Full Tumor Regression Research Articles

Velcrin molecular glues induce apoptosis in glioblastomas with high PDE3A and SLFN12 expression.

Velcrins are molecular glues that kill cells by inducing the formation of a protein complex between the RNase SLFN12 and the phosphodiesterase PDE3A. Formation of the complex activates SLFN12, which cleaves tRNALeu(TAA) and induces apoptosis. Velcrins such as the clinical investigational compound, BAY 2666605, were found to have activity across multiple solid tumor cell lines from the cancer cell line encyclopedia, including glioblastoma cell lines. We therefore aim to characterize velcrins as novel therapeutic agents in glioblastoma. PDE3A and SLFN12 expression levels were measured in glioblastoma cell lines, the Cancer Genome Atlas (TCGA) tumor samples, and tumor neurospheres. Velcrin-treated cells were assayed for viability, induction of apoptosis, cell cycle phases, and global changes in translation. Transcriptional profiling of the cells was obtained. Xenograft-harboring mice treated with velcrins were also monitored for survival. We identified several velcrin-sensitive glioblastoma cell lines and 4 velcrin-sensitive glioblastoma patient-derived models. We determined that BAY 2666605 crosses the blood-brain barrier and elicits full tumor regression in an orthotopic xenograft model of GB1 cells. We also determined that the velcrins BAY 2666605 and BRD3800 induce tumor regression in subcutaneous glioblastoma PDX models. Velcrins have antitumor activity in preclinical models of glioblastoma, warranting further investigation as potential therapeutic agents.

EXTH-38. VELCRIN MOLECULAR GLUES INDUCE CELL DEATH IN GLIOBLASTOMAS WITH HIGH PDE3A AND SLFN12 EXPRESSION

Abstract Velcrins are molecular glues that kill cells by inducing the formation of a protein complex between the RNase, SLFN12, and the phosphodiesterase, PDE3A. Formation of the complex activates SLFN12, which cleaves tRNALeu(TAA) and induces apoptosis. Velcrins such as the clinical investigational compound, BAY 2666605, were found to have activity across multiple solid tumor cell lines from the Cancer Cell Line Encyclopedia (CCLE), including glioblastoma cell lines. We therefore aimed to characterize velcrins as novel therapeutic agents in glioblastoma. We measured PDE3A and SLFN12 expression levels in commercially available glioblastoma cells as well as glioblastoma neurospheres from the Center for Patient Derived Models at Dana Farber Cancer Institute. We identified velcrin-sensitive glioblastoma cell lines and neurospheres, and determined that velcrin response is highly correlated with PDE3A expression. We also determined that sensitive cells exhibit a reduction in nascent polypeptide formation upon velcrin treatment, which is rescued by CRISPR-induced knockout of PDE3A. To assess velcrin efficacy in vivo, we measured central nervous system and plasma levels of the velcrin, BAY 2666605, and determined that it freely crosses the blood brain barrier. We also determined that twice daily dosing of BAY 2666605 elicits full tumor regression in an orthotopic glioblastoma xenograft model, with sustained complete responses after treatment withdrawal. Lastly, we demonstrated that the velcrins BAY 2666605 and BAY 2501293 induce tumor regression in biomarker-positive subcutaneous glioblastoma patient-derived xenograft models. In summary, our data demonstrates that velcrins have anti-tumor activity in preclinical models of glioblastoma, warranting further investigation as potential therapeutic agents.

Abstract 486: Characterization of a novel series of highly selective PARP7 inhibitors

Abstract PARP7 is a cellular stress-induced enzyme that adds mono-ADP-ribose groups to a variety of substrate proteins thereby regulating their function. One such substrate is the kinase TBK1 which regulates activity of cGAS-STING and RIG-I nucleic acid sensing pathways. Up-regulation of PARP7 expression in cancers applies a brake to cytosolic nucleic acid sensing and the Type I interferon response. This creates an immunosuppressive tumor microenvironment (TME) leading to faster tumor growth. In addition, inhibition of PARP7 has also been shown to directly arrest growth in a subset of cancer cells via the promotion of a senescence phenotype, inhibition of autophagy and regulation of metabolism. Through the use of structure-based drug design we describe the characterization of potent and selective inhibitors of PARP7. These molecules block viability of a subset of immortalized and primary cancer cells in 2D culture. We have observed potent cGAS-STING independent growth arrest in a number of cancer cell lines harboring a KRAS G12C mutation, and studies combining PARP7 inhibitors with commercially available KRAS G12C inhibitors yield additional in vitro efficacy. Recent studies have shown that inhibitors targeting oncogenic RAS pathway signaling can activate ULK1/2-mediated autophagy as an adaptive treatment resistance mechanism. Hence, our data support further studies investigating the use of PARP7 inhibitors in KRAS-driven cancers either as single agents or in combination with mutant KRAS inhibitors. Our molecules also stimulate robust induction of type I interferon (IFNβ) and downstream chemokines such as CXCL10 in a cGAS-STING pathway dependent manner in a range of cancer cells and 3D organoids. This induction is augmented by combining agonists of cGAS-STING and RIG-I pathways with PARP7 inhibitors. In addition, PARP7 inhibitors work in concert with exogenous DNA-damaging agents such as chemotherapeutics and X-ray radiation to induce the type I interferon response in cancer cells. Overall, these data highlight the opportunity to utilize PARP7 inhibitors in cancers where high genomic instability leads to aberrant cytosolic nucleic acid levels or in concert with exogenous DNA-damaging agents. Our PARP7 inhibitors exhibit highly desirable physico-chemical and in vitro ADME properties coupled with an exceptional in vitro safety profile which translates to best-in-class rodent PK. In a KRAS G12C lung cancer xenograft model (NCI-H1373), twice daily oral administration of an exemplar PARP7 inhibitor drives rapid and full tumor regression in a dose-dependent manner. The anti-tumor effects correlate with robust tumor PK/PD upon measurement of tumor IFNβ and CXCL10. In summary, we describe the characterization of a lead series of highly potent and selective PARP7 inhibitors which demonstrate excellent in vitro ADMET and in vivo PK properties leading to best-in-class anti-tumor efficacy in a KRAS-driven lung cancer xenograft model. Citation Format: Phillip M. Cowley, Barry E. McGuinness, Gillian M. Campbell, Alan Wise. Characterization of a novel series of highly selective PARP7 inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 486.

