when b cells go bad: infection, inflammation and chronic b cell stimulation
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When B Cells Go Bad: Infection, Inflammation and Chronic B Cell Stimulation. Karen S. Anderson MD PhD Associate Professor, Biodesign Institute Arizona State University Mayo Clinic Arizona. Conflicts of Interest. - PowerPoint PPT PresentationTRANSCRIPT
When B Cells Go Bad: Infection, Inflammation and Chronic B
Cell StimulationKaren S. Anderson MD PhD
Associate Professor, Biodesign InstituteArizona State University
Mayo Clinic Arizona
Conflicts of Interest• I serve on the advisory board and have received consultant
fees and stock options with Provista Dx.
• No off-label clinical diagnostics or therapeutics will be discussed.
W. Michael Kuehl & P. Leif Bergsagel, 2002
Monoclonal Gammopathy of Undetermined Signficance
• 3.2% of adults over the age of 50; 6.6% of adults over age 80• Premalignant disease that can transition to myeloma; average
2-15 years, rate 1% per year• Over 90% of patients with MM have a premalignant plasma
cell disorder• 50% Ig translocations; 50% hyperdiploid
– Errors in switch recombination and somatic hypermutation of the B cell
• Likely due to an abnormal response to antigenic stimulation
Chronic Infections and Antibodies
• EBV, HHV-8, HCV, (CMV), H. pylori are associated with B cell lymphoma and B cell chronic lymphocytic leukemia– what about myeloma?
• Ig in HCV-positive myeloma patients can target the virus
• ~20% of MGUS and myeloma Abs target infectious antigens (ASH 2013)
• Would treatment of chronic infections also prevent or treat MGUS?
Chronic Infections not Associated with MGUS
Bida and Rajkumar Mayo Clin Proc 2009
Moir and Fauci, Nat Rev Immunol 2009
How B cells Proliferate in response to infection
Antibody structure:
Ligands (antigens)
• What is the specificity of antibodies?• What is the diversity of antibodies?
When Ig genes rearrange, there are >1011 potential molecules
To facilitate research, DNASU stores over 162,000 plasmids and distributes these to researchers in 37 states and 38 countries
ProteinExpression
…
1. Print Plasmids
2. Express & capture proteins
Nucleic Acid-Programmable Protein Array (NAPPA)
Replicate arrays of candidate proteins
Find antibodies in patients’ blood
• Proteomics: the study of all of the proteins in the human body• We use plasmids to make over 10,000 human proteins• By putting these plasmids on a glass slide (NAPPA) we can make and study over 10,000
proteins at one time
NAPPA Array Production and ScreeningGene Cloning Bacterial plating
DNA preparation
Array Printing
p53
Antigen array
Add patient serum
Human papilloma virus (HPV16): ~70% of oropharyngeal cancersEmerging epidemic in US and Europe
Serologic Biomarkers for HPV+ Cancer
HPV Genome: 8 ORFsDiagnosis Prognosis
0.00
0.25
0.50
0.75
1.00
Pro
port
ion
surv
ivin
g
13 9 6 4 3 1NE2 negative84 75 66 59 33 10NE2 positive
Number at risk
0 12 24 36 48 60Follow-up time (months)
NE2 positiveNE2 negative
Kaplan-Meier survival estimates
NE2, p<0.001
Identification of the Targets of Antibodies
• Protein microarrays are now used to test >10,000 proteins for antibody targets
• Large gene collections can be leveraged for rapid protein display• We are developing pathogen-specific arrays to identify
antibodies in blood
• We need to measure the antibody IMMUNOME to understand the pathogenesis of MGUS
What is the diversity of Antibodies?Can we detect specific Ig rearrangements?
DNA Origami nanostructure design
Hao Yan and Joe Blattman, Biodesign
Overall strategy to obtain linked RNA sequences from single cells.
Conclusions
Antibodies have extraordinary sequence diversity
There are emerging technologies for quantitating that diversity
By linking single-cell RNA capture with next-gen sequencing, we may be able to: Identify early events (?pre-MGUS) of loss of diversity Rapidly generate patient-specific probes for molecular detection of
rearrangements
Acknowledgements
Mayo Oncology Don Northfelt Doug Lake Barb Pockaj Michael Barrett
ASU Biodesign Institute Joseph Blattman Hao Yan Josh LaBaer Ji Qiu Garrick Wallstrom Laura Gonzalez Jin Park Fernanda Festa
Anderson Lab Julia Cheng Ting Li Rizwan Alam Benjamin Katchman Krishna Sundaresan Diego Chowell Shay Ferdosi Hans Frykman I.Purushothaman Fernando Hernandez Robert Brown Alison Goulder Jack Resnik Peter Chang
Our PatientsNCI/Early Detection Research NetworkZicarelli Foundation