10 research outputs found
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Genomic Biomarkers Predict Response/Resistance to Lenalidomide in Non-Del(5q) Myelodysplastic Syndromes
Abstract
Lenalidomide (Len) is FDA approved for the treatment of patients (pts) with lower-risk, transfusion-dependent myelodysplastic syndromes (MDS) with deletion(5q). It is frequently used in lower-risk pts with non-del(5q) MDS, with a transfusion independence response rate of 27%. Identification of pts who may or may not respond to Len can prevent prolonged exposure to ineffective therapy, avoid toxicities, and decrease unnecessary costs. Clinical or genomic data have limited utility in predicting response/resistance to Len.
We developed an unbiased framework to study the association of several mutations/cytogenetic abnormalities in predicting response/resistance to Len in non-del(5q) pts, analogous to Netflix or Amazon's recommender system, in which customers who bought products A and B are likely to buy C: pts who have a molecular/cytogenetic abnormalities in gene A, and B are likely to respond or not respond to Len.
Clinical and genomic data from pts with MDS or other myeloid malignancies diagnosed according to 2008 WHO criteria between 02/2004 and 06/2015 were analyzed. Next generation targeted deep sequencing panel of 50 genes that are commonly mutated in MDS and myeloid malignancies was included. Association rules using an apriori algorithm were used to study the relationships among multiple genes/cytogenetic abnormalities and response/resistance to Len. Responses included complete and partial remission and hematologic improvement (CR, PR, HI) per IWG 2006 criteria. Pts with stable disease or progressive disease were considered resistant. Association rules are a machine learning algorithm used to identify the association of variables based on their relationships. Rules with the highest confidence (that an association exists) and highest lift (measuring the strength of the association) were chosen.
Of 139 pts treated with Len as monotherapy or in combination for at least 2 cycles included, 118 (85%) had MDS and 21 (15%) had other myeloid malignancies. Median age at diagnosis was 69 years (range 20-90 yrs) and 45% were female. Risk stratification by IPSS-R for MDS pts; 51.5 % had very low/low risk, 19.5% intermediate, and 29% high and very high risk disease. Most pts 100 (73%) had non-del(5q) abnormalities, others (39) had del(5q). Cytogenetic abnormalities for the non-del(5q) cohort included 58 pts with normal karyotype (NK), 19 pts with complex karyotype (CK), 4 pts with trisomy 8, 3 pts with del(7q) abnormalities, and 15 pts with other abnormalities. A total of 108 (79%) pts were treated with Len monotherapy. The median duration of treatment was 6 months (range 2- 66 m). Response rates were 46% (n=46) in the non-del(5q) cohort and 74% (n=29) in del(5q).
Association rules identified the following combinations of genomic/cytogenetic abnormalities to predict response to Len in non-del(5q): (DDX41, NK) and (MECOM, KDM6A/KDM6B). The combination of the following abnormalities predicted resistance (ASXL1, TET2, NK), (DNMT3A, SF3B1), (TP53, del(5q)+CK), (STAG2, IDH 1/2, NK), (EZH2, NK), (BCOR/ BCORL1, NK), (JAK2, TET2, NK), (U2AF1, +/- ETV6, NK). [Table 1] Only TP53/CK mutations predicted resistance to Len in del(5q) pts. These associations are present in 39% of pts with non-del(5q), and have a specificity of 77%, with a negative predictive value and sensitivity=100%. The algorithm predicted response/resistance to Len with 82% accuracy.
The median OS for non-del(5q) pts was 33.2m [95% CI: 19.9, 40.5]. The median OS for responders was 54.8 compared to 24.7 m for non-responders p=.017. The median OS for rules that predicted response was 70.3 m (95% CI: 70.3-NA). The median OS for pts with del(5q) + CK with a TP53 mutation was 9.8m. Several genomic combinations predicted very poor overall survival, including: (ETV6, U2AF1, NK), (BCOR/ BCORL1, NK), (EZH2, NK) , (JAK2, TET2, NK), with median OS of 10.7 m, 7.6 m, 10.8 m and 7.6 m, respectively. [Figure 1]
Genomic biomarkers can identify 39% of non-del(5q) MDS pts who may or may not respond to treatment with very high accuracy. Although these abnormalities are only present in a subset of pts, treatment options for these pts can be tailored, by offering alternative therapies to pts with lower-risk disease who may not respond to Len, and preferentially offering Len to those who are more likely to respond. This study highlights how advanced analytic technologies such as machine learning can translate genomic/clinic data into useful clinical tools.
Disclosures
Sekeres: Opsona: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Opsona: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees. Gerds:Celgene: Consultancy; Apexx Oncology: Consultancy; CTI Biopharma: Consultancy; Incyte: Consultancy. Carraway:Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Amgen: Membership on an entity's Board of Directors or advisory committees; Novartis: Speakers Bureau; Balaxa: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; FibroGen: Consultancy; Jazz: Speakers Bureau; Agios: Consultancy, Speakers Bureau. Santini:Novartis: Honoraria; Amgen: Membership on an entity's Board of Directors or advisory committees; Otsuka: Consultancy; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria, Research Funding; AbbVie: Membership on an entity's Board of Directors or advisory committees. Maciejewski:Ra Pharmaceuticals, Inc: Consultancy; Ra Pharmaceuticals, Inc: Consultancy; Apellis Pharmaceuticals: Consultancy; Apellis Pharmaceuticals: Consultancy; Alexion Pharmaceuticals, Inc.: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Alexion Pharmaceuticals, Inc.: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Nazha:MEI: Consultancy
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Context dependent effects of ascorbic acid treatment in TET2 mutant myeloid neoplasia
Loss-of-function TET2 mutations (
TET2
MT
) are common in myeloid neoplasia. TET2, a DNA dioxygenase, requires 2-oxoglutarate and Fe(II) to oxidize 5-methylcytosine.
TET2
MT
thus result in hypermethylation and transcriptional repression. Ascorbic acid (AA) increases dioxygenase activity by facilitating Fe(III)/Fe(II) redox reaction and may alleviate some biological consequences of
TET2
MT
by restoring dioxygenase activity. Here, we report the utility of AA in the prevention of
TET2
MT
myeloid neoplasia (MN), clarify the mechanistic underpinning of the TET2-AA interactions, and demonstrate that the ability of AA to restore TET2 activity in cells depends on N- and C-terminal lysine acetylation and nature of
TET2
MT
. Consequently, pharmacologic modulation of acetyltransferases and histone deacetylases may regulate TET dioxygenase-dependent AA effects. Thus, our study highlights the contribution of factors that may enhance or attenuate AA effects on TET2 and provides a rationale for novel therapeutic approaches including combinations of AA with class I/II HDAC inhibitor or sirtuin activators in
TET2
MT
leukemia.
