Annual Meeting Proceedings Part 1 - American Society of Clinical ...

536s Lymphoma and Plasma Cell Disorders
8106 General Poster Session (Board #39G), Mon, 1:15 PM-5:15 PM
Participation of BTK in MYD88 signaling in malignant cells expressing the
L265P mutation in Waldenstrom’s macroglobulinemia, and effect on
tumor cells with BTK-inhibitor PCI-32765 in combination with MYD88
pathway inhibitors. Presenting Author: Guang Yang, Dana-Farber Cancer
Institute, Boston, MA
Background: Bruton’s tyrosine kinase (BTK) promotes B-cell receptor
signaling along with B-cell expansion and survival through NF-�B and
MAPK. MYD88 L265P is a widely expressed somatic mutation in tumor
cells from WM patients. MYD88 L265P promotes enhanced tumor cell
survival through IRAK 1/4 mediated NF-�B and MAPK signaling. We
therefore sought to clarify the role of BTK signaling in MYD88 L265P
expressing WM cells, and the impact of BTK and MYD88/IRAK inhibition
on WM cell signaling and survival. Methods: Western blot analysis was
performed using total and phospho-specific BTK antibodies in MYD88
L265P expressing primary WM patient cells, BCWM.1 and MCWL-1 WM
cell lines following MYD88 knockdown by lentiviral transduction, and/or
use of MYD88 or IRAK signal inhibitors. Cells were also treated with the
BTK inhibitor PCI-32765, in the presence or absence of MYD88 homodimerization
or IRAK1/4 inhibitors. Annexin V/PIstaining was used to
assess cell survival, and synergism assessed with CalcuSyn software.
Results: BTK was phosphorylated in MYD88 L265P expressing WM cells.
Knockdown of MYD88 by lentiviral transduction, and/or use of MYD88 or
IRAK 1/4 kinase inhibitors led to decreased BTK phosphorylation. Phosphorylation
of BTK, TIRAP, a major TLR adapter protein for MYD88 signaling,
IRAK1, IKB�, ERK1/2 and STAT3 were significantly reduced following
treatment with PCI-32765. Treatment with PCI-32765 also induced
apoptosis of MYD88 L265P expressing WM cells, and showed synergistic
tumor cell killing in the presence of either MYD88 homodimerization or
IRAK 1/4 kinase inhibitors. Conclusions: BTK activation is facilitated by
MYD88 pathway signaling in L265P expressing WM cells, and participates
in MYD88 downstream signaling. Inhibition of BTK by PCI-327625 led to
robust tumor cell killing of MYD88 L265P expressing WM,cells, which was
potentiated by MYD88 pathway inhibitors. These studies provide the
framework for the investigation of BTK inhibitors in WM, as single agents
and in combination with MYD88 pathway inhibitors.
8108 General Poster Session (Board #40A), Mon, 1:15 PM-5:15 PM
miRNA expression profiling of CD20� plasma cell myeloma (PCM):
Upregulation of miR-155 shedding new insight into disease biology and
clinicopathologic behavior. Presenting Author: Ravindra Kolhe, Department
of Pathology, Georgia Health Sciences University, Augusta, GA
Background: Up to15-20% of patients with PCM have expression of CD20,
although the significance of this remains unclear. The prognostic significance
of CD20 expression in PCM is unclear. Recently, a class of
noncoding RNAs, miRNAs, were identified as critical gene regulators in cell
growth, disease, and development. Our study investigates importance of
miRNAs in cases of CD20� PCM and correlates them with clinicopathological
parameters. Methods: The miRNA expression profile of CD20� PCM
(n�6), diffuse large B cell lymphoma (DLBCL) (n�6), and CD20 negative
PCM (n�8) were evaluated using the Affeymertrix miRNA microarray
platform on GeneChip miRNA 2.0 array in paraffin-embedded samples.
After hybridization and data acquisition, we used Partek Genomics Suite
software for RMA normalization and to determine statistically significant
differences in miRNA expression between experimental groups by ANOVA
and pairwise comparisons (two-sided ��0.05). Results: miRNA expression
profiles of CD20� PCM, show upto �4 times upregulation of 7 miRNAs
and downregulation of 8 miRNAs. miR-155, the miRNA upregulated in
various B cell lymphomas and plays a key role in the lymphomagenesis, was
amongst the highest miRNAs that were upregulated. Conclusions: miR-155
is known to repress SH2-domain containing inositol-5-phosphatase-1
(SHIP-1), which is a critical phosphatase that negatively down modulates
AKT pathway and has functions during normal B-cell development.
Physiologically, miR-155 is upregulated during B-cell activation upon
antigen stimulation and so plays a role in antibody class switching and
plasma cell formation. We propose that this overexpression of miR-155 in
CD20� PCM unblocks AKT activity, inducing cell proliferation and may
explain some of the immunophenotypic behavior of CD20� PCM. This work
is intriguing for the new information it provides about the role of miR-155
in CD20� PCM. Furthermore, the phenotype of miR-155�ve CD20� PCM
might ultimately provide new insights into regulation of poorly understood
steps in B cell differentiation and maturation into plasma cell and more
importantly the clinicopathological behavior of this entity.
8107 General Poster Session (Board #39H), Mon, 1:15 PM-5:15 PM
Use of whole genome sequencing to identify highly recurrent somatic
mutations in Waldenström’s macroglobulinemia. Presenting Author: Zachary
R Hunter, Dana-Farber Cancer Institute, Boston, MA
Background: Waldenstrom’s Macroglobulinemia (WM) is an IgM secreting
lymphoplasmacytic lymphoma. The genetic basis for this disease remains
to be clarified. Methods: We performed whole genome sequencing (WGS)
using CD19 � selected bone marrow lymphoplasmacytic cells (LPC) from
30 WM patients. For 10 of these patients, paired CD19 � depleted
peripheral blood samples were used for WGS as normal controls. Results:
The most common somatic variants identified and validated by Sanger
sequencing included MYD88 L265P, an activating mutation for IRAK/
TRAF6/NFKB and MAPK signaling, which was observed in 27/30 (90%)
patients; the N-terminal domain of CXCR4, which included mutations
associated with WHIM syndrome and confer constitutive CXCR4 signaling
resulting from dysfunctional receptor endocytosis, a finding observed in
8/30 (27%) patients, and ARID1A (5/30; 17%), a tumor suppressor gene.
Less common somatic variants were also identified in MUC16 (4/30; 13%),
TRAF2 (3/30; 10%), TRRAP (3/30; 10%) and MYBBP1A (2/30; 7%).
Conclusions: Using WGS and confirmatory Sanger sequencing, we have
identified several somatic variants with oncogenic function, the most
common of which include MYD88 L265P, the N-terminal domain of
CXCR4, and ARID1A.
TPS8109 General Poster Session (Board #40B), Mon, 1:15 PM-5:15 PM
Phase I/II study of investigational agent MLN8237 (alisertib) plus rituximab
with or without vincristine in patients (pts) with relapsed/refractory
(rel/ref) aggressive diffuse large B-cell lymphoma (DLBCL)/transformed
follicular lymphoma (TFL). Presenting Author: Daniel Oscar Persky, University
of Arizona Cancer Center, Tucson, AZ
Background: DLBCL and TFL are aggressive subtypes of non-Hodgkin
lymphoma (NHL), with poor prognosis in the rel/ref setting. Alternatives to
standard rituximab-based treatments are needed. MLN8237 is an oral,
selective inhibitor of Aurora A kinase – a key mitotic regulator overexpressed/
amplified in various human cancers, including lymphomas (Hamada et al,
2003; Yakushijin et al, 2004; Qi et al, 2011). Emerging data from a phase
2 study suggest that MLN8237 has single agent activity in aggressive NHL
(Friedberg et al, ASH 2011). MLN8237 � rituximab (MR) � vincristine
(MRV) has also shown activity in preclinical B-cell NHL models (Mahadevan
et al, ASH 2011). Further clinical evaluation of these regimens in
rel/ref B-cell NHL is warranted. Methods: This single-arm phase 1/2 study
(ClinicalTrials.gov #NCT01397825) aims to assess the safety, efficacy,
and pharmacokinetics of, and determine a recommended phase 2 dose
(RP2D) and schedule for, MR and MRV in adults with CD20� rel/ref
DLBCL/TFL after 1–4 prior regimens (including ASCT) and ECOG PS 0–2.
Pts with other aggressive B-cell lymphomas may also enroll in phase 1.
~100 pts will be recruited at 22 sites in the US, UK, Italy, and Spain, and
study duration will be ~2 years. In part 1, a safety lead-in cohort will receive
MR (table), with the RP2D determined as when �2 dose limiting toxicities
(DLT) occur in 6 pts in cycle 1. MRV dose escalation (part 2) will then
follow a 3�3 design. Phase 2 enrollment (part 3) will follow a Simon
optimal 2-stage design, with pts receiving MRV at the RP2D determined in
part 2. The primary phase 2 objective is overall response rate by IWG
criteria. Responders may continue MLN8237 if clinical benefit is seen and
if the regimen is tolerable. Enrollment as of 20 Jan 2012 is 6 evaluable pts.
Starting doses: repeating 21-day cycles
MLN8237 Rituximab Vincristine
Phase 1 (part 1) 50 mg ECT BID, days 1–7* 375 mg/m 2 IV, day 1 –
(part 2) ~50% of MR RP2D, days 1–7 †
as above 1.4 mg/m 2 IV, days 1, 8 †
Phase 2 (part 3) RP2D from part 2
ECT, enteric coated tablet; BID, twice daily; *dose reduction permitted; † dose/schedule
adjustments based on cycle 1 DLT.
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