bims-pimaco Biomed News
on PI3K and MAPK signalling in colorectal cancer
Issue of 2020‒10‒18
twenty-two papers selected by
Lucas B. Zeiger
Beatson Institute for Cancer Research
  1. Lactate Lights up PI3K Inhibitor Resistance in Breast Cancer.
  2. Rechallenge with Anti-EGFR Therapy in Metastatic Colorectal Cancer (mCRC): Results from South Australia mCRC Registry.
  3. PIK3CA mutation enrichment and quantitation from blood and tissue.
  4. Role of Cyclooxygenase-2 in colorectal cancer patients.
  5. cGAS suppresses genomic instability as a decelerator of replication forks.
  6. Cancer research using organoid technology.
  7. A Wnt-induced phenotypic switch in cancer-associated fibroblasts inhibits EMT in colorectal cancer.
  8. SHP2 inhibition diminishes KRASG12C cycling and promotes tumor microenvironment remodeling.
  9. A small subset of Wnt-activated cells is an initiator of regrowth in colorectal cancer organoids after irradiation.
  10. Gene Expression Profiling Identifies Akt as a Target for Radiosensitization in Gastric Cancer Cells.
  11. Cyclooxygenase activity mediates colorectal cancer cell resistance to the omega-3 polyunsaturated fatty acid eicosapentaenoic acid.
  12. SYPL1 Inhibits Apoptosis in Pancreatic Ductal Adenocarcinoma via Suppression of ROS-Induced ERK Activation.
  13. Rictor promotes cell migration and actin polymerization through regulating ABLIM1 phosphorylation in Hepatocellular Carcinoma.
  14. Targeting DNA Damage Response and Replication Stress in Pancreatic Cancer.
  15. Repair of G1 induced DNA double-strand breaks in S-G2/M by alternative NHEJ.
  16. PI3K Driver Mutations: A Biophysical Membrane-Centric Perspective.
  17. Evaluation of the RAS signaling network in response to MEK inhibition using organoids derived from a familial adenomatous polyposis patient.
  18. Association of Consensus Molecular Subtypes and Molecular Markers With Clinical Outcomes in Patients With Metastatic Colorectal Cancer: Biomarker Analyses From LUME-Colon 1.
  19. Control of Glucocorticoid Receptor Levels by PTEN Establishes a Failsafe Mechanism for Tumor Suppression.
  20. Chemotherapeutic effects of MEK kinase inhibitor and BRAF kinase inhibitor on KRAS-mutated human colon cancer cell lines with different microsatellite instability.
  21. PI3K mutations detected in liquid biopsy are associated to reduced sensitivity to CDK4/6 inhibitors in metastatic breast cancer patients.
  22. Targeting KRAS(G12C): From Inhibitory Mechanism to Modulation of Antitumor Effects in Patients.
  1. Cancer Cell. 2020 Oct 12. pii: S1535-6108(20)30485-2. [Epub ahead of print]38(4): 441-443
    Lactate Lights up PI3K Inhibitor Resistance in Breast Cancer.
    Alissandra L Hillis, Alex Toker.
      Predictive biomarkers can facilitate optimal patient selection for targeted cancer therapies. In this issue of Cancer Cell, Ros et al. show the utility of noninvasive metabolic imaging of labeled carbon transfer from pyruvate to lactate to detect early response and FOXM1-mediated resistance to PI3K inhibition in estrogen-receptor-positive breast cancer.
    DOI:  https://doi.org/10.1016/j.ccell.2020.09.011
  2. Target Oncol. 2020 Oct 17.
    Rechallenge with Anti-EGFR Therapy in Metastatic Colorectal Cancer (mCRC): Results from South Australia mCRC Registry.
    Li Chia Chong, Jennifer E Hardingham, Amanda R Townsend, Cynthia Piantadosi, Gonzalo Tapia Rico, Chris Karapetis, Rob Padbury, Guy Maddern, Amitesh Roy, Timothy J Price.
      BACKGROUND: Anti-epidermal growth factor receptor (EGFR) monoclonal antibodies (cetuximab or panitumumab) are today increasingly used in the first- or second-line setting for RAS wild-type metastatic colorectal cancer (CRC) patients. Following progression beyond third- or fourth-line therapy, some patients are unsuitable for further chemotherapy because of poor performance status or patient choice. However, a significant number of patients are still candidates for further therapy despite limited standard options being available. The role of rechallenge with anti-EGFR therapy, particularly in patients who had previously responded, is often considered, but there is limited evidence in the literature to support such a strategy.OBJECTIVE: This retrospective study aims to review the outcome of metastatic CRC patients who had anti-EGFR rechallenge.
    PATIENTS AND METHODS: Patients who had been rechallenged with anti-EGFR therapy were identified from the South Australian metastatic CRC database. Patient characteristics were recorded and tumor response was retrospectively assessed using Response Evaluation Criteria in Solid Tumors (RECIST). Kaplan-Meier analysis was used to assess progression free survival (PFS) for each rechallenge and overall survival (OS).
    RESULTS: Twenty-two patients were eligible for inclusion in this analysis. Disease control rate (stable disease and partial response) was 45.4% (ten patients) for patients who received rechallenge anti-EGFR. Seven patients received a second rechallenge and disease control rate was 28.6% (two patients). The median interval time between initial anti-EGFR therapy and rechallenge was 13.5 months. The median PFS after rechallenge 1 was 4.1 months and after rechallenge 2 was 3.5 months. The median OS was 7.7 months from date of rechallenge.
