Adverse metaphase cytogenetics can be overcome by adding ... · Purpose. To determine whether a reduction in the intensity of Total Therapy (TT) reduces toxicity and maintains efficacy

Adverse metaphase cytogenetics can be overcome by adding bortezomib and thalidomide to fractionated melphalan transplants

Yogesh S Jethava1, Alan Mitchell2, Joshua Epstein1, Maurizio Zangari1, Shmuel Yaccoby1, Erming Tian1, Sarah Waheed1, Rashid Khan1, Xenofon Papanikolaou1, Monica Grazziutti1, Michele Cottler-Fox3, Nathan Petty1, Douglas Steward1, Susan Panozzo1, Clyde Bailey1, Antje Hoering2, John Crowley2, Jeffrey Sawyer3, Gareth Morgan1, Bart Barlogie1 and Frits van Rhee1

1 Myeloma Institute, University of Arkansas for Medical Sciences, Little Rock, AR; 2

Cancer Research And Biostatistics, Seattle, WA; 3 Department of Pathology, University of Arkansas for Medical Sciences.

Running title: Randomized auto-transplant trial for low-risk myeloma

Corresponding author:

Frits van Rhee, MD, PhD Myeloma Institute University of Arkansas for Medical Sciences 4301 W Markham, #816 Little Rock, AR 72205 USA Phone: 501-526-2873 Fax: 501-526-2273 Email: [email protected]

Key Point: Conditioning with melphalan, bortezomib and thalidomide confers benefit to myeloma with abnormal metaphase cytogenetics.

Abstract count: 246 words

Text Count: 2,675 words

References: 30

Conflict of Interest Disclosure: There are no relevant conflicts to disclose.

Grant Support:

Total Therapy 4 was supported by the National Cancer Institute, National Institutes of Health Program Project Grant CA55819 (to Dr. Gareth Morgan).

Research. on January 9, 2020. © 2016 American Association for Cancerclincancerres.aacrjournals.org Downloaded from

Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Author Manuscript Published OnlineFirst on November 3, 2016; DOI: 10.1158/1078-0432.CCR-15-2620

http://clincancerres.aacrjournals.org/

2

Translational Relevance

Melphalan 200mg/m2 is widely considered the standard preparatory regimenfor

autologous stem cell transplantation for myeloma. Attempts to improve conditioning with

melphalan have not been successful and included the addition of other cytotoxic agents,

skeletal targeting radioactive antibodies or radiation therapy. The present study

suggests that patients with abnormal metaphase cytogenetics, who traditionally fare

worse than their counterparts with normal metaphase cytogenetics, may benefit from

the addition of bortezomib, thalidomide and dexamethasone to a fractionated melphalan

(50mg/m2/d x 4d) conditioning regimen. Conversely, patients with a normal karyotype

are best served with preparation comprising melphalan 200mg/m2 given in a single

dose. These findings, if confirmed suggest that specific autologous stem cell

transplantation conditioning regimens can improve outcome in subgroups of myeloma

patients.

Abstract

Purpose. To determine whether a reduction in the intensity of Total Therapy (TT)

reduces toxicity and maintains efficacy.

Experimental design. 289 patients with gene expression profiling (GEP70)-

defined low-risk multiple myeloma (LRMM) were randomized between a standard arm

(TT4-S) and a light arm (TT4-L). TT4-L employed 1 instead of 2 inductions and

consolidations. To compensate for potential loss of efficacy of TT4-L, bortezomib and

thalidomide were added to fractionated melphalan 50mg/m2/d x 4.




3

Results. Grade ≥3 toxicities and treatment-related mortalities were not reduced

in TT4-L. Complete response (CR) rates were virtually identical (p=0.2; TT4-S, 59%:

TT4-L, 61% at 2 years), although CR duration was superior with TT4-S (p=0.05; TT4-S,

87%; TT4-L, 81% at 2 years. With a median follow-up of 4.5yr, there was no difference

in overall survival (OS) and progression-free survival (PFS). While metaphase

cytogenetic abnormalities (CA) tended to be an adverse feature in TT4-S, as with

predecessor TT trials, the reverse applied to TT4-L. Employing historical TT3a as

training and TT3b as test set, 51 gene probes (GEP51) significantly differentiated the

presence and absence of CA (q<0.0001), 7 of which function in DNA replication,

recombination, and repair. Applying the GEP51 model to clinical outcomes, OS and

PFS were significantly inferior with GEP51/CA in TT4-S; such difference was not

observed in TT4-L.

Conclusions. We identified a prognostic CA-linked GEP51 signature, the

adversity of which could be overcome by potentially synergizing anti-MM effects of

melphalan and bortezomib. These exploratory findings require confirmation in a

prospective randomized trial.

Introduction

With the incorporation of novel agents into auto-transplant-supported high-dose

melphalan trials, the outlook of patients with multiple myeloma (MM) has greatly

improved. In our Total Therapy (TT) program, applying all MM-active agents up-front in

order to reduce or even eliminate the development or survival of drug-resistant sub-

clones, major advances have been linked to the addition of thalidomide in TT2 and




4

bortezomib in TT3 (1, 2). However, when examined in the context of gene-expression

profiling (GEP) of CD138-purified plasma cells, such benefit was limited to the 85% of

patients presenting with GEP70-based low-risk MM (LR-MM) (3, 4). In recognition of the

lack of progress in high-risk MM (HR-MM), we decided in 2008 to assign low and high

risk patients to separate protocols. We are now reporting on clinical outcomes of 289

patients with LR-MM enrolled in a randomized phase III trial of Total Therapy 4 (TT4),

comparing a light arm (TT4-L) to a standard arm (TT4-S), with the goal of reducing

toxicity while maintaining efficacy in TT4-L.

