Hematology 2002 111

Chronic Myelogenous Leukemia

Brian J. Druker, Stephen G. O’Brien, Jorge Cortes, and Jerald Radich

The treatment options for chronic myelogenousleukemia (CML) continue to evolve rapidly. Imatinibmesylate (Gleevec, Glivec, formerly STI571) hascontinued to show remarkable clinical benefits andthe updated results with this agent are reviewed. Asrelapses using single agent imatinib have occurred,particularly in advanced phase patients, the issueof whether combinations of other antileukemicagents with imatinib may yield improved results isaddressed. In addition, data on new agents that havepotential in the treatment of CML are reviewed. Theseagents are presented in the context of their molecu-lar mechanism of action. The most recent data forstem cell transplantation, along with advances innonmyeloablative transplants, are also reviewed.

In Section I, Drs. Stephen O’Brien and BrianDruker update the current status of clinical trials

with imatinib and review ongoing investigationsinto mechanisms of resistance and combinationsof imatinib with other agents. They also presenttheir views on integration of imatinib with othertherapies.

In Section II, Dr. Jorge Cortes describes the mostrecent data on novel therapies for CML, includingfarnesyl transferase inhibitors, arsenic trioxide,decitabine, and troxatyl, among others. These agentsare discussed in the context of their molecularmechanism of action and rationale for use.

In Section III, Dr. Jerald Radich updates theresults of stem cell transplants for CML, includingemerging data on nonmyeloablative transplants. Healso presents data on using microarrays to stratifypatients into molecularly defined risk groups.

I. CURRENT STATUS OF TRIALS OF IMATINIB MESYLATE

(STI571, GLEEVEC) ALONE AND IN COMBINATION

Stephen G. O’Brien, MD, PhD,* andBrian J. Druker, MD**

Imatinib (Gleevec, Glivec, formerly STI571) is an in-hibitor of the Bcr-Abl tyrosine kinase that is central tothe pathogenesis of chronic myelogenous leukemia(CML). In preclinical studies, imatinib selectivity inhib-ited the proliferation of cells expressing Bcr-Abl in vitroand in vivo.1 The remarkable results of imatinib in clini-cal trials led to rapid Food and Drug Administration(FDA) approval of this drug for chronic phase CML pa-tients who had failed interferon and for accelerated phaseand blast crisis patients. These clinical trials will be sum-marized along with a randomized comparison of imatinibto interferon plus cytarabine in newly diagnosed chronicphase patients. Despite the impressive responses seen inchronic phase patients, numerous questions remain. Howdurable will the responses to imatinib be? Will cytoge-netic or molecular responses be useful surrogate mark-ers for survival in patients treated with imatinib? As goodas imatinib seems, is it possible to improve upon these

results? What are the mechanisms of resistance or re-lapse to imatinib and what treatments are available forrelapsed patients? How should allogeneic transplanta-tion and imatinib be integrated? These issues will beaddressed in this and subsequent articles.

* University of Newcastle, Royal Victoria Infirmary,Newcastle Upon Tyne NE1 4LP, United Kingdom

** Oregon Health Sciences University, Cancer Institute, 3181SW Sam Jackson Park Rd., L592, Portland, OR 97201-3011

Dr. Druker is supported by grants from the National CancerInstitute, The Leukemia and Lymphoma Society, BurroughsWellcome Fund, T.J. Martell Foundation, and the Doris DukeCharitable Foundation.

Dr. O’Brien has received research support from and has been aconsultant to Novartis and Schering Plough.

Acknowledgments: The authors are most grateful to JorgeCortes, Francois Guilhot, Andreas Hochhaus, and FrancoisXavier Mahon for sharing their data.

112 American Society of Hematology

Summary• Formerly known as CGP57148 or STI571, imatinib

mesylate (Gleevec, Glivec) is a 2-phenylamino-py-rimidine derivative (C

30H

35N

7SO

4, MW 589.7) that

specifically inhibits the tyrosine kinase activity ofAbl proteins, c-Abl and Bcr-Abl. Imatinib is knownto also inhibit the tyrosine kinase activity of KIT,ARG (Abl-related gene), and the platelet-derivedgrowth factor receptor (PDGF-R).

• Since June 1998, when the first patient was treated,at least 15,000 patients with chronic myeloid leuke-mia (CML) worldwide have now been treated withimatinib. Imatinib was licensed by the Food andDrug Administration (FDA) for use in the UnitedStates in May 2001. The European Medicines Evalu-ation Agency (EMEA) granted a license coveringthe UK in November 2001. The licensed indicationsinclude chronic phase CML in patients who havefailed interferon therapy (lack of response or intol-erance), accelerated phase CML, and blast crisis ofCML. A license for newly diagnosed patients is cur-rently being sought.

• In the treatment of chronic phase CML, imatinib pro-duces much better hematological and cytogeneticresponses than interferon-α (IFN-α) with most pa-tients sustaining these responses. In newly diagnosedCML the major cytogenetic response on imatinibtherapy is 83%, with 68% complete responses (com-pared with 20% and 7%, respectively, with inter-feron + Ara-C). However, there are as yet no data toconclusively demonstrate that imatinib improveslong-term survival for CML patients when comparedwith an interferon-containing regimen.

• In accelerated phase CML, response rates are infe-rior to those seen in chronic phase, but many of theseresponses are sustained.

• In blast crisis of CML (both myeloid and lymphoid),response rates compare favorably with thoseachieved with chemotherapy, but only a minority ofpatients have durable responses. The use of imatinibmay be justified in order to achieve transient con-trol of disease with minimal side effects mostly, andit may be used as a bridge to transplant. Combina-tions of imatinib with chemotherapy are being ex-plored.

• Initial data indicate that higher doses of imatinib mayproduce better responses, but these data require con-firmation in further, longer-term studies.

• Phase I/II trials with imatinib in combination withinterferon and Ara-C, which demonstrate that com-bination therapy can be safely delivered with excel-

lent response rates, have been conducted. Other com-bination studies are in progress. Whether combina-tion therapy, or higher dose imatinib monotherapy,is superior to imatinib 400 mg daily in chronic phaseis the subject of a forthcoming Phase III study.

Background and Mechanism of ActionSince the first description of the Philadelphia (Ph) chro-mosome in 1960,2 biomedical scientists have developedan impressively detailed understanding of the biologyof CML.3-7 The subsequent development of a highly suc-cessful therapeutic agent based on this knowledge isconfirmation of the value of basic biomedical researchin both an academic and industry setting. Bcr-Abl en-codes a protein, p210BCR-ABL, with dysregulated tyrosinekinase activity,8 which is necessary and sufficient forleukemogenesis.9-12 Imatinib mesylate is a potent inhibi-tor of the tyrosine kinases activity of ABL,13,14 KIT,15,16

PDGF-R,13,14,17 and ARG.18 Imatinib competitively inhib-its the interaction of adenosine triphosphate (ATP) withthese proteins,19 thereby inhibiting their ability to phos-phorylate and activate downstream target proteins.

Published Studies to Date with Imatinib

Phase I studiesA standard dose-escalation Phase I study of imatinibbegan in June 1998 at 3 centers in the United States.The study population consisted of CML patients inchronic phase, refractory or resistant to interferon-basedtherapy or intolerant of this drug.20 At later stages of thestudy, patients with CML in blast crisis and patients withPh-chromosome positive acute lymphoblastic leukemia(ALL) were also enrolled.21 Imatinib was well toleratedwith minimal side effects. Despite dose escalation from25 mg to 1000 mg in 14 cohorts of patients, a maximallytolerated dose could not be defined. Imatinib was ad-ministered once daily and pharmacokinetics showed ahalf-life of 13-16 hours. Significant clinical benefits wereobserved at daily doses above 300 mg. In chronic phasepatients who had failed therapy with interferon, 53 of54 (98%) patients treated with ≥ 300 mg per day achieveda complete hematologic response, and 96% of these re-sponses lasted beyond 1 year.20 In myeloid blast crisispatients, 21/38 (55%) patients treated at doses ≥ 300 mgper day responded, with 18% having responses lastingbeyond 1 year.21

Phase II studies of imatinibThe success of the Phase I studies prompted Phase IIstudies. Single agent imatinib was tested further in in-terferon refractory and interferon intolerant patients aswell as in accelerated phase patients and patients with

Hematology 2002 113

CML in myeloid blast crisis and Ph-chromosome posi-tive ALL. These studies accrued over 1000 patients, at30 centers in 6 countries, in 6 to 9 months. Results fromthese studies, with 18 months of follow-up, have beenpublished and are summarized in Table 1.22-25

Five hundred thirty-two chronic phase patients whowere refractory to or intolerant of interferon-α weretreated with an imatinib dose of 400 mg daily. Eligibil-ity criteria in this study allowed inclusion of patients withup to 15% blasts and 15% basophils in the marrow orperipheral blood. Median duration of disease was 34months and median duration of previous interferontherapy was 14 months. Ninety-five percent of patientsachieved a complete hematologic response (CHR), withthe median time to CHR being less than 1 month. Imatinibinduced major cytogenetic responses (≤ 35% Ph-posi-tive metaphases) in 60% of patients, with a completecytogenetic response rate of 41%. With a median fol-low-up of 18 months, the estimated progression-freesurvival was 89%. Only 2% of patients discontinuedtherapy because of adverse events.22 A major cytoge-netic response at 3 months was associated with a higherrate of progression-free survival. Baseline features thatindependently predicted a high rate of major cytogeneticresponses were the absence of blasts in the peripheralblood, a hemoglobin >12 g/dL, < 5% blasts in the mar-row, CML disease duration of less than 1 year, and aprior cytogenetic response to interferon.

Results of the Phase II study in accelerated phasepatients were equally impressive.23 Accelerated phasewas defined as 15-30% blasts or > 30% blasts pluspromyelocytes in the peripheral blood or marrow, > 20%peripheral basophils, or a platelet count less than 100 ×109/L, unrelated to therapy. Two hundred thirty-five pa-tients were enrolled in this study. Overall, 82% of pa-tients showed some form of hematologic response, with34% of patients achieving a CHR. Twenty-four percentof patients achieved a major cytogenetic response, with17% complete responses. Estimated 12-month progres-sion-free and overall survival rates were 59% and 74%,

respectively. Again, these results were achieved with-out substantial toxicity.23

Results of the Phase II study treating 260 myeloidblast crisis patients with imatinib showed an overall re-sponse rate of 52%, with sustained hematologic re-sponses lasting at least 4 weeks in 31% of patients. Eightpercent of patients achieved a complete remission (CR= < 5% blasts) with peripheral blood recovery.24 An-other 4% of patients cleared their marrows to less than5% blasts but did not meet the criteria for CR because ofpersistent cytopenias. Finally, 18% of patients either re-turned to chronic phase or had partial responses. Majorcytogenetic responses were seen in 16% of patients, with7% having complete responses. Median survival was 6.9months. Twenty percent of patients were still alive at 18months with a suggestion of a plateau on the survivalcurve. These results compare favorably with historicalcontrols treated with chemotherapy for myeloid blastcrisis in which the median survival is approximately 3months. In patients with Ph-positive ALL, 29/48 (60%)responded to single agent imatinib. However, the dura-tion of response was relatively short, with a median esti-mated time to disease progression of only 2.2 months.25

Phase III study: comparison of imatinib withIFN-ααααα in newly diagnosed patientsA Phase III randomized study, comparing imatinib at400 mg per day with interferon plus cytarabine in newlydiagnosed chronic phase CML patients, enrolled 1106patients from June 2000 to January 2001. Five hundredfifty-three patients were randomized to each treatment.Baseline characteristics were well balanced for all fea-tures evaluated, including age, WBC, Sokal and Euroscore, and time from diagnosis. With a median follow-up of 14 months, patients randomized to imatinib hadstatistically significant better results than patients treatedwith interferon plus cytarabine in all parameters mea-sured (Table 2), including rates of CHR, major and com-plete cytogenetic responses, tolerance of therapy, andfreedom from disease progression.26 Given the signifi-cant difference in the percentage of patients with dis-ease progression to accelerated phase or blast crisis, 7%with interferon versus 1.5% of patients randomized toimatinib, it seems likely that this will translate into asurvival benefit. A remaining question is the durabilityof the responses to imatinib.

Side Effects of TherapyImatinib has generally been well tolerated, with grade 3or 4 nonhematologic toxicities being uncommon. Thetoxicities observed in newly diagnosed chronic phasepatients are summarized in Table 3. The common grade1 or 2 toxicities include fluid retention, nausea, muscle

Table 1. Phase II results with imatinib.

Chronic Phase Accelerated Blast(IFN Failure)22 Phase23 Crisis24

CHR 95% 34% 8%

MCR 60% 24% 16%

CCR 41% 17% 7%

Disease progression 11% 40% 80%

Abbreviations: IFN, interferon; CHR, complete hematologicresponse; MCR, major cytogenetic response (Ph+ metaphases <35%); CCR, complete cytogenetic response.

114 American Society of Hematology

cramps, skin rashes, fatigue, and diarrhea. Practical as-pects of management of these symptoms have been re-viewed elsewhere.27 Myelosuppression is more commonin advanced phase patients than in the chronic phasepatients, as shown in Table 4, and imatinib induced pro-longed aplasia in 1% of blast crisis patients.24 In con-trast, patients with gastrointestinal stromal tumor treatedwith 400 or 600 mg per day of imatinib had rates ofgrade 3/4 neutropenia and thrombocytopenia of 5% andless than 1%, respectively, demonstrating the specificityof this side effect to leukemia patients.28 Given that 300mg seems to be a threshold dose for optimal therapeuticresponses, it is recommended that imatinib should rarely,if ever, be used at doses of less than 300 mg.20,27

Rationale for Trying to Improve uponImatinib Monotherapy at 400 mg Daily

Despite the lack of long-term follow-up data, imatinibat 400 mg daily has emerged as the preferred therapyfor newly diagnosed CML patients who do not undergoallogeneic stem cell transplant. However, only a minor-ity (5-10%) of imatinib-treated patients achieve a mo-lecular remission, that is, negative by RT-PCR for Bcr-

Table 3. Nonhematologic toxicities with imatinib in newlydiagnosed chronic phase chronic myelogenous leukemia(CML) patients.26

Adverse Event All Grades (%) Grades 3/4 (%)

Superficial edema 53.2 0.9

Nausea 42.5 0.4

Muscle cramps 35.4 1.1

Musculoskeletal pain 33.6 2.7

Rash 31.9 2.0

Fatigue 30.7 1.1

Diarrhea 30.3 1.3

Headache 28.5 0.4

Joint pain 26.7 2.2

Abdominal pain 23.4 2.0

Myalgia 20.9 1.5

Nasopharyngitis 19.2 0

Hemorrhage 18.9 0.7

Dyspepsia 15.1 0

Vomiting 14.7 0.9

Pharyngolaryngeal pain 14.2 0.2

Dizziness 13.2 0.5

Cough 12.5 0.2

URI 12.5 0.2

Pyrexia 11.8 0.5

Weight, increased 11.6 0.7

Insomnia 11.4 0

Depression 8.9 0.5

Constipation 7.6 0.7

Rigors 6.9 0

Anxiety 6.5 0.2

Dyspnea 6.5 1.3

Pruritus 6.5 0.2

Influenza-like illness 6.4 0

Night sweats 6.4 0.2

Anorexia 4.7 0

Sweating, increased 3.3 0

Alopecia 2.2 0

Table 4. Myelosuppression in the Phase II and III studies with imatinib.

Neutropenia ThrombocytopeniaGrade 3 Grade 4 Grade 3 Grade 4

ANC < 1.0 x 109/L ANC < 0.5 x 109/L ANC < 50 x 109/L ANC < 10 x 109/L

Chronic phase (newly diagnosed) 26 11% 2% 7% 0.5%

Chronic phase (IFN failure) 22 27% 8% 19% 1%

Accelerated phase 23 23% 35% 31% 12%

Blast crisis 24 16% 48% 29% 33%

*ANC indicates absolute neutrophil count; IFN, interferon.

Table 2. Phase III results of imatinib versus interferon pluscytarabine for newly diagnosed chronic phase chronicmyelogenous leukemia (CML) patients.26

Imatinib 400 mg Interferon + Ara-C

CHR 96% 67%

MCR 83% 20%

CCR 68% 7%

Intolerance 0.7% 23%

Progressive disease 1.5% 7%

All of these differences are highly statistically significant with P <0.001. Intolerance leading to discontinuation of first-line therapy;Progressive disease to accelerated phase or blast crisis.

Abbreviations: CHR, complete hematologic response; MCR, majorcytogenetic response (Ph+ metaphases < 35%); CCR, completecytogenetic response.

Hematology 2002 115

Abl transcripts (B. Druker, unpublished data). Relapsesin patients with more advanced disease have been com-mon, and it is likely that resistance to imatinib willemerge in chronic phase patients. Combinations of imat-inib with chemotherapy are an obvious choice for pa-tients with advanced phase disease. If one accepts thatchronic phase patients who do not achieve durablemolecular remissions are at risk of relapse, then it is clearthat there is much room for improvement. Options thathave shown promise thus far include higher doses of imat-inib and combinations of imatinib with IFN-α or cytarabine.The results of these treatments as compared with 400 mg/d of imatinib are summarized in Table 5.

Are Higher Doses of Imatinib More Effective?The dose of 400 mg per day of imatinib for the chronicphase studies was selected based on responses seen inthe Phase I study and a lack of sufficient safety data forhigher doses. As additional safety data from the Phase Istudy emerged, advanced phase patients were treatedwith 600 mg per day. Thus, in the Phase II study of ac-celerated phase patients, 77 patients were treated with400 mg and 158 patients with 600 mg of imatinib.23 Inthe Phase II study of myeloid blast crisis patients, 36patients were treated with 400 mg and 223 patients with600 mg.24 Retrospective analysis of prognostic factorsin the accelerated phase patients showed that the 400and 600 mg cohorts were well matched. In both studies,there was a trend toward higher hematologic and majorcytogenetic response rates in patients treated with 600mg of imatinib (Table 6). In the accelerated phase study,patients treated with 600 mg had a statistically significantimprovement in time to disease progression and survival.23

In a subsequent study of accelerated phase patients,patients were also treated with 600 mg daily of imatinib.As opposed to the accelerated protocol cited above, in

this study, patients who otherwise had chronic phasefeatures, but who had cytogenetic abnormalities besidesa single Ph-chromosome, were defined as accelerated.Fifteen patients with this definition of accelerated phasewere enrolled at Oregon Health & Science Universityand had a median disease duration of 45 months. With amedian follow-up of 12 months, the major cytogeneticresponse rate was 80% (12/15), with a complete cytoge-netic response of 67% (10/15). None of these patientshas relapsed.29 Although a small study, the results forthese relatively poor prognosis patients compare favor-ably with the 12-month results in newly diagnosed CMLpatients treated with 400 mg of imatinib.

The experience with doses higher than 600 mg perday is limited. In chronic phase patients in the Phase IIstudy who failed to achieve a cytogenetic response fol-lowing 1 year of imatinib therapy, dose escalation to 800mg per day has been allowed. Limited experience sug-gests that up to one third of patients will achieve a majorcytogenetic response with dose escalation (B. Druker,unpublished data). Investigators at M.D. Anderson haverecently reported the results of newly diagnosed chronicphase CML patients treated daily with 400 mg versus800 mg of imatinib.30 As seen in Table 5, there is a trendtoward higher cytogenetic responses in the 800-mg dosecohort. Their experience with toxicity was similar to thatreported in the Phase I study, which demonstrated thatdaily doses of 800 mg and higher were less well toler-ated than 600 mg.20 In particular, there was a higher in-cidence of fluid retention, skin rashes, and musclecramps. Daily dosing above 800 mg has generally notbeen used, as this was defined as the maximally toler-ated dose in an EORTC study of patients with gastrointes-tinal stromal tumors.31 At 1000 mg per day, the dose-limiting toxicities were nausea, vomiting, edema, andskin rashes.

Table 5. Comparison of cytogenetic responses in newlydiagnosed chronic phase chronic myelogenous leukemia(CML) patients.

Cytogenetic Responses†

6 months 9 months

Major Complete Major Complete

Imatinib 400 mg 26 63% 40% 75% 54%

Imatinib + PEG-IFN 36 76% 44% 86% 48%

Imatinib + Ara-C†† 74% 57% 80% 57%

Imatinib 400 mg30 80% 52%Imatinib 800 mg30 83% 65%

†Cytogenetic responses are reported at 6 and 9 months forcomparative purposes.†† F. Guilhot, personal communication.

Table 6. Results with 400 mg/d versus 600 mg/d in chronicmyelogenous leukemia (CML) advanced phase patients.

Accelerated23 Blast24

400 mg 600 mg 400 mg 600 mg

HR 65% 71% 9% 35%

CHR 27% 37% 3% 14%

MCR 16% 28% 6% 18%

TTP 8 months Not reached(P = 002)

12-month survival 65% 78%(P = 0.014)

Abbreviations: d, day; HR, hematologic response; CHR, completehematologic response; MCR, major cytogenetic response; TTP,time to progression.

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These data suggest that doses higher than 400 mgper day may yield improved responses. However, in theaccelerated and blast crisis studies, the main impact ofhigher doses was on time to progression and survival.As the response rates for newly diagnosed chronic phaseCML patients are already quite high, this finding im-plies that comparative studies of higher dose imatinibtherapy in this patient population will require time toprogression or survival as endpoints. Alternatively, ratesof molecular remissions, if they correlate with improvedsurvival, may be a useful early endpoint.

Combination Therapy

Imatinib in combination with interferon-αααααAs imatinib and interferon are the two most active formsof nontransplant therapy available for patients with CML,this combination is an obvious choice for testing. In vitrodata demonstrate additive or synergistic antiproliferativeeffects of the combination of imatinib with interferonusing Bcr-Abl-positive cell lines and in colony-formingassays using CML patient samples.32-34 Based on thesedata, several Phase I/II pilot studies with this combina-tion are in progress.

In a Phase I dose-escalation study using imatinibwith regular interferon, 14 chronic phase CML patientswere enrolled.35 In this study, imatinib was administeredas a single agent at 400 mg daily for 2 weeks prior toadding a specified dose of interferon. In the dose-escala-tion, no patient was able to maintain a dose of 400 mg/d ofimatinib plus 5 MU of interferon daily, primarily be-cause of hematologic toxicity. Therefore, the currentstarting dose in a Phase II study for newly diagnosedpatients is 400 mg/d of imatinib with 3 MU of interferondaily. Side effects and responses were similar to thosein a larger study described below.

In a second Phase I/II study, imatinib has been com-bined with pegylated interferon. As of December 2001,49 patients with chronic phase CML had been enteredinto PISCES (PEGIntron and Imatinib CombinationEvaluation Study); median age 52.5 years, 32 newly di-agnosed (< 6 months since diagnosis).36 Imatinib wasadministered for 2 weeks prior to adding a specified doseof interferon. After 6 months of treatment, the majorcytogenetic response rate was 73.3% (n = 22) overalland 82.4% (n = 14) in the newly diagnosed patients (com-plete response; 36.7% and 41.2%, respectively) (Table5). Of the 22 patients achieving a major cytogenetic re-sponse, 55% were taking either 200 mg/d of imatinibplus 0.25 µg/kg/wk PEGIntron (n = 8) or 200 mg/dimatinib plus 0.5 µg/kg/wk PEGIntron (n = 4). In thePhase I study, only 1/7 (14%) patients treated with 200mg/d of imatinib had a cytogenetic response, suggest-

ing that the combination of imatinib with interferon hasimproved activity over imatinib alone. Myelosuppressionwas common, with 30/49 patients experiencing grade 3/4neutropenia within the first month on study while taking400 mg/d imatinib plus 0.5 µg/kg/wk PEGIntron. Basedon this result, the recommended dose for Phase II/IIIstudies is 400 mg/d of imatinib with 0.25 µg/kg/wkPEGIntron. Grade 1/2 adverse events were common andincluded flu-like symptoms/fatigue (45%), increasedliver transaminases (transient) (55%), musculoskeletalpain (39%), edema (33%), headache (33%), and nausea(33%). Grade 3/4 adverse events were rare, with febrileneutropenia and joint/muscle pain experienced by 6%of patients. These data indicate that the combination ofimatinib plus interferon can be administered safely, butwhether improved results can be obtained will require alarge, prospective, randomized study.

