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Using CD34+ Stem Cells in Humanized Mouse Models

Feb 2, 2022 1:45:00 PM / by Champions Oncology

Cellular Therapy and Regeneration microscope of cell

Humanized immune system (HIS) mouse models have revolutionized basic and preclinical research, and these models have been refined as new mouse strains and sources of human immune cells have been evaluated. Human hematopoietic stem and progenitor cells (HSPCs) are a widely used material for engraftment of HIS mice because they can differentiate into every type of blood and immune cell, and these cells are typically defined by cell surface expression of CD34. Here we highlight what is known about CD34+ HSPCs and which applications they are best suited to in HIS models.

 

Studies in the 1990s and 2000s first defined human CD34 expression as a clinical marker for enriching bone marrow prior to transplantation. It is now known that HSPCs are CD34- immediately prior to cell division but then express CD34[1],[2], and CD34 is thought to play a role in adhesion and migration of HPSCs to the bone marrow[3],[4]. CD34+ HSPCs can be derived from different sources, including bone marrow and umbilical cord blood. Cells derived from these sources must be processed to positively select for CD34+ cells, which be done using CD34+ magnetic bead cell sorting or fluorescence-activated cell sorting. CD34+ cells can be further expanded in culture over several days or weeks using a cocktail of growth factors and cytokines like Flt-3 and stem cell factor (SCF)[5]. This enriched CD34+ cell product can be transferred into immunodeficient mouse strains that may be engineered to express human cytokines and growth factors like IL-3 and GM-CSF, which expedites engraftment and differentiation of human immune cell subsets in a matter of weeks[6].

Experimental mouse in researcher's hand

Human peripheral blood mononuclear cells (PBMC) can also be used for engraftment of immunodeficient mice, which is a rapid and straightforward method of generating HIS mice. Unfortunately, PBMC-based HIS mice typically develop graft-versus-host disease against engrafted immune cells within 4 to 6 weeks, which limits the time during which these mice can be studied in longitudinal tumor or drug studies[7]. Thus CD34+ HSPC-based HIS mice are the preferred model for preclinical oncology studies and can be transplanted with patient-derived tumor xenografts (PDX), which are well tolerated by these immunodeficient mouse strains[8]. PDX tumors in HIS mice can be measured over several weeks as tumors grow or respond to experimental therapies, and the roles of different immune cell subsets that generate or suppress anti-tumor responses can be defined.

 

Improvements in HIS mouse models continue to enhance the quality of these systems, including developing better methods for CD34+ HSPC enrichment and expansion and engineering mouse cell lines that can better tolerate HSPC and tumor transplants. Not only will this type of model continue to be the first choice of pre-clinical oncology researchers seeking to understand immune mechanisms of tumor destruction, but it will also continue to be a means of evaluating drug or biologic candidates.

 

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[1]  Sato T, Laver JH, Ogawa M. Reversible expression of CD34 by murine hematopoietic stem cells. Blood. 1999;94(8):2548-2554.

[2] Dooley DC, Oppenlander BK, Xiao M. Analysis of primitive CD34- and CD34+ hematopoietic cells from adults: gain and loss of CD34 antigen by undifferentiated cells are closely linked to proliferative status in culture. Stem Cells. 2004;22(4):556-569.

[3] Healy L, May G, Gale K, Grosveld F, Greaves M, Enver T. The stem cell antigen CD34 functions as a regulator of hemopoietic cell adhesion. Proc. Natl. Acad. Sci. USA. 1995 Dec 19;92(26):12240-4.

[4] AbuSamra DB, Aleisa FA, Al-Amoodi AS, Jalal Ahmed HM, Chin CJ, Abuelela AF, Bergam P, Sougrat R, Merzaban JS. Not just a marker: CD34 on human hematopoietic stem/progenitor cells dominates vascular selectin binding along with CD44. Blood Adv. 2017 Dec 26;1(27):2799-2816.

[5] Giassi LJ, Pearson T, Shultz LD, Laning J, Biber K, Kraus M, Woda BA, Schmidt MR, Woodland RT, Rossini AA, Greiner DL. Expanded CD34+ human umbilical cord blood cells generate multiple lymphohematopoietic lineages in NOD-scid IL2rgamma(null) mice. Exp. Biol. Med. (Maywood). 2008 Aug;233(8):997-1012.

[6] Verma B, Wesa A. Establishment of Humanized Mice from Peripheral Blood Mononuclear Cells or Cord Blood CD34+ Hematopoietic Stem Cells for Immune-Oncology Studies Evaluating New Therapeutic Agents. Curr. Protoc. Pharmacol. 2020 Jun;89(1):e77.

[7] King MA, Covassin L, Brehm MA, Racki W, Pearson T, Leif J, Laning J, Fodor W, Foreman O, Burzenski L, Chase TH. Human peripheral blood leucocyte non‐obese diabetic‐severe combined immunodeficiency interleukin‐2 receptor gamma chain gene mouse model of xenogeneic graft‐versus‐host‐like disease and the role of host major histocompatibility complex. Clin. Exp. Immunol. 2009 Jul;157(1):104-18.

[8] Meraz IM, Majidi M, Meng F, Shao R, Ha MJ, Neri S, Fang B, Lin SH, Tinkey PT, Shpall EJ, Morris J. An improved patient-derived xenograft humanized mouse model for evaluation of lung cancer immune responses. Cancer Immunol. Res. 2019 Jun 11.

 

Tags: Humanized Models