Key Drivers Of Cancer Progression Research Articles

Abstract 4133: OncoTAM, a comprehensive preclinical platform to explore macrophages as key drivers of cancer progression and develop new therapies against tumor-associated-macrophages

Abstract Tumor-Associated Macrophages (TAMs) play an important role in the development of tumors, modulation of neoangiogenesis, immune suppression, and metastasis. A high infiltration of macrophages in the tumor is also correlated with a poor prognosis in several cancer types. Therefore, they became an attractive target for cancer immunotherapies. Several macrophage-targeting approaches in anticancer therapy are under development, including TAM depletion, inhibition of new TAM differentiation, or re-education of TAM activation for cancer cell phagocytosis. In this presentation, we will share different examples of in vitro assays and in vivo models we are implementing to support preclinical development of novel TAM-targeting strategies. Antibody-dependent cellular phagocytosis (ADCP) has been used to demonstrate one crucial mechanism of action of different antibody (Ab) therapies targeting macrophages. Some of these Ab (including anti-CSF1) have then been tested in syngeneic in vivo tumor models. To reach this goal, we demonstrated that the tumor implantation site in mice (subcutaneous vs orthotopic) could impact the polarization of macrophages (M1 vs M2). Differences in the ratio of M1 and M2 subtypes infiltrating the PAN-02 pancreatic murine tumors were observed, and anti-CSF1 antibodies increased the survival of mice bearing orthotopic Renca murine kidney tumor by eliminating TAMs. Additionally, in xenograft models of human breast tumors in NOD-SCID mice, the eradication of TAMs by anti-CSF1R clearly demonstrated the importance of macrophages in the tumor progression and in the anti-tumor efficacy of Abs mediated by macrophages. By using an orthotopic Hepa1-6 murine liver cancer model, we showed high antitumor efficacy of compounds targeting the STAT6 pathway by reprogramming immunosuppressive TAMs into an M1 phenotype that promotes the induction of a cytotoxic immune response. For compounds displaying no cross-reactivity with murine target, we developed models and characterized the TAMs in breast, colon, melanoma and head & neck PDX and CDX tumors in different huCD34-engrafted mouse models. Altogether, the panel of in vitro assays and in vivo tumor models in OncoTAM should be useful to provide insights on the mechanism of action and antitumor efficacy of novel immune-oncology strategies targeting macrophages. Citation Format: Caroline Mignard, Damien France, Francis Bichat, Nicolas Legrand, Olivier Duchamp. OncoTAM, a comprehensive preclinical platform to explore macrophages as key drivers of cancer progression and develop new therapies against tumor-associated-macrophages. [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 4133.

Abstract 3878: Subcellular characterization of over 100 proteins in FFPE tumor biopsies with CosMx Spatial Molecular Imager

Abstract The spatial interactions between the immune system and tumor cells greatly influence antitumoral immunity. Characterization of immune cell composition and infiltration within the tumor niche informs prognosis, drug delivery efficiency, and therapeutic efficacy. However, few methods exist to query large numbers of immune biomarkers at subcellular spatial resolution. The CosMx™ Spatial Molecular Imager is the first platform to demonstrate simultaneous single-cell and subcellular detection of over 100 proteins on standard, biobanked, FFPE tissue samples. This high-plex protein panel detects key drivers of cancer progression and immune cell activation states. Here, we apply the CosMx 100-plex immuno-oncology assay on a set of breast cancer biopsies and demonstrate its quantitative and spatial capabilities. Key to CosMx protein technology is an antibody-oligonucleotide-conjugate 64-bit encoding method, not a cyclic exchange method. The encoding scheme is enabled by a 20nm hybridization-based optical barcode. The CosMx system uses a fully automated, cyclic microfluidics imaging system, high-resolution optics and 3D capability. The raw cyclic encoded 4-color tissue images are decoded using a robust automated decoding algorithm that detects protein sub-cellular localization and quantifies expression level. CosMx SMI produces protein localization maps for each target, which characterizes tissue microenvironment heterogeneity while providing spatial information. Additionally, accurate segmentation of individual cells enables spatial single-cell protein expression analysis, facilitating further mining and analyses of cellular subpopulations. The CosMx protein assay reagents were validated on multi-organ FFPE tissue microarrays and 35 human FFPE cell lines, including overexpression lines for key targets and cellular activation states, such as GITR, CD278, PD-L1, and PD-1. Benchmarking to multiple orthogonal datasets (e.g., the Human Protein Atlas, Cancer Cell Line Encyclopedia, and low-plex IHC) demonstrates that the assay is highly sensitive and specific. CosMx SMI protein assay can be coupled with SMI’s 1000-plex RNA-detection assay; together, such a multi-omics platform can generate an unprecedented information-rich view of spatial biology that could usher in novel discoveries about health and disease. FOR RESEARCH USE ONLY. Not for use in diagnostic procedures. Citation Format: Zachary R. Lewis, Tien Phan-Everson, Gary Geiss, Mithra Korukonda, Ruchir Bhatt, Carl Brown, Dwayne Dunaway, Joseph Phan, Alyssa Rosenbloom, Brian Filanoski, Rhonda Meredith, Kan Chantranuvatana, Yan Liang, Emily Brown, Brian Birditt, Giang Ong, Hye Son Yi, Erin Piazza, Vikram Devgan, Nicole Ortogero, Patrick Danaher, Sarah Warren, Michael Rhodes, Joseph Beechem. Subcellular characterization of over 100 proteins in FFPE tumor biopsies with CosMx Spatial Molecular Imager [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3878.