Imatinib in combination with Ara-CIn vitro data have also demonstrated additive or syner-gistic antiproliferative effects for the combination ofimatinib with Ara-C using Bcr-Abl-positive cell linesand in colony-forming assays using CML patientsamples.32,37,38 Based on these data, Phase I/II studies ofthis combination are in progress.

A Phase I study of the combination of imatinib pluslow-dose Ara-C was initiated in CML chronic phasepatients who failed interferon.39 Twenty-two patientswere enrolled in this study comprising 4 cohorts. Imatinibwas given daily at 400 or 600 mg and Ara-C was admin-istered on days 14 through 28, with cycles repeated ev-ery 28 days. In this study, the maximally tolerated dosewas 400 mg of imatinib daily, with 20 mg/m2 of Ara-Cgiven for 2 out of every 4 weeks. Myelosuppression atthe highest dose level necessitated dose reductions in allpatients such that no patient remained on treatment with600 mg plus 20 mg/m2 of Ara-C. With a median dura-tion of follow-up of 300 days, the complete hematologicresponse rate was 86%, and the major cytogenetic re-sponse rate was 32%. This compared favorably with theresults reported in Phase I and II studies of imatinib aloneand served as the impetus for a Phase II study of thiscombination in newly diagnosed patients.

A Phase II study was initiated in France to assessthe tolerability and the efficacy of imatinib in combina-tion with Ara-C. Imatinib was administered at a dailyfixed dose of 400 mg in combination with Ara-C at 20mg/m2 on days 14 through 28, with cycles repeated ev-ery 28 days. From June to August 2001, 30 previouslyuntreated CML chronic phase patients within 6 monthsof diagnosis were recruited. Median age was 48 years(range, 22-81 years). After 6 months of treatment, 24/30 (80%) patients had achieved a major cytogenetic re-

Hematology 2002 117

sponse; 17 (57%) achieved a complete cytogenetic re-sponse (Table 5; F. Guilhot, personal communication).The dose of Ara-C was reduced to a median of 77% to91% of the expected dose (range, 48%-108%), and themedian dose of imatinib was 400 mg/d, demonstratingthat the dose of 400 mg was not affected by the additionof low dose Ara-C. Grade 3/4 nonhematologic toxici-ties in this study included nausea (1.1%), skin rashes(2.1%), abdominal pain (1.5%), and weight increase(1.6%). Common grade 1/2 adverse events included nau-sea (51%), muscle cramps (38.7%), periorbital edema(27.6%), vomiting (22%), diarrhea (20.3%), skin rashes(17%), weight increase (16.7%), myalgias (13.9%),arthralgias (12.6%), and dyspepsia (13.5%). Grade 3/4neutropenia and thrombocytopenia occurred in 35% and20% of patients but was rarely associated with infec-tious or bleeding complications.

Based on the Phase II data with higher doses ofimatinib and the combinations of imatinib with inter-

feron and cytarabine, a prospective, randomized PhaseIII trial has been designed (SPIRIT). This study willcompare standard therapy for newly diagnosed chronicphase CML patients treated with 400 mg/d of imatinibto higher dose imatinib therapy versus imatinib at 400mg/d with interferon versus 400 mg/d of imatinib withlow-dose cytarabine. This study is expected to be acti-vated by the end of 2002.

Other CombinationsIn vitro combinations of imatinib with various antileu-kemic agents have been investigated and are summa-rized in Table 7. These studies used various Bcr-Abl-positive cell lines, and some studies also reported oncolony-forming assays using primary patient cells. Aconsistent observation from these studies is that increasedsynergy is observed at higher levels of Bcr/Abl kinaseinhibition.38,40 The implication of this observation is thatfull doses of imatinib might be required to achieve opti-mal therapeutic responses in combination regimens.

In addition to the compounds listed in Table 7, manynovel agents have shown activity against Bcr-Abl-ex-pressing cell lines, and their activity in combination withimatinib is summarized in Table 8. More details on theactivity of these agents are contained in Section II andin a review by La Rosee et al.41

Table 7. Summary of conventional chemotherapeutic agentsthat have been tested in combination with imatinib in vitro.*

Combined Drug Synergy Additivity Antagonism

Ara- C �

Carboplatin �

Corticosteroids �

Etoposide �

Idarubicin �

Interferon (IFN) � �

Mafosfamide �

Mitoxantrone �

Treosulfan �

Vincristine �

Nimustine hydrochloride �

Busulfan �

4-OH-cyclophosphamide �

Daunorubicin �

Doxorubicin �

Fludarabine �

Gemcitabine �

Taxotere �

Thiotepa �

Cladribine �

Hydroxyurea � �

Methotrexate �

Topotecan �

*Included are all published results in the studies of Topaly et al,38

Thiesing et al,32 and Kano et al.34 Joint action studies wereperformed using myeloid or lymphoid chronic myelogenousleukemia (CML) blast crisis cell lines or using Bcr-Abl-negative celllines with engineered Bcr-Abl expression.

Table 8. Summary of novel antileukemic agents tested incombination with imatinib in vitro.

Agent Molecular Mechanism(s) Refs.

17-AAG hsp90 chaperone function inhibition 53

Adaphostin tyrosine kinase inhibition 50

AG490 tyrosine kinase inhibition 51

As2O3 induction of mitochondrial damageBcr-Abl downregulation 55,56

Bestatin, actinonin aminopeptidase inhibition 64

Decitabine DNA hypomethylation 40

Homoharringtonine inhibition of protein synthesis 34

Leptomycin nuclear entrapment of Bcr-Abl 65

PD184352 MAPK kinase inhibition 66

Wortmannin, PI3-kinase inhibition 67LY294002

PS341 proteasome inhibition 68

SCH66336, farnesyl transferase inhibition 53,54L-744,832

TRAIL induction of apoptosis 69

Trichostatin A histone deacetylase inhibition 40

Abbreviations: As2O3, arsenic trioxide; hsp90, heat-shock protein90; PS341, proteasome inhibitor 341; MAPK, mitogen-activatedprotein kinase, 17-AAG, Allylamino-17-demethoxygeldanamycin;TRAIL, tumor necrosis factor α-related apoptosis-inducing ligand.

118 American Society of Hematology

Resistance to ImatinibAlthough many of the above combinations will be testedin clinical trials based on single agent activity, more ra-tional combinations would be based on known mecha-nisms of resistance. In the largest studies of resistanceor relapse, several consistent themes emerge. In patientswith primary resistance, that is, patients who do not re-spond to imatinib therapy, Bcr-Abl-independent mecha-nisms are most common.42 In contrast, the majority ofpatients who relapse on therapy with imatinib reactivatethe Bcr-Abl kinase. In these studies, greater than 50%and perhaps as many as 90% of patients with hemato-logic relapse have Bcr-Abl point mutations in at least 13different amino acids scattered throughout the Abl ki-nase domain (Figure 1).42-47 Other patients have ampli-fication of Bcr-Abl at the genomic or transcript level.42

It should be noted that the studies described aboverepresent a minority of CML patients. Most of the pa-tients described had advanced phase disease. The ma-jority of patients diagnosed with CML will be in thechronic phase, most will obtain a complete cytogeneticresponse with imatinib, and very few have relapsed.However, only a minority attain a molecular remission.It is assumed that if these patients relapse, similar mecha-nisms will be operative. However, the more pressingquestion involves the mechanism of molecular resistance;that is, why do residual leukemia cells persist? One pos-tulated mechanism is that quiescent stem cells may beinsensitive to imatinib,48,49 suggesting that managementof this group of patients will differ substantially frompatients with relapses.

Regardless, in relapsed patients, the Bcr-Abl kinaseremains a good target. Abl kinase inhibitors with speci-ficity that differs from imatinib have already been syn-thesized.50-52 Whether or not these compounds will in-hibit some or all of the Bcr-Abl mutations that have beendescribed needs to be determined. It is conceivable thatseveral inhibitors, analogous to cocktails of protease in-hibitors for human immunodeficiency virus (HIV),would be necessary and that the appropriate inhibitorswould be chosen based on the molecular profile of mu-tations present in individual patients. Given that Bcr-Ablkinase activity has been reactivated in relapsed patients,it might also be useful to target downstream signalingpathways, such as Raf/MEK/ERK, PI-3 kinase, AKT,or ras. For example, two groups recently reported in vitrosensitivity of imatinib-resistant Bcr-Abl-positive celllines to a farnesyl transferase inhibitor.53,54 Moreover,Hoover et al observed that this compound sensitized cellsto imatinib, even imatinib-resistant cell lines.54 Alterna-tively, strategies to decrease Bcr-Abl protein levels byusing agents such as geldanamycin, 17-AAG, or arsenictrioxide might be useful.53,55,56

What Are the Goals of Imatinib Therapy andHow Should Treatment Failure Be Defined?

The simplistic aim of all leukemia treatment is to curethe patient without undue toxicity. Whether imatinib canachieve this or indeed improve survival is, at the time ofthis writing, still uncertain. Hemopoietic stem cell trans-plantation is the only proven curative treatment for CML,but the procedure is only available to one third of pa-tients and carries significant risks.57-59 Given the unprec-edented rates of cytogenetic response that have beenobserved with imatinib, it seems reasonable to expectthat improvements in survival will be seen, but only timewill tell. The outcome of patients achieving major cyto-genetic responses on interferon therapy is certainly im-pressive, but even in patients who achieve a completecytogenetic response, long-term leukemia-free survivalis not guaranteed, and responses can be lost. Data fromthe combined European groups60 suggest that other prog-nostic factors are important in predicting outcome incomplete cytogenetic responders, and presumably thisreflects the biological heterogeneity of the disease.

Another issue is whether molecular remissions arean appropriate goal. In patients undergoing allogeneicstem cell transplantation, the majority attain this land-mark. In some patients who have low but stable levels ofresidual detectable Bcr-Abl transcripts, it is possible thatallo-immunity prevents relapse. However, for patientstreated with imatinib to minimal residual disease, thereis no allo-immunity. Unless imatinib can maintain pa-tients in this state, relapse is inevitable. Again, predic-

Figure 1. Schematic of point mutations in the Abl kinasedomain.

The Abl kinase domain, from amino acid 240 to 500, is shown withthe ATP binding domain (P), the catalytic domain (C) and theactivation loop (A). The numbers below the kinase domain areamino acids that are mutated in patients who relapsed on therapywith imatinib. The vertical lines above the kinase domain indicatethe number of times each amino acid has been found to be mutatedas compiled from Shah et al,43 Hochhaus et al,42 Branford et al,44

von Bubnoff et al,45 and Hofmann et al.47 Adapted from Shah etal.43

Hematology 2002 119

tions as to the likelihood of this occurring are impos-sible, but it seems likely that if patients have residualleukemia, they are destined to relapse if imatinib is dis-continued. Thus, if nontransplant therapy is to competewith allotransplant, then molecular remissions will haveto be the goal of therapy, unless imatinib is able to pro-long the chronic phase of the disease indefinitely. Asthe data from the initial studies mature, the answers tothese questions should be forthcoming.

So, should we be making efforts to define “failure”of imatinib in order to tailor disease management forthe individual? This decision is particularly relevant inyoung patients with viable allografting options who havechosen a trial period of imatinib to see what happens.Should such a patient be directed toward allografting ifthere has been no complete cytogenetic response after 6or perhaps 12 months, or is it reasonable to continueregardless? It is evident that rates of allografting in CMLhave dropped off recently,61 but we must be very cau-tious about abandoning a known curative therapy that, inyoung patients with a low-risk sibling option, arguably hasan acceptable toxicity, given the prospect of cure.

There is no consensus definition of “failure” or per-haps more precisely “inadequate treatment response” toimatinib therapy. As few newly diagnosed patients haverelapsed, it is perhaps too soon to develop algorithmswith parameters that consider failure of imatinib therapy.These parameters could include failure to achieve a cy-togenetic response or molecular remission after a speci-fied period of time. In examining relapses in later stagechronic phase patients who failed therapy with interferon,one group has suggested that failure to achieve a majorcytogenetic response at 6 months is a poor prognosticfactor.62 A second group has demonstrated that patientswith a major cytogenetic response are less likely to re-lapse and that most patients who will achieve a majorcytogenetic response will do so within the first 6 monthsof therapy.63 Specifically, if patients were greater than65% Ph positive at 6 months, they had a 10% chance ofobtaining a major cytogenetic response at 1 year (95%confidence interval 0-30%). However, some patients didrespond at later times, presumably because of dose es-calation of imatinib. In both studies, there were largenumbers of patients with ongoing complete hematologicresponses, but no cytogenetic response. As noted, thesepatients have a higher risk of relapse, but they are still aminority. Further, 84% of newly diagnosed patients canbe expected to achieve a major cytogenetic response at1 year. Thus, for the majority of patients, other param-eters may need to be investigated.

Attempts have been made to suggest treatment al-gorithms for newly diagnosed patients,59 but these areoften difficult to utilize in the clinic, as increasingly pa-

tients have strong views of their own that may be con-trary to an ideal algorithm. In our practice, we haveadopted somewhat of a compromise position, especiallyfor younger patients who otherwise would have beenconsidered good candidates for allo-transplants but whowish to avoid transplantation. Patients are offeredimatinib as a single agent or are enrolled in a clinicaltrial of a combination of imatinib with either interferonor cytarabine. Regardless, if a patient might be a trans-plant candidate, we recommend HLA typing of the pa-tient and appropriate family members. If no siblingmatches are found, we recommend a preliminary donorregistry search to determine if the patient will be easy ordifficult to match. This is done so that patients knowtheir options in advance, as the availability or lack ofavailability of a donor may factor into decision-making.If patients fail to achieve a complete hematologic re-sponse at 3 months or are > 65% Ph positive at 6 months,they are encouraged to reconsider transplantation. How-ever, the majority of patients are expected to achieve acomplete cytogenetic response. When a complete cyto-genetic response is achieved, we switch to monitoringBcr-Abl transcript levels with quantitative RT-PCR. Aslong as this value is stable or improving, we are com-fortable continuing therapy. However, if this value risessignificantly, patients are again encouraged to reconsiderallo-transplantation. Our hope is that by monitoring forearly relapses, we can transplant patients with minimalresidual disease. Further, we hope that transplanting pa-tients with minimal residual disease will result in a su-perior outcome compared with patients transplanted withactive disease and that this will offset any delay in per-forming the transplantation. Unfortunately, there are nodata to support this seemingly common sense approach,and close, careful monitoring of these patients will be nec-essary to determine whether this approach is advisable.

II. NOVEL THERAPIES FOR

CHRONIC MYELOGENOUS LEUKEMIA

Jorge Cortes, MD*

The introduction of imatinib mesylate to the therapeuticarmamentarium has changed the approach to CML. Aswe continue to gather more information from the ongo-ing trials with imatinib, it has become the building blockin the treatment of CML. It is also evident that there is a

* M.D. Anderson Cancer Center, 1515 Holcombe Blvd., Box428, Houston, TX 77030

Dr. Cortes has received research support from HGS, Janssen,Schering Plough, ChemGenex, and Novartis.

120 American Society of Hematology

need to continue to investigate new therapies for CMLas we discover patients who become resistant to therapywith imatinib. Molecular remissions are achieved in onlya minority of patients treated with imatinib alone. Thefocus now is to investigate therapeutic modalities withspecific targets that could eventually lead to a sequen-tial blockade of intracellular pathways and lead to in-creased rates of complete molecular remissions and,eventually, cure from CML. This section will cover novelapproaches currently being investigated in CML; anoutline of these options is presented in Table 9.

TroxacitabineTroxacitabine is a novel nucleoside analogue that dif-fers from other dideoxycytidine analogs by being in theL-configuration. L-enantiomers of nucleosides wereoriginally not considered antineoplastic agents, as theywere thought to be poor substrates for activating en-zymes. Some of these L-enantiomers (e.g., lamivudine)were found to be potent antiviral agents, and others werethen investigated as antineoplastic agents. Troxacitabinehas shown antineoplastic activity against several solidtumor cell lines and animal models.1 It has also shownsignificant activity in patients with relapsed acute my-eloid leukemia (AML). A Phase I study was recentlyconducted in patients with acute leukemias and CML inblast phase (BP). Evidence of activity was found in AMLand myelodysplastic syndrome (MDS), and the one pa-tient with CML in BP treated on this study achieved asecond chronic phase (CP).2 In a recent Phase II study,17 patients with CML in myeloid BP were treated withthe dose identified in the previous study.3 Six of thesepatients had failed prior therapy with imatinib, and 9were in the second or later relapse. Six of 16 evaluablepatients (37%) returned to a CP.3 One patient relapsedafter 20 months, 1 died in second CP from sepsis, and 4continue in second CP after a follow-up of 2 to 11months. Further investigation of troxacitabine in CMLBP is warranted.

HomoharringtoneHomoharringtone (HHT) is a plant alkaloid derived froman evergreen tree. The antileukemia activity of HHT inAML has been known for several years, and more re-cently there has been suggestion of activity also in acutepromyelocytic leukemia (APL) and MDS. Significantactivity has been reported in CML. Patients in late CPtreated with this agent after having failed IFN-α-basedtherapy and with a median time from diagnosis of 3 yearshad a CHR rate of 67% and a cytogenetic response(CGR) rate of 33% (major cytogenetic response [MCR]in 15%) (Table 10).4 Similar results were obtained us-ing a combination of HHT and low-dose cytarabine

(CHR 72%, CGR 32%, MCR 15%) with a suggestion ofimproved survival with the latter combination comparedto single-agent HHT.5 Visani et al investigated the in vitroeffect of HHT in leukemia cells from patients with CMLin different stages.6 A dose-dependent growth inhibitionand apoptosis were documented in CP samples. Syner-gistic cytotoxicity was also documented with HHT incombination with IFN-α, Ara-C, and IFN-α + Ara-C.6

On the basis of this synergy, HHT has been combinedwith IFN-α, and with IFN-α and ara-C in treating pa-tients with early CP CML. The combination of HHT andIFN-α in patients in early CP CML resulted in an MCRrate of 27% with a significantly shorter median time toachievement of an MCR compared with IFN-α alone(27% at 6 months with HHT + IFN-α vs 11% with IFN-α).7 In addition, the median daily dose of IFN-α in thefirst 12 months of therapy with the combination was 2.4MU/m2 compared to 5 MU/m2 with IFN-α alone. Theuse of a lower dose resulted in a significant reduction inIFN-α-related toxicities.7 Available now are new formu-lations of HHT that allow for subcutaneous administra-tion of this agent. HHT is also synergistic with imatinibin vitro.8 Therefore, HHT remains an attractive thera-peutic option for patients with CML after imatinib fail-ure or in combination with imatinib. Such clinical stud-ies are being developed.

DecitabineDNA methylation is an epigenetic phenomenon restrictedin mammalian DNA to CpG islands. CpG methylationresults in altered chromatin organization, leading to re-pression of gene transcription. Hypermethylation ofregulatory genes involved in cell proliferation and dif-

Table 9. New therapies in chronic myelogenous leukemia.

Family Examples

Nucleoside analogues TroxacitabineClofarabine

Plant alkaloids Homoharringtone

Hypomethylating agents Decitabine

Farnesyl transferase inhibitors R115777 SCH66336

Arsenicals Arsenic trioxide

Proteasome inhibitors PS-341

Antiangiogenic agents Bevacizumab

Peptide vaccines Bcr/Abl junction peptidesPR1

New kinase inhibitors AG957AG490Adaphostin

New formulations of Polyethylene glycol (PEG)-interferon-α interferon

PegasysAlbuferon

Hematology 2002 121

ferentiation is a common phenomenon in cancer.9 Theavailability of agents that inhibit DNA methyltransferaseactivity have made methylation an active area of trans-lational research. Hypermethylation of the Pa promoterof ABL1 has been reported, with increasing levels ofmethylation in advanced stages of CML,10,11 and in-creased methylation may have adverse prognostic im-plications.12 Although the biologic consequences of thismethylation are not well understood, hypomethylatingagents have been investigated in CML. Both 5-azacytidine (AZA) and 5-aza-2′-deoxycytidine (decita-bine, DAC) are potent DNA methylation inhibitors andhave shown significant antileukemic activity in myeloidmalignancies, including AML and MDS.9 In CML, theseagents have been mostly used in the accelerated phase(AP) and BP. AZA in combination with etoposide ormitoxantrone has resulted in responses in 25% to 60%of patients treated in several studies. DAC has been usedmostly as a single agent in CML. In 31 patients withmyeloid BP, 2 achieved a CR and 6 had objective re-sponses (hematologic improvement or back to chronicphase), for an overall response rate of 26%.13 In AP, 9 of17 (53%) patients treated had an objective response.14

These studies used a dose of DAC of 50 to 100 mg/m2

over 6 hours every 12 hours for 5 days. At these highdoses, DAC induced DNA synthesis arrest with a cyto-toxic effect. At lower doses, cytotoxicity has been mini-mal, and gene-specific hypomethylation is observed. Astudy investigating the minimal effective dose of DACwas recently reported. Significant activity with minimaltoxicity was reported in patients with AML, MDS, andCML, with the most effective dose being 15 mg/m2 dailyfor 10 days.15 This schedule is being investigated in pa-tients with imatinib-resistant CML in CP, AP, and BP.The potential synergy of DAC with imatinib will also bestudied.

Farnesyl Transferase InhibitorsOne of the best-recognized downstream events result-ing from the tyrosine kinase activity of Bcr/Abl is theactivation of Ras. Ras is synthesized as an inactive pro-tein in the cytoplasm, which is activated through post-

translational changes consisting of a prenylation processthat allows attachment to the cellular membrane. Thisprocess is mediated most prominently by farnesyl trans-ferase (FTase), and alternatively through geranylgeranylprotein transferase.16 Thus, one approach to inhibit Rasactivation is to inhibit FTase activity. Several FTase in-hibitors (FTI) are under clinical development and haveshown activity in some solid tumors and hematologicmalignancies, including AML and MDS.17-19 In CML,the activity of FTI has been demonstrated in a modelusing Bcl/Abl-transformed BaF3 cells.20 SCH66336, anFTI, induced a dose-dependent inhibition of colony for-mation and proliferation of transformed cells; colonyformation of primary cells from patients with CML wasalso inhibited. Mice treated with SCH66336 after injec-tion of Bcr/Abl-BaF3 cells survived for up to 12 months,whereas all mice treated with the vehicle only died within1 month. Similar results have been reported in a Bcr/Abl-positive ALL mouse model.21 SCH66336 has beenshown to inhibit the proliferation of imatinib-resistantBcr/Abl positive cell lines and the colony formation ofcells from imatinib-resistant CML patients.22 Further-more, SCH66336 may sensitize imatinib-resistant cellsto imatinib-induced apoptosis.22 The results of a pilotstudy using a different FTI, R115777, were recently re-ported. Twenty-two patients with CML (10 in CP, 6 inAP, and 6 in BP) were treated with R115777 at a dose of600 mg PO BID. Ninety-one percent of the patients hadreceived prior therapy with IFN-α, and 77% withimatinib. Seven patients (32%) (6 in CP and 1 in AP)had a CHR (n = 5) or partial hematologic response (n =2); 4 of these patients also had a minor CGR.23 Responseswere usually transient, with a median duration of 9.4weeks. In a Phase I study with R115777, 35 patientswith high-risk acute leukemias or CML in BP weretreated with escalating doses of R115777. Ten of 24evaluable patients (29%) responded (8 partial remission[PR], 2 CR). Among 3 patients treated for CML in BP,the 2 patients who had the Philadelphia chromosomeresponded. SCH66336 is currently being investigated inthis setting. Combination studies based on the in vitrosynergy of these agents with STI are also ongoing.