Dual activity of PD-L1 targeted Doxorubicin immunoliposomes promoted an enhanced efficacy of the antitumor immune response in melanoma murine model

BackgroundThe immunomodulation of the antitumor response driven by immunocheckpoint inhibitors (ICIs) such as PD-L1 (Programmed Death Ligand-1) monoclonal antibody (α-PD-L1) have shown relevant clinical outcomes in a subset of patients. This fact has led to the search for rational combinations with other therapeutic agents such as Doxorubicin (Dox), which cytotoxicity involves an immune activation that may enhance ICI response. Therefore, this study aims to evaluate the combination of chemotherapy and ICI by developing Dox Immunoliposomes functionalized with monovalent-variable fragments (Fab’) of α-PD-L1.ResultsImmunoliposomes were assayed in vitro and in vivo in a B16 OVA melanoma murine cell line over-expressing PD-L1. Here, immune system activation in tumor, spleen and lymph nodes, together with the antitumor efficacy were evaluated. Results showed that immunoliposomes bound specifically to PD-L1+ cells, yielding higher cell interaction and Dox internalization, and decreasing up to 30-fold the IC50, compared to conventional liposomes. This mechanism supported a higher in vivo response. Indeed, immunoliposomes promoted full tumor regression in 20% of mice and increased in 1 month the survival rate. This formulation was the only treatment able to induce significant (p < 0.01) increase of activated tumor specific cytotoxic T lymphocytes at the tumor site.ConclusionPD-L1 targeted liposomes encapsulating Dox have proved to be a rational combination able to enhance the modulation of the immune system by blocking PD-L1 and selectively internalizing Dox, thus successfully providing a dual activity offered by both, chemo and immune therapeutic strategies.Graphic

Π electron-stabilized polymeric micelles potentiate docetaxel therapy in advanced-stage gastrointestinal cancer

Gastrointestinal (GI) cancers are among the most lethal malignancies. The treatment of advanced-stage GI cancer involves standard chemotherapeutic drugs, such as docetaxel, as well as targeted therapeutics and immunomodulatory agents, all of which are only moderately effective. We here show that Π electron-stabilized polymeric micelles based on PEG-b-p(HPMAm-Bz) can be loaded highly efficiently with docetaxel (loading capacity up to 23 wt%) and potentiate chemotherapy responses in multiple advanced-stage GI cancer mouse models. Complete cures and full tumor regression were achieved upon intravenously administering micellar docetaxel in subcutaneous gastric cancer cell line-derived xenografts (CDX), as well as in CDX models with intraperitoneal and lung metastases. Nanoformulated docetaxel also outperformed conventional docetaxel in a patient-derived xenograft (PDX) model, doubling the extent of tumor growth inhibition. Furthermore, micellar docetaxel modulated the tumor immune microenvironment in CDX and PDX tumors, increasing the ratio between M1-and M2-like macrophages, and toxicologically, it was found to be very well-tolerated. These findings demonstrate that Π electron-stabilized polymeric micelles loaded with docetaxel hold significant potential for the treatment of advanced-stage GI cancers.

Bivalent therapeutic vaccine against HPV16/18 genotypes consisting of a fusion protein between the extra domain A from human fibronectin and HPV16/18 E7 viral antigens.

BackgroundIn vivo targeting of human papillomavirus (HPV) derived antigens to dendritic cells might constitute an efficient immunotherapeutic strategy against cervical cancer. In previous works, we have shown that the extra domain A from murine fibronectin (mEDA) can be used to target antigens to toll-like receptor 4 (TLR4) expressing dendritic cells and induce strong antigen-specific immune responses. In the present study, we have produced a bivalent therapeutic vaccine candidate consisting of the human EDA (hEDA) fused to E7 proteins from HPV16 and HPV18 (hEDA-HPVE7-16/18) and evaluate its potential as a therapeutic vaccine against cervical cancer.Materials and methodsRecombinant fusion proteins containing HPV E7 proteins from HPV16 and HPV18 virus subtypes fused to hEDA were produced and tested in vitro on their capacity to bind TLR4 and induce the production of tumor necrosis factor-α or interleukin (IL)-12 by human monocytes and dendritic cells. The immunogenicity and potential therapeutic activity of the vaccine in combination with cisplatin or with the TLR3 agonist molecules polyinosinic‐polycytidylic acid (Poly IC) or Poly ICLC was evaluated in mice bearing subcutaneous or genital orthotopic HPV16 TC-1 tumors.ResultshEDA-HPVE7-16/18 prototype vaccine binds human TLR4 and stimulate TLR4-dependent signaling pathways and IL-12 production by human monocyte-derived dendritic cell. Vaccination with hEDA-HPVE7-16/18 induced strong HPVE7-specific Cytotoxic T lymphocyte (CTL) responses and eliminated established tumors in the TC-1-based tumor model. The antitumor efficacy was significantly improved by combining the fusion protein with cisplatin or with the TLR-3 ligand Poly IC and especially with the stabilized analog Poly ICLC. Moreover, hEDA-HPVE7-16/18+Poly ICLC induced full tumor regression in 100% of mice bearing orthotopic genital HPV tumors.ConclusionOur results suggest that this therapeutic vaccine formulation may be an effective treatment for cervical tumors that do not respond to current therapies.