    CONCLUSIONS: Anti-EGFR rechallenge provides clinical benefit in patients with RAS wild-type metastatic CRC.
    DOI:  https://doi.org/10.1007/s11523-020-00760-8
  3. Sci Rep. 2020 Oct 13. 10(1): 17082
    PIK3CA mutation enrichment and quantitation from blood and tissue.
    Ieva Keraite, Virginia Alvarez-Garcia, Isaac Garcia-Murillas, Matthew Beaney, Nicholas C Turner, Clare Bartos, Olga Oikonomidou, Maïwenn Kersaudy-Kerhoas, Nicholas R Leslie.
      PIK3CA is one of the two most frequently mutated genes in breast cancers, occurring in 30-40% of cases. Four frequent 'hotspot' PIK3CA mutations (E542K, E545K, H1047R and H1047L) account for 80-90% of all PIK3CA mutations in human malignancies and represent predictive biomarkers. Here we describe a PIK3CA mutation specific nuclease-based enrichment assay, which combined with a low-cost real-time qPCR detection method, enhances assay detection sensitivity from 5% for E542K and 10% for E545K to 0.6%, and from 5% for H1047R to 0.3%. Moreover, we present a novel flexible prediction method to calculate initial mutant allele frequency in tissue biopsy and blood samples with low mutant fraction. These advancements demonstrated a quick, accurate and simple detection and quantitation of PIK3CA mutations in two breast cancer cohorts (first cohort n = 22, second cohort n = 25). Hence this simple, versatile and informative workflow could be applicable for routine diagnostic testing where quantitative results are essential, e.g. disease monitoring subject to validation in a substantial future study.
    DOI:  https://doi.org/10.1038/s41598-020-74086-w
  4. Front Biosci (Landmark Ed). 2021 Jan 01. 26 706-716
    Role of Cyclooxygenase-2 in colorectal cancer patients.
    Neha Nanda, Devinder K Dhawan.
      Cyclooxygenase-2 (COX-2) is an inducible enzyme which triggers the biosynthesis of prostaglandins. COX-2 is activated in response to inflammatory stimuli and is one of the major molecules that is involved in the development and progression of colorectal cancer (CRC). Consistent with such a conceptual framework, it has been shown that COX-2 inhibitors prevent the carcinogenesis of CRC and aid in the treatment of sporadic or familial cases of CRC as shown by an overall increase in the survival rate. However, prolonged use of these inhibitors is associated with increase risk of development of cardiovascular complications, implying that further research is required to identify COX-2 inhibitors that are associated with lower risks of such complications.
  5. Sci Adv. 2020 Oct;pii: eabb8941. [Epub ahead of print]6(42):
    cGAS suppresses genomic instability as a decelerator of replication forks.
    Hao Chen, Hao Chen, Jiamin Zhang, Yumin Wang, Antoine Simoneau, Hui Yang, Arthur S Levine, Lee Zou, Zhijian Chen, Li Lan.
      The cyclic GMP-AMP synthase (cGAS), a sensor of cytosolic DNA, is critical for the innate immune response. Here, we show that loss of cGAS in untransformed and cancer cells results in uncontrolled DNA replication, hyperproliferation, and genomic instability. While the majority of cGAS is cytoplasmic, a fraction of cGAS associates with chromatin. cGAS interacts with replication fork proteins in a DNA binding-dependent manner, suggesting that cGAS encounters replication forks in DNA. Independent of cGAMP and STING, cGAS slows replication forks by binding to DNA in the nucleus. In the absence of cGAS, replication forks are accelerated, but fork stability is compromised. Consequently, cGAS-deficient cells are exposed to replication stress and become increasingly sensitive to radiation and chemotherapy. Thus, by acting as a decelerator of DNA replication forks, cGAS controls replication dynamics and suppresses replication-associated DNA damage, suggesting that cGAS is an attractive target for exploiting the genomic instability of cancer cells.
    DOI:  https://doi.org/10.1126/sciadv.abb8941
  6. J Mol Med (Berl). 2020 Oct 14.
    Cancer research using organoid technology.
    Kai Kretzschmar.
      Organoid technology has rapidly transformed basic biomedical research and contributed to significant discoveries in the last decade. With the application of protocols to generate organoids from cancer tissue, organoid technology has opened up new opportunities for cancer research and therapy. Using organoid cultures derived from healthy tissues, different aspects of tumour initiation and progression are widely studied including the role of pathogens or specific cancer genes. Cancer organoid cultures, on the other hand, are applied to generate biobanks, perform drug screens, and study mutational signatures. With the incorporation of cellular components of the tumour microenvironment such as immune cells into the organoid cultures, the technology is now also exploited in the rapidly advancing field of immuno-oncology. In this review, I discuss how organoid technology is currently being utilised in cancer research and what obstacles are still to be overcome for its broader use in anti-cancer therapy.
    Keywords:  3D culture; Cancer; Drug screening; Immuno-oncology; Organoids; Personalised medicine; Pre-clinical models
    DOI:  https://doi.org/10.1007/s00109-020-01990-z
  7. Cancer Res. 2020 Oct 14. pii: canres.0263.2020. [Epub ahead of print]
    A Wnt-induced phenotypic switch in cancer-associated fibroblasts inhibits EMT in colorectal cancer.
    Mohammed H Mosa, Birgitta E Michels, Constantin Menche, Adele M Nicolas, Tahmineh Darvishi, Florian R Greten, Henner F Farin.
