Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Abstract 1551: Identification of plasma biomarker concentration changes resulting from the administration of the Vascular Disrupting Agent BNC105 across 3 clinical trials in mesothelioma, ovarian and renal cancer

Abstract BNC105 is a tubulin depolymerisation agent. Its activity includes effects on both cancer cells and on solid tumor microvasculature. BNC105 shows evidence of strong anti-cancer efficacy in vitro and in animal models. In solid tumors its efficacy is driven by selective destruction of tumor vasculature (Vascular Disrupting Agent - VDA) and direct action on tumor cells through suppression of their proliferation. In non-proliferating blood cancers (e.g. Chronic Lymphocytic Leukaemia) BNC105 activates pro-apoptotic proteins, which mediate cancer cell death. BNC105P may be useful in the treatment of human cancers both as a monotherapy and also in combination therapies. To date BNC105 has been evaluated in mesothelioma as a monotherapy, in ovarian cancer in combination with gemcitabine/carboplatin and in renal cancer in combination with everolimus. Seventy nine (79) patients receiving intravenous administrations of BNC105 in these trials were blood sampled at baseline and following BNC105 administration. The plasma concentrations of a panel of 83 plasma analytes were investigated for changes resulting from the administration of BNC105. Here, we report data across the three clinical trials on plasma biomarkers that consistently change in response to the administration of BNC105. Our findings include a number of biomarkers that have not been previously associated with the action of a VDA. Significant increases within 3 hours following BNC105 administration were observed across all studies regardless of monotherapy or combination. These included Ferritin, IL-8, IL-10, IL-16, MIP1B, MCP-1, TNFR2, and MMP9. Decreases in several markers were also observed that included Alpha-2 Macroglobulin, Myoglobin, PAI-1, TIMP-1. Correlation of biomarker changes to 6 month Progression free survival (6MPFS) was undertaken in 44 patients treated with BNC105+everolimus in the phase II renal cancer trial. This analysis resulted in the identification of 4 plasma analytes where changes from baseline were associated with progression free survival at 6 months. Taken together, the biomarker program that has accompanied the clinical development of BNC105 has allowed the identification of exploratory plasma biomarkers associated with treatment with a VDA that can potentially identify patient populations that respond better to treatment. These results will support the continued development of BNC105 in biomarker-guided clinical trial designs. Citation Format: Gabriel Kremmidiotis, Annabel Leske, Jeremy Simpson, Elizabeth Doolin, Jose Iglesias. Identification of plasma biomarker concentration changes resulting from the administration of the Vascular Disrupting Agent BNC105 across 3 clinical trials in mesothelioma, ovarian and renal cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1551. doi:10.1158/1538-7445.AM2015-1551</jats:p

Abstract 662: Evaluation of the anti-cancer effects of the tumor selective vascular disruption agent BNC105 in preclinical renal cancer models

Abstract The disruption of blood vessels that feed tumors represents one of the most promising therapeutic strategies for treating cancer. BNC105 is a tubulin targeting dual acting vascular disruption agent with cytotoxic ability in solid tumors. A prodrug formulation, BNC105P, is under evaluation in phase II clinical trials for malignant mesothelioma and metastatic renal cell carcinoma as a 10min IV infusion. We have conducted a number of preclinical evaluations that provide a strong rationale for combining BNC105P treatment with agents that target mTOR signalling. BNC105 activity operates through the selective disruption of tumor blood vessels. Over 95% of blood flow is disrupted in tumors grown in xenograft animal models using human cancer cell lines representing a number of different cancer types, including breast, lung, colon, brain and mesothelioma. In the renal cancer setting we evaluated the anti-cancer effects of BNC105 in a Caki-1 xenograft model and a syngeneic murine renal cancer orthotopic model. Animals carrying Caki-1 solid tumors were treated with a single dose of BNC105P. Disruption of blood flow within the tumors was observed as early as 3 hr post-treatment. Similarly, blood flow disruption was seen in mice carrying solid tumors orthotopically inoculated in the kidney capsule using the RENCA mouse renal cancer cell line. Interestingly, BNC105P treatment also caused blood flow disruption in lung metastatic lesions seen in a number of the animals orthotopically inoculated with RENCA. The vasculature in non-cancerous portions of the kidney and all normal tissues examined, remained intact. Tumor re-vascularisation following BNC105P treatment was observed 2 days following administration. Immunohistochemical and Western blot analyses in BNC105P treated tumors revealed that a number of proteins involved in the mTOR signalling pathway exhibit expression changes consistent with activation of this pathway possibly a result of BNC105P induced tumor hypoxia. Up-regulation in phosphorylated mTOR, Hif1a and down-regulation of phosphorylated 4EBP1 were observed. The consequences of targeting mTOR in combination with BNC105P are currently under investigation in these preclinical models. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 662. doi:10.1158/1538-7445.AM2011-662</jats:p

