Search

XLSX file - 768K, Nanostring CRC codeset.

PDF file - 674K, Histological characterization of patients' carcinomas and the correspondent xenograft tumors.

Supplementary Figures S1-S11. Figure S1. Apoptosis of CRC patient-derived cells upon treatment with API-2 or NVP-BKM120 in combination with XAV939. Figure S2. Evaluation of apoptosis in primary sphere cell cultures by immunofluorescent quantification of cleaved-CASPASE3. Figure S3. Frequent mutations in CRC do not determine the response to API2, NVP-BKM120 or NVP-TNKS656. Figure S4. NVP-TNKS656 is not toxic and reduces nuclear β-catenin in the skin and small intestine in NOD-SCID mice. Figure S5. Analyses of pharmacodynamic markers at endpoint of in vivo experiments with API2 and NVP-TNKS656. Figure S6. Nuclear FOXO3A content is preserved in PDX and primary sphere cultures. Figure S7. Nuclear FOXO3A in paired primary tumors and liver metastases. Figure S8. Immunohistochemical staining of β-catenin in CRC tumors. Figure S9. β-catenin in single-agent treatments, relevant mutations, TNM stage or tumor site do not associate with a differential response to PI3K/AKT/mTOR inhibitors. Figure S10. Gene expression clustering and mutational status of baseline tumor samples from patients treated with PI3K/AKT/mTOR inhibitors. Figure S11. Molecular prescreening scheme to select CRC patients eligible for the treatment with PI3K/AKT and/or Wnt/β-catenin inhibitory drugs.

Purpose: PI3K pathway activation occurs in concomitance with RAS/BRAF mutations in colorectal cancer, limiting the sensitivity to targeted therapies. Several clinical studies are being conducted to test the tolerability and clinical activity of dual MEK and PI3K pathway blockade in solid tumors.Experimental Design: In the present study, we explored the efficacy of dual pathway blockade in colorectal cancer preclinical models harboring concomitant activation of the ERK and PI3K pathways. Moreover, we investigated if TP53 mutation affects the response to this therapy.Results: Dual MEK and mTORC1/2 blockade resulted in synergistic antiproliferative effects in cell lines bearing alterations in KRAS/BRAF and PIK3CA/PTEN. Although the on-treatment cell-cycle effects were not affected by the TP53 status, a marked proapoptotic response to therapy was observed exclusively in wild-type TP53 colorectal cancer models. We further interrogated two independent panels of KRAS/BRAF- and PIK3CA/PTEN-altered cell line– and patient-derived tumor xenografts for the antitumor response toward this combination of agents. A combination response that resulted in substantial antitumor activity was exclusively observed among the wild-type TP53 models (two out of five, 40%), but there was no such response across the eight mutant TP53 models (0%). Interestingly, within a cohort of 14 patients with colorectal cancer treated with these agents for their metastatic disease, two patients with long-lasting responses (32 weeks) had TP53 wild-type tumors.Conclusions: Our data support that, in wild-type TP53 colorectal cancer cells with ERK and PI3K pathway alterations, MEK blockade results in potent p21 induction, preventing apoptosis to occur. In turn, mTORC1/2 inhibition blocks MEK inhibitor–mediated p21 induction, unleashing apoptosis. Clin Cancer Res; 21(24); 5499–510. ©2015 AACR.

PDF file - 115K, 18-FDG is the best radiotracer for tumor growth evaluation.

Supplementary Tables S1-S15. Table S1. Genes and positions analyzed by amplicon sequencing. Table S2. Genes sequenced by Haloplex Target Enrichment System. Table S3. Antibodies. Table S4. Treatment-induced apoptosis in 10 primary patient-derived sphere cell cultures. Table S5. Genotype of patient-derived colorectal cancer models. Table S6. Exome sequencing of colorectal cancer patient-derived models. Table S7. Genes regulated by NVP-TNKS656 in PDX-P2. Table S8. Genes regulated by NVP-TNKS656 in PDX-P30. Table S9. Genes regulated by NVP-TNKS656 in PDX-P5. Table S10. Intestinal β-catenin/TCF signature in human CRC. Table S11. Genes regulated by β-catenin/FOXO3A signaling in human CRC. Table S12. Molecular profiling of 130 CRC tumors. Table S13. Clinicopathological data of 40 CRC patients treated with PI3K/AKT/mTOR inhibitors and their corresponding baseline tumor samples analyzed for mutations and nuclear β-catenin content. Table S14. Nuclear β-catenin content but not relevant mutations or TNM stage determine response of CRC patients to PI3K/AKT/mTOR inhibitors. Table S15. Multivariate analysis of baseline tumors from patients treated with PI3K/AKT/mTOR inhibitors.