      Tumor progression is recognized as a result of an evolving crosstalk between tumor cells and their surrounding non-transformed stroma. Although Wnt signaling has been intensively studied in colorectal cancer (CRC), it remains unclear whether activity in the tumor-associated stroma contributes to malignancy. To specifically interfere with stromal signals, we generated Wnt-independent tumor organoids that secrete the Wnt antagonist Sfrp1. Subcutaneous transplantation into immunocompetent as well as immunodeficient mice resulted in a strong reduction of tumor growth. Histological and transcriptomic analyses revealed that Sfrp1 induced an EMT (epithelial-mesenchymal transition) phenotype in tumor cells without affecting tumor-intrinsic Wnt signaling, suggesting involvement of non-immune stromal cells. Blockage of canonical signaling using Sfrp1, Dkk1, or fibroblast-specific genetic ablation of β-catenin strongly decreased the number of cancer-associated myofibroblasts (myCAFs). Wnt activity in CAFs was linked with distinct subtypes, where low and high levels induced an inflammatory-like CAF (iCAF) subtype or contractile myCAFs, respectively. Co-culture of tumor organoids with iCAFs resulted in significant upregulation of EMT markers, while myCAFs reverted this phenotype. In summary, we show that tumor growth and malignancy are differentially regulated via distinct fibroblast subtypes under the influence of juxtacrine Wnt signals.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-20-0263
  8. J Exp Med. 2021 Jan 04. pii: e20201414. [Epub ahead of print]218(1):
    SHP2 inhibition diminishes KRASG12C cycling and promotes tumor microenvironment remodeling.
    Carmine Fedele, Shuai Li, Kai Wen Teng, Connor J R Foster, David Peng, Hao Ran, Paolo Mita, Mitchell J Geer, Takamitsu Hattori, Akiko Koide, Yubao Wang, Kwan Ho Tang, Joshua Leinwand, Wei Wang, Brian Diskin, Jiehui Deng, Ting Chen, Igor Dolgalev, Ugur Ozerdem, George Miller, Shohei Koide, Kwok-Kin Wong, Benjamin G Neel.
      KRAS is the most frequently mutated human oncogene, and KRAS inhibition has been a longtime goal. Recently, inhibitors were developed that bind KRASG12C-GDP and react with Cys-12 (G12C-Is). Using new affinity reagents to monitor KRASG12C activation and inhibitor engagement, we found that an SHP2 inhibitor (SHP2-I) increases KRAS-GDP occupancy, enhancing G12C-I efficacy. The SHP2-I abrogated RTK feedback signaling and adaptive resistance to G12C-Is in vitro, in xenografts, and in syngeneic KRASG12C-mutant pancreatic ductal adenocarcinoma (PDAC) and non-small cell lung cancer (NSCLC). SHP2-I/G12C-I combination evoked favorable but tumor site-specific changes in the immune microenvironment, decreasing myeloid suppressor cells, increasing CD8+ T cells, and sensitizing tumors to PD-1 blockade. Experiments using cells expressing inhibitor-resistant SHP2 showed that SHP2 inhibition in PDAC cells is required for PDAC regression and remodeling of the immune microenvironment but revealed direct inhibitory effects on tumor angiogenesis and vascularity. Our results demonstrate that SHP2-I/G12C-I combinations confer a substantial survival benefit in PDAC and NSCLC and identify additional potential combination strategies.
    DOI:  https://doi.org/10.1084/jem.20201414
  9. Cancer Sci. 2020 Oct 11.
    A small subset of Wnt-activated cells is an initiator of regrowth in colorectal cancer organoids after irradiation.
    Hiroko Endo, Jumpei Kondo, Kunishige Onuma, Masayuki Ohue, Masahiro Inoue.
      Most colorectal cancers (CRCs) are differentiated adenocarcinomas, which maintain expression of both stemness and differentiation markers. This observation suggests that CRC cells could retain a regeneration system of normal cells upon injury. However, the role of stemness in cancer cell regeneration after irradiation is poorly understood. Here, we examined the effect of radiation on growth, stemness, and differentiation in organoids derived from differentiated adenocarcinomas. Following a sub-lethal dose of irradiation, proliferation and stemness markers, including WNT target genes, were drastically reduced, but differentiation markers remained. After a static growth phase after high dose of radiation, regrowth foci appeared; these consisted of highly proliferating cells that expressed stem cell markers. Radio-sensitivity and the ability to form foci differed among the cancer tissue-originated spheroid (CTOS) lines examined and showed good correlation with in vivo radiation sensitivity. Pre-treating organoids with histone deacetylase inhibitors increased radiation sensitivity; this increase was accompanied by the suppression of Wnt signal-related gene expression. Accordingly, Wnt inhibitors increased organoid radio-sensitivity. These results suggested that only a small subset of but not all cancer cells with high Wnt activity at the time of irradiation could give rise to foci formation. In conclusion, we established a radiation sensitivity assay using CRC organoids that could provide a novel platform for evaluating the effects of radiosensitizers on differentiated adenocarcinomas in CRC.
    Keywords:  Wnt; colorectal cancer; organoid; radiation sensitivity assay; radiosensitizer
    DOI:  https://doi.org/10.1111/cas.14683
  10. Front Oncol. 2020 ;10 562284
    Gene Expression Profiling Identifies Akt as a Target for Radiosensitization in Gastric Cancer Cells.
    Kyung Hwan Kim, Han Sang Kim, Sang Cheol Kim, DooA Kim, Yong Bae Kim, Hyun Cheol Chung, Sun Young Rha.
      Background: Despite the important role of radiotherapy in cancer treatment, a subset of patients responds poorly to treatment majorly due to radioresistance. Particularly the role of radiotherapy has not been established in gastric cancer (GC). Herein, we aimed to identify a radiosensitivity gene signature and to discover relevant targets to enhance radiosensitivity in GC cells.Methods: An oligonucleotide microarray (containing 22,740 probes) was performed in 12 GC cell lines prior to radiation. A clonogenic assay was performed to evaluate the survival fraction at 2 Gy (SF2) as a surrogate marker for radiosensitivity. Genes differentially expressed (fold change > 6, q-value < 0.025) were identified between radiosensitive and radioresistant cell lines, and quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) was performed for validation. Gene set and pathway analyses were performed using Ingenuity Pathway Analysis (IPA).
    Results: Radiosensitive (SF2 < 0.4) and radioresistant cell lines (SF2 ≥ 0.6) exhibited a marked difference in gene expression. We identified 68 genes that are differentially expressed between radiosensitive and radioresistant cell lines. The identified genes showed interactions via AKT, HIF1A, TGFB1, and TP53, and their functions were associated with the genetic networks associated with cellular growth and proliferation, cellular movement, and cell cycle. The Akt signaling pathway exhibited the highest association with radiosensitivity. Combinatorial treatment with MK-2206, an allosteric Akt inhibitor, and radiotherapy significantly increased cell death compared with radiotherapy alone in two radioresistant cell lines (YCC-2 and YCC-16).
    Conclusion: We identified a GC-specific radiosensitivity gene signature and suggest that the Akt signaling pathway could serve as a therapeutic target for GC radiosensitization.
    Keywords:  Akt; MK-2206; gastric cancer; gene signature; radiosensitivity
    DOI:  https://doi.org/10.3389/fonc.2020.562284
  11. Cancer Chemother Pharmacol. 2020 Oct 11.
    Cyclooxygenase activity mediates colorectal cancer cell resistance to the omega-3 polyunsaturated fatty acid eicosapentaenoic acid.
    Milene Volpato, Nicola Ingram, Sarah L Perry, Jade Spencer, Amanda D Race, Catriona Marshall, John M Hutchinson, Anna Nicolaou, Paul M Loadman, P Louise Coletta, Mark A Hull.
      PURPOSE: The naturally-occurring omega-3 polyunsaturated fatty acid eicosapentaenoic acid (EPA) is safe, well-tolerated and inexpensive, making it an attractive anti-cancer intervention. However, EPA has only modest anti-colorectal cancer (CRC) activity, when used alone. Both cyclooxygenase (COX) isoforms metabolise EPA and are over-expressed in CRC cells. We investigated whether COX inhibition increases the sensitivity of CRC cells to growth inhibition by EPA.METHODS: A panel of 18 human and mouse CRC cell lines was used to characterize the differential sensitivity of CRC cells to the growth inhibitory effects of EPA. The effect of CRISPR-Cas9 genetic deletion and pharmacological inhibition of COX-1 and COX-2 on the anti-cancer activity of EPA was determined using in vitro and in vivo models.
    RESULTS: Genetic ablation of both COX isoforms increased sensitivity of CT26 mouse CRC cells to growth inhibition by EPA in vitro and in vivo. The non-selective COX inhibitor aspirin and the selective COX-2 inhibitor celecoxib increased sensitivity of several human and mouse CRC cell lines to EPA in vitro. However, in a MC38 mouse CRC cell tumour model, with dosing that mirrored low-dose aspirin use in humans, thereby producing significant platelet COX-1 inhibition, there was ineffective intra-tumoral COX-2 inhibition by aspirin and no effect on EPA sensitivity of MC38 cell tumours.
    CONCLUSION: Cyclooxygenase inhibition by non-steroidal anti-inflammatory drugs represents a therapeutic opportunity to augment the modest anti-CRC activity of EPA. However, intra-tumoral COX inhibition is likely to be critical for this drug-nutrient interaction and careful tissue pharmacodynamic profiling is required in subsequent pre-clinical and human studies.
    Keywords:  Aspirin; Cancer pharmacology; Celecoxib; Colorectal cancer; Cyclooxygenase; Drug metabolism; Eicosapentaenoic acid
    DOI:  https://doi.org/10.1007/s00280-020-04157-2
  12. Front Oncol. 2020 ;10 1482
    SYPL1 Inhibits Apoptosis in Pancreatic Ductal Adenocarcinoma via Suppression of ROS-Induced ERK Activation.
    Yunda Song, Xuesong Sun, Fangting Duan, Chaobin He, Jiali Wu, Xin Huang, Kaili Xing, Shuxin Sun, Ruiqi Wang, Fengxiao Xie, Yize Mao, Jun Wang, Shengping Li.