Abstract 2768: Development of a tubulin fractionation assay for the evaluation of “on target” activity of tubulin targeting agents in clinical PBMC samples

Abstract The novel agent, BNC105, is a tubulin polymerization inhibitor which acts as a vascular disruption agent (VDA). BNC105 exhibits 100-fold selectivity for activated endothelial cells compared to quiescent endothelial cells, providing a large differential between effects on cancer vasculature and normal vasculature. We developed a tubulin fractionation assay that enables evaluation of tubulin polymerisation changes in peripheral blood mononuclear cell samples (PBMCs) obtained from patients treated with BNC105. The assay capitalizes on the mass differences between polymerized and depolymerised tubulin to separate each fraction in a density gradient by ultracentrifugation. Experimental conditions during the extraction process were optimised to maintain the polymerisation state of tubulin. This optimisation was carried out in cell lines and PBMCs from healthy volunteers. Critical parameters include temperature, GTP concentration, DMSO concentration, pH, and buffer density. Tubulin monomer and polymer extracts were resolved using SDS-PAGE and transferred onto nitrocellulose membranes followed by detection using βI tubulin specific antibodies. The tubulin bands were analysed by densitometry and normalized to an actin loading control. Following exposure of cell lines or PBMCs to tubulin polymerisation inhibitors for a period of 60 minutes, the depolymerised tubulin fraction increased, while the polymerised tubulin fraction decreased. Conversely, exposure to tubulin polymerizing agents caused a greater proportion of tubulin to appear in the polymerized fraction. We used this method to evaluate the tubulin polymerisation status in PBMC samples obtained from cancer patients treated with BNC105. Samples were obtained prior to dosing and at 1, 2, 3-5, 7, and 24 hours post-dosing. Based on the data obtained, BNC105 treatment caused an 80% reduction in the polymerised tubulin fraction at the 1, 2 and 3-5 hour sampling time points. The amount of tubulin in the polymerised fraction returned to pre-dose levels by the 7 hour post-dose time point. The pharmacokinetic profile of BNC105 shows a steep curve reaching Cmax at around 20 minutes post-administration, followed by plasma clearance with no BNC105 detectable after the 7 hour time point. The pharmacokinetics of BNC105 supports our tubulin depolymerisation data with respect to response time and the subsequent recovery period. These results were reproducible, and have shown a similar pattern in all patients tested to date. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2768.</jats:p

Abstract 3543: Anti-tumor activity of the vascular disruption agent BNC105 in models of mesothelioma and lung cancer

Abstract BNC105 is a small molecule that exerts anti-cancer activity through disruption of tubulin polymerisation. BNC105 is unique in its ability to selectively target activated endothelial cells usually found in tumor blood vessels. A prodrug formulation, BNC105P, is under evaluation in phase II clinical trials for mesothelioma and renal cancer as a 10min IV infusion. In a xenograft model utilizing the human lung cancer cell line Calu-6, BNC105P caused significant tumor vascular disruption following a single dose. Similar levels of vascular disruption were observed in xenograft tumors arising from subcutaneous injection of the human mesothelioma cell line MSTO-211H. In addition, near complete vascular disruption was observed in lung metastatic lesions in a syngeneic kidney orthotopic model using the murine renal cancer cell line RENCA. We conducted preclinical evaluations that provide evidence of the activity of BNC105 in animal models of mesothelioma and lung cancer as a monotherapy and in combination with current standard of care therapies. Tumor doubling time, tumor growth delay and animal survival were assessed. Animal survival end points were defined in terms of tumor volume, with animals bearing tumors &amp;gt;2000mm3 being euthanised. Cisplatin and Gemcitabine are currently used in first line treatment of non small cell lung cancer. The effect of BNC105P treatment as a monotherapy or in combination with these two agents was investigated in a xenograft model of lung cancer. Mice carrying subcutaneous Calu-6 tumors were treated with two BNC105P doses 1 week apart as a monotherapy or in combination with Cisplatin or Gemcitabine. Combined therapy with BNC105P and Cisplatin resulted in reduced tumor growth. Tumor doubling time for the combination was 27 days, whereas the doubling time for treatment with BNC105P or Cisplatin monotherapies was 20 days and 10 days respectively. The tumor growth delay seen with the combination was 17 days greater compared to cisplatin alone with all animals in the combination arm showing tumor regression during this period. Significant increases in survival were also observed with 100% of animals in the combination group surviving, compared to 33% in the BNC105P and 0% in the Cisplatin treatment groups. In combination with Gemcitabine, BNC105P conferred tumor growth delay that was 3 days greater than BNC105P monotherapy and 17 days greater than Gemcitabine monotherapy. The survival rate in the combination group was 55% compared to 33 % in BNC105P monotherapy group and 0% in the Gemcitabine only group. Pemetrexed in combination with Cisplatin is the currently approved first line treatment for mesothelioma. The therapeutic potential of BNC105P was compared to Cisplatin and Pemetrexed in a xenograft model of the human mesothelioma cell line MSTO-211H. Compared to control, BNC105P increased survival rate by 40% with Cisplatin and Pemetrexed monotherapies increasing survival rates by 20%. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3543. doi:10.1158/1538-7445.AM2011-3543</jats:p

Abstract 2774: Anti-cancer activity of the tumor-selective, hypoxia-inducing, agent BNC105 in platinum resistant ovarian cancer

Abstract BNC105 is a small molecule that exerts anti-cancer activity through disruption of tubulin polymerization. BNC105 is unique in its ability to selectively block tumor blood flow resulting in hypoxia and cancer cell necrosis. A prodrug formulation, BNC105P, is under evaluation in a phase I/II clinical trial for metastatic renal cell carcinoma in combination with the mTOR inhibitor Everolimus. With the aim of expanding the development of BNC105 in the clinic, we investigated its activity in preclinical models of ovarian cancer. Despite modest improvements in patient outcomes as a result of surgery or platinum based chemotherapy, the majority of ovarian cancer patients relapse and die of their disease. There is a clear unmet medical need for more effective systemic therapy. The activity of BNC105 on the human ovarian carcinoma cell line A2780 and a derived cisplatin-resistant sub-line, A2780cis, was investigated using in vitro proliferation assays. BNC105 was a potent inhibitor of proliferation in both the A2780 and A2780cis cell lines with an IC50 of 0.3 and 0.1 nM respectively. In comparison, cisplatin and carboplatin were considerably less potent against A2780 with an IC50 of 330 nM and 7822 nM respectively. Both cisplatin and carboplatin were inactive against the sub-line A2780cis (IC50 &amp;gt;5 uM). BNC105 was also a potent disruptor of tumoral blood flow in A2780cis solid tumors grown subcutaneously in Balbc nu/nu mice. Tumors in mice treated with BNC105P exhibited a clear pattern of cavitation, with the center being completely deprived of blood perfusion. A small amount of blood perfusion was evident in the periphery of these tumors. In contrast, tumors obtained from animals treated with the vehicle control displayed dense blood perfusion throughout. Based on the observation of direct proliferative activity against A2780cis cells and the disruption of blood circulation in A2780cis tumors, it was reasoned that BNC105P treatment should induce tumor growth inhibition and increase survival in animals bearing A2780cis tumors. Mice bearing A2780cis solid tumors were treated with two weekly doses of BNC105P at 24mg/kg. For comparison purposes, separate animal groups bearing A2780cis tumors were treated with either cisplatin at 4mg/kg or carboplatin at 50mg/kg. Consistent with the lack of activity exhibited by platinum agents for these cells in vitro, there was no therapeutic benefit seen in animals treated with either platinum agent. In contrast, BNC105P treatment resulted in a robust inhibition of tumor growth (81% reduction compared to control) and improved overall survival. Our data demonstrate the potential therapeutic utility of BNC105 in suppressing the growth of ovarian tumors that respond poorly to platinum based therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2774. doi:1538-7445.AM2012-2774</jats:p

Abstract 4039: Harnessing the tumor adaptive response to hypoxia to identify novel combinations of the vascular disrupting agent BNC105 with targeted therapeutics.