1200P - Tumour stroma targeting and modulation by OMTX705 ADC, a novel and potent immunotherapeutic treatment of solid tumours

BackgroundTumor stroma represents 20-60% of solid tumor mass and plays a key role in promotion, invasiveness and metastasis. FAP-expressing CAFs, the predominant stroma cell type, are involved in tumor immune response. A novel antibody-drug conjugate, OMTX705, was generated through CYS-based conjugation of a new anti-FAP humanized antibody to a novel cytolysin using an optimized vcPABA linker. MethodsIn vivo studies were performed in patient-derived xenograft models for NSCL cancer in humanized mice. Tumor volume and animal weight were monitored over 4-week treatment with OMTX705 administered intravenously at different doses, alone or combined with Pembrolizumab. FACS and IHC analysis of markers for immune cells, CAFs, and cell proliferation or apoptosis, were performed on tumor samples to study OMTX705 effect on tumor stroma and elucidate its mechanism of action. In vitro assays were performed to support in vivo data. OMTX705 stability in blood was determined after i.v administration in CD1 mice. ResultsOMTX705 showed 100% tumor growth inhibition and higher activity than Pembrolizumab in the humanized PDX model for NSCL cancer without weight loss. Full tumor regression was found in 10% of mice treated with OMTX705 alone and 20% in combination with Pembrolizumab. A significant delay in tumor recurrence was observed. OMTX705 was 100% stable in bloodstream after 7 days. FACS analysis evidenced a significant increase in CD8(+) T cell tumor infiltration. Results from in vitro studies and IHC analysis of tumors identified CAFs as highly specific reservoir cells for OMTX705 from which the high potent cytolysin payload is released to induce apoptosis of adjacent tumor cells and CD8(+) T cell immunomodulation. ConclusionsOMTX705 alone induces tumor shrinkage, and full regression with Pembrolizumab, through highly specific, CAF-dependent and long-lasting modulation of tumor stroma. This novel mechanism of action makes OMTX705 a potent and innovative strategy for NSCL cancer treatment. Considering FAP is expressed in a wide array of carcinomas, OMTX705 represents a highly promising and well-tolerated candidate to treat solid tumors with low response to anti-PD1 immunotherapies. Legal entity responsible for the studyOncomatryx Biopharma, S.L. FundingOncomatryx Biopharma, S.L. DisclosureM. Fabre: Full / Part-time employment: Oncomatryx Biopharma, S.L. C. Ferrer: Full / Part-time employment: Oncomatryx Biopharma, S.L. S. Domínguez-Hormaetxe: Full / Part-time employment: Oncomatryx Biopharma, S.L. R. Kontermann: Advisory / Consultancy: Oncomatryx Biopharma, S.L.; Advisory / Consultancy: SunRock Biopharma, S.L.; Advisory / Consultancy: Roche. K. Pfizenmaier: Advisory / Consultancy: Oncomatryx Biopharma, S.L.; Advisory / Consultancy: SunRock Biopharma, S.L. O. Seifer: Research grant / Funding (institution): Oncomatryx Biopharma, S.L. M.D. Vivanco: Research grant / Funding (institution): Oncomatryx Biopharma, S.L. S. Lee: Research grant / Funding (institution): Oncomatryx Biopharma, S.L. P. López-Casas: Full / Part-time employment: Bioncotech Therapeutics, S.L. M. Abbas: Full / Part-time employment: Tube Pharmaceuticals GmBH. W. Richter: Advisory / Consultancy: Oncomatryx Biopharma, S.L.; Shareholder / Stockholder / Stock options, Full / Part-time employment: Tube Pharmaceuticals GmBH. L. Simon: Honoraria (self), Leadership role, Travel / Accommodation / Expenses, Shareholder / Stockholder / Stock options, Licensing / Royalties, Full / Part-time employment, Patents: Oncomatryx Biopharma, S.L.; Honoraria (self), Leadership role, Travel / Accommodation / Expenses, Shareholder / Stockholder / Stock options, Licensing / Royalties, Full / Part-time employment, Patents: Patia Biopharma, SA de CV; Honoraria (self), Leadership role, Travel / Accommodation / Expenses, Shareholder / Stockholder / Stock options, Full / Part-time employment: Patia Europe, S.L.; Shareholder / Stockholder / Stock options, Full / Part-time employment: PatiaCan, SA de CV; Honoraria (self), Leadership role, Shareholder / Stockholder / Stock options, Full / Part-time employment: Permedika Entrepeneurs; Travel / Accommodation / Expenses, Shareholder / Stockholder / Stock options: SunRock Biopharma, S.L.; Licensing / Royalties, Patents: Quimatryx, S.L.; Shareholder / Stockholder / Stock options: Stemcell Therapeutics, S.L.; Shareholder / Stockholder / Stock options: Helenes Ventures, S.L.; Shareholder / Stockholder / Stock options: Nellum Corp. M. Hidalgo: Advisory / Consultancy: Oncomatryx Biopharma, S.L.; Honoraria (self), Advisory / Consultancy, Shareholder / Stockholder / Stock options: Champions Oncology; Honoraria (self), Advisory / Consultancy, Shareholder / Stockholder / Stock options: Pharmacite Biotech; Shareholder / Stockholder / Stock options: BioOncotech; Shareholder / Stockholder / Stock options: Nelum; Honoraria (self), Advisory / Consultancy, Research grant / Funding (self): Pfizer; Honoraria (self), Advisory / Consultancy: Novartis; Honoraria (self): MSD Oncology; Honoraria (self), Advisory / Consultancy, Research grant / Funding (self), Travel / Accommodation / Expenses: Celgene; Honoraria (self), Advisory / Consultancy, Research grant / Funding (self), Travel / Accommodation / Expenses: BiolineRX; Honoraria (self), Advisory / Consultancy: Roche/Genentech; Honoraria (self), Advisory / Consultancy: SOBI; Honoraria (self), Advisory / Consultancy: Agenus; Honoraria (self), Advisory / Consultancy: Erytech Pharma; Advisory / Consultancy: Merck; Advisory / Consultancy, Research grant / Funding (institution): AstraZeneca/MedImmune; Advisory / Consultancy, Travel / Accommodation / Expenses: Menarini; Advisory / Consultancy, Travel / Accommodation / Expenses: Bayer; Research grant / Funding (self): Bicycle Therapeutics; Research grant / Funding (self): Asana Biosciences.