Table 10. Response criteria in chronic myelogenous leukemia (CML).

Response Definition

Complete hematologic response Normalization of the peripheral blood with WBC <10 x 109/L with no immature forms, normalplatelet count, and disappearance of all signs and symptoms of the disease

Partial hematologic response As above, but persistent immature peripheral cells (blasts, promyelocytes, myelocytes), persistentsplenomegaly or thombocytosis, but ≥ 50% reduction.

Complete cytogenetic response 0% Ph+ metaphases on cytogenetic analysis

Partial cytogenetic response 1-35% Ph+ metaphases on cytogenetic analysis

Minor cytogenetic response 35-95% Ph+ metaphases on cytogenetic analysis

122 American Society of Hematology

Arsenic TrioxideArsenicals were used in the treatment of CML manyyears ago. Newer and safer formulations of arsenicalshave resurrected interest in these agents. Arsenic triox-ide (As

2O

3) is an agent with significant clinical activity

in APL. Although the mechanism of action is still beinginvestigated, there is evidence suggesting downregulationof antiapoptotic proteins in APL cell lines after expo-sure to As

2O

3. A similar effect has been reported in many

other hematologic malignancy models.24,25 Incubation ofCML cell lines with concentrations of As

2O

3 that are

clinically achievable resulted in dose-dependent growthinhibition and induction of apoptosis.26 A significant re-duction in the expression of the Bcr-Abl protein was alsoobserved but did not appear to interfere with the kinaseactivity of Bcr-Abl.26,27 As

2O

3 and imatinib mesylate are

synergistic in growth inhibition and induction ofapoptosis in CML cell lines.28,29 Because of this reportedsynergy, clinical trials investigating this combination areongoing.

Proteasome InhibitionThe ubiquitin-proteasome pathway is the principal in-tracellular pathway responsible for the degradation ofproteins.30 Multiple proteins with a variety of cellularfunctions are substrate for this pathway, including manythat participate in cell cycle regulation and tumor growth.Thus, proteasome inhibitors are being investigated as anew approach for anticancer therapy. Although the spe-cific mechanism through which these agents exert theirantineoplastic activity is unclear, inactivation of NF-κBappears to play a prominent role. NF-κB is inhibited inthe cytoplasm through binding to IκB, a substrate forproteasomes.31 In CML, Bcr-Abl activates NF-κB-de-pendent transcription, and NF-κB may be required forBcr-Abl-mediated transformation.32,33 Proteasome inhi-bition with N-carbobenzoxy-L-leucyl-L-leucyl-norvalinal (LLnV) decreased the expression of the Bcr/Abl protein in K562 cells with no change in the expres-sion of p145Abl.34 The molecular mechanisms leading tothis decreased expression of Bcr/Abl protein are unclear,but the reduction led to caspase activation and apoptosis.PS-341 is a potent and selective inhibitor of proteasomesthat has shown activity against a broad range of humantumor cells and is currently being investigated in vari-ous types of malignancies. Significant clinical activityhas been demonstrated in multiple myeloma. In vitro,PS-341 induced significant growth inhibition andapoptosis of several Bcr/Abl positive cell lines, as wellas both imatinib-sensitive and -resistant cell lines.35 Clini-cal studies in imatinib-resistant patients are ongoing.

Antiangiogenic AgentsThere is growing evidence of the significance of angio-genesis in leukemia. Elevations of plasma vascular en-dothelial growth factor (VEGF) levels have been reportedin patients with CML compared to controls (76.3 vs 26.7pg/mL) and were the highest among all leukemias tested(including chronic lymphocytic leukemia [CLL], MDS,AML, ALL, and chronic myelomonocytic leukemia[CMML]). Plasma levels of bFGF, HGF, and TNFα werealso elevated.36 There is also a significant increase inbone marrow vascularity as determined by the numberof blood vessels and area of vascularity. Patients withCML had a median of 21.4 blood vessels (11.2 for con-trols, P = 0.003), and the relative vascular area was 6.2%(compared to 2.8%, P = 0.02). These values were thehighest among all leukemias.36 VEGF has also beenshown to suppress the function of dendritic cells,37 whichhave been shown to stimulate autologous antileukemiaT-cell response, particularly in CML. Antibodies toVEGF may enhance the function of dendritic cells.38

Recently, increased cellular expression of VEGF hasbeen associated with shorter survival in a multivariateanalysis in 148 CML patients in CP.39 Therefore, sup-pression of VEGF might prove beneficial in CML, pos-sibly through enhancing a specific anti-CML immunereaction. Specific anti-VEGF monoclonal antibodies(i.e., bevacizumab) are currently in clinical trials in pa-tients with CML.

Peptide VaccinesThe development of a specific immune therapy has beenof significant interest for some time. Evidence of theantileukemia effect from the graft-versus-leukemia ef-fect has triggered investigations into approaches to de-velop specific antileukemic immunity in a more specificand less toxic way. One such approach currently in clini-cal trials is the use of leukemia-specific peptides to stimu-late a T-cell-mediated antitumor effect. The chimericp210bcr-abl protein generated by the fusion of the Abl andBcr genes is a candidate peptide because it is tumor spe-cific and it contains a sequence of amino acids not ex-pressed in normal cells. Because T cells do not recog-nize intact proteins but rather peptide fragments of 8 to12 amino acids in length presented through the majorhistocompatibility complex (MHC), small peptides de-rived from the fusion region of p210bcr-abl have been in-vestigated. The immunogenicity of such peptides hasbeen documented in animal models, where immuniza-tion of mice can elicit a peptide-specific CD4+ T-cellresponse.40 These T cells recognized and proliferated inresponse to the intact p210bcr-abl protein but not other pro-teins with a different junction sequence (e.g., p185bcr-abl)or unrelated proteins. Several peptides have been iden-

Hematology 2002 123

tified that can bind to class I and II HLA molecules andelicit HLA-restricted cytotoxicity in vitro.41 Using a mix-ture of these peptides, Pinilla-Ibarz et al vaccinated pa-tients with Ph-positive CML in CP in PR or CR aftertherapy with IFN-α or hydroxyurea.42 Patients received5 vaccinations over 10 weeks with 1 of 4 different doses.While none of the 12 patients treated had delayed-typehypersensitivity (DTH) before therapy, 2 developed sig-nificant reactivity after vaccination. Similarly, 3 patientsdeveloped peptide-specific T-cell proliferation not ob-served before therapy. All responses occurred at 1 ofthe 2 highest doses tested. No significant cytotoxic orantibody response was obtained in any patient. No sig-nificant toxicity was observed. One patient had a tran-sient molecular response and one a transient partial cy-togenetic response.42 In a Phase II trial using this ap-proach, all 12 evaluable patients developed a DTH andCD4 proliferative response after vaccination and 6 hadIFN-γ release by CD4 ELISPOT.43 Three patients (allreceiving IFN-α) had a decrease in the percentage ofPh-positive metaphases, and 1 patient vaccinated for re-currence after bone marrow transplant (BMT) becametransiently PCR negative.43

Another peptide being investigated is PR1. Protein-ase 3 is a serine protease that is induced during differen-tiation, stored in azurophilic granules, and overexpressedin myeloid leukemias. PR1 is a 9-amino acid peptidederived from proteinase 3 with a binding motif for HLA-A2.1.44 PR1 was used to generate primary cytotoxic Tlymphocytes (CTL) in vitro, and these CTL demonstratedsignificant lytic activity against fresh leukemia cells frompatients with HLA-A2.1+ CML or AML.44 CTL gener-ated through stimulation with PR1 inhibited colony-form-ing units from HLA-A2.1+ CML patients but not HLA-A2.1+ bone marrows or HLA-A2.1– CML, demonstrat-ing the specificity of these lymphocytes.45 PR1-specificCTL have been identified in 11 of 12 patients who re-sponded to IFN-α and in 6 of 8 who responded to allo-geneic BMT but were undetectable in untreated patients,those treated with chemotherapy, or those who failed torespond to IFN-α.46 In a Phase I study, patients have beenreceiving escalating doses of PR1 together with granu-locyte-macrophage colony-stimulating factor (GM-CSF). Preliminary data showed the development of asignificant increase in PR1-specific CTL that correlatedwith a molecular response in 1 patient with APL, but noresponse was observed in 1 patient with CML.47 Thisvaccine is currently being investigated in CML and othermyeloid leukemias.

New Kinase InhibitorsThe major clinical success of imatinib in CML has trig-gered increased enthusiasm for the investigation of other

agents that could inhibit tyrosine kinases. An alternativeapproach to tyrosine kinase inhibition is the use of mol-ecules that prevent the binding of peptide substratesrather than ATP binding. A family of such agents, calledtyrphostins, has been found to inhibit the Bcr/Abl ty-rosine kinase. A member of this family, AG957, can in-hibit Bcr/Abl autophosphorylation but is not specific forBcr/Abl and is not as active as imatinib.48,49 Adaphostinis the adamantyl ester of AG957 with greater in vitropotency than AG957.48 Imatinib-resistant cell lines aresensitive to adaphostin, and a combination of these 2agents may be synergistic.50 Inhibitors of other kinasesare also being investigated. AG490 is a potent inhibitorof JAK2, another tyrosine kinase, and is synergistic withSTI571. Src kinase inhibitors are also being investigated.Although these molecules are earlier in their develop-ment, they may prove valuable in the near future eitherin combination with imatinib or alone for patients whohave developed resistance to imatinib.

IFN-αααααThe efficacy of IFN-α in CML is unquestionable. Pa-tients treated in early chronic phase CML with IFN-αachieved MCR rates of 20% to 40% and complete cyto-genetic response (CCR) rates of 5%-30%. CombiningIFN-α with cytarabine may increase the rate of CCR to25%-35%. The introduction of imatinib has changed thetherapeutic algorithm of CML. Thus, the use of IFN-αas first-line therapy in CML is likely to disappear soon.However, it is still a valuable option and in coming yearsthere will be an increasing number of patients whoseCML may have become resistant to imatinib and whowould have not been exposed to IFN-α. New formula-tions of IFN-α may improve tolerance and efficacy. At-tachment of IFN-α to polyethylene glycol (PEG) pro-longs the half-life of IFN-α, allowing a weekly admin-istration. In a Phase I study on patients with CML thatfailed to respond or were intolerant to IFN-α, signifi-cant activity was observed. Seven of 19 patients withactive disease achieved a CHR and 2 (11%) had a CCR.Seven of 8 patients treated in CHR improved their re-sponse to complete (n = 4) or partial cytogenetic response(n = 3). All 6 patients intolerant to IFN-α tolerated PEG-IFN, and 4 improved their cytogenetic response. A dif-ferent approach at sustained release formulations of IFN-α is Albuferon, composed of recombinant human albu-min fused to recombinant human IFN-α. Because themedian half-life of albumin is approximately 20 days, itprolongs the half-life of IFN-α and allows for adminis-tration every 2 weeks. This drug is currently being in-vestigated in CML.

124 American Society of Hematology

SummaryMultiple new agents are currently being developed inCML. Most of these agents are now being investigatedin early clinical trial in patients who have developed re-sistance to imatinib. Their mechanisms of action are di-verse, and many may be synergistic with imatinib. Theseagents will soon be used in different combinations, mostlikely including imatinib, with the hope of obtaining acomplete blockade of the intracellular pathways that aretriggered by Bcr/Abl. If this is successful, complete eradi-cation of disease may become a reality for the majorityof patients with CML.