Abstract BNC105 is a small molecule Vascular Disrupting Agent (VDA) that exerts anti-cancer activity through selective shut-down of tumor blood vessels. A single dose of BNC105 causes a very high degree of hypoxia resulting in &amp;gt;95% necrosis 24hrs post treatment in rodent tumor models. Tumor recovery from the hypoxic stress caused by BNC105 occurs by day 2 post-treatment. We have conducted immunohistochemical analysis to identify the molecular basis driving this tumor recovery. Using the mouse renal cancer orthotopic tumor model RENCA, we demonstrated that BNC105 causes significant vascular shutdown within the tumor mass and is followed by activation of a number of proteins that have been previously shown to be involved in tumor adaptive responses to hypoxia. These responses included: upregulation of GLUT1, a potential reflection of increased cancer cell dependency on glucose metabolism, increased expression of the angiogenic drivers HIFα and VEGFA and increased phosphorylation of mTOR and 4EBP1. Furthermore, there was upregulation of PERK and phosphorylation of eIF2α, reflecting activation of the Unfolded Protein Response (UPR). Similar observations were recorded when these proteins were studied in BNC105 treated mice bearing tumors of the human renal cancer cell lines Caki-1 (VHL wild type) and A-498 (VHL mutant), suggesting that these changes are likely to be generic to renal tumors being exposed to BNC105 induced hypoxia. These observations led us to investigate the potential therapeutic benefit of combining BNC105 with agents inhibiting the function of proteins being upregulated as a result of BNC105 induced hypoxia. Rapamycin was used to inhibit mTOR signalling. Treatment of mice bearing A498 tumors with BNC105, Rapamycin or the combination demonstrated greater tumor inhibition when the combination was used. Tumor growth inhibition with Rapamycin alone was 18% and BNC105 alone 12%. The Rapamycin + BNC105 combination treatment resulted in 47% tumor growth inhibition and was statistically significant compared to the inhibition seen with the monotherapies (p&amp;gt;0.05). The potential benefit of combining BNC105 with the proteasome inhibitor Bortezomib was also investigated. Upregulation of PERK has been shown to be associated with ER stress caused by accumulation of unfolded proteins. We hypothesised that concurrent inhibition of the proteasome with Bortezomib will increase cell stress resulting in increased tumor necrosis. Treatment of RENCA tumors with BNC105 + Bortezomib resulted in increased tumor necrosis compared to animals treated with Bortezomib or BNC105 alone. This suggests that the tumor is more susceptible to these agents when in combination. These data demonstrate that BNC105 can be combined with therapies targeting tumor adaptive responses to hypoxia to yield greater anti-tumor efficacy. Citation Format: Tina C. Lavranos, Daniel J. Inglis, Donna M. Beaumont, Annabell F. Leske, Chloe K. Brown, Gabriel Kremmidiotis. Harnessing the tumor adaptive response to hypoxia to identify novel combinations of the vascular disrupting agent BNC105 with targeted therapeutics. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4039. doi:10.1158/1538-7445.AM2013-4039</jats:p

Abstract B92: The tubulin-targeting agent BNC105 potentiates the efficacy of immune checkpoint inhibitors in preclinical models of colorectal cancer