Daratumumab displays in vitro and in vivo anti-tumor activity in models of B-cell non-Hodgkin lymphoma and improves responses to standard chemo-immunotherapy regimens.

CD38 is expressed in several types of non-Hodgkin lymphoma (NHL) and constitutes a promising target for antibody-based therapy. Daratumumab (Darzalex) is a first-in-class anti-CD38 antibody approved for the treatment of relapsed/refractory (R/R) multiple myeloma (MM). It has also demonstrated clinical activity in Waldenström macroglobulinaemia and amyloidosis. Here, we have evaluated the activity and mechanism of action of daratumumab in preclinical in vitro and in vivo models of mantle cell lymphoma (MCL), follicular lymphoma (FL) and diffuse large B-cell lymphoma (DLBCL), as monotherapy or in combination with standard chemo-immunotherapy. In vitro, daratumumab engages Fc-mediated cytotoxicity by antibody-dependent cell cytotoxicity and antibody-dependent cell phagocytosis in all lymphoma subtypes. In the presence of human serum, complement-dependent cell cytotoxicity was marginally engaged. We demonstrated by Selective Plane Illumination Microscopy that daratumumab fully penetrated a three-dimensional (3D) lymphoma organoid and decreased organoid volume. In vivo, daratumumab completely prevents tumor outgrowth in models of MCL and FL, and shows comparable activity to rituximab in a disseminated in vivo model of blastic MCL. Moreover, daratumumab improves overall survival (OS) in a mouse model of transformed CD20dim FL, where rituximab showed limited activity. Daratumumab potentiates the antitumor activity of CHOP and R-CHOP in MCL and FL xenografts. Furthermore, in a patient-derived DLBCL xenograft model, daratumumab anti-tumor activity was comparable to R-CHOP and the addition of daratumumab to either CHOP or R-CHOP led to full tumor regression. In summary, daratumumab constitutes a novel therapeutic opportunity in certain scenarios and these results warrant further clinical development.

Abstract 1558: Therapeutic targeting of cell surface plectin induces anti-cancer immune response and pancreatic cancer regression

