Pediatric Preclinical Testing Initiative

Rally Foundation funds 'Next Steps' Undifferentiated Sarcoma Research

 

Partnership with JAX creates new childhood cancer models

We are excited to partner with Jackson Laboratory on the creation of a series of mouse models of childhood cancers.  Our first successful patient-derived xenograft model is now established for an embryonal rhabdomyosarcoma, with many more models in development.  The sample came from an autopsy-derived Legacy Gift.  This model accompanied by the creation of a primary cell culture model (pictured), with genetic characterization.  These resources are intended to be broadly available to all laboratories in North America and beyond.

Because pediatric cancers are rare, few established cell lines are available and many of these cell lines are very old.  For embryonal rhabdomyosarcoma, the RD cell line was established in 1968 and reported in the literature in 1969.  We are grateful to the family that made their child's Legacy Gift possible in order to advance the field.  
 

Moving the field, One cell line at a time

An interesting idea of the Chordoma Foundation is to create a $10,000 prize for new chordoma cell lines.  Not sure what chordoma is?  It wouldn't be surprising... this is a very rare cancer of the cranial base and spinal column.  Few cell lines or tissue resources existed before the Chordoma Foundation initiatives.  Now, though, a great deal is known and the path towards treatment is ever closer.    
 

Open Science Forum: DIPG Preclinical Consortium

Check back frequently to this blog post, which gives a week to week account of the project entitled, "Rapid Preclinical Development of a Targeted Therapy Combination for DIPG" described here and here. This study is made possible by a $100,000 grant from TheCureStartsNow. An additional $28,000 supplement from The Lyla Nsouli Foundation for Children’s Brain Cancer Research has allowed our project to expand to our 2 participating European collaborators.


The full international team is:


Charles Keller MD, Kellie Nazemi MD, Nate Selden MD, PhD and Dan Guillaume MD, PhD at the Oregon Health & Science University
(cell lines:  OHSU-DIPG#1) 
Oren Becher MD, Duke University Medical Center
(cell line, from mouse:  11.1003.2, which is GFAP tva:floxed PDGF p53) 
Michelle Monje MD, PhD, Stanford University
(cell lines:  SU-DIPG-I, SU-DIPG-II, SU-DIPG-III, SU-DIPG-IV, SU-DIPG-V, SU-DIPG-VI)
Maryam Fouladi MD, Cincinnati Children’s Hospital Medical Center
Cynthia Hawkins, MD, PhD, University of Toronto
(cell lines: HSC-DIPG27, HSC-DIPG58)
 Xiao-Nan Li MD, PhD, Baylor College of Medicine
(cell lines:  IBs-1215DIPG, IBs-A1204DIPG, IBs-AB10206DIPG, IBs-C1220DIPG, IBs-J1204DIPG, IBs-W0128DIPG)
Dannis G. van Vuurden MD, MSc, & Esther Hulleman, VU Cancer Center Amsterdam, 
(cell lines:  VUMC-DIPG-A, VUMC-DIPG-B)
Jacques Grill, Institut Gustave-Roussy, Villejuif, France
(cell line names not specified; samples not to be received for sequencing) 
and


The ultimate goal is to move the most effective single agent or combination therapy forward to early phase clinical trials in the next 18-24 months. The clinical trial would be available from the Children's Oncology Group, and/or collaborating institutions. This is the first time that a group of basic and translational scientists and physicians from eight institutions throughout North America and Europe have come together as a consortium to focus on DIPGs and to focus on a bench-to-bedside approach to rationally target therapy for children with DIPGs.


Project Progress:
11/01/2011: The CureStartsNow and affiliated foundations provide $100,000 for project for labs in North America.
11/21/2011: Prototype drug plates sent to Becher, Li and Monje labs (shipping to Hawkins lab tba).
11/23/2011: The Lyla Nsouli Foundation provides $28,000 to add participating European labs.
11/23/2011: Prototype drug plates sent to Grill and Hulleman labs.
11/28/2011: Concept of exon sequencing of paired cell lines, primary tumors and normal DNA (via tertiary funding source) considered.
12/08/2011: Subcontract paperwork to each site has been initiated.
12/08/2011: Plastic tips for robotic drug plating on order (needed to create more plates).
12/08/2011: (Keller lab) Plan initiated for biomarker characterization of OHSU-DIPG#1 cell culture.
12/08/2011: (Keller lab) Sequenom MassArray screen completed for OHSU-DIPG#1 cell culture.
12/15/2011:  Plastic tips for drug plating were backorder from Eppendorf but may arrive in the coming days.  Plan to create/send 384-well drug plates to Cynthia Hawkins group once tips are in. Also sending additional plates to Oren Becher lab at that time.
12/15/2011:  After extensive review, a generation #2 drug plate set has been designed.  60 agents across 3 plates, dozens of molecular targets.  Incorporates the consensus preferred compounds from labs of Jim Olson, Maryam Fouladi, Robin Jones, Darren Hargrave, Jacques Grill and Esther Hulleman.
12/21/2011:  Plastic tips were received this week and new drug plates have been printed and will ship to the Li lab and Hawkins lab soon.  Experience with the plates thus far (Becher lab) suggests that a new plate types (non-clear/black plastic) with or without gelatin undercoating may be needed.  Assessment will be made once each of the other labs has experience with the generation #1 plates.
12/23/2011:  In an amazing development, ABC2 and CureSearch for Children's Cancer have jointly funded a supplement to the Preclinical Consortium project (with $76,000 and $25,000, respectively).  This new supplement will allow us to exon sequencing of all 16 cell lines, their primary tumor and paired normal DNA via the functional genomics laboratory of Dr. Paul Spellman (OHSU). From this data we will prioritize combination drug testing on the most clinically ‘representative’ samples (i.e., samples that have classical DIPG findings which are in order: c‐MET amplification, p53 deletion, PDGFRA amplification, IGF1R/IGF2 amplification, CDK4/CDK6 gain or Shh pathway mutation, EGFR amplification or KRAS/NRAS mutation) or cell lines that are most similar overall to one another (i.e., not outliers).
01/05/2012:  Each lab was requested to organize samples for DNA sequencing; correspondence from Stanford and Amsterdam was received; further more active correspondence expected as the new year ensues.
01/12/2012:  Most of the cell cultures from participating centers are now verified by name, and in some cases new cell cultures are available.  
01/12/2012:  The first drug screen on a generation#1 drug plate set has been performed by the Becher lab using a mouse DIPG culture.  A logistical success, and at least 8 interesting cytocidal drugs have been identified. 
01/19/2012:  Our first probabilistic boolean model (PBM) of the data above is from the mouse DIPG cells is being generated by our collaborators at Texas Tech, Dr. Ranadip Pal and graduate student Noah Berlow.  
01/26/2012:  PBM data still pending (results soon); DNA or RNA samples from DIPG cultures yet to be received from each institution (for sequencing; target date to receive all samples is Feb 9); 24 new sets of generation#1 plates printed for sending out tomorrow to Toronto (for use with potential new primary culture) and other sites; interesting spherical cell culture beads with different coatings being explored at OHSU as a potential way to accelerate DIPG cell culture growth ;official press release now available here. 
01/28/2012:  First (draft) PBM tumor maintenance circuit now created for mouse DIPG cells.  Thank you, Noah!  Many known targets were affirmed, but new ones were found, and co-dependencies were identified.  Validation experiments will follow. 
02/02/2012:  New generation#1 drug plates sent to Toronto and Duke; orders submitted for 55 of the 60 drugs to be used to create generation#2 drug plates; DNA/RNA samples received from Stanford and Toronto.  Many public questions asked about the bioinformatics approach.  For clarification of PBM tumor circuits a movie is available here.  
02/09/2012:  samples for deep sequencing received from Amsterdam, Stanford and Toronto.  Additional samples from Stanford and Baylor to arrive today.  Second run of raw data for mouse DIPG culture drug plates  received from Duke, with analysis now underway.  
02/16/2012:  Second run of drug screen from Duke mouse model consistent with first run; Partial data for 30 drugs screened received for SU-DIPG-II from Stanford.  One strong drug 'hit' found in common with Duke mouse model; DNA and RNA sequencing of samples is to start shortly.  
IMPORTANT UPDATE:  Another cell line has been added as an with an new affiliate investigator & participating institution.  A DIPG primary cell culture being studied at Nationwide Children's Hospital in Columbus, OH, by Dr. Mariko DeWire.  Characterization will include focused genetic characterization and drug screening.  This additional affiliate is being made possible by the generous $8800 sponsorship of the Team Julian Foundation.  
02/23/2012:  DNA and RNA for sequencing can be summarized as:  we have samples related to 19 independent cultures:
  
DNA from the primary tumor from 5; 
RNA from the primary tumor from 0;
DNA from normal tissue from 12;
RNA from normal tissue from 5;
DNA from xenograft tissue for 7;
RNA from xenograft tissue for 0;
DNA from tumor cell cultures from 12;
RNA from tumor cell cultures from 8.
  
Complete sets of DNA & RNA from tumor cultures, the original primary tumor and normal tissue were not available from all samples; however, we are moving forward in the interest of time (and the goal of getting the clinical trial designed in the near term).
02/23/2012:  The results for the second half of the first SU-DIPG-I drug screen are received and are being processed.  Many thanks to Michelle (and Oren) for being among the best and most proactive members of the consortium early on.  
03/01/2012:  Common target(s) found in human SU-DIPG-I cell line and mouse 11.1003.2 cell line drug screens; preliminary results to be presented Monday March 5 at PBTC meeting.  Results of SU-DIPG-II cell line now being analyzed.
03/05/2012:  Organization and early results of the Consortium presented at the Pediatric Brain Tumor Consortium (PBTC) closed meeting in Dallas.  Very well received.  
03/08/2012:  In processing RNA and DNA for sequencing, some samples were not of sufficient quantity.  We have contacted each collaborator to send new aliquots for these samples. 
03/14/2012:  Please see our guest blog entry from Nettie here.  
03/15/2012:  The DNA and RNA sequencing continues to be delayed.  Each lab is actively creating supplemental DNA or RNA so that we can begin sequencing April 1 (our new deadline). 
03/22/2012:  there is nothing new to report this week (sadly).  We are just waiting for the replacement DNA's and RNA's to be sent in from the consortium members.  
03/29/2012:  Preliminary results (similar to those presented at the PBTC meeting) were presented at the COG Brain Tumor committee meeting in Minneapolis.  Discussions were active to extend the same approach to other pediatric brain tumors, including high risk medulloblastoma, high grade gliomas, and intracranial germ cell tumors.  
04/02/2012:  At AACR a key new presentation from the Raabe lab was today, "High-level activation of the Notch pathway in diffuse intrinsic pontine glioma" by Hutt et al (a Johns Hopkins - NCI collaboration that generated a new cell line/mouse model of DIPG).  Also presented by Dr. Ian Pollack are the promising early results of a DIPG vaccine trial (caveats not withstanding).  Photo to right is Dr. Eric Raabe and staff scientist, Marianne Hutt.  
04/05/2012:  subcontract financial paperwork now in place (completed) for Institut de Cancérologie Gustave Roussy.  
04/12/2012 (late entry):  DNA samples still be processed for sequencing.
04/19/2012:  Most samples submitted pass quality control to move on to DNA exon sequencing (see table to right).  Only 2 samples overall failed, and these were the normal tissue controls.  We are thus moving on to exon sequencing.  However, our OHSU sequencing core has an 8-10 week backlog.  Therefore we are looking at sending out the samples for sequencing elsewhere.  In addition, RNA from the samples have not been fully processed/analyzed because the manufacturer of a key reagent are backlogged at least several weeks.  There appears to be no way around this reagent backlog.  Meanwhile, "version 2.1" drug plates are nearing ready for printing on the robot.  We have 5 of the 60 ordered drugs yet to receive before we start printing.
ps.  a special thanks to Keith Desserich for organizing yesterday's DIPG Collaborative webinar.  
04/26/2012:  Manufacturer kits to pre-process RNA for sequencing came in this week, and the samples will be prepared next week.  We are weighing options of sending DNA samples for sequencing at OHSU (a longer wait but cost will decrease in that period) versus a secondary service provider outside of OHSU.  We also have an informal participant in the Raabe laboratory, who received v1.0 drug plates for their new DIPG primary cell culture.  This participant and the related expenses are unfunded, and we would welcome a community gift to bring this work more formally into the Consortium.
nb.  Another new DIPG primary cell culture has been established at OHSU, but not in time for sequencing.  Thanks as always to Michelle Monje for the culturing advice that makes this possible for OHSU and other institutions to successfully grow this type of tumor cells.  
05/05/2012:  DNA samples now submitted for sequencing at OHSU core facility.  If we were a regular customer, it might take 2 months.  We've been moved up the queue somewhat with the understanding that our project is time-sensitive.  RNA sample pre-processing is still not performed, but might be in the next 1-2 weeks, with sequencing results likely to follow in another ~ 2 weeks.      
05/10/12:  DNA sequencing *might* get run in the next week if we are lucky.  There is no change in status of RNA sequencing.  No progress on drug testing pilot from Toronto thus far.  Amsterdam labs are working out how to evenly plate spheres into drug plates (this is an issue for all labs; breaking apart cells leads to poor survival, but quantifying cells in an intact sphere for even plating is not easy).  Villejuif lab has several cell lines, but has requested more plates.  The prior shipment of plates was held in customs and thawed out too long, which may have impacted the usefulness of the first set of plates that were sent. 
05/17/2012:  RNA processing for sequencing to be done in the coming week.  DNA final quality check shows that 3 formalin-fixed, paraffin-embedded samples failed (38 of 41 samples passed).  Raw DNA sequencing data may be available as early as 5/27.   Dr. Monje sent (yet) another set of drug screen results from a human DIPG culture (SU-DIPG-VI)... overall similar to past results with other cultures, which is good to see. Also, Dr. Kristin Schroeder's abstract from the Becher lab has been accepted for an oral presentation at the ISPNO meeting in Toronto next month.  Finally, we may have (accidentally) identified a way to improve propagation of early DIPG cultures.  Validation experiments are ongoing.       
 

