1

Single agent laromustine (VNP40101M), a novel alkylating agent, has significant activity in older patients with previously untreated poor-risk acute myeloid leukemia.

1Gary J. Schiller, 2Susan M. O'Brien, 3Arnaud Pigneux, 4Daniel J DeAngelo, 5Norbert Vey, 6Jonathan Kell, 7Scott Solomon, 8Robert K. Stuart, 9Verena Karsten, 9Ann L. Cahill, 10Maher X. Albitar, 11Francis J. Giles

1David Geffen School of Medicine at UCLA, Los Angeles, CA; 2The University of Texas M.D. Anderson Cancer Center, Houston, TX; 3Hopital Haut Leveque, Bordeaux, France; 4Dana-Farber Cancer Institute, Boston, MA; 5Institut Paoli-Calmettes, Marseille, France; 6University Hospital of Wales, Cardiff, UK; 7Northside Hospital, BMT Group, Atlanta, GA; 8MUSC Hollings Cancer Center, Charleston, SC; 9Vion Pharmaceuticals, Inc., New Haven, CT; 10Quest Diagnostics Nichols Institute, San Juan Capistrano, CA; 11 CTRC at The University of Texas Health Science Center, San Antonio, TX

This study received research support from Vion Pharmaceuticals, Inc, New Haven, CT

Address reprint requests to Francis J. Giles, MD, CTRC at The UT Health Science Center, 7979 Wurzbach Road, Mail code 8026, Urschel Tower, Suite 600, San Antonio, TX 78229 Phone: 210 450 3838. Fax 210 450 9823 Email: frankgiles@aol.com

Running head: Laromustine in elderly with previously-untreated poor-risk AML

Presented in part at ASCO Annual Meeting 2008 – May 20 2008: Abs 7026

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Abstract

Purpose: An international phase II study of laromustine (VNP40101M), a

sulfonylhydrazine alkylating agent, was conducted in patients ≥60 years with previously

untreated poor-risk acute myeloid leukemia (AML).

Patients and Methods: Laromustine 600 mg/m2 was administered as a single 60-

minute intravenous infusion. Patients were ≥ 70 years or ≥ 60 years with at least one

additional risk factor – unfavorable AML karyotype, ECOG Performance Score (PS) of

2, and/or cardiac, pulmonary or hepatic comorbidities.

Results: Eighty-five patients (median age 72 years (60 to 87)) were treated. Poor-risk

features included: Age ≥ 70, 78%; adverse karyotype, 47%; PS 2, 41%; pulmonary

disease 77%; cardiac disease 73%; hepatic disease 3%. 96% of patients had ≥ 2 risk

factors, 39% ≥ 4. The overall response rate (ORR) was 32%: 20 patients (23%)

achieving complete response (CR), 7 (8%) CR with incomplete platelet recovery. ORR

was 20% in patients with adverse cytogenetics; 32% in those ≥70 years; 32% in those

with PS of 2; 32% and 34%, in patients with baseline pulmonary or cardiac dysfunction

respectively, and 27% in 33 patients with ≥4 risk factors. Twelve (14%) patients died

within 30 days of receiving laromustine therapy. Median overall survival was 3.2

months, with a 1-year survival of 21%; the median duration of survival for those who

achieved CR/CRp was 12.4 months, with a 1-year survival of 52%.

Conclusion: Laromustine has significant single-agent activity in elderly patients with

poor-risk AML. Adverse events are predominantly myelosuppressive and/or respiratory.

Response rates are consistent across a spectrum of poor-risk features.

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Introduction

Elderly patients with AML have a very poor prognosis, attributable to having both more

resistant disease and relatively poor tolerance of cytotoxic agents.1In contrast to

younger patients, treatment for elderly patients with AML has not improved over recent

decades.1-3 The majority of older patients with AML are not offered current standard

induction regimens and survival for those who are treated is very poor.4-6 Laromustine

({VNP40101M; Cloretazine, OnriginTM}, Vion Pharmaceuticals Inc, New Haven, CT) is a

1,2-bis(sulfonyl)hydrazine with broad spectrum antitumor activity in preclinical models.7-

20 Laromustine is initially activated to yield 90CE [1,2-bis(methylsulfonyl)-1-(2-

chloroethyl)hydrazine] and methylisocyanate. 90CE rapidly produces an alkylating,

chloroethylating species, similar to the species generated by BCNU

[carmustine]).11,20BCNU and laromustine produce different decomposition products as

laromustine does not yield hydroxyethylating, vinylating, or aminoethylating species.