Abstract Background: Pancreatic ductal adenocarcinoma (PDA) is the 3rd deadliest cancer, diagnosed typically in advanced stages, with only an 8% 5-year survival rate, thus demonstrating the need for novel therapeutic approaches that significantly enhance chemo- and/or immune-therapy. Our team previously identified a promising functional target for cancer therapy in PDA, cell surface plectin 1 (CSP1) that is aberrantly expressed on PDA cells and thus a cell surface-associated biomarker of cancer. CSP1 expression first becomes apparent in high grade dysplasias, remaining high in early and advanced cancers and in metastases. Our first-in-human imaging trial in PDA patients using a CSP1-targeted imaging agent revealed that CSP is an available target and accessible for binding, a potentially a target for cancer therapy. We hypothesized that a monoclonal antibody (mAb) against CSP1 could lead to novel pancreatic cancer treatment options, thus, we developed a therapeutic mAb, e.g., ZB131, representing a first-in-class antibody selectively targeting CSP1. Methods: ZB131 is a humanized mAb targeted against human plectin 1 (rhSec8) that also binds murine CSP1. ZB131 affinity and its effect on cancer cells including proliferation, cytotoxicity, and migration were tested in vitro using saturation binding, SRB-based survival assays, flow cytometry, and migration assays on various pancreatic cell types and homeostatic “normal” controls. In vivo validation was performed using two nu/nu mouse models bearing subcutaneous MiaPACA2 or Yapc PDA cells, and a syngeneic KPC-derived tumor model to also evaluate immune responses to tumors treated with ZB131 or IgG control. Results: ZB131 exhibits high specificity and high affinity (0.4±0.1nM) to CSP1, and functionally induces G0 growth arrest followed by necrotic cell death of PDA cells in culture, and is synergistic with gemcitabine resulting in a 50-fold decrease in IC50. In vivo, in subcutaneous xenograft models, ZB131 monotherapy decreased PDA tumor volume 5-fold as compared to controls, and in combination with cisplatin resulted in sustained tumor reduction with greater than 85% tumor necrosis. In subcutaneous syngeneic PDA models, ZB131 induced complete tumor regression within 35 days mediated by an anti-tumor immune response as upon tumor rechallenge, full tumor regression was again achieved without additional ZB131 therapy. Leukocyte complexity analysis of regressing PDA tumors versus controls revealed an ~3-fold increase in effector and central memory T cells. Conclusion: CSP1 is a first in class anti-cancer target expressed on the cell surface of PDA, as well as other cancers including ovarian, esophageal and head neck. ZB131, an anti-CSP1 mAb, induces tumor cell intrinsic cell death, as well as a robust anti-tumor T cell response leading to complete tumor regression indicating the potential therapeutic efficacy of ZB131 in late-stage cancers. Citation Format: Julien Dimastromatteo, Amanda Poisonnier, Samantha Perez, Lisa Coussens, Kimberly Kelly. Therapeutic targeting of cell surface plectin induces anti-cancer immune response and pancreatic cancer regression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1558.

Abstract 4479: BT8009: A bicyclic peptide toxin conjugate targeting Nectin-4 (PVRL4) displays efficacy in preclinical tumor models

Abstract We have identified a Nectin-4 targeting peptide for delivery of cytotoxic agents to Nectin-4 expressing tumors. Nectin-4 is a cell adhesion molecule, that is highly expressed in certain tumor types, including bladder, TNBC and NSCLC, but has a restricted distribution in normal tissue. Bicycles® are small (1.5kDa) fully synthetic, structurally constrained, peptide drugs that combine the affinity of antibodies with the pharmacokinetic properties of small molecules. The Bicycle phage display platform was utilised to rapidly identify a high affinity (18 nM) and highly selective bicyclic Nectin-4 binding peptide. Synthetic modification of the initial lead peptide improved affinity (0.3 nM), hydrophilicity and stability. The optimised peptide is conjugated through an inert sarcosine spacer chain and a cleavable linker to the toxin MMAE, to form the Bicycle Toxin Conjugate (BTC) BT8009. Once bound to cell surface Nectin-4, the linker system is cleaved by peptidases (e.g. cathepsin B) upregulated in the tumor micro environment. Full structure of BT8009 will be disclosed within the presentation. Fluorescence polarisation and surface plasmon resonance show BT8009 has low nanomolar affinity (3nM) for Nectin-4 and high selectivity (&amp;gt;1000 fold) over Nectins 1-3, and the 5 nectin-like, family members. Good affinity for the cognate native protein is maintained across the preclinical safety species. High content imaging of cultures of MDA-MB-468 cells, demonstrates binding of the BTC to cell membrane. In vivo, BT8009 is well tolerated in mouse and rat and shows regressions across a range of cell and patient derived xenograft models. Efficacy correlates with the expression level of Nectin-4 on the tumor cells, and the dose delivered. In the MDA-MB-468 (TNBC) CDX model full tumor regression was seen with weekly i.v. administration of 3 mg/k g i.v. (4 doses), with no tumor regrowth out to 70 days post last dose. Similar efficacy was seen in two NSCLC PDX models (adenocarcinoma and squamous cell carcinoma) with full regression attained in both. Near full regression was seen in a squamous cell esophageal cancer PDX. Larger tumors show a similar degree of sensitivity to the BTC, with rapid tumor regression from a volume of 800 mm3. As a small peptide BT8009 undergoes rapid clearance from the plasma with minimal exposure to non-targeted organs. After a single dose, MMAE has been shown to be retained within tumor tissue in excess of 60 h, at exposure levels significantly greater than corresponding plasma and other tissue levels. Bicycle Toxin Conjugates represent a novel treatment modality for nectin-4 expressing tumors with excellent efficacy in several mouse xenograft models. Citation Format: Mike Rigby, Paul Beswick, Gemma Mudd, Katerine Van Rietschoten, Liuhong Chen, Sophie M. Watcham, Heather Allen, Amy Brown, Helen Harrison, Gavin Bennett, Phil Jeffrey, Peter U. Park, Maria Koehler, Nicholas Keen. BT8009: A bicyclic peptide toxin conjugate targeting Nectin-4 (PVRL4) displays efficacy in preclinical tumor models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4479.

Primary tumor regression patterns in esophageal squamous cell cancer treated with definitive chemoradiotherapy and implications for surveillance schemes.