      Synaptophysin-like 1 (SYPL1) is a neuroendocrine-related protein. The role of SYPL1 in pancreatic ductal adenocarcinoma (PDAC) and the underlying molecular mechanism remain unclarified. Here, after analyzing five datasets (GSE15471, GSE16515, GSE28735, TCGA, and PACA-AU) and 78 PDAC patients from Sun Yat-sen University Cancer Center, we demonstrated that SYPL1 was upregulated in PDAC and that a high level of SYPL1 indicated poor prognosis. Bioinformatics analysis implied that SYPL1 was related to cell proliferation and cell death. To validate these findings, gain-of-function and loss-of-function experiments were carried out, and we found that SYPL1 promoted cell proliferation in vitro and in vivo and that it protected cells from apoptosis. Mechanistic studies revealed that sustained extracellular-regulated protein kinase (ERK) activation was responsible for the cell death resulting from knockdown of SYPL1. In addition, bioinformatics analysis showed that the expression of SYPL1 positively correlated with antioxidant activity. Reactive oxygen species (ROS) were upregulated in cells with SYPL1 knockdown and vice versa. Upregulated ROS led to ERK activation and cell death. These results suggest that SYPL1 plays a vital role in PDAC and promotes cancer cell survival by suppressing ROS-induced ERK activation.
    Keywords:  ERK; ROS; SYPL1; apoptosis; pancreatic ductal adenocarcinoma
    DOI:  https://doi.org/10.3389/fonc.2020.01482
  13. Int J Biol Sci. 2020 ;16(15): 2835-2852
    Rictor promotes cell migration and actin polymerization through regulating ABLIM1 phosphorylation in Hepatocellular Carcinoma.
    Xin Dong, Mei Feng, Hui Yang, Hengkang Liu, Hua Guo, Xianshu Gao, Yucun Liu, Rong Liu, Ning Zhang, Ruibing Chen, Ruirui Kong.
      As one of the most ominous malignancies, hepatocellular carcinoma (HCC) is frequently diagnosed at an advanced stage, owing to its aggressive invasion and metastatic spread. Emerging evidence has demonstrated that Rictor, as a unique component of the mTORC2, plays a role in cell migration, as it is dysregulated in various cancers, including HCC. However, the underlying molecular mechanism has not been well-characterized. Here, evaluation on a tissue-array panel and bioinformatics analysis revealed that Rictor is highly expressed in HCC tissues. Moreover, increased Rictor expression predicts poor survival of HCC patients. Rictor knockdown significantly suppressed cell migration and actin polymerization, thereby leading to decreased nuclear accumulation of MKL1 and subsequent inactivation of SRF/MKL1-dependent gene transcription, i.e. Arp3 and c-Fos. Mechanistically, we identified ABLIM1 as a previously unknown phosphorylation target of Rictor. Rictor interacts with ABLIM1 and regulates its serine phosphorylation in HCC cells. We generated ABLIM1 knockout cell lines of HCC, in which dominant negative mutations of Ser 214 and Ser 431 residues inhibited the ABLIM1-mediated actin polymerization and the MKL1 signaling pathway. Overall, ABLIM1 phosphorylation induced by Rictor plays an important role in controlling actin polymerization in HCC cells.
    Keywords:  ABLIM1; Rictor; actin polymerization; hepatocellular carcinoma; phosphorylation
    DOI:  https://doi.org/10.7150/ijbs.46285
  14. Gastroenterology. 2020 Oct 08. pii: S0016-5085(20)35229-X. [Epub ahead of print]
    Targeting DNA Damage Response and Replication Stress in Pancreatic Cancer.
    Stephan B Dreyer, Rosie Upstill-Goddard, Viola Paulus-Hock, Clara Paris, Eirini-Maria Lampraki, Eloise Dray, Bryan Serrels, Giuseppina Caligiuri, Selma Rebus, Dennis Plenker, Zachary Galluzzo, Holly Brunton, Richard Cunningham, Mathias Tesson, Craig Nourse, Ulla-Maja Bailey, Marc Jones, Kim Moran-Jones, Derek W Wright, Fraser Duthie, Karin Oien, Lisa Evers, Colin J McKay, Grant A McGregor, Aditi Gulati, Rachel Brough, Ilirjana Bajrami, Stephan Pettitt, Michele L Dziubinski, Juliana Candido, Frances Balkwill, Simon T Barry, Robert Grützmann, Lola Rahib, , , Amber Johns, Marina Pajic, Fieke E M Froeling, Phillip Beer, Elizabeth A Musgrove, Gloria M Petersen, Alan Ashworth, Margaret C Frame, Howard C Crawford, Diane M Simeone, Chris Lord, Debabrata Mukhopadhyay, Christian Pilarsky, David A Tuveson, Susanna L Cooke, Nigel B Jamieson, Jennifer P Morton, Owen J Sansom, Peter J Bailey, Andrew V Biankin, David K Chang.
      BACKGROUND AND AIMS: Continuing recalcitrance to therapy cements pancreatic cancer (PC) as the most lethal malignancy, which is set to become the second leading cause of cancer death in our society. The study aim was to investigate the association between DNA damage response (DDR), replication stress and novel therapeutic response in PC to develop a biomarker driven therapeutic strategy targeting DDR and replication stress in PC.METHODS: We interrogated the transcriptome, genome, proteome and functional characteristics of 61 novel PC patient-derived cell lines to define novel therapeutic strategies targeting DDR and replication stress. Validation was done in patient derived xenografts and human PC organoids.
    RESULTS: Patient-derived cell lines faithfully recapitulate the epithelial component of pancreatic tumors including previously described molecular subtypes. Biomarkers of DDR deficiency, including a novel signature of homologous recombination deficiency, co-segregates with response to platinum (P < 0.001) and PARP inhibitor therapy (P < 0.001) in vitro and in vivo. We generated a novel signature of replication stress with which predicts response to ATR (P < 0.018) and WEE1 inhibitor (P < 0.029) treatment in both cell lines and human PC organoids. Replication stress was enriched in the squamous subtype of PC (P < 0.001) but not associated with DDR deficiency.