Materials and Methods

The protocol schema is portrayed in supplementary table 1. Briefly, eligible

patients with GEP70-defined LR-MM were randomized between TT4-S and TT4-L. TT4

induction in both arms was similar to TT3b except for the addition of melphalan

10mg/m2 test-dosing 48hr following bortezomib 1.0mg/m2 test-dosing for the purpose of

pharmacogenomic investigations (5, 6). TT4-L differed from TT4-S by reduction from 2

cycles to 1 cycle each of induction with M-VTD-PACE prior to and consolidation with

dose-reduced VTD-PACE after tandem transplants. The transplant regimen in L-TT4

was altered from a single melphalan dose of 200mg/m2 (MEL200) in TT4-S to a

fractionated 50mg/m2/d x 4d (MEL50x4) schedule with the aim to avoid peak MEL dose

levels and reduce mucosal toxicity. To compensate for the potentially reduced efficacy

of this strategy, bortezomib, thalidomide, and dexamethasone (VTD) were added to the

fractionated MEL to exploit the observed synergism between MEL and VTD (7, 8).

Maintenance therapy was with bortezomib, lenalidomide and dexamethasone (VRD) in

both arms for 3 years. Two-hundred eighty-nine patients were randomized (TT4-S,




5

n=145; TT4-L, n=144) and stratified by ISS stage and the presence of metaphase

cytogenetic abnormalities (CA). Metaphase abnormalities were deemed to be present if

at least 2 cells with the same karyotype were present. Protocol eligibility included age

>18yr and =<75yr and normal cardio-pulmonary function; liver function tests could be up

to twice normal; and creatinine levels up to 3mg/dL were acceptable. Patients had to

have active and measurable MM fulfilling CRAB criteria (9). All patients had to sign a

written informed consent in keeping with institutional, national and Helsinki Declaration

guidelines. The protocol and its revisions had been approved by the institutional review

board, which received annual progress reports. As TT4 was supported with a grant from

the National Cancer Institute, a Data Safety and Monitoring Board (DSMB) reviewed

toxicities and efficacies at least annually. We also had independent data audits every 6

to 8 months to verify adherence to protocol stipulations, examine pharmacy records and

informed consents, and verify recorded toxicities especially causes of death (COD) and

clinical efficacy in terms of complete response (CR) and CR duration (CRD),

progression-free survival (PFS) and overall survival (OS).

Patient work up was per protocol and was similar to our practice in TT3 (2).

Special studies included GEP of plasma cells (GEP-PC) and whole bone marrow

biopsies GEP-BMBX), procured from both random iliac crest sites and from magnetic

resonance imaging (MRI)-defined focal lesions under computer-assisted tomography

guidance. Random bone marrow samples were submitted for metaphase cytogenetic

analysis and interphase fluorescence in situ hybridization (FISH) for the detection of

deletion 17 p and amplification of 1q21. The protocol called for serial short-term and

longer-term studies of GEP and MRI as well as 18-fluoro-deoxyglucose positron




6

emission tomography (PET). Pharmacogenomic studies called for repeat GEP sampling

from random iliac crest bone marrow and MRI-defined focal lesion(s) 48 hours after

test-dosing with bortezomib 1.0mg/m2 and again 48 hours after test-dosing with

melphalan 10mg/m2 prior to starting the full VTD-PACE program. PET follow-up studies

were scheduled on day 5 after commencing VTD-PACE and prior to first

transplantation. These correlative studies will be the subject of a separate report. Most

patients were treated in the outpatient setting and were checked daily by MM-

experienced nursing staff and weekly by their physicians. Hematopoietic progenitor

collection was started when CD34 collection criteria were met with the goal to procure at

least 20 million CD34 cells per kilogram body weight.

The primary clinical endpoint was reduction in toxicity in TT4-L versus TT4-S.

Secondary endpoints included complete response (CR) defined according to IMWG

criteria, the duration of which was measured from its onset to progression or death (10).

Overall survival (OS) was calculated from registration until the date of death.

Progression-free survival (PFS) was similarly calculated, but also incorporated

progressive disease as an event. Time to progression (TTP) was measured from

registration until the date of progression or relapse, whereas time to relapse (TTR)

focused on the subset of patients achieving CR. Death without prior progression or

relapse was included as a competing risk in analyses of TTP and TTR. Post-relapse

survival (PRS) was measured from the date of relapse or progression until death. For

all time-to-event analyses, patients were censored at the date they were last known to

be alive. Causes of death (COD) included treatment-related mortality (TRM), myeloma-

related mortality (MRM), and a third category capturing other or indeterminate causes




7

(OIM) such as fatal car accident, stroke and other causes that could not be attributed to

treatment or disease. Toxicities were graded according to Version 3 of the NCI

After accrual and randomization of 289 patients, the DSMB recommended

closure of the TT4-L arm due to lack of reduction in toxicity vis-à-vis TT4-S. Two

patients randomized to TT4-L, were treated according to the standard arm after the

DSMB recommend closure of TT4-L.These patients are accounted for in the 289

patients. Higher mortality at 1 year in patients ≥65yr on both arms (10.6% and 10.2%

for TT4-L and TT4-S) prompted subsequent accrual of 74 such patients to Total

Therapy 6 (TT6) designed for previously treated patients, because its less dose-intense

therapies had resulted in TRM of 0% (11). Younger patients continue enrollment to

TT4-S but only the 289 randomized patients are the subject of this report.

