment. 4. ts who have disease progression (biochemical, clinical or radiographic) while receiving docetaxel, or within 120 days of completing docetaxel-based chemotherapy and in the opinion of the investigator is unlikely to derive icant t from additional docetaxel-based therapy, or was intolerant to therapy with this agent.

. Patients must have evidence of progressive disease defined as at least one of the following: a. Progressive measurable disease: using tional solid tumour criteria RECIST 1.1. b. Bone scan progression: at least two new lesions on bone scan. c. Increasing PSA: at least two consecutive rising PSA values over a reference value (PSA #1) taken at least 1 week apart. A third PSA (PSA #3) is required to be greater than PSA #2; if not, a fourth PSA (PSA #4) is required to be greater than PSA #2. 6. Patients with a PSA � 2 ng/ml.

PCT /EP2014/054300 7. Patients with prior surgical or medical tion with a serum testosterone of <50 ng/ml. If the method of castration is luteinizing hormone releasing level hormone (LHRH) agonists, the patient must be willing to continue the use of LHRH ts during protocol treatment. 8. Eastern Cooperative Oncology Group performance status (ECOG PS) 0, 1 or 2. 9. Patients have te hematologic function (absolute neutrophil count [ANG] �1500/ul, obin �9 g/dl, and platelets �100,000/ul).

. Patients have adequate hepatic function (bilirubin::;; 1.5 times the upper limit of normal (ULN)], aspartate transaminase [AST] and e transaminase [ALT]::;; 3 times the ULN, or::;; 5 times the ULN if liver metastases are present). 11. Adequate renal function (creatinine ::;; 1.5 x ULN or calculated creatinine clearance> 40 ml/min). 12. A urinary protein of::;; 1+ on ck or routine urinalysis (UA). If urine ck or routine analysis indicates � 2+ proteinuria, then a r urine must be collected and must demonstrate < 1000 mg of protein in 24 hours to allow participation in the study. 13. Adequate coagulation function (an international normalized ratio [INR] ::;; 1.5 and a partial thromboplastin time [PTT] ::;; 5 seconds above the ULN [unless on oral anticoagulant therapy]). Patients receiving full-dose anticoagulation therapy are eligible provided they meet all other criteria, are on a stable dose of oral anticoagulant or low molecular weight heparin (except warfarin, which is not permitted). 14. Fasting plasma glucose < 8.9 mmol/L (< 160 mg/dl) or HbA1c < 8.0%.

Exclusion criteria 1. Patients that have received more than two prior taxane based cytotoxic chemotherapy regimen for metastatic disease. Patients who have had a ent break from docetaxol ed by a second or third docetaxel-based regimen, with subsequent disease progression, are eligible. 2. ts that have received prior enzalutamide in any setting will not be eligible.

PCT /EP2014/054300 3. Patients who have received abiraterone, or cabazitaxel treatment, within 4 weeks before start of study treatment. 4. Patient that have received prior therapy with mitoxantrone for advanced prostate cancer (prior adjuvant therapy with mitoxantrone is permitted). 5. Patients that have been treated with any of the following within 4 weeks of starting trial medication: chemotherapy, immunotherapy, biological therapies, molecular targeted, hormone therapy, radiotherapy (except in case of localized radiotherapy for analgesic purpose or for lytic lesions at risk of fracture which can then be completed within 2 weeks prior to study treatment). 6. Use of any investigational drug within 4 weeks before start of trial treatment or concomitantly with this trial. 7. ts that have been treated with strong CYP2C8 inhibitors; strong or moderate CYP3A4 or CYP2C8 inducers; CYP3A4, CYP2C9 and CYP2C19 substrates with a narrow therapeutic index, within 4 weeks of starting the trial. 8.Patients with a history of symptomatic tive heart failure or has a prestudy echocardiogram or ated acquisition (MUGA) scan with left ventricular ejection on (LVEF) that is � 10% below the LLN. 9. QTcF prolongation> 450 ms or QT prolongation deemed clinically relevant by the investigator (e.g., congenital long QT syndrome).The QTcF will be calculated as the mean of the 3 ECGs taken at ing.

. Patients with small cell or neuroendocrine tumours. 11. Patients with known or suspected leptomeningeal ases. 12. Uncontrolled or poorly controlled ension. 13. Patients with poorly controlled diabetes mellitus. Patients with a history of diabetes are allowed to participate, provided that their blood glucose is within normal range (fasting < 160 mg/dl or below ULN) and that they are on a stable dietary or therapeutic regimen for this condition. 14. Known human immunodeficiency virus infection or acquired immunodeficiency me-related illness. 15. Patients with epilepsy, seizures, or posing factors for seizure as judged by the investigator. 16. ts unable to comply with the ol as judged by the investigator.

PCT /EP2014/054300 17. Active alcohol or active drug abuse as judged by the investigator. 18.A history of allergy to human monoclonal antibodies. 19. Prior therapy with agents targeting IGF and/or IGFR pathway.