    CONCLUSIONS: Replication stress and DDR deficiency are independent of each other, creating opportunities for therapy in DDR proficient PC, and post-platinum therapy.
    Keywords:  DNA damage response; Pancreatic cancer; personalized medicine; replication stress
    DOI:  https://doi.org/10.1053/j.gastro.2020.09.043
  15. Nat Commun. 2020 Oct 16. 11(1): 5239
    Repair of G1 induced DNA double-strand breaks in S-G2/M by alternative NHEJ.
    Wei Yu, Chloé Lescale, Loelia Babin, Marie Bedora-Faure, Hélène Lenden-Hasse, Ludivine Baron, Caroline Demangel, José Yelamos, Erika Brunet, Ludovic Deriano.
      The alternative non-homologous end-joining (NHEJ) pathway promotes DNA double-strand break (DSB) repair in cells deficient for NHEJ or homologous recombination, suggesting that it operates at all stages of the cell cycle. Here, we use an approach in which DNA breaks can be induced in G1 cells and their repair tracked, enabling us to show that joining of DSBs is not functional in G1-arrested XRCC4-deficient cells. Cell cycle entry into S-G2/M restores DSB repair by Pol θ-dependent and PARP1-independent alternative NHEJ with repair products bearing kilo-base long DNA end resection, micro-homologies and chromosome translocations. We identify a synthetic lethal interaction between XRCC4 and Pol θ under conditions of G1 DSBs, associated with accumulation of unresolved DNA ends in S-G2/M. Collectively, our results support the conclusion that the repair of G1 DSBs progressing to S-G2/M by alternative NHEJ drives genomic instability and represent an attractive target for future DNA repair-based cancer therapies.
    DOI:  https://doi.org/10.1038/s41467-020-19060-w
  16. Cancer Res. 2020 Oct 12. pii: canres.0911.2020. [Epub ahead of print]
    PI3K Driver Mutations: A Biophysical Membrane-Centric Perspective.
    Mingzhen Zhang, Hyunbum Jang, Ruth Nussinov.
      Ras activates its effectors at the membrane. Active PI3Kα and its associated kinases/phosphatases assemble at membrane regions enriched in signaling lipids. By contrast, the Raf kinase domain extends into the cytoplasm and its assembly is away from the crowded membrane surface. Our structural membrane-centric outlook underscores the spatiotemporal principles of membrane and signaling lipids which helps clarify PI3Kα activation. Here we focus on mechanisms of activation driven by PI3Kα driver mutations, spotlighting the PI3Kα double (multiple) activating mutations. Single mutations can be potent, but double mutations are stronger: their combination is specific, a single strong driver cannot fully activate PI3K, and two weak drivers may or may not do so. By contrast, two strong drivers may successfully activate PI3K, where one, e.g. H1047R, modulates membrane interactions facilitating substrate binding at the active site (km) and the other, e.g. E542K and E545K, reduces the transition state barrier (ka), releasing autoinhibition by nSH2. Although mostly unidentified, weak drivers are expected to be common, so we ask here how common double mutations are likely to be and why PI3Kα with double mutations responds effectively to inhibitors. We provide a structural view of hotspot and weak driver mutations in PI3Kα activation, explain their mechanisms, compare these with mechanisms of Raf activation, and point to targeting cell-specific, chromatin-accessible, and parallel (or redundant) pathways to thwart the expected emergence of drug resistance. Collectively, our biophysical outlook delineates activation and highlights the challenges of drug resistance.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-20-0911
  17. Sci Rep. 2020 Oct 15. 10(1): 17455
    Evaluation of the RAS signaling network in response to MEK inhibition using organoids derived from a familial adenomatous polyposis patient.
    Hiroki Osumi, Atsushi Muroi, Mizuho Sakahara, Hiroshi Kawachi, Takuya Okamoto, Yasuko Natsume, Hitomi Yamanaka, Hiroshi Takano, Daisuke Kusama, Eiji Shinozaki, Akira Ooki, Kensei Yamaguchi, Masashi Ueno, Kengo Takeuchi, Tetsuo Noda, Satoshi Nagayama, Naohiko Koshikawa, Ryoji Yao.
      RAS signaling is a promising target for colorectal cancer (CRC) therapy, and a variety of selective inhibitors have been developed. However, their use has often failed to demonstrate a significant benefit in CRC patients. Here, we used patient-derived organoids (PDOs) derived from a familial adenomatous polyposis (FAP) patient to analyze the response to chemotherapeutic agents targeting EGFR, BRAF and MEK. We found that PDOs carrying KRAS mutations were resistant to MEK inhibition, while those harboring the BRAF class 3 mutation were hypersensitive. We used a systematic approach to examine the phosphorylation of RAS effectors using reverse-phase protein array (RPPA) and found increased phosphorylation of MEK induced by binimetinib. A high basal level of ERK phosphorylation and its rebound activation after MEK inhibition were detected in KRAS-mutant PDOs. Notably, the phosphorylation of EGFR and AKT was more closely correlated with that of MEK than that of ERK. Transcriptome analysis identified MYC-mediated transcription and IFN signaling as significantly correlated gene sets in MEK inhibition. Our experiments demonstrated that RPPA analysis of PDOs, in combination with the genome and transcriptome, is a useful preclinical research platform to understand RAS signaling and provides clues for the development of chemotherapeutic strategies.