Statistical Analysis

The data set was created on January 16, 2015 on 289 patients randomized

between TT4-S and TT4-L. In accordance with the intent-to-treat principle, all patients

were analyzed according to their randomized arm. The Kaplan-Meier method was used

to estimate the distributions of OS, PFS and CR duration (12). Cumulative incidences

for COD, TTP and TTR were calculated using the method of Gooley, et al. (13). Group

comparisons for survival endpoints and cumulative incidence were performed using the

log-rank test (14). Multivariate models of prognostic factors were carried out using Cox

regression (15). P-values <0.05 were considered statistically significant.

Results

Clinical Outcomes




8

Patient characteristics are summarized in Table 1 and were not different between

the 2 arms. The consort flow diagram (supplemental figure 1) portrays the progression

of patients through the various protocol steps and lists the off-study reasons. Grade ≥ 3

toxicities occurred with similar frequencies in the 2 arms of TT4 (supplemental Table 2),

with no significant differences observed between major toxicity categories. With a

median follow-up of 4.5 years in both arms, 112 and 108 patients are alive at the time of

analysis while 100 and 92 are progression-free on TT4-S and TT4-L, respectively

(Figure 1). Two-year OS estimates are similar in both arms, 90% and 87% (Figure 1A);

the corresponding PFS estimates of 84% and 79% were also comparable (Figure 1B).

At the time of analysis, 90 and 92 had achieved CR status on TT4-S and TT4-L,

respectively, for 2-yr estimates of 59% and 61% (Figure 1C); 2-year CRD estimates are

87% and 81% (P=0.05) (Figure 1D). Median TTP was similar in both arms, with 2-yr

estimates of 8% and 11% , while there was a trend towards earlier TTR from CR on the

TT4-L arm (12% vs 8% at 2 years, p=0.07) (data not shown). COD were divided into

MRM (p=0.70), TRM (p=0.61) and OIM (p=0.25) and were not different between both

arms (Supplemental Figure 2A). Post-relapse survival (PRS) was similar in the 2 arms

(Supplemental Figure 2B).

As in previous TT trials, the presence of CA affected clinical outcomes (Figure 2).

Surprisingly, the presence of CA had opposite prognostic connotations in the 2 study

arms. In TT4-S, CA showed a strong trend toward inferior OS (Figure 2A) while the

reverse applied to TT4-L (Figure 2B). Non-significant trends in opposite directions were

noted for PFS (Figure 2C, D). CRD tended to be inferior in patients with CA-type MM in

TT4-S (Figure 2E) with an opposite trend in TT4-L (Figure 2F), though neither




9

comparison was statistically significant. TTR was significantly steeper in the presence of

CA in TT4-S (Figure 2G) and did not affect this outcome variable in TT4-L (Figure 2H).

Next we examined the impact of baseline factors on outcomes (Table 2 and

Table 3). The univariate cox analysis of baseline features associated with OS and PFS

across TT4 arms are summarized in Table 2. OS and PFS were inferior in the case of

older age ≥65yr, high beta-2-microglobulin (B2M) (both ≥3.5mg/L and >5.5mg/L),

presence on PET-CT of more than 3 fluoro-deoxyglucose avid focal lesions (FL), low

albumin levels (<3.5g/dL) posed a significant hazard for OS but not for PFS, and the

reverse applied to CRP elevation (≥8mg/L). There was a trend for better OS and PFS

for the CD1 and CD2 molecular subgroups (Table 2). In the presence of CA, in

comparison to its absence (no CA), OS tended to be shorter in TT4-S (p=0.08) and

longer in TT4-L (p=0.07). On multivariate analysis (Table 3), older age, B2M >5.5mg/L,

and PET-FL >3 independently imparted inferior OS and PFS. Low albumin adversely

affected OS and was of borderline significance for PFS. Patients with CA enjoyed

superior OS when randomized to TT4-L with a strong trend apparent also for PFS. In

the case of TT4-S, differences were in the opposite direction, though not significant. The

different effect of CA on outcomes by treatment arm (interaction) was statistically

significant for both OS and PFS (p = .0035 and .0491, respectively).

As we had previously reported on prognostic implication of imaging parameters,

we sought to determine whether the adverse clinical impact of PET-FL>3 pertained to

both TT4 arms (16). Regardless of treatment arm, OS and PFS were inferior for PET-FL

>3 (Supplementary Figure 3). The timing of onset of CR was not affected by PET-FL,

while CRD was superior in case of PET-FL =<3 for the TT4-S arm but not for TT4-L. A




10

trend for more rapid TTP with PET-FL >3 was observed in TT4-S whereas TTP was

similar in TT4-L. TTR was significantly faster with PET-FL >3 only in case of TT4-S.

GEP probes linked to the presence of cytogenetic abnormalities (CA)

The observation of CA’s favorable OS impact in TT4-L was unexpected and at

variance with findings in all previous TT protocols (17). We therefore analyzed whether

the presence or absence of metaphase CA could be linked to certain gene probes

which might explain the superior performance in TT4-L of fractionated melphalan with

added bortezomib and thalidomide. A training set of patients with LR-MM enrolled in

TT3a was chosen for this endeavor. Among 266 untreated patients with available

baseline GEP studies, 90 (34%) exhibited CA. Among the test set of 164 patients with

baseline GEP accrued to TT3b, 67 (41%) qualified as having CA. Using a false

discovery rate of 0.0001, 51 probes significantly distinguished patients with and without

CA (Supplementary Table 3). Seven of the 51 genes function in DNA replication,

recombination, and repair; five in nucleic acid metabolism, and 4 in RNA post-

translational modification and RNA damage and repair. Ingenuity pathway analysis

identified a network of eight interrelated genes that were overexpressed in the CA

group, indicating that these MM cells have a higher proliferative activity (Supplementary

Figure 4). We then examined clinical outcomes by the GEP51-CA prediction model in

the 2 arms of TT4 (Figure 3). In TT4-S, GEP51/no-CA had superior OS and PFS

compared to GEP51/CA (Figure 3A, B), which was not observed in TT4-L (Figure 3C,

D).

