LUKAS WYATT PFANNENSTIEL, Ph.D.

***** ******** **, *** ****, Cleveland, OH 44118

443-***-****

acako2@r.postjobfree.com

SUMMARY:

A dedicated and motivated immunologist with an interest in immunoassay and immunotherapy

development.

Experienced in planning and troubleshooting scientific research projects and capable of working

both independently and as part of a team to achieve common goals.

Broad experience in molecular biology and cellular immunology methods including human

specimens and animal models.

Successfully wrote and published scientific manuscripts as well as assisted in preparation of

grant applications.

Lab management experience including supervising technicians and students to achieve group

goals.

EXPERIENCE:

Cleveland Clinic Lerner Research Institute, Cleveland, OH 12/2009 – Present

Postdoctoral Research Fellow, Department of Immunology

● Characterizing and investigating mechanisms of tumor -induced T cell dysfunction and

senescence in mouse and human CD8+ T cells using in vitro co-cultures, in vivo mouse tumor

models, FACS, and in vitro suppression assays.

● Investigating use of antibody blockade of inhibitory T cell signaling molecules in vivo by isolating

and purifying antibody from hybridoma cultures.

● Characterized tumor-induced T cell phenotype changes in human patient peripheral blood and

tumor-infiltrating lymphocytes using flow cytometry.

● Characterized IL-7 induced signaling pathways in human T cells using western blotting.

● Investigating the role of Mcl-1 in chemotherapy-induced senescence in tumor cell lines using in

vitro senescence assays, western blotting, and histological staining of xenograft tumors in mice.

● Coordinated laboratory and mouse colony relocation from the University of Maryland Medical

School to the Cleveland Clinic.

● Planned and purchased lab equipment for starting a new lab at the Cleveland Clinic.

● Obtained institutional approval for, and currently maintaining three transgenic mouse colonies.

● Prepared and submitted publication manuscripts and assisted in prepa ration of grant

applications.

● Supervised graduate students and technicians to achieve project goals and troubleshooting

scientific assays.

Johns Hopkins University, Baltimore, MD 05/2002 – 11/2009

Graduate Student, School of Medicine, Program in Cellular and Molecular Medicine

● Investigated potential CD4+ T cell epitopes of Her-2/neu using libraries of overlapping peptides

and T cells purified from in vitro cultures or lymph node tissue from vaccinated mice.

● Characterized enhanced maturation of in vitro and in vivo derived dendritic cells after treatment

with Taxol by FACS and intracellular cytokine staining.

● Characterized enhanced activation of naive CD8+ T cells after exposure to Taxol -treated dendritic

cells in vitro and in vivo by intracellular staining and chromium-release assays.

● Characterized reduced tumor growth in Her-2/neu transgenic mice after treatment with Taxol

and Cyclophosphamide.

● Characterized immune response changes after regulatory cell depletion using antibody isolated

and purified from hybridoma cultures.

● Produced tetramer to study Her-2/neu specific T cells generated in vaccinated mice.

● Supervised training of undergraduate and high school students in various molecular

biology techniques.

University of Kansas, Lawrence, KS 06/1999 – 05/2001

Undergraduate Research Assistant, Department of Molecular Biosciences

● Investigated gene regulation of flagellar growth in green alga Chlamydemonas reinhartii through

random mutagenesis and Southern blot analysis.

● Tutored and proctored undergraduate students taking biology courses.

EDUCATION:

Ph.D., Cellular and Molecular Medicine, 2009

Johns Hopkins University School of Medicine, Baltimore, MD

Advisor: Elizabeth M. Jaffee, M.D.

Thesis title: Mechanistic Pathways to Augment Responses to the Her2/neu Whole-Cell Tumor Vaccine

B.S., Cell Biology, 2001

University of Kansas, College of Liberal Arts and Sciences, Lawrence, KS

Advisor: William Dentler, Ph.D.

Graduated with departmental honors.

MEMBERSHIPS:

American Association of Immunologists

Department of Immunology Students and Fellows Committee

SKILLSETS:

Immunological Assays

o In vitro T cell expansion, function, and suppression assays.

o In vitro culture of dendritic cells.

o Ex vivo monitoring of immune treatments via cytokine ELISA, immune cell phenotyping,

and tetramer staining.

o Flow cytometry including use of FACS Calibur and LSR II instruments as well as data

analysis.

o Tissue dissociation and purification of immune cell subsets including dendritic cells and

tumor-infiltrating lymphocyte populations.

Molecular Biology

o Growth and maintenance of cell lines.

o Assaying protein expression and phosphorylation via western blotting.

o Antibody production and purification from hybridoma cultures.

o Tetramer production and purification.

o Cloning and verification of DNA plasmids.

o Expression of exogenous protein constructs by lipofectamine and electroporation

transfection.

Animal Models

o Maintenance of transgenic mouse colonies and creation of novel genotypes through

breeding.

o Injection and monitoring of tumor growth.

o Administration of peptide and whole-cell vaccines, adoptive transfer of transgenic T

cells, and antibody-based immunoadjuvants.

Human Specimens

o Processing of blood to isolate lymphocyte populations.

o Digestion and dissociation of tumor specimens to study infiltrating T cell populations.

o Cryopreservation of purified cells.

Staining of tissue sections for immunohistochemical and immunofluorescence assays.

o

PUBLICATIONS:

1. Pfannenstiel LW, Demelash A, Gastman BR. Raiding the pharmacy: genomic screening identifies

known chemotherapies as negative regulators of Mcl-1. Genome Med. 2012 Jun 27;4(6):53.

2. Bolesta E, Pfannenstiel LW, Demelash A, Lesniewski M, Tobin M, Schlanger S, Nalar SC,

Papadimitriou JC, Kalvakolanu DV, Gastman BR. Inhibition of Mcl -1 promotes senescence in cancer

cells: implications for preventing tumor growth and chemotherapy resistance. Mol Cell Biol. 2012

May;32(10):1879-92.

3. Pfannenstiel LW*, Zhang Y*, Bolesta E, Montes C, Zhang X, Chapoval AI, Gartenhaus R, Strome SE,

Gastman BR. Interleukin-7 CD27-CD28- suppressor T-cells: implications for cancer immunotherapy.

Clin Cancer Res. 2011 Aug 1; 17(15): 4975 -86. 2011 Jun 28. (*co-first author)

4. Pfannenstiel LW, Lam SS, Emens LA, Jaffee EM, Armstrong TD. Paclitaxel enhances early dendritic

cell maturation and function through TLR4 signaling in mice. Cell Immunol. 2010;263(1):79 -87.

5. Ercolini AM, Ladle BH, Manning EA, Pfannenstiel LW, Armstrong TD, Machiels JP, Bieler JG, Emens

LA, Reilly RT, Jaffee EM. Recruitment of latent pools of high -avidity CD8 T cells to the antitumor

immune response. Journal of Experimental Medicine, 2005; 201(10): 1591 -1602.

REFERENCES- Available Upon Request

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