    DOI:  https://doi.org/10.1038/s41598-020-74530-x
  18. Clin Colorectal Cancer. 2020 Sep 15. pii: S1533-0028(20)30127-4. [Epub ahead of print]
    Association of Consensus Molecular Subtypes and Molecular Markers With Clinical Outcomes in Patients With Metastatic Colorectal Cancer: Biomarker Analyses From LUME-Colon 1.
    Heinz-Josef Lenz, Guillem Argiles, Takayuki Yoshino, Sabine Tejpar, Fortunato Ciardiello, Jürgen Braunger, Alexey V Salnikov, Ogsen Gabrielyan, Ramona Schmid, Josef Höfler, Thomas Kitzing, Eric Van Cutsem.
      INTRODUCTION: LUME-Colon 1 (NCT02149108) was a global, placebo-controlled phase III study of nintedanib in advanced colorectal cancer (CRC). Pre-specified biomarker analyses investigated the association of CRC consensus molecular subtypes (CMS) and tumor genomic and circulating biomarkers with clinical outcomes.MATERIALS AND METHODS: Archival tumor tissue, cell-free DNA (cfDNA), and plasma samples were collected for genomic, transcriptomic, and proteomic analyses to investigate potential associations between CRC CMS and other biomarkers with nintedanib response and clinical outcomes.
    RESULTS: Of the 765 treated patients, 735, 245, and 192 patient samples were analyzed in the circulating protein, tumor tissue, and cfDNA datasets, respectively. Patients were classified as CMS1 (1.7%), CMS2 (27.7%), CMS3 (0.9%), CMS4 (51.5%), or unclassified (18.2%). Unclassified/mixed CMS was associated with longer overall survival (OS) with nintedanib vs. CMS2 or CMS4 (interaction P-value = .0086); no association was observed for CMS4. Gene expression-based pathway analysis revealed an association between vascular endothelial growth factor-related signaling and OS for nintedanib (P = .0498). The most frequently detected somatic mutations were APC (72.0% [tumor tissue] vs. 56.8% [cfDNA]), TP53 (47.1% vs. 34.9%), KRAS (40.8% vs. 28.6%), and PIK3CA (16.6% vs. 11.5%); concordance rates were > 80%. Median OS differences were observed for APC and TP53 mutations vs. wild-type in cfDNA, indicating a potential prognostic value. Circulating ANG-2, CA-9, CEACAM1, collagen-IV, IGFBP-1, ICAM-1, IL-8, and uPAR were potentially prognostic for both OS and progression-free survival.
    CONCLUSION: We demonstrated the feasibility of large-scale biomarker analyses and CMS classification within a global clinical trial, and identified signals suggesting a potential for greater nintedanib treatment response in the unclassified/mixed CMS subgroup, despite these tumors showing heterogeneous patterns of CMS mixtures. Our results revealed a high degree of concordance in somatic mutations between tumor tissue and cfDNA. Associations with prognosis for cfDNA somatic mutations, as well as several protein-based biomarkers, may warrant further investigation in future trials.
    Keywords:  Circulating biomarkers; Genomic biomarkers; Nintedanib; Predictive marker; Prognostic marker
    DOI:  https://doi.org/10.1016/j.clcc.2020.09.003
  19. Mol Cell. 2020 Oct 15. pii: S1097-2765(20)30657-2. [Epub ahead of print]80(2): 279-295.e8
    Control of Glucocorticoid Receptor Levels by PTEN Establishes a Failsafe Mechanism for Tumor Suppression.
    Hon Yan K Yip, Annabel Chee, Ching-Seng Ang, Sung-Young Shin, Lisa M Ooms, Zainab Mohammadi, Wayne A Phillips, Roger J Daly, Timothy J Cole, Roderick T Bronson, Lan K Nguyen, Tony Tiganis, Robin M Hobbs, Catriona A McLean, Christina A Mitchell, Antonella Papa.
      The PTEN tumor suppressor controls cell death and survival by regulating functions of various molecular targets. While the role of PTEN lipid-phosphatase activity on PtdIns(3,4,5)P3 and inhibition of PI3K pathway is well characterized, the biological relevance of PTEN protein-phosphatase activity remains undefined. Here, using knockin (KI) mice harboring cancer-associated and functionally relevant missense mutations, we show that although loss of PTEN lipid-phosphatase function cooperates with oncogenic PI3K to promote rapid mammary tumorigenesis, the additional loss of PTEN protein-phosphatase activity triggered an extensive cell death response evident in early and advanced mammary tumors. Omics and drug-targeting studies revealed that PI3Ks act to reduce glucocorticoid receptor (GR) levels, which are rescued by loss of PTEN protein-phosphatase activity to restrain cell survival. Thus, we find that the dual regulation of GR by PI3K and PTEN functions as a rheostat that can be exploited for the treatment of PTEN loss-driven cancers.
    Keywords:  failsafe mechanism
    DOI:  https://doi.org/10.1016/j.molcel.2020.09.027
  20. J Chemother. 2020 Oct 14. 1-8
    Chemotherapeutic effects of MEK kinase inhibitor and BRAF kinase inhibitor on KRAS-mutated human colon cancer cell lines with different microsatellite instability.
    Kyung-Ok Kim, Woo-Jae Park, YunJae Jung, Won-Suk Lee.
