Background
Developed by the Broad Institute of MIT and Harvard, PRISM (Profiling Relative Inhibition Simultaneously in Mixtures) is a novel DNA barcoding technology that allows for rapid, viability screening of more than 900 human cancer cell-line models in mixtures. These 900 cell lines represent more than 45 major lineages of cancer.
Traditional cell line viability screening is done one cell line at a time, requiring lots of time and reagents, which can slow down drug discovery and development efforts. PRISM’s unique approach to barcoding and pooling cell lines for high-throughput screening overcomes the limitations of single cell line studies, helping researchers rapidly collect cancer cell line drug-sensitivity data on an unprecedented scale.
The true power of PRISM is not only viability screening of a large number of cell lines, it is their genomic characterization. Through the Cancer Cell Line Encyclopedia efforts and now the Dependency Map efforts we have genomically profiled and identified genetic dependencies for the PRISM cell set. We compare the viability profile of your drug with these baseline genomic features and genetic dependencies to identify the patient population and target of your drug. PRISM screening can be used to:
- Confirm the mechanism of action of your drug
- Uncover novel drug mechanisms of action
- Identify unexpected off-target effects and toxicities
- Establish which patient populations might benefit from treatments
Our original efforts have been published by Channing Yu in 2016, to prove that mixing barcoded cell lines together is feasible and by Steven Corsello in 2020 showing that screening a large number of known drugs across over 500 barcoded cell lines identified new indications for non-oncology drugs.
PRISM has been collaborating with researchers in academia and industry since 2015. Through two large pharmaceutical collaborations we have screened over 30,000 compounds in 500 cell lines and dozens of compounds with academic researchers.
Our current cell set is over 900 cancer cell lines, that is available for anyone to screen three times per year. We now offer single agent small molecule, combination and aqueous screening. We have collaborated with over 150 academic and industrial collaborators in the last couple of years, many of them have screened multiple compounds with us.
First publication of the PRISM method from 2016. 102 cell lines were labeled with unique 24-nucleotide barcodes and used to screen 8400 compounds.
An updated panel of 578 barcoded cancer cell lines were used to profile the activity of 4518 existing drugs. Results were used to begin the PRISM drug repurposing resource.
Cell Lines Collection
Our cell line collection, contains over 900 DNA barcoded cell lines that represent over 45 major types of cancer and the genomic diversity of existing two dimensional cell line models. We have a large number of pediatric cancer cell lines as well as over 100 hematopoietic cell lines. Our cell lines are our most important asset and we therefore take great pride in the quality of our cell set. All of our cell lines are:
- Grown in antibiotic-free RPMI 1640 media
- Mycoplasma negative
- STR fingerprinted to verify each cell lines identity
- Sequence verified to confirm each cell lines’ unique 24-nucleotide barcode is correct
- Mixed together by doubling time into pools of 20-25 cell lines. Pools of cell lines are then frozen in Assay-ready pools so they can be easily thawed for screening.
The full collection of PRISM cell lines is screened for all standard PRISM experiments. This PR1000 collection is divided into two cell sets:
- PR500: 488 solid tumor adherent cell lines
- PR300+: 431 hematopoietic, solid tumor, and pediatric cell lines
See a full annotated list of our PRISM barcoded cell lines.
Assay
5-Day viability assay
Our standard assay, is a 5-day viabilty assay where 384-well plates are prepared by seeding adherent pools at 1,250 cells per well and suspension pools at 2,000 cells per well.
Test agents are added to 384-well plates at 8 doses with 3-fold dilutions and screened in triplicate. At the end of incubation, cells are lysed, and mRNA or gDNA is isolated. Barcode sequences are amplified by PCR and quantified using a Luminex FlexMap 3D® scanner or NextGen sequencing. Relative barcode abundance (compared to vehicle control) is then used to generate cell line sensitivity profiles for each test agent and concentration tested. Once we have the PRISM sensitivity profiles, we do univariate and multivariate modeling to identify the most correlated features to your drug.
Single-agent screening (DMSO)
Our most popular screen is the single agent screen, which is called Medium Throughput Screening or MTS. We screen standard DMSO compounds at a top dose of your choice that we dilute threefold over 8 dilutions. Collaborators send us 150uL of a 1000X stock of your top dilution that must be soluble. We do not require the structure of your compound and do not quality control it with mass spectrometry.
Combination screening (DMSO)
We offer DMSO-based combination screening in PRISM. Combination screening in PRISM requires careful selection of drug doses which can be especially difficult in a pooled context. Therefore, it is only recommended to use this assay for test agents that have been screened in PRISM before as single agents. When selecting an anchor dose our recommendation is to select a dose that does not broadly affect cell viability but gives a reproducible phenotypic effect in a specific cell line or set of cell lines. All combination screens are run in the following format with DMSO-based test agents:
- Drug A at dose (7 pt. dose with 3-fold dilutions) alone
- Drug B at one anchor dose alone
- Drug A at dose (7 pt. dose with 3-fold dilutions) plus Drug B at one anchor dose
This screen of Drugs A and B counts as two ‘slots’ in the PRISM assay.
Aqueous screening
In addition to the standard DMSO-based screening format, PRISM now has the ability to screen aqueous antibodies, antibody-drug conjugates, and growth-inhibiting cytokines. The main difference between our aqueous screen and the DMSO-based screen is that we plate the cells first in the aqueous screen followed by the aqueous agents by ECHO transfer. This method gives us the highest quality data and does not freeze the aqueous reagents. We have written a White Paper on our ADC screening, which you can access here, as well as the data for the ADC’s in the screen and the most common payloads.
Data
Data analysis and predictive modeling
PRISM sensitivity profiles are compared to deep multi-omic cell line characterization information in univariate analyses as well as multivariate predictive modeling algorithms to identify features that correlate with sensitivity.
The Broad Institute feature sets used to understand the differences between sensitive and insensitive cell lines are from the Cancer Dependency Map include:
Cell line lineage and subtype, mutation status, gene expression, protein expression, copy number, methylation, and metabolomics
Loss-of-function genetic perturbation data using CRISPR and shRNA
PRISM viability data from over 6,000 annotated commercial drugs (Drug Repurposing Hub)
A current list of datasets used for each of the feature analyses can be found on our GitHub repository, and are available for download in the DepMap portal.
Data deliverables
We provide your data via a secure data portal. We share detailed analysis of the a detailed report for the quality of the assay, viability of all 900 cell lines, feature set correlation analysis, and data from our validation compounds reports as well as all the raw data from your test agents. See below for more details.
Standard data release includes:
Collaborator data
- Report files
- Summary of analyzed data for each test agent
- Log-fold change
- Viability of each cell line at each drug dose (displayed as a heatmap)
- Individual dose data
- Summary of analyzed data for each test agent
- Dose-response curves
- Computed for each cell line that displays sensitivity to drug treatment
- Used to calculate parameters like AUC and IC50
- Raw data
- Raw and normalized fluorescence values
- Viability data (and replicate-collapsed viability data)
- Detailed cell line quality control data for all test agents
- Feature set correlations
- Interactive plots and tables for univariate (continuous and discrete associations) and multivariate (Random Forest model) analyses▪ Lineage, mutation, gene expression, CRISPR knockout, repurposing, etc.
- Matrix versions
- Data correctly formatted for upload into the DepMap data explorer tool
Combination screen-specific deliverables
- Analysis of the test agent at dose and anchor separately
- Analysis of the combination
- Analysis of the differential response between single agent and combination
Validation data
Single-agent DMSO screens
- 3 validation compounds run on each compound plate. Each screen will have multiple instances of our 3 validation compounds for your reviewin parallel:
- AZ-628, a pan-RAF inhibitor
- Imatinib, a BCR-ABL1 kinase, KIT, and PDGFR receptor tyrosine kinase inhibitor
- Nutlin-3a, an inhibitor of the MDM2-p53 interaction
Combination screens
- 1 known synergistic combination run in parallel:
- Temozolomide, a DNA alkylating agent + O6-benzylguanine, an alkyltransferase inhibitor
Aqueous screens
- 2 validation compounds run in parallel:
- Imatinib, a BCR-ABL1 kinase, KIT, and PDGFR receptor tyrosine kinase inhibitor
- Palbociclib, a CDK4/6 inhibitor
PRISM includes validation agents in each screen with known mechanisms of action (MOA). These well-characterized, commercial reagents produce robust, reproducible sensitivity profiles and feature correlations which are used to ensure high cross-screen reproducibility.
Here we provide an example of PRISM data for two frequently screened compounds:
- AZ628, a BRAF inhibitor
- Imatinib, an inhibitor of PDGFRA and the BCR-ABL1 fusion protein
Melanoma cell lines containing the hotspot BRAF V600E mutation are more sensitive to AZ628 than BRAF wild-type cell lines. CML cell lines, which contain BCR-ABL1 fusions, and lines that have high expression of PDGFRA are selectively killed by imatinib, as expected.
Collaborating with Us
Consortium-style screens
PRISM invites all collaborators to submit test agents to three consortium-style screens each year. Multiple collaborators will submit their test agents during our 3-week submission windows. Submission windows are announced about 3 weeks prior to the opening of the submission window. These screens use the standard PRISM 5-day assay study design described HERE and may include:
- DMSO-based single-agents- 1 slot
- DMSO-based combinations (1 anchor dose compound A X 7 doses of compound B and 7 doses of compound B alone) 2 slots
- Aqueous-based test agents including antibodies, antibody drug conjugates and growth-inhibitory cytokines- 1 slot
Data for each screen is returned in approximately 4 months. [timeline graphic]
To receive information about our upcoming screen and notifications for future screens, please subscribe to our mailing list.
Industry collaborations
The Broad Institute and PRISM’s mission is to help improve cancer patients’ lives. To that end, we are excited to collaborate with you to advance the understanding of cancer therapeutics and accelerate the drug discover process. Our working model is that of a scientific collaboration to further cancer research.
Some key points of our Sponsored Research agreement include:
Data Ownership – PRISM retains the right to use collaboratively generated data to validate and improve the PRISM platform. The structure and identity of your compound will not be shared with us. Unique identifiers are given to all test agents.
Intellectual Property – non-exclusive royalty free licenses are granted
Publication – data may be jointly published when both partners agree
Publishing PRISM Data
Our collaborators may use PRISM data for publication at any time. As authorship should reflect each party’s contributions, we would appreciate inclusion of PRISM team members as authors. Please include: Matthew G. Rees, Melissa M. Ronan, Jennifer A. Roth of Broad Institute of MIT and Harvard, 415 Main Street, Cambridge, MA, 02142, USA.
We kindly request 30 days’ notice for our team to review manuscripts and 10 days’ for posters. Please email prism@broadinstitute.org for reviews.
Other PRISM activities
Making data for the community
In collaboration with the Broad’s Dependency Map team, public PRISM data is being generated every year. All of this data will be shared on the DepMap portal, including data from the Corsello et al Drug Repurposing study, which features 6000 drugs with diverse mechanisms of action and indications screened across 500 cell lines. Our newest dataset the Oncology Reference screen which is the latest targeted oncology therapies screened across our 900 PRISM cell lines will be available to the public in 2024. Check back for updates!.
PRISM Research & Development
PRISM is committed to further developing and improving our cutting-edge screening technologies. One area of interest is modeling the tumor microenvironment. We know that two-dimensional cell lines are very limited in what answers they can provide and therefore are working hard on new assays to better mimic the tumor microenvironment. We are working on 3D spheroid and co-culture assays. Please email us if you are interested in collaborating with us to use the cell lines in novel ways.