Table S1. MiSeq 57-gene panel list

Purpose: Oncogenic mutations in the KRAS/PI3K/AKT pathway are one of the most frequent alterations in cancer. Although PI3K or AKT inhibitors show promising results in clinical trials, drug resistance frequently emerges. We previously revealed Wnt/β-catenin signaling hyperactivation as responsible for such resistance in colorectal cancer. Here we investigate Wnt-mediated resistance in patients treated with PI3K or AKT inhibitors in clinical trials and evaluate the efficacy of a new Wnt/tankyrase inhibitor, NVP-TNKS656, to overcome such resistance.Experimental Design: Colorectal cancer patient-derived sphere cultures and mouse tumor xenografts were treated with NVP-TNKS656, in combination with PI3K or AKT inhibitors.We analyzed progression-free survival of patients treated with different PI3K/AKT/mTOR inhibitors in correlation with Wnt/β-catenin pathway activation, oncogenic mutations, clinicopathological traits, and gene expression patterns in 40 colorectal cancer baseline tumors.Results: Combination with NVP-TNKS656 promoted apoptosis in PI3K or AKT inhibitor-resistant cells with high nuclear β-catenin content. High FOXO3A activity conferred sensitivity to NVP-TNKS656 treatment. Thirteen of 40 patients presented high nuclear β-catenin content and progressed earlier upon PI3K/AKT/mTOR inhibition. Nuclear β-catenin levels predicted drug response, whereas clinicopathologic traits, gene expression profiles, or frequent mutations (KRAS, TP53, or PIK3CA) did not.Conclusions: High nuclear β-catenin content independently predicts resistance to PI3K and AKT inhibitors. Combined treatment with a Wnt/tankyrase inhibitor reduces nuclear β-catenin, reverts such resistance, and represses tumor growth. FOXO3A content and activity predicts response to Wnt/β-catenin inhibition and together with β-catenin may be predictive biomarkers of drug response providing a rationale to stratify colorectal cancer patients to be treated with PI3K/AKT/mTOR and Wnt/β-catenin inhibitors. Clin Cancer Res; 22(3); 644–56. ©2015 AACR.

Purpose: Within the aim of advancing precision oncology, we have generated a collection of patient-derived xenografts (PDX) characterized at the molecular level, and a preclinical model of colon cancer metastasis to evaluate drug-response and tumor progression.Experimental Design: We derived cells from 32 primary colorectal carcinomas and eight liver metastases and generated PDX annotated for their clinical data, gene expression, mutational, and histopathological traits. Six models were injected orthotopically into the cecum wall of NOD-SCID mice in order to evaluate metastasis. Three of them were treated with chemotherapy (oxaliplatin) and three with API2 to target AKT activity. Tumor growth and metastasis progression were analyzed by positron emission tomography (PET).Results: Patient-derived cells generated tumor xenografts that recapitulated the same histopathological and genetic features as the original patients' carcinomas. We show an 87.5% tumor take rate that is one of the highest described for implanted cells derived from colorectal cancer patients. Cecal injection generated primary carcinomas and distant metastases. Oxaliplatin treatment prevented metastasis and API2 reduced tumor growth as evaluated by PET.Conclusions: Our improved protocol for cancer cell engraftment has allowed us to build a rapidly expanding collection of colorectal PDX, annotated for their clinical data, gene expression, mutational, and histopathological statuses. We have also established a mouse model for metastatic colon cancer with patient-derived cells in order to monitor tumor growth, metastasis evolution, and response to treatment by PET. Our PDX models could become the best preclinical approach through which to validate new biomarkers or investigate the metastatic potential and drug-response of individual patients. Clin Cancer Res; 19(24); 6787–801. ©2013 AACR.

Supplementary Figure and Table Legends. The text describes the panels and tables included as Supplementary Figure and Table Legends

Figure S2. Immunohistochemistry of p53 in tumors from CRC patients treated with MEK- plus PI3K-inhibitors

PDF file - 1113K, Patient-derived cells preserve pluripotency.