III. TRANSPLANT APPROACHES FOR CML

Jerald Radich, MD*

The exciting success of imatinib has dramatically re-vised the way clinicians think about treating CML. Al-though blood or marrow transplant (BMT) is thus farthe only “curative” approach in CML, the prolongedcytogenetic remissions seen with imatinib therapy inchronic phase patients make medical therapy a tempt-ing first-line approach, even in patients who may be finetransplant candidates. Inevitably, physicians and patientsmust now consider the option of imatinib against theoption of a transplant. In making this comparison, theymust weigh what is known about the curative potential

for BMT and imatinib with consideration of the poten-tial toxicities of each therapy. Complicating the issue isthat while the impressive results of imatinib have de-servedly seized the headlines in CML therapy, transplan-tation results for CML have also improved considerablyover the years. This brief review will attempt to definethe current strategies and outcomes following transplan-tation (Table 11).

The widespread application of BMT has been lim-ited by donor availability and the high toxicity of theprocedure in older patients, which limits the age of eli-gibility to less than 55-65 years at many centers. Ongo-ing advances in alternative donor sources (including un-related donors and, to a lesser degree, cord blood), moreaccurate molecular HLA typing, and less toxic regimens,including reduced intensity or nonmyeloablative trans-plants, are broadening the potential use of this treatmentmodality.

Matched Related Allogeneic TransplantsSeveral factors influence the outcome of allogeneic re-lated transplants. The principal determinant of survivalis the phase of disease at the time of transplantation. Asa rule of thumb, survival decreases by half as one mi-grates from chronic phase, through accelerated phase,to blast crisis (Figure 2).1,2 Patients transplanted in re-mission after blast phase (also known as the secondchronic phase) tend to have survival outcomes similarto accelerated phase cases. Survival in chronic phase isquite good, ranging from 60-80% at 5 years.1-7 The useof chemotherapy preparative regimens (as opposed tototal body irradiation–based [TBI-based] regimens) andimproved supportive therapy (including cytomegalovi-rus and fungal prophylaxis) may have improved survivaleven more. For example, both an initial and a follow-upreport of chronic phase CML patients randomized to apreparative regimen of TBI and cytoxan or busulfan andcytoxan (BU/CY) demonstrated similar effectiveness ofboth regimens.3,8 Subsequently, a pharmocologic assayfor blood busulfan levels revealed that patients with lev-els > 900 ng/mL had better survival, and less relapse,compared to patients with a level < 900 ng/mL.9 This

* Fred Hutchinson Cancer Research Center, 1100 FairviewAve., N., #D4-100, Seattle, WA 98104

Dr. Radich is supported by grants from the National CancerInstitute, the Friends of Jose Carreras Foundation, and contractlaboratory support from Novartis.

Acknowledgements: I thank Dr. Derek Stirewalt and ReginaldClift for their careful reading and helpful comments. Becauseof the size constraints of this review, important work frommany colleagues may have been omitted. I apologize that Icould not mention all of the contributions they have made tothis field.

Table 11. Survival following different transplant approaches.*

Chronic, % Accelerated, % Blast, % Comments

Related 60-80 40-50 10-20 Age and Dx→BMT interval less important than in URD setting

URD 50-80 20-40 5-20 GVHD the biggest hurdle to success and quality of life

Autologous 40-70 NA NA Limited role may be resurrected in the imatinib era

Non-myeloablative NA NA NA Investigational

*Five-year estimates.

Abbreviations: BMT, blood or marrow transplant; Dx, diagnosis; GVHD, graft-versus-host disease; NA, not applicable; URD, unrelated donor.

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fact has led to subsequent targeting of busul-fan in all patients to a level > 900 ng/mL (thisstrategy also allows correction for patientswith a high busulfan level to prevent poten-tial toxicity). This change in preparative regi-mens may have contributed to the apparentimprovement of survival in CML chronicphase patients transplanted using targeted BU/CY. Indeed, in 131 consecutive chronic phaseCML patients treated at our institution usinga preparative regimen of pharmacologicallytargeted BU/CY, the 3-year survival was 86%,with a relapse rate of 8%. Unfortunately, ad-vances that may have pushed progress inchronic phase patients have not been easilytranslated to accelerated and blast crisis trans-plants. Survival rates in these phases have re-mained relatively static over the past decade,with survival of accelerated phase approxi-mately 40% and blast crisis approximately 10-20%. Thus, phase continues to be the drivingforce in any decision making, weighing themorbidity of the transplant against the risk ofprogression of disease with the associatedpoorer transplant outcomes seen with ad-vanced disease.

Advancing age has been the major con-straint in transplants; in general, the rate ofregimen-related toxicity climbs with patientage.2,4 Age limits have gradually increasedover time with the introduction of new pre-parative regimens and improved supportivetherapies. The effect of age, however, maybe less now than in previous times, especiallywhen applied to chronic phase patients.Chronic phase patients transplanted with TBI-based regimens generally demonstrated anage effect, with superior outcomes in youngpatients (< 21 years of age), with steady dropoff in survival by decade of age.2 This ageeffect may have been mitigated by the use ofnon-TBI regimens (e.g., targeted BU/CY), assome recent data no longer show an age ef-fect in patients up to 65 years of age (Figure3). For patients in accelerated phase and blastcrisis there are no similar data, but given thehigh regimen-related mortality seen in thesepatients (especially in those with blast cri-sis) one would suspect that increasing agewould continue to have an untoward effecton survival.

The time from first diagnosis to trans-plantation is a risk factor for poor outcome,

Figure 2. Survival of CML patients in chronic, accelerated and blast phase.

All patients were transplanted from matched related donors.

Figure 3. The effect of age on survival.

Figure 3A shows the influence of age on matched transplants for CML chronicphase after receiving a preparative regimen of total body irradiation/ (TBI/CY).

Figure 3B shows the survival after targeted busulfan/CY (BU/CY). There is nosignificant difference in the survival of these age cohorts up to age 65.

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with longer time intervals being associated with moretoxicity and perhaps greater relapse rates.1,2,4,5 The criti-cal cutoff time is unclear. Recent data from the Interna-tional Bone Marrow Transplant Registry support a sig-nificant outcome advantage for patients transplanted <1 year from diagnosis in both the related and unrelatedsetting (Figure 4); recent data from Seattle suggest thatfor matched related donor transplant the cutoff may beat 2 years postdiagnosis.2 The etiology of this effect hasnever been entirely clear, but it is likely mainly due tothe cumulative toxicity of the conventional therapy, lead-ing to an increase in non-leukemic mortality. It is clearthat prolonged therapy with oral busulfan was associ-ated with a poor outcome,1,10 and results have been con-troversial as to the effects of prolonged interferon therapyand subsequent transplant toxicity.11-15 However, the in-teraction of time from diagnosis and outcomes has beendifficult to study adequately in the “modern age” of trans-plantation since so few patients eligible for transplanta-tion wait longer than 1-2 years to undergo the proce-dure. The long-term effect of imatinib on transplant out-comes is unknown but obviously must be determined inthe future.

Unrelated Donor TransplantsThe use of unrelated donor (URD) transplants is limitedby both the availability of donors and the increased tox-icity of the procedure compared to related transplants,largely owing to the effects of graft-versus-host disease(GVHD). Fully matched unrelated donors are now avail-able for over 50% of Caucasian patients, but unfortu-nately, the percentage of available donors remains sub-stantially lower for patients from other ethnic groups.The advent of molecular DNA assessment of HLA typ-ing has made for a rigorous and stringent selection ofunrelated matched donors, and this improvement in typ-

Figure 5. The effect of age on survival in unrelated donor(URD) transplants for chronic myelogenous leukemia (CML).

The results show a significant age effect after the age of 40. Allpatients were transplanted from a single center (Fred HutchinsonCancer Research Center) and received a standard total bodyirradiation/cytoxan (TBI/CY) preparative regimen.

Figure 4. The IBMTR experience of chronic myelogenousleukemia (CML) chronic phase patients.

The curves demonstrate both the effect of a related versusunrelated donor, and the benefit of early transplantation on survival(Figure 4 reprinted with permission from the IBMTR).

ing has translated into greatly improved transplant out-comes. Thus, in many centers, the success after URDtransplantation is comparable to that of matched relateddonor transplants.16-18

As with related donor transplants, disease phase isthe major determinant in URD transplant outcome, with3-year survival in chronic phase patients at least doublethat of patients transplanted in accelerated phase.18,19

Survival following a URD transplant for blast crisis isunusual, and indeed, most clinicians feel that patients inblast crisis should be re-induced into a chronic phasebefore a transplant is attempted, if possible. Age contin-ues to be a major determinant in the outcome of URDtransplantation, although a clear-cut breakpoint in de-termining “good” and “bad” risk groups has not beendemonstrated (Figure 5)16,17,19 In addition, time from di-agnosis to transplant is strongly associated with outcome,with a significant increase in mortality occurring whentransplant is delayed for greater than a year (Figures 5and 6). For “younger” patients, the survival results inchronic phase CML are similar for fully matched, unre-lated matched, and related transplants, especially forpatients in “good risk” groups.18 In a multicenter analy-sis of National Marrow Donor Program (NMDP) data,disease-free survival of unrelated and related transplantsfor CML chronic phase patients aged 30-40 years, trans-planted within 1 year of diagnosis, was 67% versus 57%,respectively. In addition, 2 other studies have observeda near-equivalence of disease-free survival for chronicphase CML using either a fully matched unrelated orrelated donor.16,17 Estimates of disease-free survival of >70% were found for “younger” patients (≤ 50 years ofage) transplanted within a year of diagnosis (Figure 7).17

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the chronic phase setting. The concern here is that giventhe excellent results of conventional bone marrow trans-plantation in chronic phase CML, any possible benefitsfrom PBSC may be offset by potential increase in GVHD.

A few studies suggest an advantage of PBSC in theURD transplant setting. Two studies from the Essengroup evaluating the role of PBSC in the unrelated trans-plant setting have demonstrated the superiority of PBSCversus BMSC. The first study showed that patients re-ceiving PBSC had the bcr-abl fusion transcript detectedless often compared to BMSC patients, suggesting a moredramatic graft-versus-leukemia effect in the PBSC pa-tients.21 The second study compared PBSC with BMSCin chronic phase CML and found that PBSC were asso-

Figure 7. A comparison of related and unrelated transplantsfor chronic myelogenous leukemia (CML) chronic phase.Figure 7A shows the experience from the Fred Hutchinson CancerResearch Center, and Figure 7B the experience from the Universityof Minnesota. There were no significant differences between theunrelated and related survivals.

Figure 7B reprinted with permission from Davies SM, DeFor TE,McGlave PB, et al. Equivalent outcomes in patients with chronicmyelogenous leukemia after early transplantation of phenotypicallymatched bone marrow from related or unrelated donors. Am J Med.2001;110(5):339-346..

Bone Marrow Versus Peripheral Blood Stem CellsPeripheral blood stem cells (PBSC) have emerged as apreferred source of stem cells in autologous transplan-tation, where their use has been associated with quickerengraftment and less regimen-related toxicity comparedto stem cells derived from bone marrow (BMSC). In theallogeneic transplant setting; however, the use of PBSCis more controversial. A randomized trial showed im-proved early overall survival and disease-free survivalin PBSC compared to bone marrow recipients in patientswith “high risk” leukemias (which included CML inaccelerated phase and blast crisis).20 It is unclear whetherPBSC are superior to, equal to, or worse than BMSC in

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Figure 6. Comparison of matched related and unrelatedtransplants.Chronic myelogenous leukemia (CML) chronic phase patientstreated at National Marrow Donor Program (NMDP) Centers from1988-1999. Figure 6A shows the disease-free survival for matchedrelated transplants, divided by time from diagnosis to transplant.Figure 6B shows the same effect for URD transplants.

Figure 6A reprinted with permission from Weisdorf DJ, Anasetti C,Antin JH, et al. Allogeneic bone marrow transplantation for chronicmyelogenous leukemia: comparative analysis of unrelated versusmatched sibling donor transplantation. Blood. 2002;99(6):1971-1977.

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ciated with improved survival for the PBSC group (94%vs 66%), owing mostly to a decrease in acute transplant-related mortality (5% vs 30%). Overall incidence of acuteand chronic GVHD appeared similar in the two groups.22

Last, a retrospective study of PBSC transplants comparedto a matched set of BMSC URD transplants suggestedthat PBSC and BMSC transplants had similar rates ofGVHD, relapse, and survival.23 The effects appearedsimilar in all phases of CML.