. Patients who are sexually active and unwilling to use a lly acceptable method of contraception (e.g. such as implants, injectables, combined oral contraceptives, some intrauterine s or omized partner for participating females, condoms for participating males) during the trial and for at least three months after end of active therapy. Men unwilling to agree to not donate sperm while on trial drug and up to 6 months following the last dose of trial drug.

Additional ion criteria for part II: 21. For patients that are to undergo the optional tumour biopsy, a history of a hereditary bleeding disorder, or clinically relevant major bleeding event in the past 6 months, as judged by the investigator.

Treatments to be stered nce: IGF mAb_ 1 human monoclonal antibody Pharmaceutical form: Liquid formulation Source: Boehringer lngelheim Pharma GmbH & Co. KG Unit strength: 10 mg/ml of IGF mAb_1 supplied in 20 ml vials.

Appropriate dose of IGF mAb_1 will be diluted in logical sodium chloride solution (0.9%).

Duration of use: One hour at the start of each week (Day 1, 8, 15 and 22) of a 28 day cycle of treatment until disease progression or undue toxicities. Infusion duration may be extended to over one hour in case of infusion reaction or adverse events.

Route of administration: Intravenous Starting dose: 750 mg (up to 1000 mg) total dose by one hour i.v. infusion PCT /EP2014/054300 Additional information: Dose will be ed during part I tolerability/safety and dose finding phase nce: Enzalutamide (Xtandi®) Pharmaceutical form: Liquid-filled soft gelatin capsule Source: Astellas Unit strength: 40 mg Duration of use: 160 mg once daily during each cycle of treatment Route of administration: Oral Starting dose: 160 mg once daily Additional information: Dose will be adjusted during part I tolerability /safety and dose finding phase from that stated in the summary of product teristics (SPC).

Table 1 gives an overview of the ons, protein expression and effects of androgen and IGF signaling inhibition observed in the 15 different tested prostate cancer cell lines.

Table 1 ., c c � :,,: Q. .... u C ..0 ::, u �� � a: � ;e'.j:i E oi :c u !:2 a: E: Q. E: m :i: .29 :c E 00 .c < ;;: f- � "' ·= � .29 .c"' ·= LJ.J < "iii ·-C E 0 0u <.J LJ.J LJ.J 22Rvl + ~ + - + ~ ~ (Q546R) BM-1604 - + ~ wt - - ~ ~ derived from DU-145 Bob - ~ ~ - - - - Spontaneously immortalized CRPC (1391H) C4-2 + + - wt - + - - LNCaP d cell lines (xenograft in castrated mice) C4-2B + + - wt - + - - PCT /EP2014/054300 DU 145 - + - wt - - ~ - Isolated from ent metastases * DUCaP + + + wt ERG + + +++ from same PCa patient as VCaP LNCap.FGC + + - wt - + - - *MDAPCa 2b + + + wt - + + ++ NCI-H660 - ~ - n.d. ERG - n.d. n.d.

P4E6 - + ~ wt - - - - PC-3 - + - wt - - - - Shmac4 - + ~ wt - - - - Shmac5 - + ~ wt - - - - Isolated from different metastases *VCaP + + + wt ERG + + +++ from same PCa patient as DUCaP Cell lines labeled with an asterisk represent responsive cell lines expressing wt AR, wt Pl3K, PTEN and .

Abbreviations: AR= androgen receptor; IGF-1R = Insulin-like growth factor 1 receptor; mut = mutated; n.d. = not determined; wt= wild type

Claims (8)

Claims 1.

1. Use of an insulin-like growth factor (IGF) receptor antagonist in the manufacture of a ment for the treatment of prostate neoplasia, wherein the medicament is formulated for administration in combination with an en or antagonist selected from enzalutamide and abiraterone acetate, wherein the IGF receptor antagonist is an antibody having heavy chain complementary determining regions of SEQ ID NO: 1 (HCDR1), SEQ ID NO: 2 (HCDR2), and SEQ ID NO: 3 (HCDR3) and light chain determining regions of SEQ ID NO: 4 (LCDR1), SEQ ID NO: 5 (LCDR2), and SEQ ID NO: 6 (LCDR3), or an antibody having heavy chain complementary determining regions of SEQ ID NO: 11 (HCDR1), SEQ ID NO: 12 (HCDR2), and SEQ ID NO: 13 (HCDR3) and light chain determining regions of SEQ ID NO: 14 (LCDR1), SEQ ID NO: 15 (LCDR2), and SEQ ID NO: 16 (LCDR3), or an antibody having heavy chain complementary ining regions of SEQ ID NO: 21 (HCDR1), SEQ ID NO: 22 (HCDR2), and SEQ ID NO: 23 (HCDR3) and light chain determining regions of SEQ ID NO: 24 (LCDR1), SEQ ID NO: 25 (LCDR2), and SEQ ID NO: 26 (LCDR3), ), or an antibody having heavy chain complementary determining s of SEQ ID NO: 31 (HCDR1), SEQ ID NO: 32 (HCDR2), and SEQ ID NO: 33 (HCDR3) and light chain determining s of SEQ ID NO: 34 (LCDR1), SEQ ID NO: 35 (LCDR2), and SEQ ID NO: 36 (LCDR3), or an antibody having a heavy chain variable region of SEQ ID NO: 7 and a light chain variable region of SEQ ID NO: 8, or an antibody having a heavy chain variable region of SEQ ID NO: 17 and a light chain le region of SEQ ID NO: 18, ), or an antibody having a heavy chain variable region of SEQ ID NO: 27 and a light chain variable region of SEQ ID NO: 28, ), or an antibody having a heavy chain variable region of SEQ ID NO: 37 and a light chain le region of SEQ ID NO: 38, ), or an antibody having a heavy chain variable region of SEQ ID NO: 41 and a light chain variable region of SEQ ID NO: 42, ), or an antibody having a heavy chain variable region of SEQ ID NO: 43 and a light chain variable region of SEQ ID NO: 44, or an dy having a heavy chain of SEQ ID NO: 9, and a light chain of SEQ ID NO: 10, or an antibody having a heavy chain of SEQ ID NO: 19, and a light chain of SEQ ID NO: 20, or an antibody having a heavy chain of SEQ ID NO: 29, and a light chain of SEQ ID NO: 30, or an antibody having a heavy chain of SEQ ID NO: 39, and a light chain of SEQ ID NO: 40.

2. The use of claim 1 wherein the prostate neoplasia, is benign prostatic lasia (BPH) or prostate cancer.

3. The use of claim 2 wherein the prostate cancer, is castration resistant prostate cancer.

4. The use of any one of claims 1 to 3, wherein the antibody has a heavy chain of SEQ ID NO: 39, and a light chain of SEQ ID NO: 40.

5. Use of an androgen or antagonist selected from enzalutamide and erone acetate in the manufacture of a ment for the treatment of prostate neoplasia, wherein the medicament is ated for administration in combination with an IGF receptor antagonist, wherein the IGF receptor antagonist is an antibody having heavy chain complementary determining regions of SEQ ID NO: 1 (HCDR1), SEQ ID NO: 2 (HCDR2), and SEQ ID NO: 3 (HCDR3) and light chain determining regions of SEQ ID NO: 4 (LCDR1), SEQ ID NO: 5 (LCDR2), and SEQ ID NO: 6 (LCDR3), or an antibody having heavy chain complementary determining regions of SEQ ID NO: 11 (HCDR1), SEQ ID NO: 12 (HCDR2), and SEQ ID NO: 13 (HCDR3) and light chain determining regions of SEQ ID NO: 14 (LCDR1), SEQ ID NO: 15 (LCDR2), and SEQ ID NO: 16 (LCDR3), or an antibody having heavy chain complementary ining regions of SEQ ID NO: 21 ), SEQ ID NO: 22 ), and SEQ ID NO: 23 (HCDR3) and light chain determining regions of SEQ ID NO: 24 (LCDR1), SEQ ID NO: 25 (LCDR2), and SEQ ID NO: 26 (LCDR3), ), or an dy having heavy chain complementary determining regions of SEQ ID NO: 31 (HCDR1), SEQ ID NO: 32 (HCDR2), and SEQ ID NO: 33 (HCDR3) and light chain determining regions of SEQ ID NO: 34 (LCDR1), SEQ ID NO: 35 (LCDR2), and SEQ ID NO: 36 (LCDR3), or an antibody having a heavy chain variable region of SEQ ID NO: 7 and a light chain variable region of SEQ ID NO: 8, or an antibody having a heavy chain variable region of SEQ ID NO: 17 and a light chain variable region of SEQ ID NO: 18, ), or an antibody having a heavy chain variable region of SEQ ID NO: 27 and a light chain variable region of SEQ ID NO: 28, ), or an antibody having a heavy chain variable region of SEQ ID NO: 37 and a light chain variable region of SEQ ID NO: 38, ), or an antibody having a heavy chain variable region of SEQ ID NO: 41 and a light chain variable region of SEQ ID NO: 42, ), or an antibody having a heavy chain variable region of SEQ ID NO: 43 and a light chain variable region of SEQ ID NO: 44, or an antibody having a heavy chain of SEQ ID NO: 9, and a light chain of SEQ ID NO: 10, or an antibody having a heavy chain of SEQ ID NO: 19, and a light chain of SEQ ID NO: 20, or an antibody having a heavy chain of SEQ ID NO: 29, and a light chain of SEQ ID NO: 30, or an dy having a heavy chain of SEQ ID NO: 39, and a light chain of SEQ ID NO: 40.

6. The use of claim 5 wherein the te neoplasia, is benign prostatic hyperplasia (BPH) or prostate cancer.

7. The use of claim 6 wherein the prostate cancer, is castration resistant prostate cancer.

8. The use of any one of claims 5 to 7, wherein the antibody has a heavy chain of SEQ ID NO: 39, and a light chain of SEQ ID NO: 40. Boehringer Ingelheim International GmbH By the Attorneys for the Applicant SPRUSON & FERGUSON Per: