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Unprecedented progress in our understanding of the biologic and
molecular genetic underpinnings of human plasma-cell neoplasms (PCN) has
led to the development of novel targeted therapeutic agents – including
proteasome blockers (bortezomib), immunomodulatory agents
(lenalidomide), therapeutic antibodies, and a variety of emerging
small-drug inhibitors of cellular signal transduction pathways – that
are now beginning to produce tangible benefits for patients with PCN.
Nonetheless, the prognosis and outcome of PCN remain grim, caused in
large measure by serious limitations in our knowledge of the genetic
pathways of malignant plasma-cell (PC) transformation.
The research project, entitled, "Defining genetic pathways of plasma-cell neoplasia," seeks to address outstanding questions on the genetics of PCN in order to facilitate ongoing efforts by the clinical research community and the pharmacological industry to treat and prevent these malignancies more effectively. Using two powerful, complementary, unbiased approaches to genetic forward screening in a newly developed mouse model of human PCN, designated iMycΔΕμ, we will first identify candidate driver genes involved in PC tumor progression in mice, and then evaluate these genes across the mouse-human species barrier to identify and validate the involvement of orthologous human driver genes in human PCN. To accomplish this goal, we propose three innovative, closely interrelated, specific research aims.
Aim 1 is to identify PCN driver genes in a virus-based forward genetic
screen in mice. The rationale for this aim is based in part on findings
from cancer screens, showing that the Moloney-based murine leukemia
virus, MOL4070LTR, accelerates PC tumors in strain C.iMycΔΕμ
mice – a gene-insertion model of the Myc-activating T(12;15)(Igh-Myc)
translocation. Aim 2 is to identify PCN driver genes in a
transposon-based forward genetic screen in mice. The rationale for this
aim develops from evidence that the Sleeping Beauty (SB)
transposon-based cancer screen provides a flexible, efficient gene
discovery tool that complements virus-dependent screens; e.g., by its
applicability to quiescent cells such as PCs. Aim 3 is to validate the
involvement of human orthologs of the putative driver genes identified
in Aims 1 and 2 in human PCN, and the potential of these genes as new
targets for cancer therapy and prevention (Fig. 1).
Figure 1: Premalignant conditions
in MM, WM and CLL (panel A) and experimental strategy to elucidate
genetic tumor progression pathways in Myc-transgenic mice of great
relevance for human MM/WM (panel B).
(A) MGUS (IgG- or IgA-producing in the vast majority of cases), IgM MGUS and MBL represent the earliest detectable stages in the manifestation of MM, WM and B-CLL (chronic lymphocytic leukemia), respectively. The three premalignant conditions can be viewed as suspended states in the continuum from premalignant alteration to malignant disease. Alternatively, it has recently been proposed that MGUS should be viewed as a malignant tumor that is kept at bay by an ongoing immune response.
(B) We use iMyc-transgenic mice, which are prone to malignant B-cell tumors sharing important features with human MM and WM, in two complementary forward-genetic screens to unravel the genetic pathways of tumor progression.
Transgenic mouse models of human cancer are experimental model systems
that rely on laboratory mice that have been genetically manipulated to
render them prone to neoplasms that accurately recapitulate important
features of their human cancer counterparts. Model systems of this
sort: enable researchers to study the onset and progression of cancer in
ways that cannot be pursued in human beings; advance our understanding
of the molecular genetic and biological events that contribute to the
development and spread of cancer cells; and provide a valuable
preclinical platform for evaluating new approaches to treat and prevent
cancer in patients. The latter is particularly important in
circumstances in which drug testing requires an intact, immunocompetent
animal that is able to produce the same kind of tumor microenvironment
and recruit the same types of tumor bystander cells commonly found in
human patients. To give but one example, therapeutic antibodies target
cancer cells by recruiting normal immune cells to the site of attack;
thus, the preclinical testing of these antibodies requires strains of
laboratory mice that have a normal, fully functioning immune system.
The development of an immunocompetent, transgenic mouse model of human WM that will be useful for preclinical testing of WM drug candidates
is the main objective of this research. To that end, we
are generating a designer model of human WM designated C.IL6/BCL2/AIDnull.
This model combines three crucial pathogenetic factors of human WM –
namely the B-lymphocyte growth, differentiation and survival factor
IL-6, the cellular oncoprotein BCL-2, and the inability of WM cells to
perform immunoglobulin isotype switching (AIDnull) – on the
genetic backgroud of BALB/c (abbreviated as C). Strain C mice are
highly susceptible to malignant B-lymphocyte transformation (Fig. 2).