Purpose: The primary tumor regression patterns of patients with esophageal squamous cell carcinoma (ESCC) treated with definitive chemoradiotherapy (CRT) were investigated to determine an optimal surveillance scheme.Method: The clinical data and radiology images of patients before CRT, at completion of CRT and every 1–3 months for the subsequent 12 months or until disease progression were retrospectively reviewed to define the patterns of primary tumor regression after CRT. Survival rates were analyzed statistically in order to determine an optimal surveillance scheme.Results: A total of 82 patients were enrolled in the present study for analysis. At the first surveillance visit date at the end of CRT, a total of 21 patients achieved complete response (early-CR), 29 patients reached incomplete response (IR), 25 patients maintained stable disease (SD) and 7 patients encountered progression of disease (PD). During subsequent surveillance, a total of 14 IR patients regressed continuously to CR (later-CR), 15 patients maintained IR (early-IR) and 9 SD patients gradually regressed to IR (later-IR). At full tumor regression (FTR), a total of 21, 14, 15, 9, 16 and 7 patients were defined as early-CR, later-CR, early-IR, later-IR, SD and PD, respectively. The median FTR time for later-CR and later-IR was 7.5 and 7 weeks, respectively. The 3-year overall survival rate of the early-CR group was 85.7% (P<0.001), which was higher compared with the later-CR (16.7%), early-IR (20%), later-IR (11.1%), SD (6.3%) and PD (0%) groups.Conclusion: The early-CR following CRT is a robust prognostic predictor in patients with ESCC. To optimize the determination of tumor regression, ≥7 weeks after CRT is an optimal initial surveillance visit date. The surveillance of non-CR patients should concentrate on symptoms, nutrition and psychosocial support, rather than screening for recurrence of the disease.

Efficacy of a Covalent ERK1/2 Inhibitor, CC-90003, in KRAS-Mutant Cancer Models Reveals Novel Mechanisms of Response and Resistance.

As a critical signaling node, ERK1/2 are attractive drug targets, particularly in tumors driven by activation of the MAPK pathway. Utility of targeting the MAPK pathway has been demonstrated by clinical responses to inhibitors of MEK1/2 or RAF kinases in some mutant BRAF-activated malignancies. Unlike tumors with mutations in BRAF, those with mutations in KRAS (>30% of all cancers and >90% of certain cancer types) are generally not responsive to inhibitors of MEK1/2 or RAF. Here, a covalent ERK1/2 inhibitor, CC-90003, was characterized and shown to be active in preclinical models of KRAS-mutant tumors. A unique occupancy assay was used to understand the mechanism of resistance in a KRAS-mutant patient-derived xenograft (PDX) model of colorectal cancer. Finally, combination of CC-90003 with docetaxel achieved full tumor regression and prevented tumor regrowth after cessation of treatment in a PDX model of lung cancer. This effect corresponded to changes in a stemness gene network, revealing a potential effect on tumor stem cell reprograming. IMPLICATIONS: Here, a covalent ERK1/2 inhibitor (CC-90003) is demonstrated to have preclinical efficacy in models of KRAS-mutant tumors, which present a therapeutic challenge for currently available therapies.

СРАВНИТЕЛЬНАЯ ОЦЕНКА МОДИФИКАЦИЙ РАЗЛИЧНЫХ МЕТОДОВ ЛУЧЕВОЙ ТЕРАПИИ У БОЛЬНЫХ РАКОМ ШЕЙКИ МАТКИ

The aim of the research is to make a comparative evaluation of the results of treatment of cervical cancer in patients who had the radiation therapy with the application of the modes of sequential and parallel methods of bringing the dose to the pathological focus. The results of combined radiation therapy in 120 patients with cervical cancer were analyzed. Depending on the treatment tactics, the patients were divided into 2 groups of 60 patients who underwent a sequential or parallel method of bringing the dose to the lesion and regional metastasis routes, when the total focal dose in both cases matched the optimally required parameters – 80-85 Gr in the primary tumor and points "A" and 60 Gr in points "B". Distance radiation therapy was carried out 5 times a week, followed by a break of 3-5 weeks and a further application of the intracavitary method. In the second case, a daily distance therapy was carried out to a total focal dose of 18 Gr, after which the number of sessions per week decreased to 3 times and in some days (2 times a week) there was done an intracavitary treatment. The following results were obtained: an objective therapeutic effect was observed in 108 (90%) patients. Full tumor regression was recorded in 95 (87.9%), partial – in 12.1%, and failures were detected in 10% of women. During the three-year follow-up period, 20.0% of patients of the first group and 10.9% of the second group showed progression of the disease manifested by recurrence in the irradiation zones, recurrence+metastases and metastases of the primary tumor, usually in the inguinal and paraaortic lymph nodes, pelvic bones, liver, lungs. The median of remission duration in the first group was 20 months with a three-year survival rate of 68.0%, and in the second it was 26 and 82.0%, respectively. Complications in the form of mucous response to radiation exposure, radiation cystitis, proctitis, enterocolitis in the second group of patients were significantly less common than in the first. Thus, for the treatment of patients with cervical cancer, the most effective is the use of combined radiation therapy in parallel irradiation.