Reduced Intensity or Nonmyeloablative TransplantsInnovative approaches using reduced intensity or non-myeloablative transplants have recently been pioneeredto engender a graft-versus-leukemia effect without ex-posing the patient to the toxicity of the preparative regi-men. The first candidates for these transplants have beenpatients deemed at high risk of transplant complica-tions—that is, relatively older patients or those with com-promised organ function. These studies were performedon a variety of hematological malignancies. The proce-dure seems best suited for patients with slow-growingtumors, such as low-grade lymphomas, chronic leuke-mias, or acute leukemia in remission. This is becausethe nonmyeloablative approach is essentially a race be-tween the reconstitution of the immune system and theassociated antileukemic effect, versus the growth of themalignancy. Major, durable responses were documentedin chronic lymphocytic leukemia, myeloma, low-gradelymphomas, acute leukemia, and renal cell carcinoma.24,26

The data in CML are limited.25,26 The M.D. Andersongroup reported on 27 CML patients, of which 21 werein “transformed” state.25 Overall survival for these CMLcases at 1 year was 32%. A study by McSweeney et alused a less intense regimen and included 9 patients withCML.26 Of the 6 patients transplanted in chronic phase,4 achieved not only a cytogenetic remission but also a“molecular remission” studied by RT-PCR. Not surpris-ingly, these molecular responses lagged behind cytoge-netic remissions by several months. In the 3 acceleratedphase patients, 2 had progressive disease, yet 1 achieveda molecular remission. Graft rejection has been observedmore frequently in non-ablative than in ablative trans-plant, especially in patients with CML and MDS. This ismost likely the result of a relatively active donor im-mune system, which in CML has been spared exposureto cytotoxic and immunosuppressive therapy. Nonethe-less, the early results are promising and suggest that thenonmyeloablative approach may be quite attractive inCML, especially in chronic phase. This is not entirelysurprising, as many lines of evidence suggest that CMLis especially sensitive to immunologic effects (includ-ing the high relapse rates in syngeneic and T-cell-de-pleted transplants; the beneficial effects of donor leuko-

cyte infusions [DLI]; and the effects of interferon). How-ever, the question of whom to apply the nonmyeloablativeapproach to is problematic. Restricting the procedure toonly those patients who fail imatinib or those who havea related or unrelated donor but are too old or too ill forconventional transplantation would confine the treatmentto a very small niche, indeed. One possible approachmight be to add non-myeloablative transplants as an im-munologic adjuvant after initial “debulking” with imatinib,followed by the addition to imatinib after transplant to “mopup” any residual disease.

Autologous TransplantsAutologous transplants have the attraction of circum-venting the toxicities associated with GVHD, but alas,the curative potential may be limited by both the lack ofimmunologic effect and potential problems of contami-nation of the graft with residual leukemia cells. None-theless, several studies have suggested a potential ex-tension of the natural history of chronic phase CML af-ter autologous transplantation, with long-term survivalof > 40% after 5 years for those patients transplanted inchronic phase. As with allogeneic transplants, the suc-cess of autologous transplants is most influenced by thestage of disease at the time of transplantation. In a re-view of the experience in 8 centers, McGlave et al foundthe 3-year survival for patients transplanted in chronic,accelerated, and blast phase to be approximately 60%,30%, and 0%, respectively.27 More recently, the Hammer-smith group reported on the results of 56 patients whoreceived a busulfan-only preparative regimen.28 The 3-year survival was 76% for patients in chronic phase and30% for patients with advanced phase disease (acceler-ated, blast, or second chronic phase). However, only 2of 45 patients studied 12 months post-transplant achieveda CCR.

Although it is logical to prefer a stem cell product“purged” of CML cells, is it essential? On one hand,Carella et al treated 30 patients with high-dose chemo-therapy and collected PBSC; after transplantation, pa-tients received a short course of interleukin-2 and re-mained on interferon.29 Only 2 patients progressed toblast crisis. Of the 16 patients who received Ph-negativePBSC, 8 remained in CCR (thus, overall 8/30 patientsachieved CCR). No patient who received Ph-bearingPBSC products achieved a CCR. On the other hand,Meloni et al published results on 26 CML chronic phasepatients who had unpurged PBSC collected at diagnosisafter granulocyte colony-stimulating factor (G-CSF)mobilization, followed by autologous transplantation.30

All patients received interferon post-transplant. The 10-year survival of this cohort was 55%, and remarkably, 8of 26 patients achieved a CCR at some time during their

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posttransplant course. Thus, the role of purging stem cellsis still unclear, although the concept is appealing. Thecommon denominator of post-transplant immunotherapyseems important.

Given that imatinib can often produce cytogeneticremissions, the question of harvesting PBSC at the timeof cytogenetic remission, with potential use at the timeof progression, has come to the forefront. Anecdotal re-ports suggest that stem cells collected after imatinibtherapy can engraft, but the issue of engraftment is stillunanswered. After engraftment comes the issue of re-lapse. Given that interferon post-transplant may preventor delay relapse, the addition of imatinib after autolo-gous transplant may be a logical next step for study.

Minimal Residual DiseaseRelapse continues to be a major obstacle to cure in trans-plantation, increasing in frequency with advancing phaseof disease. As such, relapse occurs in ~10-20% of chronicphase transplants and up to 50% for patients transplantedin blast crisis. Risk increases with the use of T-cell deple-tion and in twin and autologous transplants, presumablyfrom a lessening of the graft-versus-leukemia effect. Itis now well established that the detection of the chimericbcr-abl mRNA transcript by RT-PCR is a powerful pre-dictor of subsequent relapse.31-44 In general, the qualita-tive test (yes versus no) is predictive of relapse “early”(6-12 months) posttransplant, and quantitative assaysincrease the precision of predicting relapse.33,35,37,40,43,44

With effective monitoring techniques and interventionstrategies, one can envision a day when grafts are ma-nipulated to decrease the risk of GVHD, and then an inter-vention (e.g., T cells, interferon) is added at the point ofdetectable residual disease. Such a strategy decreases thetoxicity of the transplant (by decreasing GVHD) whilepreserving the anti-leukemic effect of the transplant.

The early detection of minimal residual disease(MRD) allows for treatment to be initiated when the dis-ease burden is far less than at frank cytogenetic or patho-logical relapse. The underlying assumption is that suchearly treatment will translate into a greater response rate.Several examples demonstrate this concept. Cytogeneticrelapse following allogeneic transplant has effectivelybeen treated with a low dose of interferon (1-3 × 106 U/m2/d).45 In this study 14 patients were treated, and re-markably, 12/14 achieved a CCR. Of 9 patients in CCRtested for bcr-abl by RT-PCR, 4 also had evidence of amolecular remission. Eight patients (57%) remained ina durable CCR lasting up to 5 years. The treatment ofMRD with T-cell immunotherapy has been demonstratedby van Rhee et al, who treated 14 relapsed CML pa-tients (hematologic, cytogenetic, or molecular) withDLI.46 Of the 7 patients with hematologic relapse, 3 had

a CCR to DLI; however, 2 of these patients developedaplasia. Three patients with hematologic relapse and 2patients with only bcr-abl molecular positivity weretreated; all had a complete response, and there were noaplastic events. There was no clear difference in the in-cidence of severe GVHD among the 3 groups. Thus,patients treated earlier in the course of relapse had im-proved response and fewer complications. Further stud-ies of DLI administration posttransplant have also sug-gested the potential increased efficacy of treating a lowerdisease burden afforded by MRD testing.47,48

In the near future, novel therapeutic approaches maybe used to treat MRD in the post-transplant setting. Giventhe promise of early trials in treating cytogenetic ormolecular relapse, the potential use of less toxic agents(imatinib) would seem quite promising. However, it can-not be emphasized enough that any treatment of MRDpost-transplant is investigational and should take placeonly in the context of a research trial.

Molecularly Defined Risk GroupsTreatment response in CML is variable, to both non-transplant and transplant modalities. For example, inaccelerated phase CML, only a small percentage of pa-tients will obtain a complete response to imatinib or in-terferon, and approximately one half will relapse aftertransplantation. Why is there such heterogeneity in re-sponse and relapse? Why do some patients in acceler-ated phase behave like chronic phase patients (survivewithout relapse), while others behave like blast crisispatients (relapse shortly after transplant)? Presumablypart of this heterogeneity is based on the individual ge-netics of the particular leukemia. The genetic basis ofthese differences in gene expression may reside in thecontrols of the cell cycle or apoptotic pathways, or al-ternatively, genes involved in drug metabolism and drugresistance. Regardless of the specific mechanism, we hy-pothesize that different patterns of gene expression de-tected prior to transplant will be associated with differ-ent outcomes. If this proves true, we would be able topredict high- and low-risk patients prior to transplantand tailor therapy appropriately.

Much is said about the promise of tailoring indi-vidual therapy to the genetic specifics of each patient’sleukemia, but in most diseases, the effective therapeuticmodalities are so limited as to make such a promise atad optimistic. Not so in CML, as patients may enjoy aconsiderable long-term benefit, if not cure, with severalapproaches (interferon, imatinib, transplantation). Howdoes one pick the best treatment? Prognostic scores, suchas those proposed by Hasford et al49 and Sokal et al,50

are useful but do not disclose information about thepathogenesis of disease progression or disease response.

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Several studies have used DNA microarrays to studythe differences in gene expression seen from differenttypes of leukemia or different subsets of leukemia.51,52

Comparisons may be made between disease types, suchas acute lymphoblastic versus acute myeloid leukemia,or among different phenotypes within a specific disease,such as treatment responders versus non-responders.Such an approach should be quite informative in CML.For example, a comparison of gene expression differ-ences between chronic phase and accelerated phase orblast crisis may detect genes involved in the progressionof the disease; these genes could then be studied in bio-logical context, as well as serve as targets for more sen-sitive assays of disease state. In comparing a pool ofblast crisis samples to a pool of chronic phase samples,we have found approximately 500 genes significantlydifferent between the two disease states.53 Blast crisisand chronic phase patients demonstrate clear differencesin gene expression, and occasional cases occur in whichthe clinical and pathological diagnosis is quite discor-dant from the gene expression pattern (Figure 8, seeColor Figures, page 514). In addition, gene expressionstudies may give insights into the biology of response(e.g., by defining a subset of genes associated withimatinib response). Patients could be screened for thepresence of these genes, prior to therapy, to determinethe best treatment strategy. Studies in Ph+ ALL have sug-gested a defined subset of genes associated with the un-responsive imatinib phenotype.54

Of course, gene expression studies are only part ofthe biological picture, but the rapidly evolving technol-ogy to document chromosomal and DNA changes (bycomparative genomic hybridization or single nucleotidepolymorphism studies) and the rapidly evolving field ofproteomics are bound to make a mighty contribution toboth our biological understanding and our treatment al-gorithms.

ConclusionPatients and physicians face a difficult choice for treat-ment of early CML. Imatinib is a very exciting drug, buta number of unknown elements should at least give theclinician pause. First, it will take several years to estab-lish whether imatinib and its predictable spinoffs (newtyrosine kinase inhibitors, and combinations of imatinibwith old drugs) are curative. Second, it is not known ifimatinib treatment failures will return to a chronic phaseor will evolve with a more aggressive disease difficult totreat with transplantation. With frequent monitoring (e.g.,quantitative RT-PCR) one would expect to detect patientsbound to relapse early. It is also unclear what, if any,long-term side effects from imatinib might become un-masked with transplantation. As part of this reflection,

it is important to report that success in transplantationhas also improved over the past years; it remains an im-portant treatment option that should not be neglected.

While many dwell on the question of “which is bet-ter” (usually pitting imatinib versus transplantation), weshould rather rejoice that there are several effective treat-ment options in CML. One possible approach for deci-sion making (Figure 9, see Color Figures, page 515)revolves around the presence or absence of a stem celldonor. In the near future, treatment of CML may beguided by molecular studies (Figure 10, see Color Fig-ures, page 515). After initial therapy would come mo-lecular monitoring of the disease, with early detectionof disease recurrence triggering the appropriate next levelof therapeutic intervention. CML has always been theparadigm for “bench to bedside” translational research.Now, once again, the collision of new therapeutic op-tions with advances in molecular biology offers a “nextgeneration” of effective therapy for the vast majority ofCML patients, and may show the way for the future treat-ment of other malignancies.