Discussion




11

The results of this phase-3 TT4 trial failed to show that TT4-L was less toxic and

was, in fact, inferior to TT4-S in terms of CRD with a trend for inferiority in case of TTR.

One-year mortality was similar, 7.6% in TT4-L and 6.9% in TT4-S (Fisher’s Exact Test

p=0.75), but low in both arms in younger patients (TT4-S, 6.2%; TT4-L, 5.2%).

Unanticipated were the opposite implications of the presence of CA on OS: adverse in

TT4-S (p=0.08) as in all previous TT trials, and favorable in TT4-L (p=0.07), with

significant opposite prognostic implications in the 2 study arms. The other clinical

endpoints were only marginally affected except for significantly faster TTR in case of CA

in TT4-S. When analyzed in the context of all competing variables, TT4-L impacted OS

of patients with CA-type MM favorably, with a strong trend apparent also for PFS. T It

seems reasonable to speculate that CA-type MM derived benefit from conditioning with

fractionated MEL VTD since T4-L employed 1 less cycle of induction and consolidation

treatment. This combination has also been found to be synergistic when used to treat

patients with VMP ineligible for stem cell transplantation in the VISTA study (18).

We previously reported that bortezomib and VTD could be added to fractionated

EL in doses up to 250mg/m2 in an advanced patient population with acceptable toxicity

and high response rates (7, 8). Others have also explored the use of bortezomib and

MEL as a conditioning regimen prior to transplantation and found that both agents can

be safely combined and possibly produce increased response rates (19-22). The

present paper is the first randomized study comparing single dose melphalan 200mg/m2

with a fractionated MELVTD schedule. The results of TT4 indicate that CA-type

myeloma benefits from MELVTD, whilst non-CA MM fares better with application of a

single high dose MEL 200mg/m2. Prior attempts to improve conditioning with MEL have




12

not been successful and included the addition of other drugs, skeletal targeting

radioactive antibodies or radiation therapy (23-28).

Mono- or poly ubiquination of DNA-repair enzymes such H2AX, BRCA1, and

FANCD2 is required for recruitment to sites of DNA double stranded breaks and is

essential to the DNA damage response (29). Bortezomib reduces the availability of

nuclear ubiquitin and may thereby impair homologous recombination mediated DNA-

repair. It has been suggested that bortezomib renders myeloma cells more vulnerable

to alkylator-mediated DNA-damage by essentially inducing a BRCAness type state (30).

CA-type MM is likely to harbor more genomic chaos and may be more dependent on

efficient DNA-repair explaining the sensitivity to fractionated MEL-VTD conditioning.

Availability of GEP led us to examine whether CA-linked gene probes could be

identified. Indeed 51 genes were identified that distinguished no-CA from CA

subgroups. When examined for its clinical relevance, the GEP51 model was prognostic

in TT4-S so that GEP51/CA prediction was associated with inferior OS and PFS. In

contrast, such Kaplan-Meier plots were superimposable between these 2 groups in

TT4-L. Ingenuity pathway analysis suggests that CA-type MM has overexpression of

genes critical to proliferation. The ability to examine metaphase cytogenetics inherently

implies that the myeloma cells are able to survive and proliferate in vitro without the

support of the myeloma micro-environment. Increased ability to proliferate may be a

favorable evolutionary trait, but on the other hand stress requirements for DNA-repair to

prevent fatal mutations and cell death. In this context, one could speculate that

exposure to DNA-damaging alkylators such as MEL in the setting of bortezomib-

induced reduced DNA-repair could lead to fatal “mitotic catastrophes”.




13

This is the first clinical trial, which suggests that tailoring the transplant

conditioning regimen to myeloma biology may further improve outcome. However, the

study was not designed to specifically study the impact of conditioning on outcome in

the context of metaphase CA. These preliminary findings therefore require confirmation

in future phase III studies.

Author Contributions

FvR, AH, JC and BB designed the research. JE, MZ, SY, ET, SW, RK, YJ, XP, MG, NP,

DS, SP, CB, MCF, FvR and BB performed the research. FvR, AM, AH, JC and BB

analyzed the data. FvR, AM, JC, BB and GM wrote the manuscript.




14

References

1. Barlogie B, Tricot G, Anaissie E, Shaughnessy JD, Jr, Rasmussen E, van Rhee

F, et al. Thalidomide and hematopoietic-cell transplantation for multiple myeloma.

N Engl J Med 2006;354:1021-1030.

2. Barlogie B, Anaissie E, van Rhee F, Haessler J, Hollmig K, Pineda-Roman M, et

al. Incorporating bortezomib into upfront treatment for multiple myeloma: Early

results of Total Therapy 3. Br J Haematol 2007;138:176-185.

3. Shaughnessy JD, Jr., Zhan F, Burington BE, Huang Y, Colla S, Hanamura I, et

al. A validated gene expression model of high-risk multiple myeloma is defined

by deregulated expression of genes mapping to chromosome 1. Blood

2007;109:2276-2284.

4. Barlogie B, Mitchell A, van Rhee F, Epstein J, Morgan GJ, Crowley J. Curing

myeloma at last: Defining criteria and providing the evidence. Blood

2014;124:3043-3051.

5. Shaughnessy JD, Jr., Qu P, Usmani S, Heuck CJ, Zhang Q, Zhou Y, et al.

Pharmacogenomics of bortezomib test-dosing identifies hyperexpression of

proteasome genes, especially PSMD4, as novel high-risk feature in myeloma

treated with Total Therapy 3. Blood 2011;118:3512-3524.