Abstract B031: FAK/PYK2 inhibition enhances antitumor efficacy of anti-PD-1 and anti-4-1BB antibodies

Abstract Although durable responses to single agent immune checkpoint inhibitors have been reported, additional approaches are needed to extend this therapeutic benefit to a greater proportion of patients. Combination of immunotherapy agents with tumor microenvironment modulators has the potential to overcome barriers that tumor cells develop to evade the immune system, and provide benefit to a greater proportion of patients. Focal Adhesion Kinase (FAK) and the family member PYK2 are potentially valuable targets due to their roles in regulating key cellular populations in the tumor microenvironment. The FAK/PYK2 dual inhibitors, VS-6063 and VS-4718, have previously been shown to inhibit monocyte-derived macrophages, reduce tumor-associated macrophages in xenograft models, and promote a CD8+ T cell-mediated anti-tumor response in squamous cell carcinoma models. We now report the combination of VS-4718 with an anti-PD-1 mAb shows substantially improved efficacy over anti-PD-1 mAb alone in MC38 syngeneic tumor-bearing animals. Analysis of MC38 tumors at day 12 of treatment revealed a significant increase in the CD8+ T cell/Treg ratio in tumors in the VS-4718 + anti-PD-1 combination group, providing a mechanistic understanding for the enhanced efficacy of this combination. To explore additional combination options, we tested the combination of VS-4718 with anti-4-1BB in the MC38 model. Consistent with what was observed with the anti-PD-1 combination, addition of VS-4718 also enhanced the efficacy of an anti-4-1BB mAb inducing full tumor regression. To further delineate the effect of FAK inhibition, an in vitro T cell proliferation assay was conducted. VS-6063 and VS-4718 dose-dependently stimulated proliferation of CD8+ cytotoxic T cells isolated from healthy donors. This is in distinct contrast to other protein kinase inhibitors, such as the SRC inhibitor dasatinib which potently impaired the proliferation of CD8+ cytotoxic T cells. In addition, both VS-4718 and VS-6063 decreased CD8+ T cell exhaustion markers (LAG3 and PD-1), and increased T cell-mediated tumor cell killing in vitro. These data provide additional rationale for ongoing clinical trials testing the FAK/PYK2 inhibitor VS-6063 in combination with pembrolizumab (anti-PD-1) plus gemcitabine in pancreatic cancer or in combination with avelumab (anti-PD-L1) in ovarian cancer. Citation Format: Jonathan A. Pachter, David T. Weaver. FAK/PYK2 inhibition enhances antitumor efficacy of anti-PD-1 and anti-4-1BB antibodies [abstract]. In: Proceedings of the Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016 Sept 25-28; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(11 Suppl):Abstract nr B031.

EVALUATION OF THE CLINICAL USE OF HYPOXIC CELL SENSITIZERS IN RADIATION THERAPY OF MALIGNANT EPITHELIAL SKIN TUMORS

Aim: To increase the efficacy of radiation therapy of malignant epithelial cell skin neoplasms with the use of radiation sensitizers of hypoxic tumor cells. Materials and methods: The study was performed in 517 patients with basal cell (n = 361) and squamous cell (n = 156) skin cancer, 274 (53%) of whom had T2 and 243 (47%), T3 tumors. Patients with locally advanced and metastatic tumors were excluded from the study. The following treatment modalities were used: distant gamma-therapy, short-distance radiation therapy and combined radiation therapy with the use of non-conventional dose fractioning at total local doses equal to 72–73 Gr. The sensibilization of hypoxic tumor cells to radiation therapy with metronidazole was done by targeted delivery of the drug to the tumor by means of topical application of Coletex-M drapes impregnated with metronidazole in a high concentration (up to 20 mcg/cm²). The second method of radiosensibilization of hypoxic tumor cells was based on a preliminary use of low intensity laser radiation onto the tumor. As a source this radiation, a helium neon laser was used with the power of up to 12 mVt and the wave length of 0.63 to 0.89 mcm, duration of sessions from 3 to 15 minutes. The control group comprised 192 skin cancer patients who underwent radiation therapy without the use of radiation sensitizers. Results: The use of metronidazole and low intensity laser radiation within the radiation therapy of T3 skin cancer patients, compared to the treatment without the radiation modifiers, significantly improved the immediate cure rates (full tumor regression at 1 to 1.5 months after completion of radiation) from 75.5 ± 3.1% to 89.2 ± 1.9% (р < 0.05). In the group with basal cell skin cancer that underwent radiation therapy combined with metronidazole, there was an association of its radio-modifying effect and tumor size. Short-distance roentgenotherapy of patients with T2 basal cell skin cancer and tumor size of < 4 cm was equally effective, irrespective of the use of metronidazole: the immediate cure rate was 94.8 ± 2.2% (92/97 patients) in the metronidazole group and 89.8 ± 3.9% (53/59) in the control group. With the combined radiation therapy of T2 basal skin cancer patients with the tumor size of 4–5 cm, the immediate cure rate was increased significantly from 73.2 ± 6.9% (30/42) in the control group to 88.2 ± 3.7% in the metronidazole group (67/76) (р < 0.05). With distant gamma-therapy of T3 basal cell skin cancer, this parameter was increased from 70.5 ± 6.8% (31/44) in the control group to 88.4 ± 4.8% (38/43) in the metronidazole group (р < 0.05). The radiation-sensitizing effect of metronidazole and low intensity laser combined with the distant gamma-therapy of T3 basal cell skin cancer was significant and similar: the immediate cure rate with the use of metronidazole was 85.9 ± 4.6% (49/57), and with the use of low intensity laser, 84.6 ± 5.0% (44/52), compared to 66.0 ± 6.9% (31/47) in the control group (р < 0.05). Although the relapse rate after radiation therapy with metronidazole or low intensity laser was lower by 5–11%, compared to that after the radiation without modifiers, there was no statistically significant difference after a 3-year follow-up. Conclusion: A significant benefit of radiation therapy combined with radiation sensitizers on the immediate cure rate of locally-advanced basal cell skin cancers was confirmed. It seems that radiation sensitizers of hypoxic cells do improve immediate results of cure without any significant effect on tumor relapse rate in the future.