The Impact of Obesity, Adipose Tissue, and Tumor Microenvironment on Macrophage Polarization and Metastasis

Simple SummaryThe inflammatory adipose microenvironment in obesity plays a crucial role in cancer development and metastases. By focusing on adipocytes and macrophages, as well as the extracellular matrix, the cellular and molecular mechanisms that link inflammation, obesity, and cancer will be addressed by this review. After describing the tumor microenvironment and extracellular matrix, the influence of M1, M2, and tumor-associated macrophages will be explored through their origin, classification, polarization, and regulatory networks, including their potential role in angiogenesis, invasion, metastasis, and immunosuppression, with a specific focus on the roles of adipocytes in this process.Tumor metastasis is a major cause of death in cancer patients. It involves not only the intrinsic alterations within tumor cells, but also crosstalk between these cells and components of the tumor microenvironment (TME). Tumorigenesis is a complex and dynamic process, involving the following three main stages: initiation, progression, and metastasis. The transition between these stages depends on the changes within the extracellular matrix (ECM), in which tumor and stromal cells reside. This matrix, under the effect of growth factors, cytokines, and adipokines, can be morphologically altered, degraded, or reorganized. Many cancers evolve to form an immunosuppressive TME locally and create a pre-metastatic niche in other tissue sites. TME and pre-metastatic niches include myofibroblasts, immuno-inflammatory cells (macrophages), adipocytes, blood, and lymphatic vascular networks. Several studies have highlighted the adipocyte-macrophage interaction as a key driver of cancer progression and dissemination. The following two main classes of macrophages are distinguished: M1 (pro-inflammatory/anti-tumor) and M2 (anti-inflammatory/pro-tumor). These cells exhibit distinct microenvironment-dependent phenotypes that can promote or inhibit metastasis. On the other hand, obesity in cancer patients has been linked to a poor prognosis. In this regard, tumor-associated adipocytes modulate TME through the secretion of inflammatory mediators, which modulate and recruit tumor-associated macrophages (TAM). Hereby, this review describes the cellular and molecular mechanisms that link inflammation, obesity, and cancer. It provides a comprehensive overview of adipocytes and macrophages in the ECM as they control cancer initiation, progression, and invasion. In addition, it addresses the mechanisms of tumor anchoring and recruitment for M1, M2, and TAM macrophages, specifically highlighting their origin, classification, polarization, and regulatory networks, as well as their roles in the regulation of angiogenesis, invasion, metastasis, and immunosuppression, specifically highlighting the role of adipocytes in this process.

Remodeling the ECM: Implications for Metastasis and Tumor Dormancy.

Simple SummaryIn this perspective article we discuss the importance of investigating the role of the ECM on tumor metastasis and tumor dormancy. First, we discuss the importance of the ECM in cancer progression. Then, we revise the parallelisms between dormant cells and stem cells and the role of the ECM in stem cell regulation. Finally, we comment on the evidence describing the role of the ECM in tumor dormancy and highlight key outstanding questions in the tumor dormancy field.While most primary tumors can be effectively treated, therapeutics fail to efficiently eliminate metastases. Metastases arise from cancer cells that leave the primary tumor and seed distant sites. Recent studies have shown that cancer cells disseminate early during tumor progression and can remain dormant for years before they resume growth. In these metastatic organs, cancer cells reside in microenvironments where they interact with other cells, but also with the extracellular matrix (ECM). The ECM was long considered to be an inert, non-cellular component of tissues, providing their architecture. However, in recent years, a growing body of evidence has shown that the ECM is a key driver of cancer progression, and it can exert effects on tumor cells, regulating their metastatic fate. ECM remodeling and degradation is required for the early steps of the metastatic cascade: invasion, tumor intravasation, and extravasation. Similarly, ECM molecules have been shown to be important for metastatic outgrowth. However, the role of ECM molecules on tumor dormancy and their contribution to the dormancy-supportive niches is not well understood. In this perspective article, we will summarize the current knowledge of ECM and its role in tumor metastasis and dormancy. We will discuss how a better understanding of the individual components of the ECM niche and their roles mediating the dormant state of disseminated tumor cells (DTCs) will advance the development of new therapies to target dormant cells and prevent metastasis outgrowth.

Neutrophil Heterogeneity in Cancer: From Biology to Therapies.

Neutrophils have been extensively described in the pathophysiology of autoimmune and infectious diseases. Accumulating evidence also suggests the important role of neutrophils in cancer progression through their interaction with cancer and immune cells in blood and in the tumor microenvironment (TME). Most studies have described neutrophils as key drivers of cancer progression, due to their involvement in various tumor promoting functions including proliferation, aggressiveness, and dissemination, as well as in immune suppression. However, such studies were focusing on late-stages of tumorigenesis, in which chronic inflammation had already developed. The role of tumor-associated neutrophils (TANs) at early stages of tumor development remains poorly described, though recent findings indicate that early-stage TANs may display anti-tumor properties. Beyond their role at tumor site, evidence supported by NLR retrospective studies and functional analyses suggest that blood neutrophils could also actively contribute to tumorigenesis. Hence, it appears that the phenotype and functions of neutrophils vary greatly during tumor progression, highlighting their heterogeneity. The origin of pro- or anti-tumor neutrophils is generally believed to arise following a change in cell state, from resting to activated. Moreover, the fate of neutrophils may also involve distinct differentiation programs yielding various subsets of pro or anti-tumor neutrophils. In this review, we will discuss the current knowledge on neutrophils heterogeneity across different tissues and their impact on tumorigenesis, as well as neutrophil-based therapeutic strategies that have shown promising results in pre-clinical studies, paving the way for the design of neutrophil-based next generation immunotherapy.

Curation of the Pancreatic Ductal Adenocarcinoma Subset of the Cancer Genome Atlas Is Essential for Accurate Conclusions about Survival-Related Molecular Mechanisms.

Purpose: Publicly available databases, for example, The Cancer Genome Atlas (TCGA), containing clinical and molecular data from many patients are useful in validating the contribution of particular genes to disease mechanisms and in forming novel hypotheses relating to clinical outcomes.Experimental Design: The impact of key drivers of cancer progression can be assessed by segregating a patient cohort by certain molecular features and constructing survival plots using the associated clinical data. However, conclusions drawn from this straightforward analysis are highly dependent on the quality and source of tissue samples, as demonstrated through the pancreatic ductal adenocarcinoma (PDAC) subset of TCGA.Results: Analyses of the PDAC-TCGA database, which contains mainly resectable cancer samples from patients in stage IIB, reveal a difference from widely known historic median and 5-year survival rates of PDAC. A similar discrepancy was observed in lung, stomach, and liver cancer subsets of TCGA. The whole transcriptome expression patterns of PDAC-TCGA revealed a cluster of samples derived from neuroendocrine tumors, which have a distinctive biology and better disease prognosis than PDAC. Furthermore, PDAC-TCGA contains numerous pseudo-normal samples, as well as those that arose from tumors not classified as PDAC.Conclusions: Inclusion of misclassified samples in the bioinformatic analyses distorts the association of molecular biomarkers with clinical outcomes, altering multiple published conclusions used to support and motivate experimental research. Hence, the stringent scrutiny of type and origin of samples included in the bioinformatic analyses by researchers, databases, and web-tool developers is of crucial importance for generating accurate conclusions. Clin Cancer Res; 24(16); 3813-9. ©2018 AACR.

Intracellular hypoxia measured by 18F-fluoromisonidazole positron emission tomography has prognostic impact in patients with estrogen receptor-positive breast cancer

BackgroundHypoxia is a key driver of cancer progression. We evaluated the prognostic impact of 18F-fluoromisonidazole (FMISO) prior to treatment in patients with breast cancer.MethodsForty-four patients with stage II/III primary breast cancer underwent positron emission tomography/computed with 18F-fluorodeoxyglucose (FDG-PET/CT) and FMISO. After measurement by FDG-PET/CT, the tissue-to-blood ratio (TBR) was obtained using FMISO-PET/CT. FMISO-TBR was compared for correlation with clinicopathological factors, disease-free survival (DFS), and overall survival (OS). Multiplex cytokines were analyzed for the correlation of FMISO-TBR.ResultsTumors with higher nuclear grade and negativities of estrogen receptor (ER) and progesterone receptor had significantly higher FMISO-TBR than other tumors. Kaplan-Meier survival curves showed that patients with a higher FMISO-TBR (cutoff, 1.48) had a poorer prognosis of DFS (p = 0.0007) and OS (p = 0.04) than those with a lower FMISO-TBR. Multivariate analysis indicated that higher FMISO-TBR and ER negativity were independent predictors of shorter DFS (p = 0.01 and 0.03). Higher FMISO-TBR was associated with higher plasma levels of angiogenic hypoxic markers such as vascular endothelial growth factor, transforming growth factor-α, and interleukin 8.ConclusionsFMISO-PET/CT is useful for assessing the prognosis of patients with breast cancer, but it should be stratified by ER status.Trial registrationUMIN Clinical Trials Registry, UMIN000006802. Registered on 1 December 2011.

Loss of Forkhead box M1 promotes erythropoiesis through increased proliferation of erythroid progenitors

Forkhead box M1 (FOXM1) belongs to the forkhead/winged-helix family of transcription factors and regulates a network of proliferation-associated genes. Its abnormal upregulation has been shown to be a key driver of cancer progression and an initiating factor in oncogenesis. FOXM1 is also highly expressed in stem/progenitor cells and inhibits their differentiation, suggesting that FOXM1 plays a role in the maintenance of multipotency. However, the exact molecular mechanisms by which FOXM1 regulates human stem/progenitor cells are still uncharacterized. To understand the role of FOXM1 in normal hematopoiesis, human cord blood CD34+ cells were transduced with FOXM1 short hairpin ribonucleic acid (shRNA) lentivirus. Knockdown of FOXM1 resulted in a 2-fold increase in erythroid cells compared to myeloid cells. Additionally, knockdown of FOXM1 increased bromodeoxyuridine (BrdU) incorporation in erythroid cells, suggesting greater proliferation of erythroid progenitors. We also observed that the defective phosphorylation of FOXM1 by checkpoint kinase 2 (CHK2) or cyclin-dependent kinases 1/2 (CDK1/2) increased the erythroid population in a manner similar to knockdown of FOXM1. Finally, we found that an inhibitor of FOXM1, forkhead domain inhibitor-6 (FDI-6), increased red blood cell numbers through increased proliferation of erythroid precursors. Overall, our data suggest a novel function of FOXM1 in normal human hematopoiesis.

Macrophages as Key Drivers of Cancer Progression and Metastasis.

Macrophages are one of the most abundant immune cells in the tumour microenvironment of solid tumours and their presence correlates with reduced survival in most cancers. Macrophages are present at all stages of tumour progression and stimulate angiogenesis, tumour cell invasion, and intravasation at the primary site. At the metastatic site, macrophages and monocytes prepare for the arrival of disseminated tumour cells and promote their extravasation and survival by inhibiting immune-mediated clearance or by directly engaging with tumour cells to activate prosurvival signalling pathways. In addition, macrophages promote the growth of disseminated tumour cells at the metastatic site by organising the formation of a supportive metastatic niche. The development of agents inhibiting the recruitment or the protumorigenic effector functions of macrophages in both the primary tumour and at the metastatic site is a promising strategy to improve cancer survival in the future.

Structural Mechanisms and Drug Discovery Prospects of Rho GTPases.

Rho GTPases regulate cellular morphology and dynamics, and some are key drivers of cancer progression. This superfamily offers attractive potential targets for therapeutic intervention, with RhoA, Rac1 and Cdc42 being prime examples. The challenges in developing agents that act on these signaling enzymes include the lack of obvious druggable pockets and their membrane-bound activities. However, progress in targeting the similar Ras protein is illuminating new strategies for specifically inhibiting oncogenic GTPases. The structures of multiple signaling and regulatory states of Rho proteins have been determined, and the post-translational modifications including acylation and phosphorylation points have been mapped and their functional effects examined. The development of inhibitors to probe the significance of overexpression and mutational hyperactivation of these GTPases underscores their importance in cancer progression. The ability to integrate in silico, in vitro, and in vivo investigations of drug-like molecules indicates the growing tractability of GTPase systems for lead optimization. Although no Rho-targeted drug molecules have yet been clinically approved, this family is clearly showing increasing promise for the development of precision medicine and combination cancer therapies.

Loss of FoxM1 Promotes Erythroid Differentiation through Increased Proliferation of Erythroid Progenitors

FoxM1 belongs to the fork head/winged-helix family of transcription factors and regulates a network of proliferation-associated genes including the G2/M transition, chromosome segregation, and spindle assembly. FoxM1 expression is commonly upregulated in a number of human cancers such as liver, ovarian, breast, prostate, colon, and brain tumors. Its abnormal upregulation has been shown to be a key driver of cancer progression and an initiating factor of oncogenesis. In normal cells, FoxM1 is highly expressed in multipotent progenitor cells and inhibits differentiation of the progenitors, suggesting that FoxM1 plays in a role in the maintenance of multipotent progenitor cells. However, the exact molecular mechanism by which FoxM1 regulates stem/progenitor cells is still uncharacterized. In this study, we have examined the role of FoxM1 in normal hematopoiesis using human cord blood CD34+ cells.To investigate the role of FoxM1 in normal hematopoiesis, we infected human cord blood CD34+ cells with FoxM1 shRNA lentivirus and observed blood cell differentiation using FACS analysis with a range of cell surface markers. We found that knockdown of FoxM1 resulted in an increase of the erythroid population (CD71+/GlyA+), a decrease of the myeloid population (CD11b+), and an unchanged megakaryocyte population (CD41a+) in two phase liquid culture system. Overall, we found a 2-fold increase in the erythroid population compared to the myeloid population. Importantly, methylcellulose colony assays also demonstrated increased numbers of CFU-E colonies (2-2.5 fold increase compared to control) and decreased numbers of CFU-GM colonies in FoxM1 knockdown cells. Taken together, these findings imply a role for FoxM1 in normal erythropoiesis.To better define the function of FoxM1 in hematopoietic cells, we sorted distinct populations of cells based on their cell surface marker expression and quantitated FoxM1 expression level by RT-qPCR. FoxM1 had a 3-fold increased expression in CD71+ (erythroid) cells compared to CD11b+ (myeloid) cells. Additionally, we found FoxM1 expression was particularly elevated in the BFU-E and CFU-E stages of erythropoiesis, suggesting a functional role for FoxM1 in erythroid progenitor proliferation.Finally, to study the potential molecular mechanism of FoxM1 in normal hematopoiesis, we analyzed cell cycle progress in FoxM1 knockdown cells with DAPI staining. We found increased S and G2/M phases in FoxM1 knockdown cells, which was significant only in the CD71+ (erythroid) population and not in the unsorted cell populations. We also detected an increase of BrdU+ cells in FoxM1 knockdown CD71+ population by BrdU incorporation assay, indicating faster proliferation of erythroid progenitors with FoxM1 knockdown.These findings suggest a novel function of FoxM1 in normal human hematopoiesis, in which FoxM1 deficiency leads to increased proliferation of erythroid progenitors resulting in increased erythroid differentiation. Our data indicate that FoxM1 inhibitors, such as Thiostrepton or FDI-6, may be beneficial in treating patients with anemia. DisclosuresNo relevant conflicts of interest to declare.