6. Nair B, van Rhee F, Shaughnessy JD, Jr., Anaissie E, Szymonifka J, Hoering A,

et al. Superior results of Total Therapy 3 (2003-33) in gene expression profiling-

defined low-risk multiple myeloma confirmed in subsequent trial 2006-66 with

VRD maintenance. Blood 2010;115:4168-4173.




15

7. Hollmig K, Stover J, Talamo G, Zangari M, Thertulien R, van Rhee F, et al.

Addition of bortezomib (VelcadeTM) to high dose melphalan (Vel-Mel) as an

effective conditioning regimen with autologous stem cell support in multiple

myeloma (MM). Blood 2004;104:929.

8. Pineda-Roman M, Cottler-Fox M, Hollmig K, Anaissie EJ, van Rhee F, Tricot G,

et al. Retrospective analysis of fractionated high-dose melphalan (F-Mel) and

bortezomib-thalidomide-dexamethasone (VTD) with autotransplant (AT) support

for advanced and refractory multiple myeloma (AR-MM). Blood 2006;108:3102.

9. International Myeloma Working Group. Criteria for the classification of

monoclonal gammopathies, multiple myeloma and related disorders: A report of

the International Myeloma Working Group. Br J Haematol 2003;121:749-757.

10. Durie BG, Harousseau JL, Miguel JS, Bladé J. Barlogie B, Anderson K, et al.

International uniform response criteria for multiple myeloma. Leukemia

2006;20:1467-1473.

11. van Rhee F, Giralt S, Barlogie B. The future of autologous stem cell

transplantation in myeloma. Blood 2014;124:328-333.

12. Kaplan EL, Meier P. Nonparametric estimation from incomplete observations. J

AM Stat Assoc 1958; 53:457-481.

13. Gooley TA, Leisenring W, Crowley J, Storer BE. Estimation of failure probabilities

in the presence of competing risks: New representations of old estimators. Stat

Med 1999;18:695-706.

14. Mantel N. Evaluation of survival data and two new rank order statistics arising in

its consideration. Cancer Chemother Rep 1966;50:163-170.




16

15. Cox DR. Regression models and life-tables. J R Stat Soc 1972;34:187-220.

16. Waheed S, Mitchell A, Usmani S, Epstein J, Yaccoby S, Nair B, et al. Standard

and novel imaging methods for multiple myeloma: correlates with prognostic

laboratory variables including gene expression profiling data.

Haematologica 2013;98:71-8.

17. Arzoumanian V, Hoering A, Sawyer J, van Rhee F, Bailey C, Gurley J, et al.

Suppression of abnormal karyotype predicts superior survival in multiple

myeloma. Leukemia 2008;22:850-5.

18. San Miguel JF, Schlag R, Khuageva NK, Dimopoulos MA, Shpilberg O, Kropff M,

et al. Bortezomib plus melphalan and prednisone for initial treatment of multiple

myeloma. N Engl J Med 2008;359:906-917.

19. Roussel M, Moreau P, Huynh A, Mary JY, Danho C, Caillot D, et al. Bortezomib

and high-dose melphalan as conditioning regimen before autologous stem cell

transplantation in patients with de novo multiple myeloma: A phase 2 study of the

Intergroupe Francophone du Myelome (IFM). Blood 2010;115:32-37.

20. Lonial S, Kaufman J, Tighiouart M, Nooka A, Langston AA, Heffner LT, et al. A

phase I/II trial combining high-dose melphalan and autologous transplant with

bortezomib for multiple myeloma: A dose- and schedule-finding study. Clin

Cancer Res 2010;16:5079-5086.

21. Thompson PA, Prince HM, Seymour JF, Ritchie D, Stokes K, Burbury K, et al.

Bortezomib added to high-dose melphalan as pre-transplant conditioning is safe

in patients with heavily pre-treated multiple myeloma. Bone Marrow Transplant.

2011;46:764-765.




17

22. Lee SR, Kim SJ, Park Y, Sung HJ, Choi CW, Kim BS et al. Bortezomib and

melphalan as a conditioning regimen for autologous stem cell transplantation in

multiple myeloma. Korean J Hematol. 2010;45:183-187.

23. Shimoni A, Smith TL, Aleman A, Weber D, Dimopoulos M, Anderlini P, et al.

Thiotepa, busulfan, cyclophosphamide (TBC) and autologous hematopoietic

transplantation: An intensive regimen for the treatment of multiple myeloma.

Bone Marrow Transplant. 2001;27:821-828.

24. Anagnostopoulos A, Aleman A, Ayers G, Donato M, Champlin R, Weber D, et al.

Comparison of high-dose melphalan with a more intensive regimen of thiotepa,

busulfan, and cyclophosphamide for patients with multiple myeloma. Cancer

2004;100:2607-2612.

25. Christoforidou AV, Saliba RM, Williams P, Qazilbash M, Roden L, Aleman A, et

al. Results of a retrospective single institution analysis of targeted skeletal

radiotherapy with (166) holmium-DOTMP as conditioning regimen for autologous

stem cell transplant for patients with multiple myeloma. Impact on transplant

outcomes. Biol Blood Marrow Transplant 2007;13:543-549.

26. Giralt S, Bensinger W, Goodman M, Posoloff D, Eary J, Wendt R, et al. 166ho-

DOTMP plus melphalan followed by peripheral blood stem cell transplantation in

patients with multiple myeloma: Results of two phase 1/2 trials. Blood

2003;102:2684-2691.

27. Desikan KR, Tricot G, Dhodapkar M, Fassas A, Siegel D, Vesole DH, et al.

Melphalan plus total body irradiation (MEL-TBI) or cyclophosphamide (MEL-CY)

as a conditioning regimen with second autotransplant in responding patients with




18

myeloma is inferior compared to historical controls receiving tandem transplants

with melphalan alone. Bone Marrow Transplant 2000;25:483-487.

28. Moreau P, Facon T, Attal M, Hulin C, Michallet M, Maloisel F, et al. Comparison

of 200 mg/m(2) melphalan and 8 Gy total body irradiation plus 140 mg/m(2)

melphalan as conditioning regimens for peripheral blood stem cell transplantation

in patients with newly diagnosed multiple myeloma: Final analysis of the

Intergroupe Francophone du Myélome 9502 randomized trial. Blood

2002;99:731-735.

29. Neri P, Ren L, Gratton K, Stebner E, Johnson J, Klimowicz A, et al. Bortezomib-

induced "BRCAness" sensitizes multiple myeloma cells to PARP inhibitors. Blood

2011;118:6368-6379.

30. Fonseca R. Innovation in myeloma treatments PARP excellence! Blood

2011;118:6234-6235.




Table 1: Patient characteristics

Factor All

Patients S-TT4 L-TT4 Median Age (yr) 61.4

(N=289)(30.4 - 75.9)

60.4 (N=145)(34.2 - 75.2)

62.2 (N=144) (30.4 - 75.9)

Age >= 60yr 157/289 (54%)

74/145 (51%)

83/144 (58%)

Female 111/289 (38%)

60/145 (41%)

51/144 (35%)

IgA 46/287 (16%)

22/143 (15%)

24/144 (17%)

IgG 179/287 (62%)

93/143 (65%)

86/144 (60%)

ISS Stage 1 86/289 (30%)

44/145 (30%)

42/144 (29%)

ISS Stage 2 124/289 (43%)

61/145 (42%)

63/144 (44%)

ISS Stage 3 79/289 (27%)

40/145 (28%)

39/144 (27%)

Albumin < 3.5 g/dL

123/289 (43%)

58/145 (40%)

65/144 (45%)

B2M >= 3.5 mg/L

153/289 (53%)

77/145 (53%)

76/144 (53%)

B2M > 5.5 mg/L 79/289 (27%)

40/145 (28%)

39/144 (27%)

Creatinine >= 1.5 mg/dL

31/289 (11%)

15/145 (10%)

16/144 (11%)

Hemoglobin < 10 g/dL

109/289 (38%)

55/145 (38%)

54/144 (38%)

Baseline PET FL > 0

166/261 (64%)

82/126 (65%)

84/135 (62%)

Baseline PET FL > 3

95/261 (36%)

45/126 (36%)

50/135 (37%)

Baseline FL-SUV > 3.9

79/168 (47%)

40/84 (48%)

39/84 (46%)

Cytogenetic abnormalities

112/283 (40%)

53/142 (37%)

59/141 (42%)

amp1q21 (FISH)

63/259 (24%)

31/127 (24%)

32/132 (24%)




2

Factor All

Patients S-TT4 L-TT4 delTP53 (FISH) 19/259

(7%) 11/127 (9%)

8/132 (6%)

GEP 70 High Risk

2/286 (1%)

0/143 (0%)

2/143 (1%)

GEP CD-1 subgroup

16/286 (6%)

6/143 (4%)

10/143 (7%)

GEP CD-2 subgroup

56/286 (20%)

23/143 (16%)

33/143 (23%)

GEP HY subgroup

102/286 (36%)

49/143 (34%)

53/143 (37%)

GEP LB subgroup

45/286 (16%)

25/143 (17%)

20/143 (14%)

GEP MF subgroup

8/286 (3%)

6/143 (4%)

2/143 (1%)

GEP MS subgroup

35/286 (12%)

22/143 (15%)

13/143 (9%)

GEP PR subgroup

24/286 (8%)

12/143 (8%)

12/143 (8%)

n/N (%): n- Number with factor, N- Number with valid data for factor ND: No valid observations for factor Bold print signifies Fisher’s exact test p-value < 0.05




Table 2: Univariate Cox analysis of baseline features associated with overall survival (OS) and progression-free survival (PFS) across TT4 arms

Overall Survival Progression-Free

Survival

Variable n/N (%) HR (95%

CI) P-value HR (95%

CI) P-value Univariate TT4-L 144/289

(50%) 1.14

(0.71, 1.82)

0.593 1.24 (0.83, 1.85)

0.290

Age >= 65 yr 96/289 (33%)

2.27 (1.41, 3.64)

<.001 1.89 (1.26, 2.82)

0.002

Albumin < 3.5 g/dL

132/289 (46%)

1.90 (1.17, 3.08)

0.010 1.43 (0.96, 2.14)

0.077

B2M >= 3.5 mg/L

153/289 (53%)

2.40 (1.44, 4.01)

<.001 1.79 (1.18, 2.70)

0.006

B2M > 5.5 mg/L

79/289 (27%)

1.93 (1.18, 3.14)

0.009 1.88 (1.24, 2.85)

0.003

Creatinine > 1.5 mg/dL

31/289 (11%)

1.30 (0.65, 2.62)

0.461 1.70 (0.98, 2.94)

0.060

CRP >= 8 mg/L 80/288 (28%)

1.60 (0.98, 2.61)

0.058 1.56 (1.03, 2.36)

0.037

Hb < 10 g/dL 109/289 (38%)

1.42 (0.88, 2.28)

0.151 1.31 (0.87, 1.96)

0.193

Baseline PET FL > 0

166/261 (64%)

1.49 (0.86, 2.57)

0.153 1.35 (0.86, 2.12)

0.187

Baseline PET FL > 3

95/261 (36%)

2.72 (1.63, 4.52)

<.001 2.13 (1.40, 3.26)

<.001

GEP CD-1 subgroup

16/285 (6%)

0.22 (0.03, 1.57)

0.131 0.30 (0.07, 1.21)

0.090




2


Survival


CI) P-value HR (95%

CI) P-value GEP CD-2

subgroup 56/285 (20%)

1.47 (0.85, 2.54)

0.171 1.49 (0.94, 2.37)

0.090

GEP HY subgroup

102/285 (36%)

0.84 (0.50, 1.39)

0.489 0.74 (0.48, 1.14)

0.175

GEP LB subgroup

44/285 (15%)

1.26 (0.69, 2.30)

0.454 1.47 (0.90, 2.40)

0.126

GEP MF subgroup

8/285 (3%)

1.13 (0.28, 4.61)

0.867 1.12 (0.35, 3.53)

0.853

GEP MS subgroup

35/285 (12%)

0.75 (0.34, 1.64)

0.470 0.69 (0.34, 1.36)

0.280

GEP PR subgroup

24/285 (8%)

1.25 (0.57, 2.72)

0.583 1.33 (0.69, 2.56)

0.393

TT4-S only

Any CA vs No CA

53/142 (37%)

1.84 (0.93, 3.64)

0.081 1.42 (0.79, 2.56)

0.244

TT4-L only

Any CA vs No CA

59/142 (42%)

0.51 (0.25, 1.07)

0.074 0.71 (0.40, 1.26)

0.248

HR- Hazard Ratio, 95% CI- 95% Confidence Interval, P-value from Wald Chi-Square Test in Cox Regression NS2- Multivariate results not statistically significant at 0.05 level. All univariate p-values reported regardless of significance. Multivariate model uses stepwise selection with entry level 0.1 and variable remains if meets the 0.05 level. A multivariate p-value greater than 0.05 indicates variable forced into model with significant variables chosen using stepwise selection.




Table 3: Multivariate Cox regression analysis of baseline features associated with overall survival (OS) and progression-free survival (PFS) across TT4 arms


Survival


CI) P-value HR (95%

CI) P-value Multivariate

Age >= 65 yr 86/257 (33%)

2.01 (1.19, 3.38)

0.0086 1.76 (1.14, 2.72)

0.0110

Albumin < 3.5 g/dL

118/257 (46%)

1.90 (1.12, 3.20)

0.0167 1.46 (0.95, 2.24)

0.0838

B2M > 5.5 mg/L

67/257 (26%)

2.40 (1.35, 4.29)

0.0030 2.08 (1.30, 3.35)

0.0024

Baseline PET FL > 3

93/257 (36%)

2.94 (1.73, 5.01)

<.0001 2.24 (1.44, 3.49)

0.0003

Any CA vs No CA (TT4-L) *

58/133 (44%)

0.36 (0.16, 0.79)

0.0113 0.56 (0.30, 1.05)

0.0698

Any CA vs No CA (TT4-S) *

50/124 (40%)

1.73 (0.81, 3.71)

0.1575 1.35 (0.71, 2.56)

0.3565

HR- Hazard Ratio, 95% CI- 95% Confidence Interval, P-value from Wald Chi-Square Test in Cox Regression P-value for the CA by Treatment arm interactions, OS: p = 0.0035; PFS: p = 0.0491 * Denominators represent the total number of patients relevant to this hazard ratio rather than the total number of patients in the model.




Figure Legend

Figure 1: Survival Outcomes in TT4 by arm (A: OS, B: PFS, C: CR, D: CRD)

Figure 2: Clinical outcomes according to the presence of cytogenetic abnormalities (no

CA vs CA) by TT4 arm

Figure 3: Clinical outcomes according to 51-gene model predicting CA versus no-CA




Figure 1: Survival outcomes in TT4 by arm (A: OS, B: PFS, C: CR, D: CRD)

A

B

C

D

Overall Survival: UARK 2008-01Randomized Patients Only

By Treatment Arm

0%

20%

40%

60%

80%

100%

0 1 2 3 4 5 6Years from Registration

S-TT4L-TT4

Events / N33 / 14536 / 144

2-YearEstimate

90% (85, 95)87% (81, 92)

Logrank P-value = .59

Progression-Free Survival: UARK 2008-01Randomized Patients Only

By Treatment Arm

0%

20%

40%

60%

80%

100%


S-TT4L-TT4

Events / N45 / 14552 / 144

2-YearEstimate

84% (78, 90)79% (72, 85)


Time to Complete Response: UARK 2008-01Randomized Patients Only

By Treatment Arm

0%

20%

40%

60%

80%

100%


S-TT4L-TT4

Events / N90 / 14292 / 140

2-YearEstimate58.5%60.7%

P = .20

CR Duration: UARK 2008-01Randomized Patients Only

By Treatment Arm

0%

20%

40%

60%

80%

100%

0 1 2 3 4 5 6Years from CR

S-TT4L-TT4

Events / N14 / 9026 / 92

2-YearEstimate

87% (80, 94)81% (73, 89)





S-TT4 L-TT4

A

B

C

D

Overall Survival: UARK 2008-01Patients Randomized to S-TT4

By Presence of Cytogenetic Abnormalities

0%

20%

40%

60%

80%

100%


No CAAny CA

Events / N16 / 8917 / 53

2-YearEstimate

94% (89, 100)83% (72, 94)


Overall Survival: UARK 2008-01Patients Randomized to L-TT4


0%

20%

40%

60%

80%

100%


No CAAny CA

Events / N26 / 8310 / 59

2-YearEstimate

81% (72, 89)95% (89, 100)


Progression-Free Survival: UARK 2008-01Patients Randomized to S-TT4


0%

20%

40%

60%

80%

100%


No CAAny CA

Events / N25 / 8920 / 53

2-YearEstimate

88% (81, 95)77% (66, 89)


Progression-Free Survival: UARK 2008-01Patients Randomized to L-TT4


0%

20%

40%

60%

80%

100%


No CAAny CA

Events / N34 / 8318 / 59

2-YearEstimate

72% (63, 82)86% (77, 96)


Figure 2: Clinical outcomes according to the presence of cytogenetic abnormalities (no CA vs CA) by

TT4 arm




E

F

G

H

CR Duration: UARK 2008-01Patients Randomized to S-TT4


0%

20%

40%

60%

80%

100%


No CAAny CA

Events / N6 / 528 / 35

2-YearEstimate

92% (85, 100)79% (65, 93)


CR Duration: UARK 2008-01Patients Randomized to L-TT4


0%

20%

40%

60%

80%

100%


No CAAny CA

Events / N16 / 5110 / 39

2-YearEstimate

76% (65, 88)87% (76, 98)


Time to Relapse from CR: UARK 2008-01Patients Randomized to S-TT4


0%

20%

40%

60%

80%

100%

0 1 2 3 4 5 6Years from CR Onset

No CAAny CA

Events / N2 / 526 / 35

2-YearEstimate

3.9%15.4%

P = .03

Time to Relapse from CR: UARK 2008-01Patients Randomized to L-TT4


0%

20%

40%

60%

80%

100%

0 1 2 3 4 5 6Years from CR Onset

No CAAny CA

Events / N10 / 517 / 39

2-YearEstimate15.8%7.9%

P = .79




A B

C

D

O v e r a l l S u r v i v a l f o r P a t i e n t s w i t h G E P D a t a A v a i l a b l e i n T T 4B y P r e d i c t e d C A f o r A r m A : S - T T 3

0 %

2 0 %

4 0 %

6 0 %

8 0 %

1 0 0 %

0 2 4 6 8Y e a r s

P r e d i c t e d N o C AP r e d i c t e d a n y C A

D e a t h s / N1 6 / 9 71 7 / 4 3

3 - Y e a rE s t i m a t e

9 2 % ( 8 6 , 9 7 )7 4 % ( 5 9 , 8 8 )

L o g r a n k P - v a l u e = . 0 0 2

O v e r a l l S u r v i v a l f o r P a t i e n t s w i t h G E P D a t a A v a i l a b l e i n T T 4B y P r e d i c t e d C A f o r A r m B : L - T T 3

0 %

2 0 %

4 0 %

6 0 %

8 0 %

1 0 0 %

0 2 4 6 8Y e a r s


D e a t h s / N2 2 / 8 31 4 / 5 7


7 9 % ( 7 1 , 8 8 )8 4 % ( 7 4 , 9 4 )

L o g r a n k P - v a l u e = . 6 3

P r o g r e s s i o n F r e e S u r v i v a l f o r P a t i e n t s w i t h G E P D a t a A v a i l a b l e i n T T 4B y P r e d i c t e d C A f o r A r m B : L - T T 3

0 %

2 0 %

4 0 %

6 0 %

8 0 %

1 0 0 %

0 2 4 6 8Y e a r s


E v e n t s / N3 1 / 8 32 1 / 5 7


7 1 % ( 6 1 , 8 1 )7 7 % ( 6 6 , 8 8 )

L o g r a n k P - v a l u e = . 7 9

P r o g r e s s i o n F r e e S u r v i v a l f o r P a t i e n t s w i t h G E P D a t a A v a i l a b l e i n T T 4B y P r e d i c t e d C A f o r A r m A : S - T T 3

0 %

2 0 %

4 0 %

6 0 %

8 0 %

1 0 0 %

0 2 4 6 8Y e a r s


E v e n t s / N2 3 / 9 72 1 / 4 3


8 3 % ( 7 6 , 9 1 )6 5 % ( 4 9 , 8 0 )

L o g r a n k P - v a l u e = . 0 0 2

Figure 3: Clinical outcomes according to 51-gene model predicting CA versus no-

CA




Published OnlineFirst November 3, 2016.Clin Cancer Res Yogesh Jethava, Alan Mitchell, Joshua Epstein, et al. transplantsbortezomib and thalidomide to fractionated melphalan Adverse metaphase cytogenetics can be overcome by adding

Updated version

10.1158/1078-0432.CCR-15-2620doi:

Access the most recent version of this article at:

Material

Supplementary

http://clincancerres.aacrjournals.org/content/suppl/2016/11/03/1078-0432.CCR-15-2620.DC1

Access the most recent supplemental material at:

Manuscript

Authoredited. Author manuscripts have been peer reviewed and accepted for publication but have not yet been

E-mail alerts related to this article or journal.Sign up to receive free email-alerts

Subscriptions

Reprints and

[email protected] at

To order reprints of this article or to subscribe to the journal, contact the AACR Publications

Permissions

Rightslink site. Click on "Request Permissions" which will take you to the Copyright Clearance Center's (CCC)

.http://clincancerres.aacrjournals.org/content/early/2016/11/03/1078-0432.CCR-15-2620To request permission to re-use all or part of this article, use this link



http://clincancerres.aacrjournals.org/lookup/doi/10.1158/1078-0432.CCR-15-2620

http://clincancerres.aacrjournals.org/content/suppl/2016/11/03/1078-0432.CCR-15-2620.DC1

http://clincancerres.aacrjournals.org/cgi/alerts

mailto:[email protected]

http://clincancerres.aacrjournals.org/content/early/2016/11/03/1078-0432.CCR-15-2620


Documents

Adverse metaphase cytogenetics can be overcome by adding ... · Purpose. To determine whether a reduction in the intensity of Total Therapy (TT) reduces toxicity and maintains efficacy