Figure 2: Schematic overview of the pathogenesis of lymphoplasmacytic lymphoma (LPL)-WM (panel A) and
transgenic mouse strains that we propose to use for modeling human
LPL-WM in mice (panel B)
(A) IL6 and BCL2 have been
identified as "WM genes" – genes that confer genetic proclivity to the
disease (left). Additionally, IL-6 and BCL-2 are major player in the
LPL-WM cells (right)
(B) All strains are on the same genetic
background of BALB/c (C), an important precondition for intercrossing
the various transgenes without jeopardizing crucial practical issues of
this project, such as the ability to adoptively transfer fully
transformed tumor cells or premalignant B-lineage cells from transgenic
mice. We hypothesize that strain C.IL6/BCL2/AID-/- mice will develop IgM+ WM-like tumors.
The ongoing research project, entitled "Preclinical validation of IL-6 for translational myeloma research,"
seeks to address outstanding questions on the biology of IL-6 in order
to facilitate ongoing efforts to target IL-6 more effectively in patients with myeloma (Fig. 3).
Using transgenic mouse models of IL-6-dependent plasmacytoma (PCT) as
the principal experimental model system, we will: rigorously evaluate
the biological significance of IL-6 in Myc-driven PCT development; weigh
the relative contributions of autocrine and paracrine sources of IL-6
to plasma-cell tumor development; and assess the role that an IL-6
signaling pathway that has been largely neglected in myeloma research –
i.e., IL-6 trans-signaling – plays in plasma-cell neoplasia in
mice. Taking advantage of a set of newly developed transgenic BALB/c
(C) mouse strains available only at The University of Iowa Carver
College of Medicine, we are pursuing three important, closely
interrelated, specific research aims.
Aim 1 is to determine the global significance of IL-6 in Myc-driven PCT
in mice. The rationale for this aim includes findings that suggest IL-6
may be dispensable for plasma-cell tumors (i.e., that pathway activity
can be maintained in the absence of an IL-6/IL-6R [IL-6 receptor]
interaction). The constitutive activation of Stat3 in Abl/Myc
retrovirus-induced PCT in mice and activated B-cell like diffuse large
B-cell lymphoma in humans provides a key example to that end. Aim 2 is
to evaluate the relative importance of autocrine versus paracrine IL-6
in Myc-driven PCT in mice. The rationale for this aim includes findings
suggesting that autocrine IL-6 is important for carcinomas (such as
lung and breast cancers), whereas paracrine IL-6 is critical for blood
cancers (including lymphoma and myeloma). However, the evidence for the
latter is largely circumstantial, and the significance of paracrine
IL-6 for malignant plasma-cell transformation has never been
unequivocally proven in a genetic study. Aim 3 is to assess the role of
IL-6 trans-signaling in Myc-driven PCT in mice. The rationale for this
aim includes recent findings implicating IL-6 trans-signaling in
inflammation-dependent solid and hematopoietic cancers in humans and
mice and, more specifically, findings indicating that IL-6
trans-signaling plays a key role in the development of plasma-cell
hyperplasia in mice.
Figure 3: Targeting IL-6 for improved outcome of MM.
The scheme depicts novel approaches to inhibit IL-6 signaling in myeloma
cells (bottom) and/or bone marrow stroma cells (top): IL-6, red
squares; IL-6R/gp80, Y-shaped symbol; gp130, blue line). Numbered
yellow circles indicate active research areas, which can be categorized
(1) Antibodies to, or antagonists of, IL-6R, including
Sant7 (a modified human IL-6 that binds to gp80 with higher affinity
than normal IL-6 does; however, because Sant7 does not recruit gp130 to
the IL-6R, it blocks downstream IL-6 signaling), MRA (humanized mouse
antibody to IL-6R; ACTEMBRA 200 [Chugai Pharmaceuticals]), ERBA (IL6R
antagonist), and NRI/Tocilizumab.
(2) Antibodies to IL-6, including mouse monoclonal
antibody BE-8 and chimeric antibody CNTO 328 (Centocor), which is
currently being tested in a phase-2 clinical trial with and without
(3) Inhibitors of IL-6 trans-signaling, including
sgp130-Fc (R&D Systems), which captures circulating IL-6 and thus
prevents it from binding to membrane-bound IL-6R.
(4) Inhibitors of gp130, such as gp130-targeting peptides.
(5) Small-molecule inhibitors, including Atiprimod (Callisto).
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