Using TET2- and ascorbic acid deficient model systems Guan et al show that long term treatment with ascorbic acid delays myeloid neoplasia in mice and reveal a complex interplay of post-translational modification of lysine residues that modulate TET2 activity in neoplastic evolution
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Modulation of TET2 Activity By Ascorbic Acid and Factors Affecting Lysine Acetylation
Abstract Loss-of-function TET2 mutations (TET2mt) strongly associate with myeloid neoplasia. TET2 is an α-ketoglutarate (αKG) and Fe2+-dependent DNA-dioxygenase that hydroxylates 5-methylcytosine-DNA (5mC), producing 5-hydroxymethylcytosine-DNA (5hmC). Ascorbic acid (AA) activates TET2 and other dioxygenases via several direct and indirect mechanisms, including recharging Fe3+ to Fe2+. Recent studies, chiefly in murine models, indicated that increased AA availability may alleviate the decreased of dioxygenase activity in cells affected by hypomorphic mutations via up-modulation of the remaining functional activity, particularly in heterozygous TET2mt, but it may also act via increased compensatory activity of other TET enzymes. Beyond these observations, the mechanisms of AA action in TET2mt bone marrow cells are not well understood. Yet to be identified are factors that modulate the effects of AA. We performed in silico binding prediction validated by surface plasmon resonance assay coupled with confirmatory in vitro and in vivo assays. In cell free system we demonstrated the AA increases the production of 5mC-DNA oxidation products (TDOP, 5hmC/5fmC/5caC) in a mixture of recombinant TET2wt:TET2S1898F 1:1 mimicking heterozygous TET2mt neoplasia. Using cell-free system with recombinant TET2 protein and the aforementioned biophysicochemical analyses, we further demonstrated that AA binds to the TET2 catalytic domain adjacent to Fe2+/aKG binding site and recycle Fe2+ back into the catalysis. In vitro cultures demonstrated that AA increases TDOP and slows proliferation in Tet2+/- and Tet2-/- mouse bone marrow progenitor cells. A similar but less pronounced effect was also observed in shRNA TET2KD MOLM13 and K562 cells in which AA restored the original proliferation rate of these cells which was increased by TET2KD. Finally, using cultures of healthy human marrows (retrovirally transduced with TET2 or scrambled shRNA), we showed that AA decreased the proliferation rate of these cells similar to ectopic overexpression of TET2. When primary MDS bone marrow samples with various TET2mt (n=4) were studied, the effects of AA were less consistent. Consistent with the observation, in SIG-M5 cell line characterized by natural TET2 mutations, AA only modestly increased 5hmC 2-fold. While, in high TET2 expression CMK cell line, AA increased 5hmC 6-fold. In vivo, high-dose oral AA increased TDOP, decreased spleen size and myeloproliferation, and prolonged survival in sublethally-irradiated Tet2+/-Gulo-/- and Tet2+/- mice. However the AA treatment has only a modest effect on the overall survival of these mice. Recent reports indicated that TET2 activity is down regulated by HDAC1/2 by deacetylation at the N-terminus (Zhang et al., Molecular Cell 65, 323-335, 2017) and up regulated by Sirtuin 1 (Class III HDAC) via lysine deacetylation in the catalytic domain (Sun et al., abstract, Blood 2016 128:1053). In search of factors that modulate the effects of TET, experiments were performed to determine if acetylation also modulates the activity of TET in leukemia cells. Inhibition of class I and II but not class III HDACs by trichostatin A (TSA) increased the 5hmC level 1.5-fold. However, when combined with AA, TSA amplifies the 5hmC level nearly 4-fold. Acetylation of lysine in the catalytic site inactivates TET via structural perturbations. Therefore, activation of class III HDACs should activate TET2 and inhibition should inhibit it. Indeed, consistent with this presumption, in leukemia cells, we observed that activation of sirtuin, a class III HDAC, by small molecule SRT1720, significantly increased 5hmC while the inhibitors 3-TYP and sirtnol reduced 5hmC. Context-dependent TET2 acetylation/deacetylation significantly affects its AA-mediated activation, and thus MDS/MPN clones with dysregulation of histone acetyltransferases and/or histone deacetylases may be less responsive to AA alone. In sum, we demonstrate the mechanism and context of AA-mediated TET activation which may guide novel treatment strategies for TET2-deficient neoplasia. Our results show that TET2 activity can be modulated by various mechanisms including acetylation/deacetylation interactions with the effects of AA. Disclosures Nazha: MEI: Consultancy. Sekeres:Celgene: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Opsona: Membership on an entity's Board of Directors or advisory committees; Opsona: Membership on an entity's Board of Directors or advisory committees. Maciejewski:Alexion Pharmaceuticals, Inc.: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Apellis Pharmaceuticals: Consultancy; Apellis Pharmaceuticals: Consultancy; Ra Pharmaceuticals, Inc: Consultancy; Ra Pharmaceuticals, Inc: Consultancy; Alexion Pharmaceuticals, Inc.: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau