In Vitro and In Vivo Models of CLL–T Cell Interactions: Implications for Drug Testing

Abstract

T cells are key components in environments that support chronic lymphocytic leukemia (CLL), activating CLL-cell proliferation and survival. Here, we review in vitro and in vivo model systems that mimic CLL–T-cell interactions, since these are critical for CLL-cell division and resistance to some types of therapy (such as DNA-damaging drugs or BH3-mimetic venetoclax). We discuss approaches for direct CLL-cell co-culture with autologous T cells, models utilizing supportive cell lines engineered to express T-cell factors (such as CD40L) or stimulating CLL cells with combinations of recombinant factors (CD40L, interleukins IL4 or IL21, INFγ) and additional B-cell receptor (BCR) activation with anti-IgM antibody. We also summarize strategies for CLL co-transplantation with autologous T cells into immunodeficient mice (NOD/SCID, NSG, NOG) to generate patient-derived xenografts (PDX) and the role of T cells in transgenic CLL mouse models based on TCL1 overexpression (Eµ-TCL1). We further discuss how these in vitro and in vivo models could be used to test drugs to uncover the effects of targeted therapies (such as inhibitors of BTK, PI3K, SYK, AKT, MEK, CDKs, BCL2, and proteasome) or chemotherapy (fludarabine and bendamustine) on CLL–T-cell interactions and CLL proliferation.

NF-kB and the CLL microenvironment

“…CD4+ T cells increase STAT6 driven BCR signaling through release of IL4 ( 56 ) and in normal B cells, this has been shown to be via activation of the non-canonical NF-κB pathway ( 57 ). CD4+ T cell activation of the non-canonical NF-κB pathway in CLL is also through CD40L activation of CD40 ( 58 , 59 ) and, as a result, CD40L and IL4 are important components of laboratory-based co-culture systems which aim to mimic the tumor microenvironment ( 40 , 60 , 61 ).…”

Stromal cells engineered to express T cell factors induce robust CLL cell proliferation in vitro and in PDX co-transplantations allowing the identification of RAF inhibitors as anti-proliferative drugs

“…CLL cell birth rates in lymph nodes are 0.5-3% per day, and newborn CLL cells can exit the lymph nodes to the peripheral blood [2,3]. Lymph node niche is believed to promote CLL proliferation via microenvironmental factors such as the CD40, Toll-like receptors (TLR), and B-cell receptor (BCR) activation [3,4]. Notably, BCR crosslinking in vitro is insufficient to induce CLL proliferation [5], suggesting other microenvironmental interactions are needed to (co)drive proliferation.…”

“…Notably, BCR crosslinking in vitro is insufficient to induce CLL proliferation [5], suggesting other microenvironmental interactions are needed to (co)drive proliferation. The prime candidates for pro-proliferative signals are CLL-CD4 + T-cell interactions, since in vitro CLL cells can be stimulated to proliferate by CLL-T-cell contact [4][5][6][7][8][9]. Furthermore, in immunodeficient mice, CLL cell engraftment depends on the co-transplantation of activated T-cells [10,11].…”

“…Studies of normal B-cells point to CD40 ligand's (CD40L/CD154) crucial relevance in normal B-cell proliferation and its synchronization with BCR activation in germinal centers [16]. In CLL, CD40 stimulation alone is not sufficient to drive proliferation and must be combined with other factors, such as adding recombinant IL4, IL21, or CpG-ODN [4,5,17]. This activates downstream signaling largely shared with BCR, including NFκB, AKT, MYC, and MAPK/ERK (reviewed in [18]).…”

“…Several in vitro microenvironment model systems have been developed to study CLL biology, perform in vitro screening of chemical libraries for compounds with anti-CLL activity, and guide therapeutic decisions [19][20][21][22]. Most such co-culture models involve human or murine fibroblast-like stromal cells that provide signals to overcome spontaneous CLL cell apoptosis and/or mimic adhesion-mediated drug resistance, but these do not trigger proliferation [4,[22][23][24][25]. To induce proliferation, co-culture models typically utilize costly recombinant factors like IL4 or IL21 added to co-cultures with CD40L-expressing stromal cells; however, in such conditions, the CLL proliferation rate is relatively low and highly variable [9,26].…”