scholarly journals A statistical framework for assessing pharmacological responses and biomarkers using uncertainty estimates

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Dennis Wang ◽  
James Hensman ◽  
Ginte Kutkaite ◽  
Tzen S Toh ◽  
Ana Galhoz ◽  
...  
Keyword(s):  
Cell Lines ◽  
Dose Response ◽  
Biomarker Discovery ◽  
Statistical Framework ◽  
Curve Fit ◽  
Uncertainty Estimates ◽  
Drug Response Biomarkers ◽  
Novel Biomarkers ◽  
Response Biomarkers

High-throughput testing of drugs across molecular-characterised cell lines can identify candidate treatments and discover biomarkers. However, the cells’ response to a drug is typically quantified by a summary statistic from a best-fit dose-response curve, whilst neglecting the uncertainty of the curve fit and the potential variability in the raw readouts. Here, we model the experimental variance using Gaussian Processes, and subsequently, leverage uncertainty estimates to identify associated biomarkers with a new Bayesian framework. Applied to in vitro screening data on 265 compounds across 1074 cancer cell lines, our models identified 24 clinically established drug-response biomarkers, and provided evidence for six novel biomarkers by accounting for association with low uncertainty. We validated our uncertainty estimates with an additional drug screen of 26 drugs, 10 cell lines with 8 to 9 replicates. Our method is applicable to any dose-response data without replicates, and improves biomarker discovery for precision medicine.

scholarly journals A statistical framework for assessing pharmacological response and biomarkers using uncertainty estimates

2020 ◽  
Author(s):  
Dennis Wang ◽  
James Hensman ◽  
Ginte Kutkaite ◽  
Tzen S. Toh ◽  
Jonathan R Dry ◽  
...  
Keyword(s):  
Cell Lines ◽  
High Throughput ◽  
Drug Response ◽  
Biomarker Discovery ◽  
Cancer Cell Line ◽  
Cancer Therapies ◽  
Statistical Framework ◽  
Curve Fit ◽  
Uncertainty Estimates

AbstractDrug high-throughput screenings across large molecular-characterised cancer cell line panels enable the discovery of biomarkers, and thereby, cancer precision medicine. The ability to experimentally generate drug response data has accelerated. However, this data is typically quantified by a summary statistic from a best-fit dose response curve, whilst neglecting the uncertainty of the curve fit and the potential variability in the raw readouts. Here, we model the experimental variance using Gaussian Processes, and subsequently, leverage this uncertainty for identifying associated biomarkers with a new statistical framework based on Bayesian testing. Applied to the Genomics of Drug Sensitivity in Cancer, in vitro screening data on 265 compounds across 1,074 cell lines, our uncertainty models identified 24 clinically established drug response biomarkers, and in addition provided evidence for 6 novel biomarkers. We validated our uncertainty estimates with an additional drug screen of 26 drugs, 10 cell lines with 8 to 9 replicates. Our method is applicable to drug high-throughput screens without replicates, and enables robust biomarker discovery for new cancer therapies.

Differential response of primary cultures of breast carcinomas to lapatinib in an in vitro chemoresponse assay

2009 ◽  
Vol 27 (15_suppl) ◽  
pp. 1115-1115
Author(s):  
L. M. Hancher ◽  
P. R. Ervin ◽  
S. L. Brower
Keyword(s):  
Cell Lines ◽  
Dose Response ◽  
Response Rate ◽  
Primary Cultures ◽  
Breast Carcinomas ◽  
Human Breast ◽  
Response Curves ◽  
Her 2 ◽  
Dose Response Curves

1115 Background: Lapatinib (Tykerb) is a small molecule tyrosine kinase inhibitor that targets the intracellular domain of the epidermal growth factor receptor and HER-2, thereby inhibiting both growth and survival signaling pathways. Lapatinib is currently FDA-approved to treat HER-2-positive breast cancer previously treated with anthracycline and taxane therapies and trastuzumab. Due to the low population response rate of lapatinib, an integrated biomarker that can identify patients with an increased likelihood for response would be of great clinical utility. The current study describes an in vitro chemoresponse assay developed to predict sensitivity and resistance of primary cultures of human breast tumor specimens to lapatinib. Methods: The chemoresponse assay (ChemoFx) for lapatinib was developed using four different immortalized carcinoma cell lines (SK-OV3, BT474, MDA-MB-231, MCF7). In addition to cell lines, the chemoresponse assay was also performed on 55 first passage primary cultures of human breast carcinomas. All cultures were confirmed to contain keratin-positive epithelial cells using fluorescence immunocytochemistry. Cell lines and specimens were treated with a 10 dose concentration range of lapatinib for 72 hours and stained with DAPI; remaining live cells were counted on an inverted fluorescent imaging system. Resulting dose-response curves were analyzed and categorized as responsive, intermediate responsive, or non-responsive using a proprietary scoring algorithm. Results: All four cell lines (BT474, MDA-MB-231, MCF7, SK-OV3) were responsive to lapatinib treatment, with EC50 values of approximately 10 uM. Dose-response curves of the 55 primary breast cultures revealed that 9% of the specimens tested were responsive to lapatinib, 15% had an intermediate response, and 76% were non-responsive. These results are consistent with a reported clinical response rate of 10% for lapatinib. Conclusions: Initial results with the described integrated cell-based assay demonstrate that in vitro chemoresponse testing may be useful in predicting patient response to lapatinib. This type of in vitro biomarker is likely to increase the efficacy of the current chemotherapy decision-making process for oncologists and their patients. No significant financial relationships to disclose.

Use of an in vitro chemoresponse assay to define a subset of colorectal carcinomas responsive to cetuximab

2009 ◽  
Vol 27 (15_suppl) ◽  
pp. e15127-e15127
Author(s):  
S. D. Rice ◽  
P. R. Ervin ◽  
S. L. Brower
Keyword(s):  
Cell Lines ◽  
Dose Response ◽  
Response Rate ◽  
Primary Cultures ◽  
Colorectal Tumor ◽  
Colorectal Carcinomas ◽  
Response Curves ◽  
Drug Mechanism ◽  
Dose Response Curves

e15127 Background: Cetuximab (Erbitux) is a chimeric monoclonal antibody that binds to the extracellular domain of the epidermal growth factor receptor. This interaction interferes with ligand binding and activation of the receptor blocking the downstream signaling of EGFR and impairing cell growth and proliferation. Cetuximab also has been shown to mediate antibody dependent cellular cytotoxicity. Thus, a singular approach to drug mechanism likely would not capture the true effects of cetuximab. Cetuximab is FDA-approved to treat head and neck cancer and colorectal carcinomas and is being evaluated for use in non-small cell lung and endometrial cancer. Due to the low population response rate of cetuximab, an integrated biomarker that can identify patients with an increased likelihood for response would be of great clinical utility. The current study describes an in vitro chemoresponse assay to predict response of primary cultures of human colorectal tumor specimens to cetuximab. Methods: The chemoresponse assay (ChemoFx) was developed using four different immortalized carcinoma cell lines (NCI-H292, NCI-H522, NCI-H1666, Calu3). The chemoresponse assay was also performed on 54 first passage primary cultures of human colorectal tumor specimens. Cell lines and specimens were treated with a 10 dose concentration range of cetuximab for 72 hours, stained with a nuclear dye, and remaining post-treatment live cells were counted. Resulting dose-response curves were analyzed. Results: Two of the examined cell lines showed response to cetuximab treatment; EC50 values for NCI-H292 and NCI-H1666 were 825nM and 13nM, respectively. NCI-H522 and Calu3 were deemed non-responsive to cetuximab. Dose-response curves of the 54 primary colorectal cultures revealed that 8% of the cultures tested were responsive to cetuximab, 22% had an intermediate response, and 70% were deemed non-responsive. These results are consistent with a reported clinical response rate of 11% for cetuximab in colorectal carcinoma patients. Conclusions: Initial results demonstrate that in vitro chemoresponse testing may be useful in predicting patient response to cetuximab. This could increase the efficacy of the current chemotherapy decision-making process for oncologists and their patients. No significant financial relationships to disclose.

scholarly journals Evaluation of colorectal cancer subtypes and cell lines using deep learning

2019 ◽  
Vol 2 (6) ◽  
pp. e201900517 ◽  
Author(s):  
Jonathan Ronen ◽  
Sikander Hayat ◽  
Altuna Akalin
Keyword(s):  
Colorectal Cancer ◽  
Deep Learning ◽  
Cell Lines ◽  
Biomarker Discovery ◽  
Early Stage ◽  
Point Mutations ◽  
Disease Models ◽  
Latent Factors ◽  
Selection Of

Colorectal cancer (CRC) is a common cancer with a high mortality rate and a rising incidence rate in the developed world. Molecular profiling techniques have been used to better understand the variability between tumors and disease models such as cell lines. To maximize the translatability and clinical relevance of in vitro studies, the selection of optimal cancer models is imperative. We have developed a deep learning–based method to measure the similarity between CRC tumors and disease models such as cancer cell lines. Our method efficiently leverages multiomics data sets containing copy number alterations, gene expression, and point mutations and learns latent factors that describe data in lower dimensions. These latent factors represent the patterns that are clinically relevant and explain the variability of molecular profiles across tumors and cell lines. Using these, we propose refined CRC subtypes and provide best-matching cell lines to different subtypes. These findings are relevant to patient stratification and selection of cell lines for early-stage drug discovery pipelines, biomarker discovery, and target identification.