Supporting Information. Supplementary methods to complete the information for gene expression, genotyping, immunohistochemistry evaluation criteria and protein extraction. The file also contains legends for supplementary figures S1-S11 and supplementary tables S1-S15.

PDF file - 248K, Distant metastasis generated by cells derived from patient 5.

PDF file - 290K, Histopathological and genetic traits of CRC PDX.

Patient-derived organoids (PDOs) recapitulate tumor architecture, contain cancer stem cells and have predictive value supporting personalized medicine. Here we describe a large-scale functional screen of dual-targeting bispecific antibodies (bAbs) on a heterogeneous colorectal cancer PDO biobank and paired healthy colonic mucosa samples. More than 500 therapeutic bAbs generated against Wingless-related integration site (WNT) and receptor tyrosine kinase (RTK) targets were functionally evaluated by high-content imaging to capture the complexity of PDO responses. Our drug discovery strategy resulted in the generation of MCLA-158, a bAb that specifically triggers epidermal growth factor receptor degradation in leucine-rich repeat-containing G-protein-coupled receptor 5-positive (LGR5+) cancer stem cells but shows minimal toxicity toward healthy LGR5+ colon stem cells. MCLA-158 exhibits therapeutic properties such as growth inhibition of KRAS-mutant colorectal cancers, blockade of metastasis initiation and suppression of tumor outgrowth in preclinical models for several epithelial cancer types.© 2022. The Author(s), under exclusive licence to Springer Nature America, Inc.

Cancer; Antigen; Resistance Cáncer; Antígeno; Resistencia Càncer; Antigen; Resistència Despite the revolution of immunotherapy in cancer treatment, patients eventually progress due to the emergence of resistance. In this scenario, the selection of the tumor antigen can be decisive in the success of the clinical response. T cell bispecific antibodies (TCBs) are engineered molecules that include binding sites to the T cell receptor and to a tumor antigen. Using gastric CEA+/HER2+ MKN45 cells and TCBs directed against CEA or HER2, we show that the mechanism of resistance to a TCB is dependent on the tumor antigen. Acquired resistant models to a high-affinity-CEA-targeted TCB exhibit a reduction of CEA levels due to transcriptional silencing, which is reversible upon 5-AZA treatment. In contrast, a HER2-TCB resistant model maintains HER2 levels and exhibit a disruption of the interferon-gamma signaling. These results will help in the design of combinatorial strategies to increase the efficacy of cancer immunotherapies and to anticipate and overcome resistances. This work was supported by Asociación Española Contra el Cancer (AECC), Breast Cancer Research Foundation (BCRF-21-008), and Instituto de Salud Carlos III (PI19/01181). A.M.S. was funded by the Spanish Government (PFIS FI20/00188). B.M. was funded by a fellowship from PERIS (Departament de Salut, Generalitat de Catalunya). M.R.A. was funded by Agency for Management of University and Research Grants (AGAUR, 2022 FI_B2 00080). P.O.R. was funded by the BBVA. E.J.A. was funded by the AECC (POSTD211413AREN). VHIO acknowledges the Cellex Foundation for providing research facilities and equipment, the Centro de Investigación Biomédica en Red de Cáncer (CIBERONC) from the Institute of Health Carlos III (ISCIII), and the Department of Health (Generalitat de Catalunya, SLT008/18/00198 SLT008/18/00205) for their support on this research. The authors acknowledge financial support from the State Agency for Research (Agencia Estatal de Investigación) (CEX2020-001024-S/AEI/10.13039/501100011033) and for the Cancer Immunology and Immunotherapy (CAIMI-2) program funded by BBVA Foundation. We would like to remark the funding from B.M PERIS (Spain). The authors thank Dr. Anne Freimoser-Grundschober and Roche for helping provide the TCBs. The graphical abstract was created with BioRender.com.

Introduction: Immunotherapy has revolutionized the way many cancers are treated. Despite its success, cancer patients eventually progress due to the emergence of resistance. In this scenario, the selection of the tumor antigen is important and can be decisive in the success of the clinical response, particularly when resistance emerge. T cell bispecific antibodies (TCBs) are engineered molecules that include, within a single entity, binding sites to the T cell receptor and to a tumor-specific or a tumor-associated antigen. It is assumed that, as all TCBs have the same mechanism of action, mechanisms of resistance are independent of the tumor antigen. Objective: This study was designed to identify the mechanisms that trigger the resistance against HER2 and CEACAM5 T cell bispecific antibodies. We assessed TCB sensitivity using in vitro and in vivo assays combining human cell lines or patient-derived models with human lymphocytes. Material and Methods: Using gastric CEA+/HER2+ MKN45 cells as a model and TCBs directed against CEA or HER2, here we show for the first time that the mechanism of resistance to a T cell-based therapy is dependent on the tumor antigen. Results: Acquired resistant models to a high-affinity CEA-targeted TCB exhibit a reduction of CEA antigen levels due to epigenetic transcriptional silencing, which is reversible upon 5-AZA treatment. In contrast, a HER2-TCB resistant model maintains HER2 levels as HER2 downregulation led to an impairment of proliferation. Furthermore, using this latter model, we identified that disruption of interferon- gamma signaling confers resistance to killing by active T lymphocytes. Conclusions: Our results unveil different mechanisms of acquired resistance to TCBs depending on the selected antigen, which will help in the design of combinatorial strategies to increase the efficacy of cancer immunotherapies and to anticipate and overcome resistances. Citation Format: Alex Martínez-Sabadell, Beatriz Morancho, Irene Rius, Marta Escorihuela, Antonio Luque, Irene Chicote, Hector García Palmer, Marina Bacac, Joaquin Arribas, Enrique Javier Arenas Lahuerta. The target antigen determines the mechanism of acquired resistance to T cell based therapies [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 2044.

In the recent years has being a great expansion of new preclinical models of colorectal cancer (CRC) based on patient-derived cells, from ex vivo 2D cell lines, toward 3D tumoroids or animal xenografts. These new technologies have been key to overcome historical limitations in CRC research such as precision medicine, pharmacogenomic screenings, or investigating mechanism of drug resistance. Here we describe a method to generate metastatic CRC in mice with patient-derived cells and the evaluation of drug response with computerized tomography. CRC at this advanced stage is the most frequent situation in patients enrolled in therapies with novel drugs that in some cases are designed to target metastatic cells. Therefore, these orthotopic models could be considered the best to recapitulate advance CRC and are therefore becoming instrumental to investigate the biology behind drug-response in metastatic disease.

Notch pathway signaling is implicated in several human cancers. Aberrant activation and mutations of Notch signaling components are linked to tumor initiation, maintenance, and resistance to cancer therapy. Several strategies, such as monoclonal antibodies against Notch ligands and receptors, as well as small-molecule gamma-secretase inhibitors (GSIs), have been developed to interfere with Notch receptor activation at proximal points in the pathway. However, the use of drug-like small molecules to target the down-stream mediators of Notch signaling, the Notch transcription acti-vation complex, remains largely unexplored. Here, we report the discovery of an orally active small-molecule inhibitor (termed CB -103) of the Notch transcription activation complex. We show that CB-103 inhibits Notch signaling in primary human T cell acute lym-phoblastic leukemia and other Notch-dependent human tumor cell lines, and concomitantly induces cell cycle arrest and apoptosis, thereby impairing proliferation, including in GSI-resistant human tumor cell lines with chromosomal translocations and rearrange-ments in Notch genes. CB-103 produces Notch loss-of-function phenotypes in flies and mice and inhibits the growth of human breast cancer and leukemia xenografts, notably without causing the dose-limiting intestinal toxicity associated with other Notch inhibitors. Thus, we describe a pharmacological strategy that in-terferes with Notch signaling by disrupting the Notch transcription complex and shows therapeutic potential for treating Notch -driven cancers.

In the recent years has being a great expansion of new preclinical models of colorectal cancer (CRC) based on patient-derived cells, from ex vivo 2D cell lines, toward 3D tumoroids or animal xenografts. These new technologies have been key to overcome historical limitations in CRC research such as precision medicine, pharmacogenomic screenings, or investigating mechanism of drug resistance. Here we describe a method to generate metastatic CRC in mice with patient-derived cells and the evaluation of drug response with computerized tomography. CRC at this advanced stage is the most frequent situation in patients enrolled in therapies with novel drugs that in some cases are designed to target metastatic cells. Therefore, these orthotopic models could be considered the best to recapitulate advance CRC and are therefore becoming instrumental to investigate the biology behind drug-response in metastatic disease.

