ALBERT

Description: Background: We reported a high response rate with the combination of lenalidomide plus dexamethasone (Rev/Dex) as initial therapy for newly diagnosed multiple myeloma (Blood2005;106:4050–3). We now present new data on time to progression (TTP), progression free survival (PFS), and overall survival (OS) from this phase II trial, and also include updated response data. Patients and Methods: 34 patients (23 male and 11 female) were enrolled. Lenalidomide was given orally 25 mg daily on days 1–21 of a 28-day cycle. Dexamethasone was given orally at a dose of 40 mg daily on days 1–4, 9–12, 17–20 of each cycle. Patients were allowed to go off treatment after 4 cycles of therapy to pursue autologous stem cell transplantation (SCT), but treatment beyond four cycles was permitted at the physician’s discretion. For patients continuing therapy beyond 4 months, the dose of dexamethasone was reduced to 40 mg on days 1–4 of each cycle. Response was assessed by modified EBMT/International Myeloma Working Group Uniform Response criteria. All patients received aspirin (81 mg or 325 mg daily) as prophylaxis against DVT. Results: The median age was 64 years (range, 32–78). All patients were evaluable for response and toxicity. Median follow up is 21 months. Thirteen patients proceeded to SCT following initial therapy with Rev/Dex and were censored at that time point for purposes of calculation of response, TTP and PFS. Patients who discontinued therapy to proceed to SCT received a median of 4 cycles of therapy (range, 4–13), while those staying on Rev/Dex (n=21) received a median of 19 cycles of therapy (range, 2–30). Thirty-one of 34 patients (91%) achieved an objective response to therapy; including 6 patients (18%) achieving a complete response (CR) and 13 patients (38%) achieving very good partial response for a CR+VGPR rate of 56%. The CR+VGPR rate among the 21 patients staying on Rev/Dex as primary therapy without SCT was 67% (CR 24%, VGPR 43%). Median TTP, PFS, and OS have not been reached (Figure). By Kaplan-Meier method, the estimated 2 year progression rate was 18%. The 2-year PFS rate and OS rate were 74% and 91%, respectively. Fifty-five percent of patients experienced grade 3 or higher non-hematologic toxicity at any point during therapy, most commonly fatigue (21%), neutropenia (21%), anxiety (6%), pneumonitis (6%), muscle weakness (6%), and rash (6%). Two patients died on study: one attributed to infection unrelated to therapy, the patient had stopped all therapy for over a month before the fatal infection occurred; the other death was due to infection felt possibly related to therapy. One patient developed a pulmonary embolism (grade 4 toxicity), but recovered with therapy; no other patient developed deep vein thrombosis or pulmonary embolism. Conclusion: Rev/Dex is highly active and well tolerated in the treatment of newly diagnosed multiple myeloma with a high CR+VGPR rate of 56% for the trial, and 67% among the subset of patients receiving this regimen as primary therapy. Responses are durable with a low progression rate at 2 years.

Description: Abstract 3884 Poster Board III-820 Background and Objective Thalidomide/dexamethasone (thal/dex) combination has shown high activity in newly diagnosed multiple myeloma (MM) (Rajkumar SV. at al, J Clin Oncol 2006;24:431-436). In newly diagnosed patients, lenalidomide/dexamethasone (len/dex) has demonstrated superiority compared with high-dose dexamethasone alone (Zonder JA et al, Blood 2007;110:77). Although both thal/dex and len/dex are active in newly diagnosed MM, no randomized trial has been reported comparing these two regimens, and unfortunately none are ongoing or planned. We compared the efficacy and the toxicity of thal/dex and len/dex as primary therapy in 411 newly diagnosed MM patients treated at the Mayo Clinic. Patients and methods 411 consecutive patients seen at Mayo Clinic between 2001 and 2008, who received induction with thal/dex (n=183) or len/dex (n=288) were retrospectively studied. Thalidomide was given at a dose ranging from 100 mg/day to 400 mg/day continuously; the lenalidomide dose was 25 mg/day, days 1-21 on a 28-day cycle. All patients received dexamethasone, either at high-dose (40 mg orally on days 1-4, 9-12, and 17-20) or at low-dose (40 mg orally day 1, 8, 15, 22); each cycle was repeated every 4 weeks. In addition, a case-matched subgroup analysis that adjusted for age, gender and transplantation status was performed among patients who received high-dose dexamethasone comparing the thal/dex (n=72) and len/dex (n=72) groups. Outcome was analyzed on an intention-to-treat basis. The Chi-square or the rank sum tests were used to compare variables. Time-to-event analysis was performed using the Kaplan-Meier method and all comparisons were determined by the log-rank test and by the Cox proportional hazards model. Results On intention-to-treat analysis, of 411 patients, 80.3% versus 61.2% patients, respectively in the len/dex group and in the thal/dex group (p 〈 0.001), achieved at least a partial response. A significant difference between the 2 groups was found in terms of both very good partial response or better (34.2% vs 12.0%, p 〈 0.001) and complete response rate (13.6% vs 3.3%, p 〈 0.001). Duration of therapy was significantly longer in len/dex patients as compared to thal/dex patients: 36.7% vs 12.6% of patients who did not stop treatment to receive SCT were still receiving therapy at 1 year (p 〈 0.001).Time-to-progression was significantly better in the len/dex group than in patients receiving thal/dex (median 27.4 vs 17.2 months, HR 0.64; 95% CI 0.44-0.93; p = 0.019). Similarly, progression-free-survival was significantly higher in len/dex patients (median 26.7 vs 17.1 months, HR 0.69; 95% CI 0.48-0.98; p = 0.036). This translated into an increase in overall survival (OS) (median not reached for len/dex group compared to 57.2 months in thal/dex patients, HR 0.60; 95% CI 0.40-0.92; p = 0.018). Survival advantages were evident in patients presenting with International Staging System Stage (ISS) I/II (HR 0.57; 95% CI 0.32-1.00; p = 0.052) at diagnosis but not in patients with ISS stage III in subgroup analysis. There was a trend toward better OS in len/dex group compared to thal/dex group both for patients who underwent transplant and for patients who did not. A similar rate of patients experienced at least one grade 3 or higher adverse event (57.5% vs 54.6% in len/dex and thal/dex groups, respectively, p = 0.568). However, the toxicity profile was different in the two groups: major grade 3-4 toxicities of len/dex were hematological, in particular neutropenia (14% with len/dex vs 0.6% with thal/dex, p

Description: The use of high-resolution aCGH allows for whole genome screening for copy number changes even in tumors that rarely produce metaphase spreads. The aim of this study was to characterize the aCGH findings in B-CLL, lung MALT lymphoma, Waldenström’s Macroglobulinemia (WM), Multiple Myeloma (MM) and human myeloma cell lines (HMCLs). This study includes 301 B-cell malignancy samples (19 B-CLL, 19 MALT, 20 WM, 48 HMCLs and 195 MM) and an additional 76 melanoma samples. We found MM to have the highest number of abnormalities per karyotype, with 15.8±7.5 (mean ± SD) and 20.6±17.0 in hyperdiploid (H) and non-hyperdiploid MM (NH), respectively. The remaining diseases have a significantly lower number of abnormalities: 5.5±4.5 in B-CLL, 3.5±1.96 in MALT and 4±3.5 in WM. Aneuploidy and biallelic deletions were rare events in these three diseases. B-CLL has two highly recurrent abnormalities involving 11q22-q23 (29%) and 13q14.3 (33%). A minimal deleted region (MDR) of ∼108Kb was identified at 11q comprising ACAT1, NPAT and ATM. In 13q, a 280Kb MDR was recognized, including TRIM13, KCNRG, mir-15a and mir-16. The subgroup of MALT patients with t(11;18) had very few abnormalities and the majority are a consequence of unbalanced translocations (MALT, BIRC2 and BIRC3 deletions). In the remaining MALT samples, trisomy 3 and 18q gain were the most common abnormalities (31% each). The 6q16.3-q26 deletion was the most common abnormality in WM (45%). In 4 of 9 patients with 6q deletion there was an associated 6p gain. The same observation was found in 2 of 3 patients with 8p loss and concomitant 8q gain. Interestingly, 6p and 8q gain were not identified in patients without 6q and 8p deletions. Deletion of 17p was the most common recurrent abnormality observed in our panel of B-cell cancers, being identified in MM, WM and B-CLL. In B-CLL and WM, small mono and biallelic deletions of the NF-kB negative regulators, TRAF3 and NFKBIA, were identified, suggesting a potential involvement of this pathway in the pathogenesis of these diseases. Hierarchical clustering of the aCGH data in MM identified 3 distinct groups, NH-MM, H-MM and MM with deletions of chromosome 13 and 14. Interestingly, MGUS and SMM were more commonly clustered with H-MM. Several sub-clusters were observed in the H-MM group, the most prominent being made up exclusively of patients from TC class D1. This cluster was characterized by the presence of trisomy 11 and 21. The 11q13 TC group clustered and was highly associated with 11q amplifications due to unbalanced t(11;14). The 6q deletion defined a cluster with WM, MM and Melanoma patient with that abnormality. In the case of B-CLL and MALT, the only identifiable clusters were observed in cases with 13q loss and trisomy 3, respectively. These results highlight both, the common and unique aCGH findings in several B-cell cancers. By hierarchical clustering, MM was significantly divergent from the rest of the B-cell diseases. Due to the limited number of abnormalities in B-CLL/MALT/WM, the clustering of these groups was exclusively based on the common abnormalities. The use of aCGH has refined the MDRs of 11q and 13q in B-CLL. Additionally, the identification of deletions affecting the NF-kB pathway in B-CLL and WM has prompted us to hypothesize on the importance of constitutive NF-κB activation in these diseases.

Description: Abstract 832 We hypothesized that new therapeutic targets for multiple myeloma (MM) could be discovered through the integrative computational analysis of genomic data. Accordingly, we generated gene expression profiling and copy number data on 250 clinically-annotated MM patient samples. Utilizing an outlier statistical approach, we identified HOXA9 as the top candidate gene for further investigation. HOXA9 expression was particularly high in patients lacking canonical MM chromosomal translocations, and allele-specific expression analysis suggested that this overexpression was mono-allelic. Indeed, focal copy number amplifications at the HOXA locus were observed in some patients. Outlier HOXA9 expression was further validated in both a collection of 52 MM cell lines and 414 primary patient samples previously described. To further verify the aberrant expression of HOXA9 in MM, we performed quantitative RT-PCR, which confirmed expression in all MM patients and cell lines tested, with high-level expression in a subset. To further investigate the mechanism of aberrant HOXA9 expression, we interrogated the pattern of histone modification at the HOXA locus because HOXA gene expression is particularly regulated by such chromatin marks. Accordingly, immunoprecipitation studies showed an aberrantly low level of histone 3 lysine 27 trimethylation marks (H3K27me3) at the HOXA9 locus. H3K27me3 modification is normally associated with silencing of HOXA9 in normal B-cell development. As such, it appears likely that the aberrant expression of HOXA9 in MM is due at least in part to defects in histone modification at this locus. To determine the functional consequences of HOXA9 expression in MM, we performed RNAi-mediated knock-down experiments in MM cell lines. Seven independent HOXA9 shRNAs that diminished HOXA9 expression resulted in growth inhibition of 12/14 MM cell lines tested. Taken together, these experiments indicate that HOXA9 is essential for survival of MM cells, and that the mechanism of HOXA9 expression relates to aberrant histone modification at the HOXA9 locus. The data thus suggest that HOXA9 is an attractive new therapeutic target for MM. Disclosures: No relevant conflicts of interest to declare.

Description: Constitutive activation of NF-κB signaling pathways has been identified in a variety of malignancies, however the molecular basis has remained largely unknown. Recently, we have shown mutations in 10 genes encoding positive and negative regulators of NF-κB signaling pathways in 20% of multiple myeloma (MM) patients, with TRAF3 being the most commonly affected gene. TRAF3 is a negative regulator of NF-κB signal and its inactivation results in the constitutive activation of the alternative or noncanonical NF-κB signaling pathway. With the exception of two recent studies in MM, there is no available data regarding the mechanisms that dysregulate NF-κB in other B-cell malignancies. The aim of this study was to analyze the TRAF3 status in a cohort of Waldenström’s macroglobulinemia (WM) patients. Forty-nine WM were analyzed by using a combination of array-based comparative genomic hybridization (aCGH) and cIgM-FISH. Genome sequencing from 28 of these patients was performed on all coding exons of TRAF3 and the adjacent intron-exon junctions. A NF-κB index, as a measure of NF-κB transcription activity, was calculated from 21 patients with gene expression data. The index is the mean expression level of four probe sets differentially expressed between human myeloma cell lines with identified mutations compared to those without identified NF-κB pathway mutations (CD74, IL2RG and 2x TNFAIP3). By performing aCGH we identified a small monoallelic deletion (∼460 Kb), including the TRAF3 locus in one of 22 patients. Genome sequencing identified a missense mutation (D483N) in the remaining allele. This substitution affects a highly conserved amino acid throughout vertebrate evolution, suggesting its functional relevance. We used cIgM-FISH to screen 27 patients with no available aCGH data and we found one biallelic TRAF3 deletion and one chromosome 14 monosomy. In the later case, no DNA was available to sequence the remaining allele. Both samples with biallelic inactivation of TRAF3 possess the two highest NFKB index values of our WM cohort, suggesting NF-κB transcription activity. Moreover, the case with biallelic deletion showed the lowest level of TRAF3 gene expression. Additionally, the entire TRAF3 coding region was sequenced in 28 of these 49 patients and one homozygous nonsense mutation (K286X) was identified. No GEP was available in this sample to calculate the NF-κB index. TRAF3 was only recently implicated in a malignant process, when inactivating mutations and biallelic deletions were reported in approximately 13% of MM patients, making it the most frequently inactivated tumor suppressor in MM. Our study has identified biallelic TRAF3 inactivation in at least 3 of 49 WM patients, suggesting that the constitutive activation of NF-κB signaling pathways may be involved in the pathogenesis of WM. Recent findings showing an association between TRAF3 loss of function and good response to bortezomib in MM patients suggest that proteasome inhibitor-based therapy may be an attractive option in the treatment of the WM patients with constitutive activation of NF-κB signaling pathways. Additional mutational screening of TRAF3 and other regulators of these pathways are ongoing to determine the extent of constitutive activation present in WM and other B-cell malignancies.