scholarly journals Cannabidiol (CBD) Oil Does Not Display an Entourage Effect in Reducing Cancer Cell Viability in vitro

2020 ◽  
Vol 3 (2) ◽  
pp. 95-102
Author(s):  
Wesley M. Raup-Konsavage ◽  
Nurgul Carkaci-Salli ◽  
Kelly Greenland ◽  
Robert Gearhart ◽  
Kent E. Vrana
Keyword(s):  
Cell Viability ◽  
Cell Lines ◽  
Cancer Cell ◽  
Dose Response ◽  
Plant Material ◽  
Cannabis Sativa ◽  
Intact Plant ◽  
Response Curves ◽  
Dose Response Curves

<b><i>Introduction:</i></b> Several studies have found that cannabinoids, particularly delta-9-tetrahydrocannabinol and cannabidiol (CBD), have the ability to reduce cancer cell viability. An ongoing debate regarding the use of medical Cannabis revolves around the effectiveness of pure compounds versus intact plant material for treatment. Proponents for the use of intact plant material or botanical extracts argue that there is a synergistic effect between the different cannabinoids, terpenoids, and flavonoids; this is commonly referred to as the “entourage effect.” Our study was designed to test the validity of the proposed entourage effect in a narrow application using a cancer cell viability model. <b><i>Materials and Methods:</i></b> Six cancer cell lines, from 3 different types of human cancer were treated with 10 μM pure CBD or 10 μM CBD from hemp (<i>Cannabis sativa</i>) oil (obtained from 3 different commercial sources) for 48 h, and cell viability was measured with the MTS assay. Dose-response curves were then performed to compare the potencies of pure CBD to CBD oils. CBD concentrations were independently confirmed in the commercial oils, and cannabinoid and terpene composition were also compared. <b><i>Results:</i></b> CBD (10 μM) was able to reduce cell viability in 3 of the 6 cell lines tested, and this was found to be cell line specific and not specific to select cancers. None of the CBD oils tested were able to reduce viability to a greater extent than that of pure CBD. Additionally, dose-response curves found lower IC<sub>50</sub> values for pure CBD compared to the most potent CBD oil tested. Interestingly, some oils actually appeared to protect cancer cells from the effects of CBD. <b><i>Conclusions:</i></b> We found that pure CBD was as potent or more potent at reducing cancer cell viability as the most potent oil tested, suggesting that there is no “entourage” effect under these specific in vitro conditions.

scholarly journals Macrophage Capping Protein CapG Is a Putative Oncogene Involved in Migration and Invasiveness in Ovarian Carcinoma

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
J. Glaser ◽  
M. H. D. Neumann ◽  
Qi Mei ◽  
B. Betz ◽  
N. Seier ◽  
...  
Keyword(s):  
Cell Lines ◽  
Phosphorylation Site ◽  
Actin Binding ◽  
Breast Cancer Cell Lines ◽  
Ovarian Carcinomas ◽  
Capping Protein ◽  
Dependent Cell ◽  
Novel Biomarkers ◽  
Functional Analyses

The actin binding protein CapG modulates cell motility by interacting with the cytoskeleton. CapG is associated with tumor progression in different nongynecologic tumor entities and overexpression in breast cancer cell lines correlates with a more invasive phenotypein vitro.Here, we report a significant CapG overexpression in 18/47 (38%) of ovarian carcinomas (OC) analyzed by qRealTime-PCR analyses. Functional analyses in OC cell lines through siRNA mediated CapG knockdown and CapG overexpression showed CapG-dependent cell migration and invasiveness. A single nucleotide polymorphism rs6886 inside the CapG gene was identified, affecting a CapG phosphorylation site and thus potentially modifying CapG function. The minor allele frequency (MAF) of SNP rs6886 (c.1004A/G) was higher and the homozygous (A/A, His335) genotype was significantly more prevalent in patients with fallopian tube carcinomas (50%) as in controls (10%). With OC being one of the most lethal cancer diseases, the detection of novel biomarkers such as CapG could reveal new diagnostic and therapeutic targets. Moreover, in-depth analyses of SNP rs6886 related to FTC and OC will contribute to a better understanding of carcinogenesis and progression of OC.

Scutellaria Baicalensis, a Herbal Medicine: Antitumor Activity Against Acute Lymphocytic Leukemia, Lymphoma and Myeloma in Vitro.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4491-4491
Author(s):  
Takashi Kumagai ◽  
Claudia Muller ◽  
Julian C. Desmond ◽  
Yasufumi Imai ◽  
David Heber ◽  
...  
Keyword(s):  
Herbal Medicine ◽  
Cell Lines ◽  
Dose Response ◽  
Acute Lymphocytic Leukemia ◽  
Hematological Malignancies ◽  
Lymphocytic Leukemia ◽  
Scutellaria Baicalensis ◽  
Dependent Manner ◽  
P53 Expression

Abstract Scutellaria baicalensis is a widely used Chinese herbal medicine for anti-inflammatory and anti-cancer therapy. The anticancer activity of Scutellaria baicalensis against prostate, breast, hepatocellular, colon and squamous cell carcinomas in vitro has been reported. In this study, we initially investigated its in vitro antitumor activities against 29 cancer cell lines including prostate, breast, ovarian, endometrial and pancreatic cancers as well as myeloid and lymphocytic leukemias, lymphomas, myelomas. The cells were cultured with Scutellaria baicalensis at a dose of 50 μg/ml for 4 days. The herb showed strong anti-proliferative activities against Blin-1 and Nalm-6 acute lymphocytic leukemia cells, Daudi Burkitt′s lymphoma cells and NCI-H929 myeloma cells as measured by MTT assays. Dose-response studies showed an ED50 of 8.32 μg/ml (Blin-1), 12.3 μg/ml (Nalm-6), 4.57 μg/ml (Daudi) and 5.01 μg/ml (NCI-H929) as measured by soft agar colony assay. Treatment with Scutellaria bacalensis at 50 μg/ml for 2 days induced apoptosis of 30% of Blin-1, 17% of Nalm-6, 20% of Daudi and 39% of NCI-H929 cells as measured by Annexin V assay. It also induced G2/M cell cycle arrest of Daudi cells. Scutellaria baicalensis also induced mitchondrial damage in 4 cell lines as measured by fluorescent emissions using the JC-1 assay. Protein expressions were examined by western blot using Blin-1 cells that were most sensitive to Scutellaria baicalensis. After treatment with 50 μg/ml of Scutellaria baicalensis for 2 days, levels of p27KIP1 increased by 152 % independently of p53, expression of the pro-apoptotic gene Bax increased by 208 %, and the anti-apoptotic proteins Bcl-2 and Bcl-XL decreased by 64 % and 38 %, respectively. Notably Scutellaria baicalensis decreased the expression of c-myc oncogene in a dose-dependent manner (80 % decrease at 50 μg/ml). Scutellaria baicalensis contains 21% of baicalin as measured by HPLC. The antiproliferative dose-response assays were repeated using baicalin and calculating the percent baicalin in the herb showed that the results nearly mirrored those of Scutellaria baicalensis, suggesting baicalin is the major anticancer component of this herb. We conclude that Scutellaria baicalensis inhibited the cell growth of B cell hematological malignancies including ALL, lymphoma, and myeloma in vitro by induction of apoptosis associated with mitchondrial damage and modulation of the Bcl family of genes. Thus, Scutellaria baicalensis and its major component, baicalin, should be tested in clinical trials for these hematological malignancies.

In vitro assessment of the inflammatory breast cancer cell line SUM 149: Discovery of two single nucleotide polymorphisms in the RNase L gene.

2011 ◽  
Vol 29 (27_suppl) ◽  
pp. 196-196
Author(s):  
B. T. Nokes ◽  
H. Cunliffe ◽  
A. R. Brown ◽  
G. R. Sheth ◽  
A. Stopeck ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Line ◽  
Cell Lines ◽  
Inflammatory Breast Cancer ◽  
Dose Response ◽  
The United States ◽  
Specific Response ◽  
Rnase L ◽  
Single Nucleotide