Dormant or slow-cycling tumor cells can form a residual chemoresistant reservoir responsible for relapse in patients, years after curative surgery and adjuvant therapy. We have adapted the pulse-chase expression of H2BeGFP for labeling and isolating slow-cycling cancer cells (SCCCs). SCCCs showed cancer initiation potential and enhanced chemoresistance. Cells at this slow-cycling status presented a distinctive nongenetic and cell-autonomous gene expression profile shared across different tumor types. We identified TET2 epigenetic enzyme as a key factor controlling SCCC numbers, survival, and tumor recurrence. 5-Hydroxymethylcytosine (5hmC), generated by TET2 enzymatic activity, labeled the SCCC genome in carcinomas and was a predictive biomarker of relapse and survival in cancer patients. We have shown the enhanced chemoresistance of SCCCs and revealed 5hmC as a biomarker for their clinical identification and TET2 as a potential drug target for SCCC elimination that could extend patients' survival.

Purpose: Oncogenic mutations in the KRAS/PI3K/AKT pathway are one of the most frequent alterations in cancer. Although PI3K or AKT inhibitors show promising results in clinical trials, drug resistance frequently emerges. We previously revealed Wnt/β-catenin signaling hyperactivation as responsible for such resistance in colorectal cancer. Here we investigate Wnt-mediated resistance in patients treated with PI3K or AKT inhibitors in clinical trials and evaluate the efficacy of a new Wnt/tankyrase inhibitor, NVP-TNKS656, to overcome such resistance. Experimental Design: Colorectal cancer patient-derived sphere cultures and mouse tumor xenografts were treated with NVP-TNKS656, in combination with PI3K or AKT inhibitors.We analyzed progression-free survival of patients treated with different PI3K/AKT/mTOR inhibitors in correlation with Wnt/β-catenin pathway activation, oncogenic mutations, clinicopathological traits, and gene expression patterns in 40 colorectal cancer baseline tumors. Results: Combination with NVP-TNKS656 promoted apoptosis in PI3K or AKT inhibitor-resistant cells with high nuclear β-catenin content. High FOXO3A activity conferred sensitivity to NVP-TNKS656 treatment. Thirteen of 40 patients presented high nuclear β-catenin content and progressed earlier upon PI3K/AKT/mTOR inhibition. Nuclear β-catenin levels predicted drug response, whereas clinicopathologic traits, gene expression profiles, or frequent mutations (KRAS, TP53, or PIK3CA) did not. Conclusions: High nuclear β-catenin content independently predicts resistance to PI3K and AKT inhibitors. Combined treatment with a Wnt/tankyrase inhibitor reduces nuclear β-catenin, reverts such resistance, and represses tumor growth. FOXO3A content and activity predicts response to Wnt/β-catenin inhibition and together with β-catenin may be predictive biomarkers of drug response providing a rationale to stratify colorectal cancer patients to be treated with PI3K/AKT/mTOR and Wnt/β-catenin inhibitors. Clin Cancer Res; 22(3); 644–56. ©2015 AACR.