The chlorethylating species responsible for laromustine's alkylation is relatively specific

to the O6 position of guanine, while BCNU, unlike laromustine, also alkylates the N7

position of guanine as is more typical of most alkylating agents.11,21 Laromustine yields

over twice the molar yield of DNA cross-links than the nitrosoureas. Drugs that cause

primarily N7 alkylations exhibit one thirtieth of the anticancer activity and the same

mutagenic potential as their counterparts that form both N7 alkylations and cross-

links.14,18 Laromustine does not generate hydroxyethyl alkylations of the O6 position of

guanine – these lesions are considered to be carcinogenic but therapeutically

unimportant.14,20 Laromustine also inhibits the nucleotidyl-transferase activity of purified

4

human DNA polymerase beta (Pol beta), a principal enzyme of DNA base excision

repair (BER).22 Alkylated DNA is often repaired via BER in vivo. Inhibition of the

polymerase activity of Pol beta may account for some of the synergism between

laromustine's two reactive subspecies in cytotoxicity assays. Laromustine has

significant activity against hematologic malignancy–derived cell lines, including those

resistant to other alkylating agents, and has significant activity in animal leukemia

models.12,23-25

Myelosuppression is dose-limiting in patients with solid tumors receiving laromustine

with minimal attendant significant nonhematologic toxicity.26,27 In a phase I study of

laromustine in patients with refractory leukemia, mild reversible infusion-related

toxicities were the most frequent adverse events, occurring in 24 patients (63%) on the

first course.28 Dose escalation was terminated at 708 mg/m2 because of prolonged

myelosuppression; 600 mg/m2 was the recommended phase II study dose, with no

significant extramedullary toxicity at this dose level. The combination of laromustine

and cytarabine was also studied in patients with refractory leukemia.29 Complete

responses (CRs) were seen at laromustine dose levels 400 mg/m2 in 10 (27%) of 37

patients. Dose-limiting toxicities (gastrointestinal and myelosuppression) were seen

with 500 and 600 mg/m2 of laromustine combined with a 4-day 1.5 gm/m2/d continuous-

infusion cytarabine schedule, but not with the equivalent 3-day schedule.

A phase II study was conducted in patients with previously untreated non-favorable

karyotype AML/high-risk MDS 60 years of age at the time of therapy.30 One hundred

and thirty one patients, median age 72 years, received laromustine as a single 600

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mg/m2 intravenous infusion. Performance status was 2 in 34 (26%) of patients. Overall

response rate was 31% with 31 patients (24%) achieving CR, 9 (7%) CR with

incomplete platelet recovery (CRp). Response rates in 54 de novo AML patients was

44%, 37% in 68 patients with intermediate-risk and 26% in 54 patients with

unfavorable-risk cytogenetics. An international Phase II study of laromustine was

performed in older patients with AML and prospectively defined additional poor-risk

factors.

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Patients and methods

The study was reviewed and approved by institutional review boards at all participating

institutions. All patients provided signed informed consent indicating that they were

aware of the investigational nature of this study.

Patient Eligibility

Patients with previously untreated non–acute promyelocytic leukemia (APL) AML, age

60 years, were eligible. Patients were allowed to have active controlled infection

including chronic hepatitis and/or known CNS leukemia. Eligible patients had at least

one additional risk factor – unfavorable AML karyotype (Table 1), ECOG Performance

Score (PS) of 2, age ≥ 70 years, and/or cardiac, pulmonary or hepatic comorbidities

which were defined as per the Hematopoietic Cell Transplantation Comorbidity Index

(HCT-CI).31 Cardiac dysfunction was defined as ejection fraction≤ 50%, history of

significant coronary artery disease (one or more vessel stenosis requiring medical

treatment, stent placement or surgical bypass graft), history of CHF or MI, significant

arrhythmia including atrial fibrillation or flutter, sick sinus syndrome, or ventricular

arrhythmia, and/or valvular heart disease (excluding mitral valve prolapse). Pulmonary

dysfunction was defined as DLCO and/or FEV1 < 80% and/or dyspnea on slight activity

or at rest or requiring oxygen. Hepatic dysfunction was defined as chronic hepatitis

and/or liver cirrhosis. As the formulation of laromustine contains 30% ethanol,

concurrent treatment with disulfiram was not allowed.