Description: The success of proteasome inhibition in multiple myeloma highlights the critical role for the ubiquitin-proteasome system (UPS) in this disease. However, there has been little progress in finding more specific targets within the UPS involved in myeloma pathogenesis. In an unbiased screen of de-ubiquitinase activity, we previously found the ubiquitin hydrolase UCH-L1 to be frequently over-expressed in B-cell malignancies, including myeloma. Using a transgenic mouse model, we also found that aberrant expression of UCH-L1 is sufficient to drive the development of spontaneous B-cell lymphomas and plasmacytomas, demonstrating its oncogenic activity. Whether and how UCH-L1 affects the clinical behavior of myeloma, however, is not known. Here we show that UCH-L1 is a potent negative prognostic factor that is essential for the dissemination and progression of myeloma. We found high levels of UCHL1 to predict early progression in newly diagnosed patients; a finding reversed in studies including bortezomib. We also found high UCHL1 levels to be a critical factor in the superiority of bortezomib over high-dose dexamethasone in relapsed patients. High UCHL1 partially overlaps with, but is distinct from, known genetic risks including 4p16 rearrangement and 1q21 amplification. Using an orthotopic mouse model, we found UCH-L1 depletion delays myeloma dissemination and causes regression of established disease. We conclude that UCH-L1 is a biomarker of aggressive myeloma that may be an important marker of bortezomib response, and may itself be an effective target in disseminated disease. Disclosures Galardy: Mission Therapeutics: Research Funding.

Description: Background: IMiDs are now recognized to promote the proteasomal degradation of IKZF1/3 and the transcriptional repression of MYC and IRF4. MYC locus rearrangement is a recurrent somatic event in MM and results in MYC repositioning near superenhancers (such as IGH@, IGL@ and others). The mechanisms of IMiDs-mediated downregulation of MYC in MM cells is not well understood, in particular it is unclear how IMiDs alter the activity of the super-enhancers driving MYC transcription. Methods and results: Transcriptome analysis (RNAseq) of MM cells (OPM2 and MM1S) treated with lenalidomide or upon the silencing of IKZF3 (lentiviral delivery of dox-inducible IKZF3 shRNA) confirmed the downregulation of MYC, IRF4 and among others the upregulation of other genes of interest such as CD38, DKK1, PDL1, FOXO3 and a host of genes involved in interferon response. Gene set enrichment analysis (GSEA) confirmed the enrichment of genes signatures associated with MYC as well as the interferon response. Notably, in t(4;14) MM cell lines we observed a significant downregulation of FGFR3 while the expression of WHSC1 was not affected. This finding was confirmed in a library of t(4;14) MM cell lines by qRT-PCR analysis. As FGFR3 is driven by the 3' IGH enhancer while WHSC1 is under the control of the intronic Em enhancer in t(4;14) MM cells, and in view of the known role of IKZF1/3 in class switch recombination, we postulated that IKZF1/3 regulate the 3' IGH enhancer activity and MYC expression in cells harbouring a MYC-IGH rearrangement. To validate this hypothesis we established the genome-wide distribution of IKZF1 in OPM2 and MM1S cells (both cell lines harbour an IGH-MYC rearrangement) by ChIP-Seq. Ikaros bound to 17660 loci of which 43% were associated with gene targets. Ikaros-binding motifs and other motifs such as STAT1, E2A, RUNX1 were also identified (MEME Tomtom and Jasper motif analysis) in the vicinity of Ikaros enrichment peaks. Importantly IKZF1 peaks also mapped to the IGH 3' enhancer loci and these results were confirmed by ChIP-PCR analysis. Of interest while no Ikaros peaks mapped to MYC (within 5 kb of TSS) by ChIP-seq, modest enrichment of Ikaros at MYC promoter was identified by ChIP-PCR in MM1S and KMS11 cells, and this enrichment was significantly attenuated by lenalidomide treatment. Since inhibition of MYC occurs as a consequence of depletion of the acetylated chromatin reader BRD4 at enhancers that drive MYC expression (Delmore et al., 2011), we examined the interaction between BRD4, IKZF1 and IMiDs treatment. Chip-Seq analysis revealed that BRD4 and IKZF1 are associated with most active enhancers and promoters in MM1S tumor cells. In particular, BRD4 and IKZF1 were enriched at the IGH 3'α2 enhancer locus. Importantly, treatment with lenalidomide (10μM, 4 hours), dramatically depleted IKZF1 and BRD4 enrichment at the 3' IGH enhancers (ChIP-PCR analysis). In contrast, treatment with the bromodomain inhibitor JQ1 depleted BRD4, but not IKZF1 from the IGH 3' enhancer locus. These findings suggest that IKZF1 regulates BRD4 binding to the 3' IGH enhancers and explain the selective effect of IMiDs on MYC transcription in transformed cells harboring a MYC rearrangement. In addition, these results suggest that IMiDs will affect MYC expression only in cells were MYC is driven by an IKZF1/IKZF3 responsive enhancer. Lastly we sought to explain how IKZF1 may regulate BRD4 enrichment at H3K27Ac marks in super-enhancer loci. Since Ikaros is a known integral component of the NuRD (nucleosome remodelling and histone deacetylase) complex, we examined whether lenalidomide treatment and the ensuing Ikaros depletion lead to local gain of NuRD at MYC TSS and depletion of H3K27 acetylation. CHD4 (helicase subunit of the NURD complex) and H3K27Ac ChIP-PCR analysis revealed significant accumulation of the NuRD complex at MYC TSS as well as reduced H3K27Ac at 3'IGH enhancers after IMiDs exposure in MM1S and OPM2 cells. Importantly, CHD4 silencing abrogated lenalidomide induced MYC transcriptional repression and cell death. Conclusion: In summary, our studies suggest that Ikaros drives MYC expression in MM cells harbouring an IGH-MYC rearrangement by regulating the accumulation of the acetylated chromatin reader BRD4 at the IGH 3' enhancer and preventing the NURD complex mediated depletion of H3K27Ac marks at these enhancers. Our findings also explain the IKZF1/3 dependent mechanism of IMiDs-mediated MYC transcriptional repression. Disclosures Neri: Celgene: Research Funding. Duggan:Jansen: Honoraria; Celgene: Honoraria. Keats:Translational Genomic Research Institute: Employment. Bahlis:Celgene: Consultancy, Honoraria, Research Funding, Speakers Bureau; Johnson & Johnson: Consultancy; Johnson & Johnson: Speakers Bureau; Johnson & Johnson: Research Funding; Amgen: Consultancy.

Description: It has been suggested that errors in the repair of DNA double strand breaks (DSB) can result in gross chromosomal rearrangements (GCR), including chromosomal amplifications, deletions, inversions, and translocations. To study repair of DNA DSB in vivo, we established a subclone of the U937 cell line designated F5, in which a single copy of the rare-cutting meganuclease I-SceI recognition site has been integrated into chromosome 7. We transfected an I-SceI expression vector into these cells and analyzed misrepair events. We recovered no GCR, but instead found that most DSBs were repaired by small (

Description: Abstract 1807 We have recently demonstrated that cereblon (CRBN) mediates the direct anti-myeloma activity of immunomodulatory drugs (IMiDs). However, the genes/pathways downstream of CRBN associated with anti-myeloma activity remain unclear. We, and others, identified interferon regulatory factor 4 (IRF4) as one of the downstream targets of CRBN-associated signaling. Both lenalidomide treatment and CRBN knockdown downregulate IRF4. IRF4 levels return to baseline in IMiD resistant cells surviving CRBN silencing. To determine whether IMiD-induced IRF4 downregulation is critical to anti-MM activity, we overexpressed IRF4 in two IMiD-sensitive human MM cell lines (HMCLs), KMS11 and MM1.S, followed by lenalidomide treatment. Lenalidomide-induced cytotoxicity was greatly impaired in both HMCLs overexpressing IRF4 compared with the control virus infected cells. Further analysis indicated that IRF4 over-expression does not completely prevent lenalidomide-induced growth arrest, but reduces cell death by 70% after lenalidomide treatment. Immunoblotting analysis of KMS11 cells indicated that IRF4 over-expression blocks lenalidomide-induced activation of caspase 8, reduces up-regulation of p21waf and increases CDK6 expression but does not significantly affect lenalidomide-induced MYC down-regulation. Although cereblon and IRF4 are broadly expressed in MM, baseline levels of expression are only weakly correlated (r=0.22) in primary MM patient gene expression analysis. Gene expression studies revealed statistical changes in 1,368 genes when comparing high versus low CRBN expression in primary myeloma samples. Interestingly genes associated with high CRBN expression included cyclin D2, SOCS3 and IL4 while genes associated with low cereblon expression included cyclin D1, FRZB and CD200. In order to understand how CRBN is connected with downstream anti-myeloma signaling, a structure-function study was performed to determine which CRBN domain is required for lenalidomide-induced IRF4 down-regulation and cytotoxicity. Lentiviral constructs expressing wild-type CRBN and a series of mutated CRBN were generated, including mutations at thalidomide binding site (Y384A/W386A), deletion of DDB1 binding region (ΔMid) and truncations at N-terminal and C-terminal. Lentiviruses from these constructs were used to infect IMiDs resistant HMCLs, OCI-MY5 and MM1.S res. Both of these cell lines have very low endogenous CRBN expression and they became sensitive to lenalidomide after introduction of wild-type CRBN. Conversely, introduction of CRBN with mutated thalidomide-binding site or with DDB1 binding region depletion failed to mediate lenalidomide toxicity and down-regulation of IRF4. OCI-MY5 cells expressing either N-terminal or C-terminal truncated CRBN showed substantial reduced responses (more than 50%) to lenalidomide compared with wild-type CRBN expressing cells. Deletion of only 20–30 amino acids at either ends of CRBN greatly impaired the protein function, suggesting that protein folding might be important for CRBN-mediated IMiD response. Our data indicate that IMiD induced myeloma cytotoxicity is largely mediated by modifying CRBN associated E3 ubiqutin ligase and subsequent IRF4 downregulation, suggesting the CRBN-IRF4 axis is a potential target for development of new anti-myeloma drugs. Disclosures: Schmidt: Karyopharm: Research Funding. Stewart:Millenium: Consultancy, Honoraria, Research Funding; Onyx: Consultancy; Celgene: Consultancy.

Description: Abstract 1846 Background The low frequency of TP53 (p53) alterations (20g/l on SPEP) used for in-vivo drug combination studies. Results Preliminary results support our principle hypothesis that, a PARP inhibitor (olaparib) enhanced Melphalan sensitivity in wild type p53 MM cell lines H929 and MM1S but not in p53 mutated or homozygously deleted cell lines U266 and KMS11, respectively. Moreover, we demonstrate that p53 knock-down decreased the synergistic effect of combining a PARP inhibitor with melphalan and Nutlin-3a. Again, combining a PARP inhibitor with the DNA alkylating agent melphalan induced clear PARP cleavage, a signature of apoptosis in wild type p53 MM cell lines H929 and MM1S. No significant PARP cleavage was observed after melphalan or olaparib treatment alone in H929 and MMIS. As expected, the p53 mutant cell line U266 did not show any PARP cleavage and p21 up-regulation after both drug treatments. Here we propose that the hyper-activated p53 from PARP inhibition along with DNA alkylating agents (melphalan) and/or p53 activating agents (nutlin-3a) result in better responses. Finally, in a murine xenograft model of human MM, olaparib potentiated melphalan activity in vivo, with significant reduction in M spike level. Conclusion Our studies indicate that DNA alkylating agents activate wild type p53, but leads to DNA repair and cell survival and this can be abolished by PARP inhibition in wild type p53 expressing MM cases. Moreover, our study also found that olaparib, and ABT-888 - PARP1 inhibitors currently in clinical trials - display different dose responses in MM cell lines and in the abilities of individual PARP inhibitors to sensitize MM cell lines to melphalan varied to a great extent in a cell context- and cell line specific manner. Disclosures: No relevant conflicts of interest to declare.