Abstract 1448: Dynamic chromatin modification by tumor secreted Hsp90 modulates EMT and tumor invasion

Abstract Tumor metastasis is the main cause of prostate cancer lethality. Pathological reactivation of the developmental genetic program epithelial to mesenchymal (EMT) is associated with increased tumorigenesis and invasion, and is considered a primary culprit for tumor dissemination. Hallmarks of EMT include upregulation of the core transcription factors Zeb and Snail, concomitant with suppression of the junctional protein E-cadherin. Overexpression of the epigenetic repressor EZH2, a histone methyltransferase, is also implicated in EMT activation. Although these molecular events are well known to trend with cancer progression, a paucity of knowledge exists regarding identification of the clinically-relevant upstream triggers capable of setting these events into motion. We recently demonstrated that tumor secreted extracellular heat shock protein 90 (eHsp90) initiates EMT events in prostate cancer cells. Moreover, we reported the presence of surface Hsp90 in human prostatectomy specimens, an event highly correlated with elevated expression of a subset of EMT transcripts. The present study reveals novel mechanistic aspects of eHsp90 action. We define an epigenetic function for eHsp90 as a significant regulator of EZH2 expression and activity. This eHsp90-EZH2 axis was found to govern a majority of eHsp90's pro-EMT activity in vitro, and was essential for E-cadherin suppression and tumor invasion in vivo. Upon further investigation, it was revealed that eHsp90 sustained ERK activation. This eHsp90-ERK pathway was required for the increased expression of EZH2, as well as for its directed recruitment to the E-cadherin promoter. Moreover, eHsp90-ERK signaling augmented the epigenetic landscape of Zeb and Snail towards a profile permissive for transcriptional activation. Hence, our findings indicate that eHsp90-ERK signaling has profound effects upon key drivers of cancer progression via the concerted action of epigenetic modulation of EMT effectors and increased EZH2 expression. Collectively, these data support a model wherein tumor eHsp90 functions as an upstream rheostat for EZH2 expression and activity to orchestrate mesenchymal properties and coincident aggressive behavior. Importantly, our findings suggest that eHsp90 may be a clinically relevant instigator in the transition from localized to invasive disease. Citation Format: Krystal D. Nolan, Michael W. Hance, Omar E. Franco, Simon W. Hayward, Jennifer S. Isaacs. Dynamic chromatin modification by tumor secreted Hsp90 modulates EMT and tumor invasion. [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 1448. doi:10.1158/1538-7445.AM2015-1448

Metastasis is regulated via microRNA-200/ZEB1 axis control of tumour cell PD-L1 expression and intratumoral immunosuppression.

Immunosuppression of tumor-infiltrating lymphocytes (TIL) is a common feature of advanced cancer, but its biological basis has remained obscure. We demonstrate here a molecular link between epithelial-to-mesenchymal transition (EMT) and CD8+ TIL immunosuppression, two key drivers of cancer progression. We show that microRNA-200 (miR-200), a cell-autonomous suppressor of EMT and metastasis, targets PD-L1. Moreover, ZEB1, an EMT activator and transcriptional repressor of miR-200, relieves miR-200 repression of PD-L1 on tumor cells, leading to CD8+ T cell immunosuppression and metastasis. These findings are supported by robust correlations between the EMT score, miR-200 levels and PD-L1 expression in multiple human lung cancer datasets. In addition to revealing a link between EMT and T cell dysfunction, these findings also show that ZEB1 promotes metastasis through a heretofore unappreciated cell non-autonomous mechanism, and suggest that subgroups of patients in whom malignant progression is driven by EMT activators may respond to treatment with PD-L1 antagonists.

Systems analysis of drug-induced receptor tyrosine kinase reprogramming following targeted mono- and combination anti-cancer therapy.

The receptor tyrosine kinases (RTKs) are key drivers of cancer progression and targets for drug therapy. A major challenge in anti-RTK treatment is the dependence of drug effectiveness on co-expression of multiple RTKs which defines resistance to single drug therapy. Reprogramming of the RTK network leading to alteration in RTK co-expression in response to drug intervention is a dynamic mechanism of acquired resistance to single drug therapy in many cancers. One route to overcome this resistance is combination therapy. We describe the results of a joint in silico, in vitro, and in vivo investigations on the efficacy of trastuzumab, pertuzumab and their combination to target the HER2 receptors. Computational modelling revealed that these two drugs alone and in combination differentially suppressed RTK network activation depending on RTK co-expression. Analyses of mRNA expression in SKOV3 ovarian tumour xenograft showed up-regulation of HER3 following treatment. Considering this in a computational model revealed that HER3 up-regulation reprograms RTK kinetics from HER2 homodimerisation to HER3/HER2 heterodimerisation. The results showed synergy of the trastuzumab and pertuzumab combination treatment of the HER2 overexpressing tumour can be due to an independence of the combination effect on HER3/HER2 composition when it changes due to drug-induced RTK reprogramming.