Treatment and Study Design

Patients received laromustine 600 mg/m2 on day 1 administered by intravenous

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infusion using a polyethylene-lined administration set in a peripheral or central vein.

Patients were pretreated with antihistamines and antiemetics to prevent transient

infusion-related reactions. Patients were eligible to receive a second cycle of

laromustine induction therapy if they achieved less than CR but some marrow

improvement after the first cycle. For patients in CR after a first/second induction cycle,

or PR after a second, at least one consolidation cycle of continuous infusion cytarabine

at a dose of 400 mg/m2/day for 5 days was to be administered. Toxicity was graded

using the National Cancer Institute Common Toxicity Criteria (version 3.0). All patients

who received any therapy on study were considered assessable for toxicity and

response.

Response and Prognostic Criteria

CR was defined as normalization of the blood and bone marrow with 5% blasts, a

granulocyte count 1 x 109/L, and a platelet count 100 x 109/L. CRp was as per CR,

but with platelet counts remaining <100 x 109/L, and independence of platelet

transfusions was defined as the ability to maintain a platelet count of 20 x 109/L.

Marrow slides used for diagnosis and response evaluation were reviewed by an

independent pathologist at a central laboratory.

Statistical Analysis

A two-stage optimal minimax design was used.32 If ≤21 of 77 patients achieved CR, the

hypothesis that the ORR is ≥35% was to be rejected. Survival was measured from the

day of laromustine treatment to death as a result of any cause. Distributions of overall

survival were estimated by the method of Kaplan and Meier. Quantitative factors were

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treated as continuous variables in regression analyses, but grouped when necessary

for descriptive tables and figures.

Results

Patient Characteristics

Eighty-five eligible patients were enrolled onto the study between May 2006 and

December 2007. Sixty-two (73%) patients were classified as AML not otherwise

classified. Twenty (24%) patients had a diagnosis of AML with multilineage–dysplasia

without antecedent MDS based on morphologic findings. Two patients had evidence of

prior MDS/MPD and one patient had acute biphenotypic leukemia. Forty (47%) patients

had an unfavorable AML karyotype. Patient baseline characteristics are summarized in

Table 2. Fifty patients (59%) were male. The median patient age was 72 years (range,

60 to 87 years), sixty-six (78%) patients were older than 70 years; performance status

was 2 in 35 patients (41%). Sixty-five patients (76%) had pulmonary dysfunction, 62

(73%) cardiac dysfunction, and 3 (4%) had hepatic dysfunction at study entry (Table 3).

Among patients with cardiac disease, 30 (48%) had one cardiac risk factor, and 32

(52%) had multiple cardiac risk factors. Regardless of the number of cardiac risk

factors present, a score of one was assigned to each patient with cardiac disease.

Among patients with pulmonary dysfunction, 22 (34%) had moderate disease, with

DLCO/FEV1 >65 and <80%, whereas 43 (66%) had severe disease with FEV1 ≤65%

and/or dyspnea at rest and/or requiring oxygen. Three (4%) patients had a single poor-

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risk factor (2 with age ≥ 70 year, 1 with pulmonary dysfunction). The majority of

patients (96%) had 2 or more risk factors.

Toxicity

Infusion Reaction

Ninety-nine cycles of laromustine therapy were administered. Infusion reactions were

reported as adverse events in 21 (25%) patients during treatment with laromustine

induction cycle 1 or cycle 2. Hypotension was the most frequently reported event in 7

patients, followed by headache reported in 5 patients, nausea and vomiting in

3 patients each, and flushing, malaise and rash in 2 patients each. All of the events

were mild or moderate in intensity and resolved within 24 hours (most within a few

hours). In 3 patients interruption of laromustine infusion was required due to events of

headache, lethargy or pruritus. The events resolved and the infusions were completed

in all cases.

Myelosuppression

Grade 3/4 myelosuppression after first induction with laromustine occurred in almost all

patients with 82% and 75% of patients exhibiting Grade 4 neutropenia or

thrombocytopenia respectively. Eighteen of 27 patients who had baseline neutropenia

of grade 0 to 3 experienced grade 4 neutropenia after treatment. Forty-three of 63

patients who had baseline grade 0 to 3 thrombocytopenia experienced grade 4

thrombocytopenia after treatment. For patients who achieved CR/CRp, the median

10

time to granulocyte counts ≥0.5 x 109/L was 35 days (range 3-70) and to platelet count

≥50 x 109/L was 35 days (range 15-79). The median time to granulocyte recovery of

≥1.0 x 109/L was 38 days (range 8-79) and median time to platelet recovery ≥100 x

109/L was 35 days (range 28-70).

Adverse Events

Grade 3/4 non-hematologic adverse events that occurred within 42 days after induction

cycles 1 or 2 with laromustine and were considered potentially related to laromustine

are summarized in Table 4. Grade 3/4 febrile neutropenia occurred in 15 patients.

Gastrointestinal events such as nausea and vomiting or diarrhea were frequent but

generally mild or moderate in intensity. Only one grade 3 event of intermittent diarrhea

occurred 19 days following induction cycle 1 with laromustine. Mucositis was infrequent

(9%) and primarily of low grade; one grade 3 event of mucosal inflammation occurred

27 days after laromustine treatment. Grade 3/4 non-infectious respiratory events were

observed in 6 patients of which 3 occurred after 2 induction cycles with laromustine.

The onset of dyspnea or pleural effusions was between day 23 and 117 from the first

treatment with laromustine. The radiologic findings in 2 cases were described as

ground-glass opacities. In 2 patients the pulmonary events resolved after corticosteroid

administration. Infectious events considered possibly related to laromustine occurred in

22% of patients.

Twelve (14%) patients died within 30 days of receiving first induction with laromustine,

all deaths occurred while patients were pancytopenic. AML progression with or without

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infection was the primary cause of death in 6 patients. Two patients died with severe

pneumonia and one patient with severe neutropenic sepsis. Two patients had Grade

3/4 renal dysfunction, one died on study day 4 with tumor lysis syndrome and one, who

also had pneumonia, died on study day 29. One patient died from acute respiratory

distress syndrome on study day 17. Five responding patients died within 30 days of

receiving cytarabine for consolidation treatment, two events occurred following a

second consolidation cycle of cytarabine. These deaths also occurred in patients with

pancytopenia and were reported to be due to infection (n=2), ventricular fibrillation

(n=1), cardiac arrhythmia (n=1) and cerebral hemorrhage (n=1).

Response

Twenty-seven patients (32%) achieved CR (20, 24%) or CRp (7, 8%). The majority of

remissions were achieved after a single induction treatment with laromustine. Of 14

patients who received a second cycle of laromustine therapy, 1 achieved CR. Eighteen

and four patients received a first and second consolidation cycle with cytarabine for 5

days, respectively. Two patients received non-protocol specified consolidation therapy

– one a course of tipifarnib and a second a course of idarubicin and cytarabine. The

CR rates by individual risk group are summarized in Table 5. The ORR was 20% in

patients with adverse cytogenetics; 32% in those ≥70 years; 31% in those with PS of 2;

32% and 34% in those with pulmonary or cardiac dysfunction respectively at study

entry. The ORR was 27% in 33 patients (39%) with ≥4 risk factors.

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Treatment outcome by number of risk factors is presented in Table 5. The response

rate remained consistent as risk increased within the study population. Overall survival,

which was comparably prolonged in CR and CRp, is summarized graphically in Figure

1. The median overall survival was 3.2 months in all patients (2.1 months in non-

responders and 12.4 months in responders). The percentage of patients that were alive

12 months after receiving therapy was 21% for all patients (9% in non-responding

patients and 52% in responding patients). Among patients who did not achieve a CR

with laromustine therapy, fifteen (26%) received subsequent re-induction therapy,

which resulted in remission in 3 patients.