Abstract BNC105 is a tubulin depolymerisation agent. Its activity includes effects on both cancer cells and on solid tumor microvasculature. BNC105 shows evidence of strong anti-cancer efficacy in vitro and in animal models. In solid tumors its efficacy is driven by selective destruction of tumor vasculature (Vascular Disrupting Agent - VDA) and direct action on tumor cells through suppression of their proliferation. In non-proliferating blood cancers (e.g. Chronic Lymphocytic Leukaemia) BNC105 activates pro-apoptotic proteins, which mediate cancer cell death. BNC105P may be useful in the treatment of human cancers both as a monotherapy and also in combination therapies. Two separate in vivo studies were conducted to assess the potential therapeutic utility of combining BNC105 with antibodies that target PD-1 or CTLA4. The in vivo models utilised were xenografts of the murine colorectal cancer cell lines MC38 and CT26. C57/BL6 mice were inoculated subcutaneously with MC38 cells and treatment commenced when tumors reached a volume of approximately 100-150mm3. BNC105 was administered at 10 mg/kg i.v. on Day 1, 8 and 15 andanti-PD-1 antibody (Clone RMP1-14) was administered at 3.5mg/kg i.p. on Day 1, 4, 8, 12 and 16. Tumor growth inhibition was evident as early as Day 8 of the treatment period especially in the combination group compared to control group (p&amp;lt;0.05). On Day 17 of the treatment period, animals treated with BNC105 as a monotherapy experienced 40% inhibition of tumor growth, anti-PD-1 treated animals experienced 74% inhibition in tumor growth. Animals treated with the combination of BNC105+anti-PD-1 therapy experienced 97% inhibition in tumor growth. Balb/c mice were inoculated subcutaneously with CT26 cells. When tumors reached an average volume of approximately 100 - 150mm3 animals were randomised into 5 groups of 10 mice per group. BNC105 (10mg/kg) was administered on Days 1 and 8 and the anti-CTLA4 antibody (Clone 9D9) was administered at 10mg/kg i.p. on Days 2, 5, and 9.. Animals treated with the BNC105+anti-CTLA4 combination experienced greater inhibition of tumor growth compared to animals treated with either BNC105 or anti-CTLA4 alone. On Day 11 of the treatment period, animals treated with BNC105 as a monotherapy experienced 27% inhibition in tumor growth and anti-CTLA4 treated animals experienced 14% inhibition of tumor growth. Animals treated with the combination of BNC105+anti-CTLA4 experienced 70% inhibition in tumor growth. The synergistic relationship for each of the combinations was calculated as previously described (Cancer Biol Ther 2011; 12:837-45). BNC105 displayed synergy with both anti-PD-1 and CTLA4. These findings support the investigation of combining BNC105 with immune checkpoint inhibitors in further preclinical models with the view of progressing such combinations to clinical evaluation. Citation Format: Tina C. Lavranos, Donna Beaumont, Daniel Inglis, Michaela Scherer, Chloe Hawkins, Annabell F. Leske, Gabriel Kremmidiotis. The tubulin-targeting agent BNC105 potentiates the efficacy of immune checkpoint inhibitors in preclinical models of colorectal cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B92.</jats:p

Abstract B67: Induction of tumor hypoxia synergises with inhibitors of tumor metabolic and angiogenic adaptive responses to suppress tumor growth and enhance host survival.

Abstract BNC105 is a compound that exerts an anti-cancer action through selective destruction of tumor blood vessels. A single IV dose of BNC105 causes a very high degree of tumor hypoxia leading to &amp;gt;95% necrosis in rodent models. Despite the dramatic tumor necrosis, tumor recovery becomes evident by day 2 following BNC105 treatment. We conducted immunohistochemical, in vitro and in vivo studies to identify the cellular and molecular basis driving tumor recovery from the significant vascular destruction caused by BNC105. Renal cancer cell lines treated with BNC105 exhibited increased expression of HIFα, VEGFA and increased phosphorylation of mTOR and 4EBP1. These observations led us to investigate the potential therapeutic benefit of combining BNC105 with agents inhibiting the signalling pathways corresponding to these proteins. We hypothesised that targeting the mTOR pathway and VEGF driven revascularisation of tumors with the mTOR inhibitor Everolimus or pan-VEGFR inhibitor Pazopanib in combination with BNC105 would lead to greater therapeutic benefit. A panel of renal cancer cell lines, including VHL mutant and VHL wild type, were shown under both normoxic and hypoxic conditions to express high levels of VEGFA. Culturing these cell lines with the mTOR inhibitor Everolimus significantly reduced VEGFA expression and decreased phosphorylation of p70S6K. These findings demonstrate that Everolimus effectively curtails VEGFA signalling and is appropriate to combine with BNC105 therapy. We tested the potential combinatorial benefit of BNC105 + Everolimus in vivo using the mouse renal cancer cell line RENCA. Tumor Growth Inhibition (TGI) with Everolimus alone was 46% and BNC105 alone 18%. The BNC105 + Everolimus combination treatment resulted in 73% TGI and was statistically significant compared to the inhibition seen with the monotherapies (p&amp;lt;0.05). Similarly in a human renal carcinoma cell line xenograft (Caki-1), TGI with Everolimus alone was 23% and BNC105 25% alone and in combination increased to 46% displaying additive benefit. The potential benefit of combining BNC105 with the pan-VEGFR inhibitor Pazopanib was also investigated. We similarly hypothesised that tumor recovery from the BNC105 induced hypoxia/necrosis insult could be curtailed through inhibition of VEGF receptors. Treatment of RENCA tumors with BNC105 alone resulted in 21% TGI and Pazopanib alone 19%. In combination the TGI was 46%. Furthermore, survival was significantly increased (p=0.0001) than animals treated with monotherapies. These data demonstrate that BNC105 can be combined with Everolimus or Pazopanib, to yield greater anti-tumor efficacy in renal cancer. A randomised Phase II trial evaluating the potential benefit of combining BNC105 with Everolimus in patients with metastatic renal cancer has finished accrual and expected to yield results in the first half of 2014. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):B67. Citation Format: Daniel J. Inglis, Tina C. Lavranos, Donna M. Beaumont, Annabell F. Leske, Chloe K. Brown, Gabriel Kremmidiotis. Induction of tumor hypoxia synergises with inhibitors of tumor metabolic and angiogenic adaptive responses to suppress tumor growth and enhance host survival. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr B67.</jats:p