4112 Background: BRAF V600 mt mCRC is an aggressive disease with poor OS under standard chemotherapy. Treatment with doublet and triplet targeted combinations, such as BRAF inhibitor+ antiEGFR+/- MEK inhibitor, has been shown to improve outcomes. Prognostic factors in this targeted treated population remain to be studied. Methods: Prospective international cohort of patients who received doublet or triplet anti-BRAF combinations in clinical trials or as compassionate use. Univariate Cox models for OS were constructed and the strongest predictors in stepwise variable selection were used to develop a prognostic score. The final multivariate model with selected predictors was stratified by prior lines. Results: In total, 42 patients were enrolled. Median age 60.7 y (33-83), 61% female, 61% right-sided tumors, 26% received 2 or more prior chemotherapy lines. One patient (2.6%) achieved complete response and 36% had partial response with median follow-up of 14.3 months. Median progression-free survival was 5.5 months (CI95% 4.4-10.4) and median OS (mOS) was 10.7 months (CI95% 8.4-22.1). In univariate models, ECOG performance status (1 vs 0), CEA levels (high - > 3.5 ng/mL- vs low - < 3.5 ng/mL), CA 19.9 (high vs. low), LDH (high vs. low), number of metastatic sites and presence of liver metastasis were significant prognostic factors. On the other hand, MSI status and peritoneal or nodal metastasis did not associate with outcome. In multivariable model, strongest determinants of OS were ECOG and baseline CEA levels. If high-risk for both factors (ECOG 1 and CEA high, 46% of the patients), mOS was 5.6 months (CI95% 4.2-NA); if intermediate-risk (either ECOG 1 or CEA high, 33%), mOS was 13.5 months (CI95% 10.6-NA); if low-risk (ECOG 0 and CEA low, 21%), mOS not reached (CI95% 16.5-NA). Differences between intermediate- and high-risk prognostic groups compared to low-risk were significant (HR = 5.9, p = 0.03; and HR = 25.9, p < 0.001, respectively). Conclusions: Patients characteristics such as ECOG and surrogates of tumor burden like CEA levels remain important OS determinants in BRAF V600 mt mCRC treated with doublet or triplet targeted therapy. In fact, there are not prognostic scores regarding BRAF mt mCRC treated with targeted therapies. Our study suggests that these prognostic factors may be considered as stratification factors in future clinical trials.

Purpose: Within the aim of advancing precision oncology, we have generated a collection of patient-derived xenografts (PDX) characterized at the molecular level, and a preclinical model of colon cancer metastasis to evaluate drug-response and tumor progression. Experimental Design: We derived cells from 32 primary colorectal carcinomas and eight liver metastases and generated PDX annotated for their clinical data, gene expression, mutational, and histopathological traits. Six models were injected orthotopically into the cecum wall of NOD-SCID mice in order to evaluate metastasis. Three of them were treated with chemotherapy (oxaliplatin) and three with API2 to target AKT activity. Tumor growth and metastasis progression were analyzed by positron emission tomography (PET). Results: Patient-derived cells generated tumor xenografts that recapitulated the same histopathological and genetic features as the original patients' carcinomas. We show an 87.5% tumor take rate that is one of the highest described for implanted cells derived from colorectal cancer patients. Cecal injection generated primary carcinomas and distant metastases. Oxaliplatin treatment prevented metastasis and API2 reduced tumor growth as evaluated by PET. Conclusions: Our improved protocol for cancer cell engraftment has allowed us to build a rapidly expanding collection of colorectal PDX, annotated for their clinical data, gene expression, mutational, and histopathological statuses. We have also established a mouse model for metastatic colon cancer with patient-derived cells in order to monitor tumor growth, metastasis evolution, and response to treatment by PET. Our PDX models could become the best preclinical approach through which to validate new biomarkers or investigate the metastatic potential and drug-response of individual patients. Clin Cancer Res; 19(24); 6787–801. ©2013 AACR.

Purpose: PI3K pathway activation occurs in concomitance with RAS/BRAF mutations in colorectal cancer, limiting the sensitivity to targeted therapies. Several clinical studies are being conducted to test the tolerability and clinical activity of dual MEK and PI3K pathway blockade in solid tumors. Experimental Design: In the present study, we explored the efficacy of dual pathway blockade in colorectal cancer preclinical models harboring concomitant activation of the ERK and PI3K pathways. Moreover, we investigated if TP53 mutation affects the response to this therapy. Results: Dual MEK and mTORC1/2 blockade resulted in synergistic antiproliferative effects in cell lines bearing alterations in KRAS/BRAF and PIK3CA/PTEN. Although the on-treatment cell-cycle effects were not affected by the TP53 status, a marked proapoptotic response to therapy was observed exclusively in wild-type TP53 colorectal cancer models. We further interrogated two independent panels of KRAS/BRAF- and PIK3CA/PTEN-altered cell line– and patient-derived tumor xenografts for the antitumor response toward this combination of agents. A combination response that resulted in substantial antitumor activity was exclusively observed among the wild-type TP53 models (two out of five, 40%), but there was no such response across the eight mutant TP53 models (0%). Interestingly, within a cohort of 14 patients with colorectal cancer treated with these agents for their metastatic disease, two patients with long-lasting responses (32 weeks) had TP53 wild-type tumors. Conclusions: Our data support that, in wild-type TP53 colorectal cancer cells with ERK and PI3K pathway alterations, MEK blockade results in potent p21 induction, preventing apoptosis to occur. In turn, mTORC1/2 inhibition blocks MEK inhibitor–mediated p21 induction, unleashing apoptosis. Clin Cancer Res; 21(24); 5499–510. ©2015 AACR.