Discussion

The results of the current study confirms and extends the observation made in a prior

study, with broader patient eligibility criteria, that single agent laromustine has

significant activity in older patients with poor prognosis previously untreated AML.30

The treatment of elderly patients with AML remains a very significant challenge.1-6 A

combination of inherently biologically more resistant disease, including P-glycoprotein

overexpression and increased frequency of both adverse cytogenetic and molecular

makers, as well as an increased incidence of patient baseline comorbidities, are

associated with very poor results following current standard AML induction regimens.3,6

There are inherent difficulties in the assessment of a novel approach in a cancer

population for which no standard therapy exists and/or in which available therapies are

ineffective and/or very poorly tolerated. As a prior international Phase II study had

identified a consistent CR rate associated with single agent laromustine therapy in

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elderly patients with AML30, the current study focused on patients with disease and/or

clinical features known to be associated with a particularly poor prognosis. The study

population thus contained a large percentage of patients whose baseline cardiac,

pulmonary, and/or hepatic comorbidities rendered them unlikely to receive

anthracyclines and/or cytarabine – the common standard cytotoxic agents included in

AML induction regimens. A low dose cytarabine (LDAC) regimen was recently

randomized versus hydroxyurea in older patients with AML considered unfit for

standard induction therapy.6 LDAC, while demonstrating superior response rates and

overall survival to hydroxyurea, was not associated with achievement of CR in patients

with adverse cytogenetics and/or poor performance status. In contrast, laromustine

therapy results in CR and prolonged survival in patients prospectively selected for poor

prognosis AML. A direct comparison between LDAC and laromustine would require a

randomized study which would need to be performed in only better prognosis patients

– the issue there would be whether LDAC can be considered a reasonable regimen to

offer such patients in view of its low ability to achieve CR.

In AML, older age, adverse AML karyotype, and poor performance status are poor-risk

features.3,5,33,34 While comorbidity is acknowledged to be both an independent adverse

prognostic factor in elderly patients with AML, and to be not fully accounted for in

performance score analysis35, it is a challenge to define prospectively. The HCT-CI has

been shown to be predictive of both early mortality and overall survival in elderly

patients receiving myelosuppressive AML induction therapy.36 The HCT-CI has also

been validated in patients with myelodysplastic syndromes (MDS).37 In the currently

reported study definitions for pulmonary, cardiac and hepatic dysfunction from the

14

HCT-CI were prospectively used to define patients with baseline organ dysfunction.

Laromustine retained activity despite the presence of multiple comorbidities even when

these were present in organ systems (cardiac, pulmonary), as conditions mitigating

against the use of anthracycline/cytarabine induction regimens. The incorporation of

formal comorbidity scales in therapeutic studies in elderly AML facilitates comparisons

between approaches. While the HCT-CI has been shown to be reproducible as a

prognostic determinant in patients with hematological malignancies treated at different

institutions,38 other recently reported prognostic scales for older patients with AML also

merit further study.33,39 This study data also confirm the pattern of extramedullary

toxicity of laromustine at doses that are both myelosuppressive26,27 and capable of

significant leukemia tumor burden reduction.29,30,40,41 As anticipated with an alkylating

agent, myelosuppression and it’s consequences comprise the major toxicity of

laromustine. Medically important respiratory events marked by dyspnea that can

develop weeks to months following therapy have been observed in a few patients and

can resolve with steroid treatment. Events commonly associated with leukemia

induction treatment, such are mucositis, severe gastrointestinal toxicity and alopecia

occurred infrequently and at low grades. Cardiac events were uncommon. The

relatively constant early death rate observed in patients with multiple risk factors

indicates that the presence of these factors should not preclude administration of

laromustine therapy. Some deaths were observed following cytarabine consolidation

associated with myelosuppression suggesting that this consolidation schedule might

not be optimal for some elderly patients with poor-risk features. Optimal consolidation

approaches have not been established in the elderly population with AML due to the

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paucity of both patients in response and of nontoxic agents with anti-leukemia activity.

Such agents are now in development and approaches that may help to prolong

laromustine-induced CR significantly could include LDAC,6 hypomethylating agents,42

FLT3 inhibitors,43 vascular endothelial growth factor antagonists,44 and/or vaccines.45

The majority of patients with AML over 60 years of age do not receive the standard

AML induction therapies given to younger patients.46,47 Increasing comorbidity is

significantly associated with a reduced chance that a patient will be offered a cytotoxic

AML induction regimen.46,48 The patients treated with laromustine on this study were

chosen for multiple poor prognosis features, including the first prospective use of the