Abstract 4115: BNC420 is a novel VEGFR3 selective inhibitor, which unlike the pan-VEGFR inhibitor Sunitinib, suppresses lymphatic metastasis in a model of metastatic melanoma

Abstract The dissemination of cancer cells through the lymphatic system is a well-known route of metastasis. Studies have implicated the hypoxic tumor microenvironment as an instigator of tumor cell invasion and metastatic spread. The use of angiogenesis inhibitors targeting VEGFR2 has been reported in preclinical models to augment metastasis through increased hypoxia within the tumor microenvironment. We have sought to identify a selective VEGFR3 targeting compound that will inhibit metastatic spread through lymphatic dissemination without the concomitant hypoxia induced metastasis effect associated with VEGFR2 targeting. BNC420 is a novel small molecule inhibitor of VEGFR3 phosphorylation that displays significant selectivity over VEGFR2. In phosphorylation detection ELISA based assays BNC420 exhibited a 25 fold greater potency for VEGFR3 over VEGFR2 with IC50 of 30nM and 744nM respectively. We investigated the activity of BNC420 and the pan-VEGFR inhibitor Sunitinib in the B16F10 murine melanoma model. This model involves the intradermal administration of B16F10 melanoma cells in the mouse ear resulting in the formation of a primary lesion, metastatic satellite lesions (in-transit) and regional lymph node metastasis. While Sunitinib slowed the growth of the primary lesion, as expected, it did not inhibit the development of lymph node metastasis and appeared to enhance the development of in-transit satellite lesions by 38% compared to its corresponding vehicle control. In contrast, only 20% of animals treated with BNC420 exhibited in-transit lesions compared to 53% in the corresponding vehicle control. Furthermore, a 50% reduction in the number of metastasis to the draining lymph nodes was seen in BNC420 treated animals. The lymph node metastatic lesions were significantly smaller compared to control and in some instances only detected under microscopic examination. Melanoma lesions in animals treated with BNC420 were devoid of peritumoral lymphatic vessels. These results demonstrate that selective inhibition of VEGFR3 utilising the novel small molecule BNC420 effectively inhibits the development of both in-transit metastatic lesions and lymph node metastasis while being unencumbered by the pro-metastatic effects that can accompany VEGFR2 inhibition through the induction of tumor hypoxia. Citation Format: Annabell Leske, Tina Lavranos, Donna Beaumont, Chloe Brown, Darham Paul, Daniel Inglis, Michaela Scherer, Andrew Harvey, Gabriel Kremmidiotis. BNC420 is a novel VEGFR3 selective inhibitor, which unlike the pan-VEGFR inhibitor Sunitinib, suppresses lymphatic metastasis in a model of metastatic melanoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4115. doi:10.1158/1538-7445.AM2015-4115</jats:p