A multi-pronged approach to Immuno-Oncology

Immuno-oncology has revolutionized cancer treatment How can it be improved further? Immunotherapy + ? Combine Immune-activation with Reducing metastases Reducing relapses Synergy with chemotherapy immunotherapy

Aryl hydrocarbon Receptor (AhR) Ligand-activated transcription factor Strongly up-regulated in many cancers Constantly activated in tumor Expression of cancer-related genes Constant activation by agonist ligands Including: Tryptophan metabolites Immune suppression IDO/TDO expression Induction of Tregs Anti-inflammation Pro-metastatic Cancer Stem Cell induction Anti-apoptotic Oncogene activation Angiogenesis the environmental ligand's that activate the AHR have already been linked to increased breast cancer incidence. Now we have an idea about the mechanism how. Those chemicals would include dioxins, PCBs, and polycyclic aromatic hydrocarbons. in normal cells there are extremely low levels of AHR in the cytoplasm. Once activated by environmental ligand's it becomes an active transcription factor translocate to the nucleus and regulating transcription of probably a dozen or more genes. CYP1A1 and CYP1B1 are classic AHR responsive genes. In the cancer cells, the transcription factor is expressed at huge levels, sometimes 50 times more than in any normal cell, and it is constructively active. we now have strong evidence that the tumors produce amino acid metabolites that activate the AHR. These are the endogenous ligands. Excessive production of these metabolites has previously been associated with breast cancer. Now we know why. They constantly activate AhR.

Blocking by antagonist ligand CB7993113 - AhR inhibitor CB7993113 binding prevents: AhR from entering the nucleus Cancer-related gene transcription Hercules has developed CB7993113, a small molecule AhR inhibitor CB CB7993113 Effects Immune activation IDO/TDO inhibition Reduction of Tregs Reduction of Treg IL-10 Pro-inflammation Anti-metastatic Cancer stem cell inhibition Cancer cell apoptosis Anti-Angiogenic CB Blocking by antagonist ligand CB7993113 the environmental ligand's that activate the AHR have already been linked to increased breast cancer incidence. Now we have an idea about the mechanism how. Those chemicals would include dioxins, PCBs, and polycyclic aromatic hydrocarbons. in normal cells there are extremely low levels of AHR in the cytoplasm. Once activated by environmental ligand's it becomes an active transcription factor translocate to the nucleus and regulating transcription of probably a dozen or more genes. CYP1A1 and CYP1B1 are classic AHR responsive genes. In the cancer cells, the transcription factor is expressed at huge levels, sometimes 50 times more than in any normal cell, and it is constructively active. we now have strong evidence that the tumors produce amino acid metabolites that activate the AHR. These are the endogenous ligands. Excessive production of these metabolites has previously been associated with breast cancer. Now we know why. They constantly activate AhR. CB

CB7993113 Improves Survival in Immunodeficient Murine Cancer Model 50 mg/kg/day oral. n=12 mice per group Human oral squamous carcinoma cell line, Cal27, in an orthotopic xenograft (tongue) 90% survival AhR blocking is also effective in models without active immune system 25% survival

CB7993113 Blocks Human Breast Cancer Metastasis in a Zebrafish Model 24 Hours after injection of human RFP-MDA-MB-231 TNBC cells Metastasis/Fish 0.1% DMSO (Control) n=12 1 uM CB7993113 n=6 Treated for how long? 24 hours These results in zebrafish show metastasis blocking effect and are also indicative of lack of toxicity of CB7993113

CB7993113 reduces tumor burden in immunodeficient murine TNBC model Total Tumor Burden After Intracardiac Injection of MDA-MB-231-BO Bioluminescence Daily injection of 25mg/kg ip CB7993113 MDA-MB-231BO human TNBC tumor cells

CB7993113 Clinical Development Correlation between cancer aggression and AhR expression Focus on aggressive cancers TNBC Glioblastoma Contrary to most immuno-oncology approaches, CB7993113 may be effective in colorectal cancers CB7993113 could be combined with: Check-point inhibitors Cancer vaccines Chemotherapy Target Profile: Oral, non toxic, chronic treatment of epithelial cancers

Hercules Team Based in Amsterdam, Netherlands, since 2013 Spin off from Boston University Bart Wuurman CEO Previously: CEO Lanthio Pharma, AM-Pharma Prof David Sherr CSO Scientific founder, based in Boston Sjoerd Wadman Development Director Previously Lanthio Pharma, GSK

Finance Shareholding: DDF Ventures; 60% shareholder Current funding: Eurostars Grant funding for additional in-vivo efficacy studies pre-clinical development of CB7993113 Funding need: €10m Build team Phase I and IIa

Summary AhR inhibition with CB7993113: Anti-tumor immune activation Reduced metastatic behaviour Reduced potential for relaps Increased efficacy for chemotherapy Clinical focus on aggressive tumors

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