HCT-CI to quantify baseline comorbidity. No data has been published on the use of

standard AML induction regimens in an elderly patient cohort prospectively defined as

in the currently reported laromustine study. Recent randomized studies in potentially

similar populations have involved LDAC, hydroxyurea, or supportive care as control

arm therapies.6,49 Recent in vitro data have shown synergistic activity of laromustine

when combined with standard AML induction agents on the proliferation, viability and

apoptosis of AML cell lines and blast cells from patients.25 Future studies of the

combination of laromustine with standard induction anthracycline/cytarabine regimens

are warranted, as are those investigating whether laromustine can be substituted for

one or more of the agents within these regimens.41

Novel agents being investigated in older patients with AML, including clofarabine,

decitabine, tipifarnib, and CP-4055, warrant investigation in combination regimens with

16

laromustine.42,50-52 Laromustine offers a new therapeutic option for older patients with

poor-risk AML.

17

Table 1. Cytogenetic risk group classification

Risk group Karyotype

Intermediate Normal +6, -Y, del(12p)

Unfavorable

del (5q) / - 5q -7 /del (7q) Abnormalities involving (3q), (9q), (11q), (20q), (21q) abn (17p) t (6;9) t (9;22) complex (≥3 unrelated abnormalities)

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Table 2. Baseline characteristics of patients treated on study

No. Risk Factors

No. Patients

Unfavorable Cytogenetics

N (%)

Age ≥ 70

N (%)

ECOG PS = 2 N (%)

Cardiac Dysfunction N

(%)

Pulmonary Dysfunction

N (%)

Hepatic DysfunctiN (%)

1 3 (4%)

0 (0%)

2 (67%)

0 (0%)

0 (0%)

1 (33%)

0 (0%)

2 18 (21%)

7 (39%)

13 (72%)

2 (11%)

8 (44%)

6 (33%)

0 (0%)

≥ 3 64

(75%) 33

(52%) 51

(80%) 33

(52%) 54

(84%) 58

(91%) 3

(5%)

Total 85 40 (47%)

66 (78%)

35 (41%)

62 (73%)

65 (76%)

3 (5%)

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Table 3: Baseline HCT-CI comorbidity distribution

Comorbidity N=85 Cardiac 39 (46%) Severe Pulmonary 43 (51%) Infection 31 (36%) Arrhythmia 36 (42%) Psychiatric Disturbance 18 (21%) Diabetes 21 (25%) Moderate Pulmonary 22 (26%) Solid Tumor 11 (13%) Mild Liver 14 (16%) Heart Valve disease 17 (20%) Obesity 13 (15%) Cerebrovascular Disease 2 (2%) Moderate/Severe Liver 3 (4%) Peptic Ulcer 3 (4%) Rheumatologic 2 (2%)

HCT-CI = Hematopoietic cell transplantation-specific comorbidity index

20

Table 4. Potentially Laromustine-related adverse events occurring in > 5% of patients*

Preferred Term Grade 1-2

Grade 3

Grade 4 Overall

Febrile Neutropenia 12 14 1 27 (31.8%)

Nausea 24 24 (28.2%)

Hypotension 13 1 14 (16.5%)

Pyrexia 13 1 14 (16.5%)

Fatigue 12 1 13 (15.3%)

Diarrhea 12 1 13 (15.3%)

Vomiting 9 9 (10.6%)

Mucosal inflammation 7 1 8 (9.4%)

Headache 8 8 (9.4%)

Pleural Effusion 6 2 8 (9.4%)

Dyspnea or Hypoxia 3 3 2 6 (7.1%)

Peripheral edema 6 6 (7.1%)

Flushing 6 6 (7.1%)

Anorexia 4 2 6 (7.1%)

Confusional state 6 6 (7.1%)

* Adverse events included in this table were collected for 42 days from any treatment with laromustine (following first and/or second induction cycle).

21

Table 5 Response by risk group and by number of baseline risk factors

Risk Factors N CR/CRp Overall

Response Rate

Age ≥ 70 66 17/4 32%

ECOG PS = 2 35 7/4 31%

Unfavorable Cytogenetics 40 6/2 20%

Pulmonary Dysfunction 65 14/7 32%

Cardiac Dysfunction 62 14/7 34%

Hepatic Dysfunction 3 0/0 0%

Number of risk factors

1-2 21 8/0 38%

≥ 3 64 12/7 30%

Total 85 20/7 32%

22

Figure 1. Overall survival: All patients versus responding (---) patients

23

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