et al., SPROUTY2 (SPRY2) is an intracellular regulator of receptor tyrosine kinase signaling involved in cell growth, differentiation and tumorigenesis. Here, we show that SPRY2 is a target gene of the Wnt/β-catenin pathway that is abnormally activated in more than 90% of colon carcinomas. In human colon cancer cells, SPRY2 expression is induced by β-catenin in co-operation with the transcription factor FOXO3a instead of lymphoid enhancer factor/T-cell factor proteins. We found binding of β-catenin to the SPRY2 promoter at FOXO3a response elements. In vivo, cells marked by nuclear β-catenin and FOXO3a express SPRY2 in proliferative epithelial tissues, such as intestinal mucosa and epidermis. Consistently, inducible β-catenin deletion in mice reduced Spry2 expression in the small intestine. Moreover, SPRY2 protein expression correlated with nuclear β-catenin and FOXO3a colocalization in human colon carcinomas. Importantly, the amount of SPRY2 protein correlated with shorter overall survival of colon cancer patients. Our data reveal SPRY2 as a novel Wnt/β-catenin and FOXO3a target gene indicative of poor prognosis in colon cancer., Experiments were supported by grants from Fondo Europeo de Desarrollo Regional-Instituto de Salud Carlos III and Spanish Cooperative Research Network on Cancer (RTICC) (FIS-PI081356, RD12/0036/0001, RD12/0036/0021 and RD12/0036/0012), Plan Nacional de Biomedicina, Ministerio de Economía y Competitividad (SAF-18302) and Comunidad de Madrid (S2010/BMD-2344 Colomics2). HGP was supported by the Miguel Servet Program, Instituto de Salud Carlos III (ISCIII) and PO-M by an EMBO research fellowship.

et al., Aberrant activation of the Wnt/beta-catenin pathway is critical for the initiation and progression of most colon cancers. This activation provokes the accumulation of nuclear beta-catenin and the induction of its target genes. Apc(min/+) mice are the most commonly used model for colon cancer. They harbor a mutated Apc allele and develop intestinal adenomas and carcinomas during the first months of life. This phenotype is caused by the mutation of the second Apc allele and the consequent accumulation of nuclear beta-catenin in the affected cells. Here we describe that vitamin D receptor (VDR) is a crucial modulator of nuclear beta-catenin levels in colon cancer in vivo. By appropriate breeding of Apc(min/+) mice and Vdr(+/-) mice we have generated animals expressing a mutated Apc allele and two, one, or none Vdr wild type alleles. Lack of Vdr increased the number of colonic Aberrant Crypt Foci (ACF) but not that of adenomas or carcinomas in either small intestine or colon. Importantly, colon ACF and tumors of Apc(min/+) Vdr(-/-) mice had increased nuclear beta-catenin and the tumors reached a larger size than those of Apc(min/+) Vdr(+/+). Both ACF and carcinomas in Apc(min/+) Vdr(-/-) mice showed higher expression of beta-catenin/TCF target genes. In line with this, VDR knock-down in cultured human colon cancer cells enhanced beta-catenin nuclear content and target gene expression. Consistently, VDR depletion abrogated the capacity of 1,25(OH)(2)D(3) to promote the relocation of beta-catenin from the nucleus to the plasma membrane and to inhibit beta-catenin/TCF target genes. In conclusion, VDR controls the level of nuclear beta-catenin in colon cancer cells and can therefore attenuate the impact of oncogenic mutations that activate the Wnt/beta-catenin pathway., This work was supported by Fondo de Investigación Sanitaria-Instituto de Salud Carlos III (ISCIII, PI081356), Vall d’Hebrón Institute of Oncology, Olga Torres Foundation, Fundación de la Asociación Española de Lucha Contra el Cáncer (AECC), Ministerio de Ciencia e Innovación (SAF2010-18302) and Red Temática de Investigación Cooperativa en Cáncer (ISCIII, RD06/0020/0009).

Reference EPFL-CONF-185612View record in Web of Science Record created on 2013-03-28, modified on 2017-05-12