Effective predictive biomarkers are needed to enable personalized medicine and increase treatment efficacy and survival for cancer patients, thereby reducing toxic side effects and treatment costs. ...Patient-derived organoids (PDOs) enable individualized tumour response testing. Since 2018, 17 publications have examined PDOs as a potential predictive biomarker in the treatment of cancer patients. We review and provide a pooled analysis of the results regarding the use of PDOs in individualized tumour response testing, focusing on evidence for analytical validity, clinical validity and clinical utility. We identify future perspectives to accelerate the implementation of PDOs as a predictive biomarker in the treatment of cancer patients.
Cervical cancer is a common gynecological malignancy often caused by high-risk human papillomavirus. There is a paucity of human-derived culture systems to study the cervical epithelium and the ...cancers derived thereof. Here we describe a long-term culturing protocol for ecto- and endocervical epithelia that generates 3D organoids that stably recapitulate the two tissues of origin. As evidenced for HSV-1, organoid-based cervical models may serve to study sexually transmitted infections. Starting from Pap brush material, a small biobank of tumoroids derived from affected individuals was established that retained the causative human papillomavirus (HPV) genomes. One of these uniquely carried the poorly characterized HPV30 subtype, implying a potential role in carcinogenesis. The tumoroids displayed differential responses to common chemotherapeutic agents and grew as xenografts in mice. This study describes an experimental platform for cervical (cancer) research and for future personalized medicine approaches.
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•Establishment of long-term organoid cultures from human ecto- and endocervix•Promising platform for modeling STIs, as evidenced for HSV-1•Pap brush collection as a successful method for cervical tumoroid derivation•Cervical tumoroids display disease hallmarks, such as causative HPV infection
Human-based model systems that faithfully recapitulate cervical cancer and causative HPV infection are scarce and often inadequate. With the advances in organoid technology, Lõhmussaar et al. have now extended this knowledge to the cervix, describing a successful derivation of endo- and ectocervical organoids as well as tumoroids from the associated malignancies.
Background High dose unilobar radioembolization (also termed 'radiation lobectomy')--the transarterial unilobar infusion of radioactive microspheres as a means of controlling tumour growth while ...concomitantly inducing future liver remnant hypertrophy--has recently gained interest as induction strategy for surgical resection. Prospective studies on the safety and efficacy of the unilobar radioembolization-surgery treatment algorithm are lacking. The RALLY study aims to assess the safety and toxicity profile of holmium-166 unilobar radioembolization in patients with hepatocellular carcinoma ineligible for surgery due to insufficiency of the future liver remnant. Methods The RALLY study is a multicenter, interventional, non-randomized, open-label, non-comparative safety study. Patients with hepatocellular carcinoma who are considered ineligible for surgery due to insufficiency of the future liver remnant (< 2.7%/min/m.sup.2 on hepatobiliary iminodiacetic acid scan will be included. A classical 3 + 3 dose escalation model will be used, enrolling three to six patients in each cohort. The primary objective is to determine the maximum tolerated treated non-tumourous liver-absorbed dose (cohorts of 50, 60, 70 and 80 Gy). Secondary objectives are to evaluate dose-response relationships, to establish the safety and feasibility of surgical resection following unilobar radioembolization, to assess quality of life, and to generate a biobank. Discussion This will be the first clinical study to assess the unilobar radioembolization-surgery treatment algorithm and may serve as a stepping stone towards its implementation in routine clinical practice. Trial registration Netherlands Trial Register NL8902, registered on 2020-09-15. Keywords: Radiation lobectomy, Radioembolization, Holmium-166, .sup.166Ho, Hepatocellular carcinoma, Unilobar radioembolization
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Optimized surgical techniques and systemic therapy have increased the number of patients with colorectal liver metastases (CRLM) eligible for local treatment. To increase postoperative survival, we ...need to stratify patients to customize therapy. Most clinical risk scores (CRSs) which predict prognosis after CRLM resection were based on the outcome of studies in specialized centers, and this may hamper the generalizability of these CRSs in unselected populations and underrepresented subgroups. We aimed to externally validate two CRSs in a population-based cohort of patients with CRLM. A total of 1105 patients with local treatment of CRLM, diagnosed in 2015/2016, were included from a nationwide population-based database. Survival outcomes were analyzed. The Fong and more recently developed GAME CRS were externally validated, including in pre-specified subgroups (≤70/>70 years and with/without perioperative systemic therapy). The three-year DFS was 22.8%, and the median OS in the GAME risk groups (high/moderate/low) was 32.4, 46.7, and 68.1 months, respectively (p < 0.005). The median OS for patients with versus without perioperative therapy was 47.6 (95%CI 39.8, 56.2) and 54.9 months (95%CI 48.8, 63.7), respectively (p = 0.152), and for below/above 70 years, it was 54.9 (95%CI 49.3−64.1) and 44.2 months (95%CI 37.1−54.3), respectively (p < 0.005). The discriminative ability for OS of Fong CRS was 0.577 (95%CI 0.554, 0.601), and for GAME, it was 0.596 (95%CI 0.572, 0.621), and was comparable in the subgroups. In conclusion, both CRSs showed predictive ability in a population-based cohort and in predefined subgroups. However, the limited discriminative ability of these CRSs results in insufficient preoperative risk stratification for clinical decision-making.