196 Background: Inflammatory breast cancer (IBC) is a rare, aggressive form of breast cancer, accounting for 5% of breast cancers diagnosed annually in the United States. Understanding the distinct biology of IBC could help provide novel treatment targets. We sought to evaluate whether or not the IBC cell lines SUM 149 and SUM 190 demonstrated evidence of viral infection. Methods: We performed single nucleotide polymorphism (SNP) genotyping for 2 variants of the ribonuclease (RNase) L gene that have been correlated with the risk of prostate cancer due to a possible viral etiology. We also performed proliferation assays; developed dose response curves to assess the treatment effect of interferon-alpha (IFN-a); and assayed for evidence of the putative human mammary tumor virus (HMTV, which has been implicated but not definitively associated with IBC,) in the DNA and RNA of SUM 149 cells. Results: According to our allelic discrimination SNP assay, 2/2 IBC cell lines were homozygous for the 462 and 541 variants, whereas 0/10 non-IBC cell lines were homozygous positive for the 462 variant (p = 0.015) and 2/10 non-IBC cell lines contained homozygous alleles for the 541 variant (p = 0.52). We also found a dose and time-dependent decrease in the proliferation of SUM 149 IBC cells treated with IFN-a. In contrast, non-IBC cell lines did not show a dose-response decrease in cell proliferation. Our reverse transcriptase polymerase chain reaction (RT-PCR) and Southern blot analysis for the env/LTR and late LTR sequences of the putative HMTV revealed no evidence of the putative viral genome. Conclusions: We discovered 2 SNPs, in the 462 and 541 variants of the RNase L gene, that were homozygously mutated in IBC cell lines but the 462 variant was absent in non-IBC lines. Our discovery of these mutated SNPs present in IBC cell lines suggests a possible genetic risk factor for IBC. In our study, the IBC cell line SUM 149 demonstrated a direct and specific response to treatment with IFN-a, an antiviral agent. We noted no evidence of HMTV infection in that cell line. Further studies of the prevalence and significance of the RNase L 462 and 541 variants in human IBC tissue specimens are warranted to validate our in vitro findings.

Synergistic in Vitro Activity of MOR202, a Human CD38 Antibody, in Combination with Pomalidomide

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 5712-5712 ◽  
Author(s):  
Jan Endell ◽  
Rainer Boxhammer ◽  
Stefan Steidl
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Dose Response ◽  
Target Cells ◽  
High Potency ◽  
Response Curves ◽  
Effector Cells ◽  
Direct Cytotoxicity ◽  
Pre Treatment

Abstract Background: MOR202 is a fully human anti-CD38 antibody currently being tested in a Phase I/IIa clinical trial in multiple myeloma (MM). It mediates antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP) in MM cells with high potency (EC50 ~200 pM) representing a possible promising new therapy for MM patients. In this in vitro study, we evaluated the synergistic potential of MOR202 and pomalidomide (POM), a newly approved IMiD® immunomodulatory agent in MM therapy. Methods: Using flow cytometry analysis, POM was evaluated in relation to its effects on several parameters anticipated to be relevant for anti-tumor activity when combined with MOR202. This included the induction of direct cytotoxicity and CD38 upregulation in several MM cell lines, as well as the activation of human immune effector cells derived from peripheral blood mononuclear cells of healthy donors. On a functional level the interaction of MOR202 and POM was assessed using FACS-based ADCC assays. Different incubation schemes prior to the ADCC assays were evaluated in order to distinguish the influence of POM on ADCC activity when pre-incubated for 72 hours either on target or effector cells or on both in parallel. The observed combination effects were analyzed for synergistic potential. Experiments were carried out in triplicate and mean values (±SEM) were calculated. Results: POM as a single agent showed cytotoxic effects on MM cell lines with high potency (EC50 ~150 nM) and additionally induced an up to 2.7-fold upregulation of CD38 (EC50 ~20 nM) on CD38-expressing MM cell lines. Both effects were maximal at the last tested time point of 72 hours and strongest on cell lines with comparably lower CD38 expression levels. In combination with the observed activation of effector cells these POM-mediated mechanisms lead to a synergistically enhanced cytotoxic activity of MOR202. This synergistic benefit ranged between 1.2-fold and 3.1-fold above theoretical additivity depending on the cell line used and was most pronounced in the case of strong CD38 upregulation. Figure 1 Exemplary ADCC dose-response curves for the comparably lower CD38 expressing cell line AMO-1 after POM pre-treatment of effector cells, target cells or both. Figure 1. Exemplary ADCC dose-response curves for the comparably lower CD38 expressing cell line AMO-1 after POM pre-treatment of effector cells, target cells or both. Conclusions: The cytotoxic activity of MOR202 on MM cells was enhanced and synergized when combined with the immunomodulator agent POM via multiple mechanisms, namely direct cytotoxicity, CD38 upregulation and activation of effector cells. These results provide a mechanistic rationale for combining MOR202 and POM and warrant further evaluation in the clinical setting. Disclosures Endell: MorphoSys AG: Employment, Patents & Royalties. Boxhammer:Morphosys AG: Employment, Patents & Royalties. Steidl:MorphoSys AG: Employment, Patents & Royalties.

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