Implications of TP53 alterations for Therapy Response in Multiple Myeloma

Abstract Deletion of the tumor suppressor gene TP53is significantly associated with an unfavorable clinical course of Multiple Myeloma (MM). In addition, point mutations that abrogate p53 function similarly shorten survival. Most recently 'double hit', bi-allelic TP53inactivated MM was identified as an ultimate high risk feature of MM, affecting 3.7% of newly diagnosed MM patients (NDMM, Walker et al., Leukemia 2018). For TP53genetic analysis we combined data from two targeted sequencing M3P cohorts with targeted sequencing and FISH data, one of NDMM (n=142, Kortüm et al. Blood Cancer J. 2016), and one of multirefractory patients (rMM, n=40; Kortüm et al. Blood 2016). We also included two independent cohorts with paired NDMM and rMM, one with mutational data (n=43; Corre J et al 2017) and one with paired FISH analysis (Merz M et al. Haematologica 2017). We confirmed an increase of mutations in TP53 (8% NDMM / 16.9% rrMM), as well as del17p (12.6% / 22%). Similarly, mono- (12.7% / 22.5%) and bi-allelic events (5.6% / 17.5%) both demonstrated a significant increase over time (Figure, top). Importantly we observed deletion after mutation and vice versa in our cohorts. Next we established an AMO-1 MM cell line model (TP53+/+) mimicking mono and bi-allelic TP53-inativation. After CRISPR/Cas9-mediated TP53destruction, we introduced a modified Sleeping Beauty (SB) vector with two separate expression cassettes (p53 wt / wt and mutant p53 (R282W or R175H). Functionality of the p53 system was confirmed using nutlin-3, as inhibitor of MDM2-p53 interaction, as described elsewhere. Results: Doxorubicin and Melphalan are commonly used compounds in the treatment of MM. The IC50 of Melphalan in AMO1 naïve cells was 5µM and 5.5 µM after the reintroduction of 2 copies of wt TP53 in the KO model, with cell viability at 10µM of 30% and 31% respectively, measured by AlamarBlue Assay. Strikingly, we observed significant resistance induction in our hemizygous systems (del/wt p53; cell viability at 10µM 60% and mut/wt (62%). This furthermore increased within our homozygous models (TP53 del/del (85%); TP53 mut/del 80%) (Figure, bottom). Similar results were observed under doxorubicin treatment. Remarkably, this effect was absent against proteasome inhibition. Conclusions Here we present first evidence of TP53 inactivation impacting drug response to Melphalan and Doxorubicin, which might lead to the clonal selection of MM subclones harboring increased risk. The fact, that response to proteasome inhibition was not affected in our model might, at least in part, might explain their ability to confine high risk in MM. Figure. Figure. Disclosures No relevant conflicts of interest to declare.

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Molecular Resistance Mechanisms in Multiple Myeloma

Blood ◽

10.1182/blood-2018-99-118700 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 471-471

Author(s):

Santiago Barrio Garcia ◽

Umair Munawar ◽

Thorsten Stuehmer ◽

Hermann Einsele ◽

K. Martin Kortüm

Keyword(s):

Multiple Myeloma ◽

High Risk ◽

Disease Progression ◽

Tp53 Mutation ◽

Conflicts Of Interest ◽

Clonal Selection ◽

Tumor Therapy ◽

Genetic Alterations ◽

Detection Range

Abstract Mechanisms of drug resistance in Multiple Myeloma (MM) are poorly understood. Mutations and/or changes in the protein expression of the CRBN pathway and proteasome subunits have been identified to induce resistance to IMiDs and PIs. However, only few patients are affected by these alterations. To determine the specific genomic fingerprint of MM relapse we selected 57 MM patients from the CoMMpass trial (version IA11) that have genomic data of paired samples available (diagnosis/relapse). 35 of them have also sequential FISH-seq data. We focused on acquired mutations in first relapse and filtered all mutations and genetic alterations already present at diagnosis. Doing so, we found 1.274 mutations, representing an average of 23 new mutations/patient (range; 2-76). Of interest, 66% of the acquired mutations were present in a sub-clonal level (Variant read frequency (VRF) < 25%). Most common mutations include known hotspots of the RAS pathway (NRAS 12%, KRAS 7% and BRAF 4%). Notably, all 7 NRAS mutations in relapse were located at Q61K, suggesting a functional role of disease progression for this specific and known hotspot location. In total 5 of 35 cases (14%) with FISH-seq data developed a 17p13 deletion in relapse. Of these, three patients acquired a bi-allelic alteration in addition to a preexisting TP53 mutation and one developed a biallelic inactivation of TP53 (VRF = 100%), through parallel acquisition of del17p and TP53 mutation. Gain of 1q21 was observed in relapse in 5 of 35 (14%) cases, and one 1q gain was lost from diagnosis to relapse. Two cases (4%) presented mutations in IMiD treatment related genes, with two mutations in the CRBN pathway. One harbored a missense mutation in the Lenalidomide (LEN) degron sequence of IKZF3 (G159A) (VRF = 36%), known to be essential for the IMiD action in vitro, 45 months after continuous exposition to LEN . The other case presented two subclonal frameshift mutations in CUL4B (VRF = 5% and 32%), detected after more than three years of LEN containing therapy. We functionally validated in vitro LEN resistance through CRISPR/Cas9 knockout of CUL4B, suggesting a resistance inducing effect of the acquired CUL4B mutations. Six cases (11%) harbored acquired mutations in proteasome subunit genes (PSMC2, PSMC6, PSMD8, PSME4, PSMB9 (two mutations)), all of them had undergone prior proteasome inhibitor (PI) containing therapy. We validated earlier the 19S protein subunits PSMC6 and PSMC2 (KO and/or point mutations) as inducers of PI resistance in vitro, thus we hypothesize contribution to resistance induction / disease progression through these 19s mutations. Remarkably ubiquitin (E3, E2 and SUBs) and histone related genes (histones and histone methylases and deacetylases) were found mutated in 51% and 19% of the relapsed patients. Genes for drug transporters (ATP-binding cassette (ABC) and Solute Carrier (SLC) transporters) were hit in 32% of cases and genes for mucins (previously related with genotoxic agents and immunotherapy resistance) in 19%. Notably, RRBP1 presented 10 mutations in 6 patients (11%) with the mutations clustering within 30 amino-acids (aa) of exons 9 and 10 and 3 hotspots (2 patients each) in aa Q426P, K430R and Q436P. RRBP1 is involved in the binding of the ribosome to the endoplasmic reticulum (ER) and is related with the unfolded protein response and ER stress via GRP78. All the patients with RRBP1 mutations were pretreated with PI inhibitors and exhibited worst survival outcome affecting PFS (Pval<0.001) and OS (Pval=0.0016) in this limited dataset. The mutations were detected on average 433 days (range: 258-568) after diagnosis. Five of the 6 patients died on average 180 days after RRBP1 mutation detection (range: 18-446) further suggesting high risk features of such acquired mutations. In summary, we observe clonal selection of known high-risk related alterations like TP53 mutations, 17p deletions or 1q13 in early relapse data of the CoMMpass trial. Furthermore we identify RRBP1 mutations as a new acquired high-risk biomarker of MM. Alterations are specifically related to subclonal selection by therapy, thus we suggest that the definition of high-risk disease in MM needs to be revisited and should also include clonal selection processes under anti-tumor therapy. Figure. Figure. Disclosures No relevant conflicts of interest to declare.

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Cohort Analysis of FISH Testing of CD138+ Cells in Relapsed Multiple Myeloma: Implications for Prognosis and Choice of Therapy

Blood ◽

10.1182/blood.v124.21.3399.3399 ◽

2014 ◽

Vol 124 (21) ◽

pp. 3399-3399

Author(s):

Dean Smith ◽

Clemency Stephenson ◽

Anna Lach ◽

Steve Chatters ◽

Helena Kempski ◽

...

Keyword(s):

Multiple Myeloma ◽

High Risk ◽

Conflicts Of Interest ◽

Risk Group ◽

Cohort Analysis ◽

Fish Analysis ◽

Newly Diagnosed ◽

Interphase Fish ◽

Line Of Therapy ◽

Standard Risk

Abstract Introduction: Interphase FISH on CD138-selected bone marrow cells enables genetic risk stratification in newly diagnosed multiple myeloma (MM), however as MM remains incurable, most centres still treat newly diagnosed MM uniformly, utilising the most active regimens available. At relapse an increasing choice of regimens, coupled with co-morbidities and treatment-emergent toxicities, means no uniform approach is possible. Instead, therapy is tailored to disease and patient related risk factors. In this setting, FISH testing may be particularly useful if not done at diagnosis and to identify progression events that may alter prognosis. Aim: To evaluate the outcome of FISH analysis in consecutive patients with relapsed MM undertaken at our centre: success rate, frequency of abnormalities, incidence of progression events and correlation of FISH abnormalities with treatment outcomes. Methods: FISH analysis was performed on 192 samples from 154 relapsed patients (2012-13). Plasma cells were selected using magnetic CD138 MicroBeads and interphase FISH carried out using probes as recommended by the EMN (Ross et al, 2012). If patients had no prior results, a full FISH MM panel was performed, using probes for t(4;14), t(14;16), t(11;14), deletion 17p (17p-), Chr 1 abnormalities (1p-/1q+) and deletion 13q (13q-). If patients had been previously tested for an IgH translocation (Tx), a progression event panel was used: 1p-/1q+, 17p- and 13q-. Patients underwent FISH testing prior to starting the next line of therapy. Results: 79% of samples were successfully analysed, with analysis limited in 16% and failed in 5%. Common reasons for failure were poor quality/aged slides, insufficient material and poor hybridisation. 17% of patients had no cytogenetic abnormality. The most common abnormality was 13q- (43.1%), followed by 1q+ (41.4%), t(11;14) (18.3%), t(4;14) (12.4%), 17p- (12.0%) 1p- (8.9%), and t(14;16) (5.6%) Progression events were more common in t(14;16) and t(4;14) groups. All patients with t(14;16) and 82% with t(4;14) had an additional genetic lesion. Only 21% of patients with t(11;14) and 54% with no IgH Tx had an additional event. 80 patients (51.3%) had prior FISH results and 13 (16.3%) had developed a new abnormality on the later test. In 9 cases the progression event was 17p-, in 2 it was 1q+ and 2 cases developed 17p- and 1q+. The patients developing 1q+ were previously standard risk, so repeat testing altered risk group. Acquisition of 17p- indicates especially poor outcome, thus in all 13 cases repeat FISH analysis altered risk. Among patients with progression events none harboured t(11;14), 8 (64%) had no IgH Tx, 3 had t(14;16) and 2 had t(4;14). FISH results were correlated with clinical outcome. Patients were stratified as having high risk genetics [t(4;14), t(14,16), 17p- in ≥50% cells, 1p-/1q+] or standard risk [t(11;14), normal cytogenetics]. 63 (41%) patients were high risk, 83 (54%) standard risk, with no information available for 8 (5%). Both groups had received a median of 2 prior lines of therapy. Response rates (≥PR) to the next line of therapy were similar (60.4% standard risk vs 56.0% high risk). PFS from time of FISH was significantly longer in the standard risk group (9.8 months vs 5.9, p<0.01) as was OS (not reached vs 17.1 months, p<0.01, Fig. 1). In the high risk group, PFS was significantly longer in patients receiving a proteasome inhibitor (PI) as the next line of treatment versus those receiving other therapies (9.6 months vs 4.6, p=0.01) as was OS (not reached vs 9.7 months, p<0.01, Fig. 2). In the standard risk group, PFS was similar if patients received PI or not (9.5 months PI vs 9.8 other) as was OS (not reached both groups, Fig. 2). Conclusions: FISH analysis on MM patients at relapse was achievable. 74/154 patients had no prior results and a further 13 developed new poor prognostic markers, thus FISH at relapse provided new information in 56% of patients. Progression events were more common in patients harbouring t(4;14) or t(14;16). FISH at relapse was prognostic with high risk abnormalities associated with significantly shorter PFS and OS. The use of PI appeared to abrogate this poor prognosis, suggesting FISH at relapse could be a predictive and prognostic marker. Given the availability of second generation PI and the option of bortezomib re-treatment, results of FISH testing at relapse could directly influence clinical practice. Figure 1 Figure 1. Figure 2 Figure 2. Disclosures No relevant conflicts of interest to declare.

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Sars-Cov-2 Antibody Decay Monitoring in mRNA Vaccinated Multiple Myeloma Patients

Blood ◽

10.1182/blood-2021-154287 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 2712-2712

Author(s):

Samuel D. Stampfer ◽

Marissa-Skye Goldwater ◽

Bernard Sean S. Regidor ◽

Scott Kristian Jew ◽

Sean Elliott Bujarski ◽

...

Keyword(s):

Multiple Myeloma ◽

High Risk ◽

Healthy Subjects ◽

Conflicts Of Interest ◽

Serum Antibody ◽

Phase Iii ◽

Healthy Controls ◽

Post Vaccination ◽

Increased Risk ◽

Antibody Levels

Abstract Introduction: Amid the current COVID-19 pandemic, the highest mortality rates are among the elderly and immunocompromised. Multiple myeloma (MM) patients are immunocompromised and often are elderly. Not only do MM patients develop more frequent infections, but it is one of the leading causes of death for these patients. This population develops more severe COVID-19 due to various mechanisms that impair their ability to fight infection. This also reduces their ability to generate immunity from vaccination, as has been demonstrated by their diminished responses to vaccines for various respiratory illnesses. It follows that while phase III trial results for mRNA1273 and BNT162b2 COVID-19 vaccines showed an efficacy of 94-95% against even mild infection with this virus, the efficacy has been shown to be lower among MM patients. We recently evaluated patients' antibody responses to vaccination for COVID-19 by measuring anti-spike IgG levels in patient serum from before vaccination (baseline) and two weeks after dose 2 (D2W2) of vaccination with mRNA-1273 or BNT162b2, and found that most MM patients have impaired responses (Stampfer et al., Leukemia 2021). Patients who received mRNA-1273 vaccine had higher antibody levels than those who were vaccinated with BNT162b2, and specific clinical and myeloma-related characteristics predicted vaccine responsiveness. In the current study, we are monitoring anti-spike IgG antibody levels at Dose 2 Week 8 (D2W8) and Dose 2 Week 16 (D2W16) post-mRNA vaccination among these patients and in age-matched healthy controls to investigate antibody decay. Methods: Participants in the trial included MM patients (n=91) at the Berenson Cancer Center, and age-matched healthy controls (n=27). Healthy subjects were not known to be immunocompromised or currently receiving immunosuppressive therapy. Vaccination was done outside of the clinic and subjects provided copies of their CDC-issued COVID-19 vaccination cards to confirm dosing dates. Sera from vaccinated individuals were drawn at baseline (0-60 days prior to first vaccine dose) and at intervals following their second dose (14-21, 56-70, and 112-126 days post-vaccination). Background levels were determined from the clinic's serum bank of healthy subjects drawn pre-April 2019. Using an ELISA-based assay that detects IgG antibodies to SARS-CoV-2 spike protein, we determined Anti-SARS-CoV-2 spike ectodomain serum antibody levels and quantified them in IU/mL based on the WHO International Standard 20/136. Results: We analyzed the patient and control populations, and specifically for those classified as responders from our original published study (D2W2 >50 IU/mL). All controls but only 70% of patients (64/91) were classified as responders, so this responder-specific analysis was only relevant for patients. There was a significant decline in anti-SARS-CoV-2 spike antibodies from D2W2 to D2W8 for both patients and controls; D2W16 results are pending. Median values dropped from 283.1 IU/mL to 90.9 IU/mL among patients, and 893.6 IU/mL to 354.4 IU/mL among controls. Median antibody levels among patients classified as responders dropped from 482.9 IU/mL to 145.5 IU/mL (p <0.0001). We also found that the D2W8 value for patients was significantly lower than that of controls (90.9 IU/mL vs 354.4 IU/mL, p<0.0001), as well as among the responder group (145.5 IU/mL vs 354.4 IU/mL, p=0.0005). A cutoff of <147 IU/mL has previously been associated with an increased risk for breakthrough infections. Spike antibody levels of 50% of patients who responded and 64.8% of all patients were <147 IU/mL at D2W8, whereas only 7.4% of controls were <147 IU/mL (p<0.0001). Conclusions: The markedly lower anti-spike antibody levels among MM patients compared to healthy controls at week 8 post-vaccination indicate that they remain at high risk for breakthrough infections. Combined with their rapid decline in anti-spike antibody levels over only a 6-week period, this indicates that even MM patients who responded initially to vaccination are likely to require boosters sooner than healthy individuals. This immunocompromised population remains at high risk even following vaccination and should continue to maintain social distancing precautions during periods of high local SARS-CoV-2 transmission. Disclosures No relevant conflicts of interest to declare.

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Thrombogenic activity of doxorubicin in myeloma patients receiving thalidomide: implications for therapy

Blood ◽

10.1182/blood-2002-01-0335 ◽

2002 ◽

Vol 100 (4) ◽

pp. 1168-1171 ◽

Cited By ~ 183

Author(s):

Maurizio Zangari ◽

Eric Siegel ◽

Bart Barlogie ◽

Elias Anaissie ◽

Fariba Saghafifar ◽

...

Keyword(s):

Multiple Myeloma ◽

Multivariate Analysis ◽

High Risk ◽

Doppler Ultrasonography ◽

Standard Dose ◽

Single Agent ◽

Cytotoxic Chemotherapy ◽

Newly Diagnosed ◽

Statistical Association ◽

Increased Risk

Ten percent of newly diagnosed myeloma patients treated with any type of chemotherapy develop deep venous thrombosis (DVT). Thalidomide has proven activity in refractory multiple myeloma (MM), and although single-agent thalidomide has minimal prothrombogenic activity, its combination with cytotoxic chemotherapy is associated with a significantly increased risk of DVT. We analyzed the incidence of DVT in 232 MM patients who received a combination of chemotherapy and thalidomide on 2 protocols that differed only by the inclusion of doxorubicin in one. DT-PACE (dexamethasone/thalidomide/cisplatin/doxorubicin/cyclophos- phanide/etoposide) was offered to patients with preceding standard dose therapy, but no prior autotransplantation, while DCEP-T (dexamethasone/cyclophosphamide/etoposide/cisplatin/thalidomide) was administered for relapse after transplantation. If there were signs or symptoms suggestive of DVT, patients received additional investigations, including Doppler ultrasonography, followed by venography if indicated. Only patients on DT-PACE but not DCEP-T experienced an increased incidence of DVT. A statistical association between the incidence of DVT and combination chemotherapy including doxorubicin (P = .02) was observed; this association was confirmed on multivariate analysis. MM patients treated with thalidomide and doxorubicin have a high risk of developing DVT.

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Military Service in Vietnam/Korea and Serum Dioxin Levels Do Not Affect the Outcomes of Patients Diagnosed with Plasma Cell Dyscrasias

Blood ◽

10.1182/blood.v118.21.5098.5098 ◽

2011 ◽

Vol 118 (21) ◽

pp. 5098-5098

Author(s):

Prasanth Ganesan ◽

Emil Kuriakose ◽

Carla Smith ◽

Robert T Harris ◽

Jonathan E Dowell ◽

...

Keyword(s):

Multiple Myeloma ◽

Plasma Cell ◽

Military Service ◽

Conflicts Of Interest ◽

Military Veterans ◽

Cell Transplant ◽

Increased Risk ◽

Plasma Cell Dyscrasias ◽

Chlorinated Dioxins ◽

The Impact

Abstract Abstract 5098 Military Service in Vietnam/Korea and Serum Dioxin Levels Do Not Affect the Outcomes of Patients Diagnosed with Plasma Cell Dyscrasias. Background: Exposure to dioxin, a contaminant found in herbicides has been associated with increased risk of cancers including multiple myeloma and postulated to cause poorer survival in the exposed population. Military personnel, especially those who had served in Vietnam and Korea have an increased risk of dioxin (which contaminated the herbicide Agent Orange which was sprayed during these wars) exposure. We looked at the impact of dioxin exposure and blood levels of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) which is the most toxic of the poly-chlorinated dioxins on the survival outcomes of military veterans diagnosed with plasma cell dyscrasias (PCD). Methods: A prospective analysis of newly diagnosed and existing myeloma patients was done. Information regarding the patient and disease characteristics, the military record, and outcomes were collected. Approximately 60 ml of heparinised peripheral blood was collected and immediately frozen at −20 degrees. These samples were shipped to Eurofins Laboratory, Hamburg, Germany for dioxin level measurement. Patients' blood lipid levels were also measured and the dioxin toxic equivalent (Teq) was calculated. Overall survival (OS) was calculated from the date of diagnosis till death (Kaplan Meier method). Cox regression and log rank analysis were used to look for prognostic variables. Results: Fifty two (52) patients of PCD were available for analysis. Majority had a diagnosis of multiple myeloma. Forty one underwent treatment including stem cell transplant in 16 (Table 1 shows the patient characteristics, laboratory results and treatment outcomes). During a median follow up of 54 months (2–348), 21 patients died (progressive myeloma: 12(23%), cardiac failure: 3 (5.7%), infections: 1 (1.9%), acute myeloid leukemia: 1 (1.9%), pulmonary embolism: 1 (1.9%) and unknown: 3 (5.7%). The median OS was 111 mos (95% CI 56–155) and the estimated survival at 5 yrs was 69.5% (+/− SE 0.067). The 5 yr OS was negatively impacted by abnormal cytogenetics (40.3 % vs. 75.5%; p=0.012), and service in the army (non-army vs. army: 83% vs. 40%; p=0.032). Patients who had served in Vietnam had outcomes similar to others; Korean War veterans had a poorer OS, but this was not statistically significant (5 yr OS 68% vs. 48%; p=0.1). There was no association between TCDD levels or the Teq with OS. Abnormal cytogenetics was the only significant factor on multivariate analysis. Conclusions: We did not find an association between military service in Korea/Vietnam or serum dioxin levels and poor survival in military veterans diagnosed with Plasma cell dyscrasias. However, a study of a larger sample of myeloma patients with similar service and exposure histories maybe warranted. Disclosures: No relevant conflicts of interest to declare.

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Codon 72 Polymorphism Of TP53 Gene Is a Novel Prognostic Marker For Thalidomide Therapy In Multiple Myeloma

Blood ◽

10.1182/blood.v122.21.1891.1891 ◽

2013 ◽

Vol 122 (21) ◽

pp. 1891-1891

Author(s):

Yutaka Hattori ◽

Yurika Ikeda ◽

Yuya Suzuki ◽

Daiju Ichikawa ◽

Maiko Matsushita

Keyword(s):

Multiple Myeloma ◽

Overall Survival ◽

High Risk ◽

Prognostic Marker ◽

Mononuclear Cells ◽

Conflicts Of Interest ◽

Tp53 Gene ◽

Codon 72 ◽

Codon 72 Polymorphism ◽

Significant Difference

Abstract Backgrounds and purpose Recently, newly developed drugs such as thalidomide, lenalidomide and bortezomib have significantly improved survival of the patients with multiple myeloma (MM). However, certain group of the patients who have characteristic high-risk cytogenetic changes such as deletion of TP53 tumor suppressor gene revealed significantly shorter survival. In patients with deletion of TP53 gene, the somatic point mutation in the residual TP53 gene has been reported to be rare. Thus, the exact role of alteration of TP53 gene in high-risk myeloma remains unclear. Polymorphism of TP53 gene at codon 72 in exon 4, CGC to CCC transition, leads to arginine (Arg) to proline (Pro) amino-acid substitution. Biological analyses showed that the Arg variant more efficiently induces expression of P21 and apoptosis via Ser-6 and 20 phosphorylation of p53 protein. In contrast the Pro variant is less resistant to MDM-2-mediated degradation and more potently induced cell-cycle arrest and DNA damage repair. Recently, clinical significance of this polymorphism has been extensively studied in solid tumors as well as hematological malignancies including non-Hodgkin lymphoma and leukemia. However, consistent association of the polymorphism with cancer risk has not been elucidated. The purpose of this study is to examine codon 72 polymorphism in patients with refractory multiple myeloma (MM) treated with thalidomide, and to elucidate its association with myeloma risk and outcome of thalidomide-therapy. Patients & methods A total of 37 Japanese patients with refractory or relapsed MM who were treated with thalidomide monotherapy were included in this study. Three cases showed deletion of TP53 gene by fluorescence in situ hybridization (FISH) analyses. The codon 72 polymorphism was evaluated in the rest of 34 patients. Genomic DNA was isolated from bone marrow mononuclear cells. The genomic TP53 gene was amplified by polymerase chain reaction, and the amplified DNAs were directly sequenced. Results Direct DNA sequence showed that the Pro/Pro homozygote was observed in six patients (18%), Pro/Arg heterozygote in 12 (35%) and Arg/Arg homozygote in 16 (47%). There was no significant difference in the frequency of the polymorphism between the 34 Japanese MM patients and the healthy Japanese individuals (P=0.44). Thus, association of codon 72 polymorphism with myeloma risk has failed to be elucidated. The response rate to thalidomide therapy was 33% in the patients with Pro/Pro, 27% in Pro/Arg and 44% in Arg/Arg (P=0.49), respectively. Other clinical backgrounds including age, sex, Durie-Salomon stage, ISS stage, serum creatinine, albumin, b2M, calcium, hemoglobin levels and M-protein type were not correlated with TP53 codon 72 polymorphism, either. Progression free survival (PFS), overall survival (OS) and post-relapse survival were shown in Figure 1. The patients with Pro allele tended to show shorter PFS; 5 weeks for the patients with Pro/Pro versus 32 weeks for those with Pro/Arg plus Arg/Arg (P=0.07) (Figure 1). Overall survival (OS) of the patients with Pro/Pro, Pro/Arg and Arg/Arg allele was 18 weeks, 49 weeks and 133 weeks, respectively (P=0.027). Especially, the patients with Pro/Pro allele revealed significantly shorter OS compared with those with Pro/Arg plus Arg/Arg (18 weeks versus 100 weeks, P=0.023) (Figure 1). Significant difference of OS in patients with Pro/Pro+Pro/Arg vs Arg/Arg (P= 0.032) suggested the dominant effect of Pro allele for poorer prognosis. Post-relapse survival of the patients with Pro/Pro, Pro/Arg and Arg/Arg allele was 11, 42 and 64 weeks, respectively (P=0.054). Post-relapse survival for Pro/Pro versus Pro/Arg plus Arg/Arg were 11 weeks versus 64 weeks (P=0.029). Conclusion These results indicated that codon 72 polymorphism did not correlated with myeloma risk, but significantly associated with therapeutic outcome. Namely, the patients with Pro allele revealed earlier relapse and shorter post-relapse survival, resulted in shorter OS in thalidomide therapy. Further evaluation is needed to clarify whether the codon 72 polymorphism will be a new prognostic marker for the treatment with novel drugs. Disclosures: No relevant conflicts of interest to declare.

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Management Of Pregnancy In Patients With Antithrombin Deficiency

Blood ◽

10.1182/blood.v122.21.216.216 ◽

2013 ◽

Vol 122 (21) ◽

pp. 216-216

Author(s):

Mario von Depka ◽

Stefanie Döpke ◽

Anja Henkel-Klene ◽

Cornelia Wermes ◽

Mahnaz Ekhlasi-Hundrieser ◽

...

Keyword(s):

High Risk ◽

Pregnancy Loss ◽

Conflicts Of Interest ◽

Fetal Loss ◽

Bleeding Complications ◽

Thromboembolic Events ◽

Increased Risk ◽

History Of ◽

At Deficiency ◽

The Mean

Abstract Introduction During pregnancy women have a four- to five-fold increased risk of thromboembolism (TE) compared to women who are not pregnant. Among the most important risk factors for TE in pregnancy is the presence of thrombophilia. Multiple reports have described an association between antithrombin (AT) deficiency and an increased rate of thromboembolic events especially during pregnancy. As the placental development depends on well-balanced pro- and anticoagulant mechanisms, thrombophilia, e.g. AT deficiency may be associated with poor pregnancy outcome. Despite anticoagulation with low molecular weight heparin (LMH) during pregnancy and the postpartum period alone, women with AT deficiency are still at a high risk to develop TE, especially perinatal and during puerperium because of withheld anticoagulation to prevent bleeding complications. Therefore, several guidelines recommend the administration of antithrombin concentrates during high risk situations as pregnancy. Here, we present the results of our study on the usage of AT concentrates in pregnant women with AT deficiency who either suffered from fetal loss or thromboembolism prior inclusion. Methods In total, 22 pregnancies in 19 patients (age: 31.9±4.7; 22-41) with AT deficiency were included in this open-label, single-center study. Ten patients (53%) had a history of fetal loss, 9/19 (47%) patients hat a history of thromboembolism. During all pregnancies AT concentrate (AT-C) was administered, in 18/22 (81.8%) pregnancies LMH was given in addition. Prior pregnancy losses (21/30, 70%) occurred in all trimester (t1: n=11, t2: n=5, and in t3: n=5). Historical live birth rate (LBR) was 30%. Blood samples were collected in all trimesters and postpartum to analyze AT activity and antigen, endogenous thrombin potential (ETP), thrombin-antithrombin-complex (TAT), Fragment 1+2 (F1+2) and c-reactive protein test (CRP). A total of 114 uneventful pregnancies of 113 healthy women served as controls. Furthermore, the mean doses of AT concentrates/kg BW and the mean total number of infusions were calculated. Results In total, 21 pregnancies (95.5%) were successful. Mean total requirement of AT concentrate per pregnancy was 79.454 IU (range: 3.000-272.000 IU) during 27.8 treatment days per pregnancy (range: 1-88). Our data show an increase of F1+2 in the course of pregnancy. Mean levels of F1+2 at t1, t2 and t3 (t1= 255.9 ± 107.6, t2= 360.9 ± 117.4, t3= 545.3 ± 220.3 pmol/L) were significantly higher than in controls (t1= 82.2 ± 43, t2= 140 ± 100.2, t3= 183.5 ± 103.1, p<.001). Mean level of TAT was higher (3.1 ± 1.4 ng/mL) than in controls (1.7 ± 1.6 ng/mL, p=.001) in t1, whereas mean TAT in t2 and t3 was lower than in controls (3.8 ± 1.3 vs. 4.8 ± 1.9, p=.03; 5.0 ± 1.4 vs. 6.1 ± 3.0 ng/mL, n.s., resp.). No thromboembolic events occurred. In patients receiving AT-C, LBR increased from 30% to 95.5% (p<0.001) with a relative risk of 49.0 to develop pregnancy loss without anticoagulant treatment (5.7 – 421.8; 95% CI). Conclusion In patients with AT deficiency receiving AT concentrate and LMH we could demonstrate a significant increase of LBR from 30% to 95.5%. Furthermore, no thromboembolic events occurred, though almost half of the patients had a history of thromboembolism. There was no clear evidence of increased hypercoagulability. We conclude that combined AT concentrate and LMH are safe and efficacious for mother and child in preventing thromboembolism and pregnancy loss. Further studies to evaluate the exact mode of anticoagulation and benefit of combining AT concentrate and LMH are warranted. Disclosures: No relevant conflicts of interest to declare.

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The Frequency of Genetic Anomalies According to Clonal Plasma Cells' Phenotype in Patients with Newly Diagnosed (ND) Multiple Myeloma (MM)

Blood ◽

10.1182/blood.v126.23.5316.5316 ◽

2015 ◽

Vol 126 (23) ◽

pp. 5316-5316

Author(s):

Andrei Garifullin ◽

Irina Martynkevich ◽

Sergei Voloshin ◽

Alexei Kuvshinov ◽

Ludmila Martynenko ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Plasma Cells ◽

Conflicts Of Interest ◽

Risk Group ◽

Risk Groups ◽

Fish Analysis ◽

Newly Diagnosed ◽

Definition Of ◽

Standard Risk

Abstract Background. Genetic anomalies (GA) are primary link of pathogenesis in MM. GA lead to formation of clonal plasma cells, which has different phenotype. Aim. To estimate the incidence of GA and their correlation with clonal plasma cells' phenotype in patients with ND MM. Methods. We analysed 22 patients with ND MM (median age 57 years, range 38-80; male/female - 1:1.75). Cytogenetic analysis was performed on bone marrow samples using standard GTG-method. Metaphase FISH analysis was performed according to the manufacturer's protocol using DNA probes: LSI 13(RB1)13q14, IGH/CCND1, IGH/FGFR3, LSI TP53 (17q13.1). 8-color immunophenotypic by flow cytometry using antibody to CD45, CD38, CD138, CD56, CD19, CD20, CD27 and CD117 antigenes. Results. Translocation t(11;14) was detected in 3/14 (21.4%) patients, del(13q) - 2/14 (14.3%), t(11;14) - 3/14 (21.4%), hypodyploidy - 1/20 (5%), del(17р) - 0% patients. Clonal plasma cells' phenotype CD38+CD138+CD45- was detected in 100%. Expression CD56+ was revealed in 11/22 (50%) patients, CD19+ in 9/22 (40.9%), CD117+ in 5/22 (22.7%), CD20+ in 1/22 (4.5%), CD27+ in 1/22 (4.5%). The frequency of GA didn't depend on clonal plasma cells' phenotype and was 27.3%(3/11) in CD56+ phenotype, 23.8%(5/21) - CD20-, 23.8%(5/21) - CD27-, 23.5%(4/17) - CD117-, 23%(3/13) - CD19-, 22.2%(2/9) - CD19+, 20%(1/5) - CD117+, 18.2%(2/11) - CD56-, 0%(0/1) - CD20+, 0%(0/1) - in CD27+ phenotype. Patients of standard risk group according to mSMART 2.0 with GA had CD19-negative plasma cells' phenotype vs. CD19-positive phenotype in patients of intermediate and high-risk groups (p<0.05). 3-years overall survival in standard risk group with CD19- phenotype was 92,3%, CD19+ - 77,7% (p>0.05). Conclusion . Identification of GA, which has adverse forecast, correlates with CD19+ plasma cells phenotype. The combined definition of plasma cells phenotype and GA can improve the system of risk stratification in MM. Disclosures No relevant conflicts of interest to declare.

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Overuse of DVT Prophylaxis in Medical Inpatients

Blood ◽

10.1182/blood.v126.23.5563.5563 ◽

2015 ◽

Vol 126 (23) ◽

pp. 5563-5563 ◽

Cited By ~ 1

Author(s):

Jing Deng ◽

Lisa Thomas ◽

Huijing Li ◽

Elvin Varughesekutty ◽

Qi Shi ◽

...

Keyword(s):

Risk Assessment ◽

High Risk ◽

Conflicts Of Interest ◽

Medical Center ◽

Low Risk ◽

Risk Of Bleeding ◽

Dvt Prophylaxis ◽

Increased Risk ◽

Medical Inpatients ◽

Nonsurgical Patients

Abstract Introduction: Unfractionated heparin (UFH), or low-molecular-weight heparin (LMWH), is commonly used with mechanical prophylaxis as an anticoagulant to reduce the risk for venous thromboembolism (VTE). However, overuse of these prophylaxes can increase the risk of bleeding, heparin-induced thrombocytopenia (HIT) and associated medical cost. PURPOSE: The aim of this study is to determine the incidence of DVT prophylaxis among hospitalized nonsurgical patients in a community medical center. To evaluate the use of the prophylaxes as described above, the investigators collected data on medical inpatients and addressed how to avoid overuse. Method: A retrospective inpatient chart review of 100 general internal medicine patients analyzed data using Padua Prediction Score as the risk estimate for deep venous thrombosis (DVT). High risk for VTE was defined by a cumulative score >=4 and low risk was a score <4. Only patients at increased risk for DVT but not at high risk for bleeding qualified for heparin treatment. Results: A total of 100 patients were surveyed. 54/100 (54%) patients had low risk of DVT with score < 4, and of those 29/54 (53.7%) patients received DVT prophylaxis with SCDs and/or heparin, and 17/54 (31.5%) patients were treated with heparin. All 46 patients with score >= 4 were treated with DVT prophylaxis of which 10 patients were only treated with heparin and 36 patients were given both mechanical and chemical prophylaxis. Collectively, 53.7% of the patients received treatment with DVT prophylaxis (p < 0.001, Chi-Square test). Discussion: In hospital settings, physicians want to avoid DVT or PE so they tend to consider patients as being at moderate risk for DVT without using any method of DVT risk assessment. This leads to unnecessary overuse of DVT prophylaxis on patients and may increase the risk of bleeding and injury. Conclusion: Our data suggests that there DVT prophylaxis including UFH and LMWH was over prescribed among patients with who had marginal risk for DVT in hospitalized nonsurgical patients in a community medical center. Clinical implications: To avoid the overuse of DVT prophylaxis, physicians need to follow guidelines. Education and inclusion of the guidelines in EHRs of information on VTE risk assessment for hospitalized medical patients upon admission may reduce unneeded DVT prophylaxis and the risk of bleeding and costs associated with additional care needs. Disclosures No relevant conflicts of interest to declare.

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FISH Analysis in Multiple Myeloma - a Retrospective Study from India

Blood ◽

10.1182/blood.v128.22.5646.5646 ◽

2016 ◽

Vol 128 (22) ◽

pp. 5646-5646 ◽

Cited By ~ 1

Author(s):

Pratibha Dhiman ◽

Shalini Goel ◽

Priyanka Samal ◽

Nitin Sood ◽

Ritesh Sachdev ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

High Risk ◽

Chromosomal Aberrations ◽

Overall Response Rate ◽

Response Rate ◽

Indian Subcontinent ◽

Fish Analysis ◽

Interphase Fish

Abstract Introduction Multiple myeloma (MM) is a malignancy involving terminally differentiated plasma cells. It is characterized by a complex pattern of extensive genomic aberrations involving many chromosomes and it constitutes about 1% of all malignancies. Its exact incidence in India is not known. Based on data available from 6 population-based cancer registries in India (covering 0.3% of the population) its incidence varies from 0.3 to 1.9 per 100 000 for men and 0.4 to 1.3 per 100 000 for women. Among various prognostic markers in MM, cytogenetic abnormality detected by conventional cytogenetic and FISH studies are major factors deciding clinical outcome. Interphase FISH studies from various parts of the world have reported variable incidence (40 - 66%) of cytogenetic abberations. However, there is insufficient published data from Indian subcontinent addressing the frequency of chromosomal aberrations using interphase FISH in MM patients. Patients and Methods Sixty eight patients clinically diagnosed with multiple myeloma were studied. The retrospective period of recruitment was from Jan 2015 to June 2016. The diagnosis of MM was based on serum electrophoresis and immunofixation, bone marrow plasmacytosis, and end organ involvement. Interphase FISH analysis was performed on bone marrow samples using specific DNA probes- Del 13q14.3 (LSI D13S25) , t (4;14) ( Kreatech IgH/ FGFR3 DC-DF), t (11;14) (Zytovision directly labeled IgH/CCND1 DC-DF), t (14;16) (Kreatech IgH/ MAF DC-DF), Del 17p13.1( LSI TP53). A total of two hundred nuclei were enumerated for each FISH Panel probe and cut off for detection of deletion/ fusion signal in normal individuals was taken as 3%. An interim analysis of treatment protocols was also done. Results A total of 68 cases with MM were evaluated which included 55 males and 13 females. We report a median age of 58 years (37-86 years). Interphase FISH analysis was done in all patients. Out of sixty eight patients, 23 (33.82%) patients had one genetic abnormality. Results revealed that deletion 13q14.3 was the most frequent aberration. Out of 68 patients, 10 patients have 13q14 (14.7%) abnormality. This includes 70% males and 30% females. In addition absence of p53 at 17p13 was detected in 8/68 (11.8%) patients. Similarly 11q13 abnormality was observed in 3/68 (4.4%). IgH (14q32) aberrations were noted in 2/68 (2.94%) patients. Of which t(4;14) was detected in these patients, whereas none of them showed t(14;16). More than one chromosomal aberrations were present in 4 patients. Data for serum β2-microglobulin at the time of presentation could be evaluated in 52 patients only. Most of the patients 40(76.9%) belonged to ISS stage 3. From the available data, 4 patients with ISS stage 3 had high risk chromosomal abnormality whereas 3 patients with ISS stage 2 and none of the patient of ISS stage I disease had high risk chromosomal abnormality. A total of 40 patients received cyclophosphamide, bortezomib and dexamethasone as the primary treatment whereas 23 patients received Bortezomib, lenalidomide and dexamethasone based therapy. Post 2 cycles of cyclophosphamide based therapy showed an overall response rate (CR + VGPR) of 87.5% whereas in case of lenalidomide based therapy the overall response rate was 91%. Thirteen patients underwent autograft after durable response, out of which one had a clinical relapse within 3 months. Median survival can only be commented on further follow up. Conclusion In comparison to the west, the frequency of chromosomal aberrations are different and much less in India whereas the studies of median survival is comparable. An early age of presentation in Indian subcontinent is another issue to be addressed as we know that secondary mutations accumulate with increasing age, but a younger population presenting with same severity of disease needs exploration of additional abnormalities in India. Being a resource constraint country and non availability of molecular lab at every place, evaluation of each patient is difficult, however increasing awareness of the role of biology in the management of MM is inspiring the clinicians for detailed evaluation and close follow up of these patients. Certainly, larger trials are required to understand the biology of this disease in the country. Disclosures No relevant conflicts of interest to declare.

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