IntroductionCytoreductive surgery and hyperthermic intraperitoneal chemotherapy (CRS-HIPEC) has become standard of care for patients with peritoneal metastases of colorectal origin with a ...low/moderate abdominal disease load. In case of a peritoneal cancer index (PCI) score >20, CRS-HIPEC is not considered to be beneficial. Patients with a PCI >20 are currently offered palliative systemic chemotherapy. Previous studies have shown that systemic chemotherapy is less effective against peritoneal metastases than it is against haematogenous spread of colorectal cancer. It is suggested that patients with peritoneal metastases may benefit from the addition of intraperitoneal chemotherapy to systemic chemotherapy. Aim of this study is to establish the maximum tolerated dose of intraperitoneal irinotecan, added to standard of care systemic therapy for colorectal cancer. Secondary endpoints are to determine the safety and feasibility of this treatment and to establish the pharmacokinetic profile of intraperitoneally administered irinotecan.Methods and analysisThis phase I, ‘3+3’ dose-escalation, study is performed in two Dutch tertiary referral centres. The study population consists of adult patients with extensive peritoneal metastases of colorectal origin who have a good performance status and no extra-abdominal metastases. According to standard work-up for CRS-HIPEC, patients will undergo a diagnostic laparoscopy to score the PCI. In case of a PCI >20, a peritoneal access port will be placed in the abdomen of the patient. Through this port we will administer intraperitoneal irinotecan, in combination with standard systemic treatment consisting of 5-fluorouracil/leucovorin with oxaliplatin and the targeted agent bevacizumab. Therapy consists of a maximum of 12 cycles 2-weekly.Ethics and disseminationThis study protocol is approved by a research medical ethics committee (Rotterdam, Netherlands) and the Dutch Competent Authority (CCMO, The Hague, Netherlands). The results of this trial will be submitted for publication in a peer-reviewed scientific journal.Trail registration numberNL6988 and NL2018-000479-33; Pre-results.
The invasion-inducing T-lymphoma invasion and metastasis 1 (Tiam1) protein functions as a guanine nucleotide exchange factor (GEF) for the small GTPase Rac1. Differentiation-dependent expression of ...Tiam1 in the developing brain suggests a role for this GEF and its effector Rac1 in the control of neuronal morphology. Here we show that overexpression of Tiam1 induces cell spreading and affects neurite outgrowth in N1E-115 neuroblastoma cells. These effects are Rac-dependent and strongly promoted by laminin. Overexpression of Tiam1 recruits the α6β1 integrin, a laminin receptor, to specific adhesive contacts at the cell periphery, which are different from focal contacts. Cells overexpressing Tiam1 no longer respond to lysophosphatidic acid-induced neurite retraction and cell rounding, processes mediated by Rho, suggesting that Tiam1-induced activation of Rac antagonizes Rho signaling. This inhibition can be overcome by coexpression of constitutively active RhoA, which may indicate that regulation occurs at the level of Rho or upstream. Conversely, neurite formation induced by Tiam1 or Rac1 is further promoted by inactivating Rho. These results demonstrate that Rac- and Rho-mediated pathways oppose each other during neurite formation and that a balance between these pathways determines neuronal morphology. Furthermore, our data underscore the potential role of Tiam1 as a specific regulator of Rac during neurite formation and illustrate the importance of reciprocal interactions between the cytoskeleton and the extracellular matrix during this process.
Abstract
Background: Tumors are addicted to pathways neutralizing oxidative stress as a result of their high levels of reactive oxygen species (ROS). These ROS can oxidize nucleotides (NTs) within ...the precursor pool, which can cause cell death when incorporated into DNA. This tumor specific vulnerability might be exploited therapeutically. The mutT-homologue (MTH1) enzyme is recently identified as a non-oncogene target, which prevents incorporation of oxidized NTs into DNA (Gad et al, 2014; Huber et al, 2014). MTH1 inhibition by TH588 and (S)-Crizotinib caused tumor-specific oxidative DNA damage and cell death (Gad et al, 2014; Huber et al, 2014). Interestingly, hypoxia might enhance sensitivity to MTH1 inhibition (Qiu et al, 2015; Bräutigam et al, 2016). Thus, exploiting tumor oxidative stress via inhibition of MTH1 might be an exciting new approach to fight colorectal cancer (CRC) even in the presence of hypoxia.
Aims and Hypotheses: In the present study we aim to compare the ability of TH588 HCl and (S)-Crizotinib to inhibit CRC growth in vitro and in vivo and to assess their mechanism(s) of action, including MTH1-dependent and -independent effects. We hypothesize that both compounds have the potency to inhibit CRC growth, conceivably via different mechanisms of action, which might not dependent on MTH1 inhibition (Kettle et al, 2016).
Methods: All data were obtained using a human CRC liver metastasis-derived 3D spheroid line untreated or treated for 3 days with TH588 HCl or (S)-Crizotinib. Spheroids were pre-cultured and treated under normoxia (21% O2), hypoxia (0.1% O2) or only pre-cultured under hypoxia to mimic hypoxia and reoxygenation respectively.
Results: Using a CellTiter-Glo cell viability assay we showed that increasing concentrations of (S)-Crizotinib under normoxia reduced cell viability with 23%, 64%, and 98% for 5, 10 and 20 μM respectively. By contrast, the maximal reduction of cell viability after treatment with TH588 HCl was only 32%. Interestingly, the dose response curve of (S)-Crizotinib was steeper when spheroids were reoxygenated just before treatment compared to normoxia and hypoxia, which implies that (S)-Crizotinib is more efficient. Importantly, hypoxia completely abolished the TH588-induced reduction in cell viability seen during normoxia. While the immediate effects on cell viability were different, both drugs (10 μM) severely reduced clone-forming efficiency both under normoxia and after hypoxia-reoxygenation prior to treatment. These results suggest different mechanisms of action of both drugs. As expected, both drugs caused an increase in DNA double-strand breaks, as measured by γH2AX staining. While chemotherapy treatment usually enriches the content of cancer stem cells, so far we did not observe this with either MTH1 inhibitor.
Discussion and Conclusions: Until now, most research involving MTH1 inhibitors has been performed using cancer cell lines. In the present study we use patient-derived CRC spheroids and organoids. We previously found that organoids derived from patients with a poor-prognosis type of CRC are generally characterized by high expression of a H2O2 stress signature (Emmink et al, 2014). Exploiting this oxidative stress phenotype via inhibition of MTH1 might be an attractive new therapeutic avenue. Our results suggest that TH588 HCl and (S)-Crizotinib induce DNA damage and effectively reduce clone-forming potential of a CRC spheroid line, which was not limited by hypoxia pre-culturing. Interestingly, both compounds appear to act via different mechanisms. Further research should unravel these mechanisms of action.
Citation Format: Lizet M. van der Waals, Danielle A.E. Raats, Jennifer M.J. Jongen, Tobias B. Dansen, Inne H.M. Borel Rinkes, Onno W. Kranenburg. Exploiting oxidative stress using MTH1 inhibitors in colorectal cancer. abstract. In: Proceedings of the AACR Precision Medicine Series: Targeting the Vulnerabilities of Cancer; May 16-19, 2016; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(1_Suppl):Abstract nr A29.
Abstract
Introduction:
Three-dimensional cell culture methods are currently not widely used for cancer drug discovery due to the specific imaging demands of the systems. In particular there is a need ...for well validated robust high throughput methods for image-based assays that can be used for high content screening.
Methods:
Therefore we have developed an image-based three-dimensional high throughput toxicity assay. We used two platforms, the Thermo Scientific ArrayScan VTi and BD FACSCanto™, to characterize antiproliferative potency, cell death and cell cycle distribution. Cultures of uniform spheroids were treated with Irinotecan either alone or in combination with the ABC transporter inhibitor Valspodar, which blocks irinotecan efflux. All drugs were dispensed using a Tecan HP-D300 digital dispenser. Treatment-induced effects were analyzed after 48, 72 or 96 hours with both platforms using dihydro derivatives of fluorescent live cell dyes such as dihydrocalcein, AM and DNA-binding dyes such as DRAQ5™ and Propidium Iodide.
Results:
All cell lines tested on the two platforms showed the expected effects of Irinotecan alone or in combination with Valspodar and correlations between the Thermo Scientific ArrayScan VTi and BD FACSCanto™ were high, with Pearson tests varying from 0.76 to 0.95, p < 0.05, depending on the cell line used. All results were reproducible and showed minor variation.
Conclusion:
The demonstrated approach provides an easy-to-handle, low-cost, accurate and reproducible high content method for measuring drug efficacy in three-dimensional cell cultures. This method can be used for high throughput drug discovery screens and personalized cancer care.
Citation Format: Kari Trumpi, David A. Egan, Thomas T. Vellinga, Inne H.M Borel Rinkes, Onno W. Kranenburg. High throughput toxicity assay for three-dimensional cell cultures. abstract. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5388. doi:10.1158/1538-7445.AM2014-5388
Lysophosphatidic acid (LPA) and sphingosine-1-phosphate (S1P) are extracellular lipid mediators that signal through distinct members of the Edg/LP subfamily of G protein-coupled receptors (GPCRs). ...LPA and S1P receptors are expressed in almost every cell type and can couple to multiple G proteins (G(i), G(q) and G(12/13)) to mediate a great variety of responses, ranging from rapid morphological changes to long-term stimulation of cell proliferation. LPA serves as the prototypic GPCR agonist that activates the small GTPases Ras (via G(i)) and RhoA (via G(12/13)), leading to activation of the mitogen-activated protein kinase (MAPK) cascade and reorganization of the actin cytoskeleton, respectively. This review focuses on our current insights into how Ras-MAPK signaling is regulated by GPCR agonists in general, and by LPA in particular.
Background & Aims Stem cells of normal tissues have resistance mechanisms that allow them to survive genotoxic insults. The stem cell–like cells of tumors are defined by their tumor-initiating ...capacity and may have retained these resistance mechanisms, making them resistant to chemotherapy. We studied the relationship between resistance to the topoisomerase I inhibitor irinotecan and tumor-initiating potential in human colonosphere cultures and in mice with colorectal xenograft tumors. Methods Colonosphere cultures were established from human colorectal tumor specimens obtained from patients who underwent colon or liver resection for primary or metastatic adenocarcinoma. Stem cell and differentiation markers were analyzed by immunoblotting and fluorescence-activated cell sorting. Clone- and tumor-initiating capacities were assessed by single-cell cloning and in immune-deficient mice. Sensitivity to irinotecan was assessed in vitro and in tumor-bearing mice. The relationship between drug resistance and tumor-initiating capacity was tested by fluorescence-activated cell sorting of colonosphere cells, based on expression of ABCB1 and aldehyde dehydrogenase (ALDH) activity. Results Colonosphere cultures had a high capacity to initiate tumors in mice and were resistant to irinotecan. Inhibition of the drug-efflux pump ABCB1 by PSC-833 allowed irinotecan to eradicate tumor-initiating cells. However, ABCB1 was expressed only by a subpopulation of differentiated tumor cells that did not form clones or tumors. Conversely, tumor-initiating cells were ABCB1-negative and were identified by high ALDH activity. Tumorigenic ALDHhigh /ABCB1negative cells generated nontumorigenic ALDHlow /ABCB1positive daughter cells in vitro and in tumor xenografts. PSC-833 increased the antitumor efficacy of irinotecan in mice. Conclusions The resistance of colorectal tumors to irinotecan requires the cooperative action of tumor-initiating ALDHhigh /ABCB1negative cells and their differentiated, drug-expelling, ALDHlow /ABCB1positive daughter cells.
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