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Jill Slansky PhD

Associate Professor of Immunology


Many factors limit the adaptive immune system from killing tumors; tumors and the resident inflammatory cells have numerous ways of inhibiting productive B and T cell responses.  In addition, most antigens on tumors are “self antigens” not “neoantigens” or mutated proteins that lead to tumor development.  Therefore immune cells bind weakly to tumors and respond weakly.  Poor responses to tumor/self antigens seem disadvantageous, but actually this protects us from other autoimmune issues.  If the immune system could be better stimulated when encountering tumors, it might mediate more effective responses against the tumor.  Our goal is to make this happen.

 

Using an animal model for colon cancer, we are determining what substitutions in tumor antigen peptides improve antitumor immunity.  These so-called “mimotope” peptides (mimics of epitopes) activate T cells that respond to the tumor more effectively than the natural tumor antigen.  To understand how mimotope peptides affect T cell responses against tumors, we are characterizing the T cells responding to these peptides.  In this system, we have also started studying inflammatory monocytes and the effect of the tumor vurses peripheral expression on antitumor T cell function.  Finally, we are curious to know how the epigenome of tumor-specific T cells changes in response to effective vaccines and to the tumor.

 

We have a collaborative DoD-supported project with the labs of Drs. John Kappler (National Jewish), Peter Lee (City of Hope), and Paul Spellman (OHSU). This multi-team project combines genomics and immunotherapy to develop a new generation of therapeutic breast cancer vaccines, which improve the immune response against breast cancer. We are working on identifying the cognate antigens and mimotopes of breast cancer infiltrating T cells by using the T cells directly out of the tumors.

 

Finally, we are collaborating with the Director of the Lung Cancer Center at National Jewish Health, Dr. Jeff Kern and a number of thoracic surgeons on the immune cells in human lung cancer.  We are dissecting the function and the targets of B cells in the tumor verses tumor-adjacent lung tissue.  We hope to identify antigens that, if included in therapeutic vaccines, would eventually improve the survival of lung cancer patients.




Our Pet Crystal


 

 

Meet our current lab

 

 Brandon Moore

Brandon.Moore@UCDenver.edu

Brandon Moore
PRA

Brandon’s role in the lab is focused on contributing to the labs team of scientists through cell culture, reagent production such as antibody and tetramer, mouse colony breeding and management, in addition to other standard molecular biology procedures. Brandon is also working to complete a collaborative effort with the veterinarians at CSU to determine the DLA haplotypes of Golden Retrievers.  


 Jon Buhrman

Jonathan.Buhrman@UCDenver.edu

Jon Buhrman
Post-doc

Buhrman is investigating methods to optimize the identification and administration of mimotope vaccines. Identifying T cells, and their receptors, that result in enhanced anti-tumor immunity may allow for the identification of mimotopes that specifically target the activation of those T cell clonotypes. Furthermore, tumor-infiltrating T cells may be the optimal T cell targets of mimotope therapy and developing methods to use a polyclonal T cell population to identify potential mimotopes could improve T cell responses following immunization with mimotopes.

 
Tullia.Bruno@UCDenver.edu

Tullia Bruno, PhD
Post-doc

Tullia’s specialty is determining what lymphocytes are doing in human tumors.  She joined the lab after she received her graduate training in tumor immunology at Johns Hopkins in Baltimore, Maryland. The goal of her current project is determining the function of tumor infiltrating B cells (TIL-Bs) in lung cancer patients.  Specifically, she obtains tumor and tumor-adjacent tissue from lung cancer patients through the SPORE lung group at the University of Colorado Anschutz medical campus in order to isolate and study these TIL-Bs.  Preliminary data suggest that these TIL-Bs are anti-tumor cells that aid in eliminating tumor cells.  Further studies are underway to analyze the mechanism of this anti-tumor response.


taizo.nakano@gmail.com

​Taizo Nakano, MD
Fellow

Taizo’s focused on both making the world a better place for children with cancer and identifying the repertoire of T cells from breast cancer patients.  He is living his physician-scientist dream.


Katherine.Waugh@UCDenver.edu

​Katie Waugh
Graduate Student

Katie is analyzing the inner workings of T cells that enter tumors.  What are the reasons within the T cells on the transcriptional and epigenetic level that explain why they aren’t more effective?  Katie takes particular interest in new genomic and epigenomic technologies.


Daniel.Munson@UCDenver.edu

Dan Munson, PhD
Post-doc

Dan competed his graduate work at the University at Albany, New York where his research focused on the molecular and genetic interactions between Herpes Simplex Virus Type 1 and the host cell, specifically the generation of viral microRNAs and viral induced disruption of the cellular proteasome.  With a solid molecular background established, he moved to the world of cellular immunology and is working on determining antigenic targets of tumor infiltrating lymphocytes in breast cancer.  Following identification, he will work on developing mimotopes derived from a baculovirus peptide library to better activate host immune cells specific to cancerous tissue.


Drew.Griffiths@UCDenver.edu

​Drew Griffiths
Student

Drew Griffiths is an undergraduate student at the University of Colorado Denver who came to us from the LABCOATS program.  Drew routinely assists with a variety of projects and helps to keep the lab in order.  Drew has aspirations to become a research scientist.


 ​Jon Sprague

Jonathan.Sprague@UCDenver.edu

 

Jon Sprague, MD
Consultant

When Jon was a medical student at UCD, he developed a java-based computer program to analyze TCR genes from deep sequencing. Since he graduated, he continues to be a consultant for some of our informatics needs.

 

Former lab members

 

Protocols

 

Staining:

IHC of frozen tumors and tissues

Isolation of TILs

Splenocyte Isolation

Typing Tg Rag Mice

CD107a staining

ICC stimulation and staining

 

Baculovirus:

Baculovirus and insect cell culture

Making BV peptide libraries

 

Culture: 

CT26 culture and injection

Medium for CT26 cells

Ex vivo splenocyte culture

In Vivo Killing Assay

 

Molecular Biology & Protein:

Running a K/M lab sizing column

Making Soluble TCR (CT-TCR)

Making MHC (Ld) tetramer

Biacore Protocols

RNA protocol


Reagents
Mouse Models

gp70 KO:  Congenic mouse with BALB/c Chr 5 Mb 28-32 (encoding the MuLV that harbors the tumor antigen gp70) replaced with C57BL/6 interval which does not have gp70 in this region.  The details of the mouse construction are available in McWilliams 2008.  107 SNPs reveal BALB/c loci after 5 back-crosses, the strain has been further back-crossed to BALB/c 38 generations.

Understanding the repertoire of ab T cells that responds to different tumor vaccines has become an integral part of our research.  We wish to understand these repertoires, in part, by analyzing the gene segments that encode the TCRs of the responding cells.  We are taking advantage of the new high throughput sequencing technologies for this purpose (such as the Roche 454 technology), and it has become similarly important to update our TCR sequence analysis tools.  Here we provide a program that is capable of analyzing up to 250,000 different sequences of 400 bases each provided in a FASTA format.  The results include the frequency of different variable (V) and joining (J) gene segments and the frequency that they pair with each other.  In addition, the analysis gives the CDR3 lengths.

Mouse TCR alpha sequence analysis tool:

http://www.slanskylab.com/Sequence/MouseTCRAlphaSequenceAnalysisServlet

Mouse TCR beta sequence analysis tool:

http://www.slanskylab.com/Sequence/MouseTCRBetaSequenceAnalysisServlet

The TCR gene segment nomenclature is confusing! 

In 1995 the “WHO-IUIS Nomenclature Sub-Committee on TCR Designation” agreed on a TCR nomenclature system.[1]  For the mouse ab TCR molecules, the 75 known a chains and the 23 b chains were aligned and grouped into subfamilies; subfamilies were 75% similar.  The TCRs were named so that they could both be easily referred to in electronic files and without Greek letters, periods, hyphens, asterisks, or distinction between upper- and lower-case characters.  For example, TCRBV8S3, also referred to as Vb8.3 in text.  Many TCRs have been well-characterized in the literature using these names.

The ImMunoGeneTics (IMGT) database also developed a nomenclature (one scientist served on both committees).[2]  Most of the nomenclature does not overlap.  At one point, the gene segments in the IMGT database were numbered in the same order as they appear on the chromosome, but new alpha genes were identified and it was discovered that the entire alpha locus has been duplicated in some inbred strains of mice (129) and triplicated in others (C57BL/6)!! (K/M discussions.)  Thus, the numbering is no longer in order.  In addition, investigators have identified differences in sequences that were assumed to be the same gene segment.  It is unknown if these differences are sequencing errors, polymorphisms, or different genes resulting from duplication. The Vb8.3 gene using the first nomenclature is referred to as TRVB13-1 by IMTG, leading to mistakes and confusion.

IMTG provides a program on their website that will analyze one sequence at a time according to their nomenclature.  For different purposes, we have tried to incorporate the different nomenclatures, to include the corrections that we are aware of (navigate to the specific V and J sequences from the frequently asked questions), and this tool also has the ability to compare many sequences to each other.  Currently the program includes the mouse TCR gene segments, but depending on the interest and the direction of our research, we may include human gene segments and immunoglobulin in the future.

This program was developed by Jonathan Sprague, M.D., while he was a student at the University of Colorado School of Medicine.  He received help and input from Kimberly Jordan, Ph.D., who provided most of the data to be analyzed during development.  We also received significant input from Philippa Marrack, Ph.D. and John Kappler, Ph.D., particularly deciphering the alpha genes, and James Scott-Brown, Ph.D, in determining what features should be incorporated into the program.



[1] WHO-IUIS Nomenclature Sub-Committee on TCR Designation: AF Williams, JL Strominger, J Bell, TW Mac, J Kappler, P Marrack, B Arden, MP LeFranc, L Hood, S Tonegawa, and M Davis. Nomenclature for T-cell receptor (TCR) gene segments of the immune system.  Immunogenetics 42(6): 451-453 (1995).  Mouse T-cell receptor variable gene segment families, same journal pp. 501-530.

[2] IMGT, the international ImMunoGeneTics database. MP Lefranc , V Giudicelli , C Ginestoux , Bodmer J, W Müller, R Bontrop, M Lemaitre, A Malik, V Barbié, D Chaume. Nucleic Acids Res. 27(1):209-12 (1999).