      We analyzed responsiveness of KRAS-mutated CRC cell lines with distinctive MSI status against mitogen-activated protein kinase (MEK) inhibitor (selumetinib; AZD) and/or B-raf proto-oncogene (BRAF) kinase inhibitor (vemurafenib; PLX). The viability of MSI-high (MSI-H) KRAS-mutated LS174T cells treated with AZD or PLX was 24.5 ± 0.9% or 71.4 ± 3.6%, respectively, and the viability of microsatellite stable (MSS) KRAS-mutated SW480 cells for AZD or PLX was 57.4 ± 3.1% or 43.1 ± 1.8%, respectively. These observations imply that the therapeutic efficacy of MEK kinase inhibitors or BRAF kinase inhibitors against KRAS-mutated colon cancer cells may differ between MSI-H and MSS. However, a combination of both inhibitors synergistically inhibits the proliferation of KRAS-mutated colon cancer cells regardless of MSI status. The underlying synergistic cytotoxic efficacy of AZD/PLX combination on KRAS-mutated colon cancer cells with different MSI status was further substantiated by markedly decreased phosphorylation of ERK in both LS74T and SW480 cell lines upon AZD and PLX treatment. Based on these collective data, we propose that MSI status should be considered when MEK kinase inhibitor or BRAF kinase inhibitor is treated for KRAS-mutated colon cancer, and that combination of both inhibitors synergistically inhibit proliferation of KRAS-mutated colon cancer cells independent of MSI status.
    Keywords:   KRAS-mutation; BRAF kinase inhibitor; Colorectal cancer; MEK kinase inhibitor; chemotherapy; microsatellite instability
    DOI:  https://doi.org/10.1080/1120009X.2020.1829326
  21. Pharmacol Res. 2020 Oct 10. pii: S1043-6618(20)31549-8. [Epub ahead of print] 105241
    PI3K mutations detected in liquid biopsy are associated to reduced sensitivity to CDK4/6 inhibitors in metastatic breast cancer patients.
    Marzia Del Re, Stefania Crucitta, Giulia Lorenzini, Claudia De Angelis, Lucrezia Diodati, Diletta Cavallero, Irene Bargagna, Paola Cinacchi, Beatrice Fratini, Barbara Salvadori, Matteo Ghilli, Manuela Roncella, Andrea Fontana, Romano Danesi, Federico Cucchiara.
      BACKGROUND: PI3K pathway hyperactivation due to PIK3CA mutations contributes to endocrine resistance, and PIK3CA is one of the most frequently mutated genes in breast cancer (BC), occurring approximately 40% of HR+, HER2- advanced BC (ABC). Cyclin-dependent kinase 4 and 6 inhibitors (CDK4/6i) have changed the treatment landscape of HR+, HER2- ABC. Putative mechanisms of resistance to CDK4/6i have been identified, but limited data are available on PI3K deregulation. The present study evaluates the impact of PIK3CA mutations on CDK4/6i plus hormone therapy treatment and evaluates characteristics that may suggest PI3K screening in patients with ABC.METHODS: ABC patients were enrolled, and 12 ml of blood were collected in EDTA tubes at baseline prior to CDK4/6i plus hormone therapy treatment. Plasma was separated and circulating free DNA (cfDNA) was extracted. PIK3CA mutation analysis was performed on a ddPCR. Selected and analyzed mutations included: p.C420R, p.E542 K, p.E545A, p.E545D, p.E545 G, p.E545 K, p.Q546E, p.Q546R, p.H1047 L, p.H1047R, p.H1047Y. Statistical analysis were performed to investigate the predictive power of such mutations and any association with clinical factors.
    RESULTS: Thirty patients were enrolled. PIK3CA mutation status at baseline was independently associated with shorter median PFS (7.44 vs 12.9 months, p = 0.01) in subject receiving CDK4/6i plus hormone therapy treatment. PIK3CA mutations were found to be associated to Ki67 expression in primary lesions (p = 0.006). Moreover, the probability to find a PI3K mutation improved considering also the therapeutic management in previous lines of treatment (McFadden's R2 = 0.415, p = 0.004; AUC of the ROC curve = 0.914).
    CONCLUSION: The findings of this pilot study suggest that the presence of a PI3K mutation in liquid biopsy correlates with a worse PFS in patients with ABC receiving CDK4/6i, and that liquid biopsy is a useful tool to suggests a better tailored pharmacological intervention.
    Keywords:  CDK4/6 inhibitors; PI3K; advanced breast cancer; liquid biopsy; personalized medicine
    DOI:  https://doi.org/10.1016/j.phrs.2020.105241
  22. Cell. 2020 Oct 14. pii: S0092-8674(20)31241-1. [Epub ahead of print]
    Targeting KRAS(G12C): From Inhibitory Mechanism to Modulation of Antitumor Effects in Patients.
    Dongsung Kim, Jenny Yaohua Xue, Piro Lito.
      KRAS mutations are among the most common genetic alterations in lung, colorectal, and pancreatic cancers. Direct inhibition of KRAS oncoproteins has been a long-standing pursuit in precision oncology, one established shortly after the discovery of RAS mutations in human cancer cells nearly 40 years ago. Recent advances in medicinal chemistry have established inhibitors targeting KRAS(G12C), a mutation found in ∼13% of lung adenocarcinomas and, at a lower frequency, in other cancers. Preclinical studies describing their discovery and mechanism of action, coupled with emerging clinical data from patients treated with these drugs, have sparked a renewed enthusiasm in the study of KRAS and its therapeutic potential. Here, we discuss how these advances are reshaping the fundamental aspects of KRAS oncoprotein biology and the strides being made toward improving patient outcomes in the clinic.
    DOI:  https://doi.org/10.1016/j.cell.2020.09.044
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