Children's Cause Cancer Advocacy and Survivorship

Exciting news comes from the CCCA for the quality of life for childhood cancer survivors! The excerpt below comes directly from the CCCA's recent news feed, click here to read more.

CCCA Supports Introduction of Childhood Cancer Survivorship Legislation

On September 22, 2011, Senators Jack Reed (D-RI) and Kay Bailey Hutchison (R-TX) and Representatives Jackie Speier (D-CA) and Michael McCaul (R-TX) introduced bipartisan legislation to improve the care and quality of life of childhood cancer survivors. The Pediatric, Adolescent and Young Adult Cancer Survivorship Research and Quality of Life Act of 2011 marks a critical step forward in the delivery of medical and psychosocial care to survivors of childhood cancer.
 

Free podcast!

Highlighting: Solving Kids’ Cancer – FREE podcast!

We would like to introduce you to the This Week in Pediatric Oncology podcast (thanks to the ped-onc-surv for the hat tip!)

Solving Kids' Cancer is a website that hosts podcasts and so far 17 episodes have been published:

1. 4/11/2011 Epidemiology of Childhood Cancer (landmark paper)
2. 4/13/2011 Interview with Greg Reaman (outgoing chair of COG)
3. 4/25/2011 Vincristine Pharmacogenetics, Irinotecan/Temozolomide for Relapsed Neuroblastoma
4. 5/8/2011 Meetings Recap and Immunotherapy for Synovial Cell Sarcoma
5. 5/19/2011 Hedgehog Signaling and Itraconazole
6. 5/26/2011 Interview with Dr Archie Bleyer (was head of Children's Cancer Group for 10 years)
7. 6/2/2011 Interview with E. Anders Kolb and Andrew Napper on drug development
8. 7/9/2011 Seneca Valley virus and medulloblastoma
9.7/16/11 Interview with Dr Peter Adamson
10. 7/7/2011 Interview with Dr Robert Seeger
11.7/12/2011 BuMel SIOP results and MIBG transplant with Dr Brian Weiss
12. 8/5/2011 ALL: Anti-CD19 BiTE and Genetic Risk Groups
13.8/19/2011 Updates, epidemiology of CNS tumors, birth order, and cell phone risks
14.8/24/2011 Interview with Dr. Kate Matthay
15.9/28/2011 MicroRNAs and hereditary cancer
16.10/7/2011 Genetic Underpinnings of Ewing Sarcomas: Interview with Dr. Stephen L. Lessnick
17.10/13/2011 Personalized medicine: Interview with Giselle Sholler

You can subscribe by email, RSS or iTunes on the SKC site (see the right side of their webpage)!