Lymphoma of the ocular adnexa.

Over the last decades, the incidence of ocular adnexal lymphomas has been increasing. Over the last decade advances in the imaging of ocular adnexal lymphomas have enabled precise biopsies of the tumors. Biopsies are necessary for immunophenotyping, correct molecular classification and the immediate start of the appropriate treatment. In a retrospective study, we collected data from 16 patients (19 eyes) treated for lymphoma of the ocular adnexa between 2006 and 2011 with various tumor sites. We evaluated ocular symptoms of the patients in correlation to the tumor localization on imaging and determined the access of biopsy. Follow-up period (13.7 ± 15.4 months) and therapy were analyzed. The mean age of study patients was 67.4 ± 13.6 years. Seven patients presented with exophthalmos, 6 with upper lid swelling, 5 with double vision, 5 with reduced visual acuity and one patient with retinal detachment. None of the patients complained of pain. In 7 cases the location of the OAL was superior, in 6 retrobulbar, in 4 patients in the lacrimal gland (1 bilaterally affected), one case with lacrimal sac infiltration and another with isolated subconjunctival lymphoma. Patients with retrobulbar tumors complained of visual loss, whereas patients with anterior orbital tumors showed localized nodular swelling. In 11 cases (58%) the diagnosis was marginal zone B-cell lymphoma (MALT), in 5 (26%) follicular lymphoma. 11 patients (69%) received curative and one HIV-positive patient palliative radiation. Three patients were treated with systemic rituximab due to generalized lymphoma and one of them was complemented with CHOP therapy. One patient received chemotherapy alone (CHOP). During the follow-up the HIV-positive patient died. All our lymphoma patients showed full tumor regression, without side effects from radiotherapy. In this study we emphasize the need to perform a diagnostic biopsy. The risk of biopsy by a trained surgeon is small. Early and accurate diagnosis is crucial for proper treatment. In unclear situations repeated biopsies might be necessary.

Effect of a Vitamin D3 derivative (B3CD) with postulated anti-cancer activity in an ovarian cancer animal model

The objective of the present study was to test the hypothesis that Calcidiol derivative B3CD qualifies as a potential anti-cancer drug in vivo employing an ovarian cancer xenograft model in mice. In addition, the selectivity of B3CD on viability and proliferation of platinum-resistant human ovarian cancer cell lines in comparison to control cell lines was analyzed in vitro. B3CD displayed cell line-specific cytotoxicity screened against a panel of ovarian and other carcinoma cell lines, endothelial and control cells. B3CD, at sub-cytotoxic concentrations, revealed stronger effects on the proliferation of SKOV-3 ovarian cancer cells vs. primary fibroblasts as determined by BrdU incorporation analysis. Treatment with B3CD at 0.5 microM resulted in highly condensed chromatin and fragmented nuclei in SKOV-3 cells but not in primary fibroblasts. B3CD induced cell death at low drug concentrations (< or = 0.5 microM) in SKOV-3 ovarian cancer cells is mediated by the p38 MAPK signaling pathway: B3CD induced p38 MAPK expression and activation in SKOV-3 cells and inhibition of p38 signaling counteracted B3CD induced cell death in vitro. An ovarian cancer cell animal model (human SKOV-3 cell derived xenografts in nude mice) revealed that tumor growth in few B3CD treated mice accelerated while the majority of B3CD treated mice displayed delayed tumor growth or full tumor regression. B3CD possesses anti-ovarian cancer properties in vitro and in vivo. We propose the further development of non-calcemic bromoacetoxy derivatives of vitamin D(3) as potential anti-cancer therapeutics.

Treatment with ozone/oxygen‐pneumoperitoneum results in complete remission of rabbit squamous cell carcinomas

Head and neck squamous cell carcinomas (HNSCC) represent a group of metastasizing tumors with a high mortality rate in man and animals. Since the biomolecule ozone was found to inhibit growth of various carcinoma cells in vitro we here applied the highly aggressive and lethal VX2 carcinoma HNSCC tumor model of the New Zealand White rabbit to test whether ozone exerts antitumorous effects in vivo. Therapeutic insufflation of medical ozone/oxygen (O(3)/O(2)) gas mixture into the peritoneum (O(3)/O(2)-pneumoperitoneum) at an advanced stage of tumor disease led to a survival rate of 7/14 rabbits. Six of the seven surviving rabbits presented full tumor regression and the absence of local or distant lung metastases. Insufflation of pure oxygen (O(2)) resulted in a survival rate of 3/13 animals accompanied by full tumor remission in 2 of the 3 surviving animals. Of the 14 sham-treated animals only 1 had spontaneous tumor remission and survived. No adverse effects or changes in standard blood parameters were observed after repeated intraperitoneal insufflations of the O(3)/O(2) or O(2) gas. Animals with O(3)/O(2)-induced tumor eradication developed tolerance against reimplantation of the VX2 tumor. This could be reversed by immune suppression with a combination of dexamethasone and cyclosporin A suggesting an antitumorous effect of O(3)/O(2)-mediated activation of the body's own immunosurveillance. Although the exact mechanisms of action are still unclear the present data point to O(3)/O(2)-pneumoperitoneum as a promising new strategy in anticancer therapy.