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40. La Rosée P, Johnson K, Moseson EM, O’Dwyer ME, DrukerBJ. Preclinical evaluation of the efficacy of STI571 incombination with a variety of novel anticancer agents [ab-stract]. Blood. 2001;98:839a.

41. La Rosee P, O’Dwyer ME, Druker BJ. Insights from pre-clinicalstudies for new combination treatment regimens with the Bcr-Abl kinase inhibitor imatinib mesylate (Gleevec/Glivec) inchronic myelogenous leukemia: a translational perspective.Leukemia. 2002;16:1213-1219.

42. Hochhaus A, Kreil S, Corbin AS, et al. Molecular andchromosomal mechanisms of resistance to imatinib (STI571)therapy. Leukemia. In press.

43. Shah NP, Nicoll JM, Nagar B, et al. Multiple BCR-ABL kinasedomain mutations confer polyclonal resistance to the tyrosinekinase inhibitor imatinib (STI571) in chronic phase and blastcrisis chronic myeloid leukemia. Cancer Cell. 2002;2:117-125.

44. Branford S, Rudzki Z, Walsh S, et al. High frequency of pointmutations clustered within the adenosine triphosphate-bindingregion of BCR/ABL in patients with chronic myeloid leukemia

132 American Society of Hematology

or Ph-positive acute lymphoblastic leukemia who developimatinib (STI571) resistance. Blood. 2002;99:3472-3475.

45. von Bubnoff N, Schneller F, Peschel C, Duyster J. BCR-ABLgene mutations in relation to clinical resistance of Philadelphia-chromosome-positive leukaemia to STI571: a prospective study.Lancet. 2002;359:487-491.

46. Roche-Lestienne C, Soenen-Cornu V, Grardel-Duflos N, et al.Several types of mutations of the Abl gene can be found inchronic myeloid leukemia patients resistant to STI571, and theycan pre-exist to the onset of treatment. Blood. 2002;100:1014-1018.

47. Hofmann WK, Jones LC, Lemp NA, et al. Ph(+) acutelymphoblastic leukemia resistant to the tyrosine kinaseinhibitor STI571 has a unique BCR-ABL gene mutation. Blood.2001;99:1860-1862.

48. Graham SM, Jorgensen HG, Allan E, et al. Primitive, quiescent,Philadelphia-positive stem cells from patients with chronicmyeloid leukemia are insensitive to STI571 in vitro. Blood.2002;99:319-325.

49. Holtz MS, Slovak ML, Zhang F, Sawyers CL, Forman SJ, BhatiaR. Imatinib mesylate (STI571) inhibits growth of primitivemalignant progenitors in chronic myelogenous leukemiathrough reversal of abnormally increased proliferation. Blood.2002;99:3792-3800.

50. Mow BM, Chandra J, Svingen PA, et al. Effects of the Bcr/ablkinase inhibitors STI571 and adaphostin (NSC 680410) onchronic myelogenous leukemia cells in vitro. Blood.2002;99:664-671.

51. Sun X, Layton JE, Elefanty A, Lieschke GJ. Comparison ofeffects of the tyrosine kinase inhibitors AG957, AG490, andSTI571 on BCR-ABL—expressing cells, demonstrating synergybetween AG490 and STI571. Blood. 2001;97:2008-2015.

52. Dorsey JF, Jove R, Kraker AJ, Wu J. The pyrido[2,3-d]pyrimidine derivative PD180970 inhibits p210Bcr-Abltyrosine kinase and induces apoptosis of K562 leukemic cells.Cancer Res. 2000;60:3127-3131.

53. Topaly J, Schad M, Zeller WJ, Ho AD, Fruehauf S. Strongsynergism of different signal transduction inhibitors in chronicmyelogenous leukemia. [abstract] Blood. 2001;98:617a.

54. Hoover RR, Mahon FX, Melo JV, Daley GQ. OvercomingSTI571 resistance with the farnesyl transferase inhibitorSCH66336. Blood. 2002;100:1068-1071.

55. La Rosee P, Johnson K, O’Dwyer ME, Druker BJ. In vitrostudies of the combination of imatinib mesylate (Gleevec) andarsenic trioxide (Trisenox) in chronic myelogenous leukemia.Exp Hematol. 2002;30:729-737.

56. Porosnicu M, Nimmanapalli R, Nguyen D, Worthington E,Perkins C, Bhalla KN. Co-treatment with As2O3 enhancesselective cytotoxic effects of STI-571 against Bcr-Abl-positiveacute leukemia cells. Leukemia. 2001;15:772-778.

57. Sawyers CL. Chronic myeloid leukemia. N Engl J of Med.1999;340:1330-1340.

58. Silver RT, Woolf SH, Hehlmann R, et al. An evidence-basedanalysis of the effect of busulfan, hydroxyurea, interferon, andallogeneic bone marrow transplantation in treating the chronicphase of chronic myeloid leukemia: developed for theAmerican Society of Hematology. Blood. 1999;94:1517-1536.

59. Goldman JM, Druker BJ. Chronic myeloid leukemia: currenttreatment options. Blood. 2001;98:2039-2042.

60. Bonifazi F, de Vivo A, Rosti G, et al. Chronic myeloid leukemiaand interferon-alpha: a study of complete cytogenetic respond-ers. Blood. 2001;98:3074-3081.

61. Gratwohl A, Baldomero H, Urbano-Ispizua A. Transplantationin chronic myeloid leukaemia. Lancet. 2002;359:712.

62. Marin D, Bua M, Marktel S, et al. The combination of

cytogenetic response after 6 months treatment with STI571 andthe presence of cytopenias in patients with CML in chronicphase resistant to or intolerant of interferon-alfa defines fourdifferent prognostic groups. [abstract] Blood. 2001;98:846a.

63. O’Dwyer ME, Mauro MJ, Blasdel C, et al. Lack of cytogeneticresponse is an adverse prognostic factor for relapse of chronicphase CML patients treated with imatinib mesylate (STI571)[abstract]. Blood. 2001;98:137a

64. Sawafuji K, Miyakawa Y, Weisberg E, Griffin JD, Ikeda Y,Kizaki M. Effects of aminopeptidase inhibitors on STI571resistant cell lines. [abstract] Blood. 2001;98:309a

65. Vigneri P, Wang JY. Induction of apoptosis in chronic myelog-enous leukemia cells through nuclear entrapment of BCR-ABLtyrosine kinase. Nat Med. 2001;7:228-234.

66. Yu C, Krystal G, Varticovksi L, et al. Pharmacologic mitogen-activated protein/extracellular signal-regulated kinase kinase/mitogen-activated protein kinase inhibitors interact synergisti-cally with STI571 to induce apoptosis in Bcr/Abl-expressinghuman leukemia cells. Cancer Res. 2002;62:188-199.

67. Klejman A, Rushen L, Morrione A, Slupianek A, Skorski T.Phosphatidylinositol-3 kinase inhibitors enhance the anti-leukemia effect of STI571. Oncogene. 2002;21:5868-5876.

68. Gatto SR, Scappini B, Verstovsek S, et al. In vitro effects of PS-341 alone and in combination with STI571 in Bcr-Abl positivecell lines both sensitive and resistant to STI571. [abstract]Blood. 2001;98:101a.

69. Nimmanapalli R, Porosnicu M, Nguyen D, et al. Cotreatmentwith STI-571 enhances tumor necrosis factor alpha-relatedapoptosis-inducing ligand (TRAIL or apo-2L)-inducedapoptosis of Bcr- Abl-positive human acute leukemia cells. ClinCancer Res. 2001;7:350-357.

II. Novel Therapies forChronic Myelogenous Leukemia

1. Grove KL, Guo X, Liu SH, et al. Anticancer activity of beta-L-dioxolane-cytidine, a novel nucleoside analogue with theunnatural L configuration. Cancer Res. 1995;55:3008-3011.

2. Giles FJ, Cortes JE, Baker SD, et al. Troxacitabine, a noveldioxolane nucleoside analog, has activity in patients withadvanced leukemia. J Clin Oncol. 2001;19:762-771.

3. Giles FJ, Garcia-Manero G, Cortes JE, et al. Phase II study oftroxacitabine, a novel dioxolane nucleoside analog, in patientswith refractory leukemia. J Clin Oncol. 2002;20:656-664.

4. O’Brien S, Kantarjian H, Keating M, et al. Homoharringtoninetherapy induces responses in patients with chronic myelog-enous leukemia in late chronic phase. Blood. 1995;86:3322-3326.

5. Kantarjian HM, Talpaz M, Smith TL, et al. Homoharringtonineand low-dose cytarabine in the management of late chronic-phase chronic myelogenous leukemia. J Clin Oncol.2000;18:3513-3521.

6. Visani G, Russo D, Ottaviani E, et al. Effects ofhomoharringtonine alone and in combination with alphainterferon and cytosine arabinoside on ‘in vitro’ growth andinduction of apoptosis in chronic myeloid leukemia and normalhematopoietic progenitors. Leukemia. 1997;11:624-628.

7. O’Brien S, Kantarjian H, Koller C, et al. Sequentialhomoharringtonine and interferon-alpha in the treatment ofearly chronic phase chronic myelogenous leukemia. Blood.1999;93:4149-4153.

8. Scappini B, Onida F, Kantarjian H, et al. In vitro effects ofSTI571-containing drug combinations on growth of Ph-positivemyelogenous leukemia-derived cells [abstract]. Blood.2000;96:99a.

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9. Santini V, Kantarjian HM, Issa JP. Changes in DNA methylationin neoplasia: pathophysiology and therapeutic implications.Ann Intern Med. 2001;134:573-586.

10. Issa JP, Kantarjian H, Mohan A, et al. Methylation of the ABL1promoter in chronic myelogenous leukemia: lack of prognosticsignificance. Blood. 1999;93:2075-2080.

11. Zion M, Ben-Yehuda D, Avraham A, et al. Progressive de novoDNA methylation at the bcr-abl locus in the course of chronicmyelogenous leukemia. Proc Natl Acad Sci U S A.1994;91:10722-10726.

12. Ben-Yehuda D, Krichevsky S, Rachmilewitz EA, et al.Molecular follow-up of disease progression and interferontherapy in chronic myelocytic leukemia. Blood. 1997;90:4918-4923.

13. Sacchi S, Kantarjian HM, O’Brien S, et al. Chronic myelog-enous leukemia in nonlymphoid blastic phase: analysis of theresults of first salvage therapy with three different treatmentapproaches for 162 patients. Cancer. 1999;86:2632-2641.

14. Kantarjian HM, O’Brien SM, Keating M, et al. Results ofdecitabine therapy in the accelerated and blastic phases ofchronic myelogenous leukemia. Leukemia. 1997;11:1617-1620

15. Issa JP, Garcia-Manero G, Mannari R, et al. Minimal effectivedose of the hypomethylating agent decitabine in hematopoieticmalignancies [abstract]. Blood. 2001;98:594a.

16. Beaupre DM, Kurzrock R. RAS and leukemia: from basicmechanisms to gene-directed therapy. J Clin Oncol.1999;17:1071-1079.

17. Karp JE, Lancet JE, Kaufmann SH, et al. Clinical and biologicactivity of the farnesyltransferase inhibitor R115777 in adultswith refractory and relapsed acute leukemias: a phase 1clinical-laboratory correlative trial. Blood. 2001;97:3361-3369.

18. Kurzrock R, Sebti SM, Kantarjian HM, et al. Phase I study of afarnesyl transferase inhibitor, R115777, in patients withmyelodysplastic syndrome [abstract]. Blood. 2001;98:623a.

19. Kurzrock R, Cortes JE, Ryback ME, et al. Phase II study ofR115777, a farnesyltransferase inhibitor, in myelodysplasticsyndrome [abstract]. Blood. 2001;98:848a.

20. Peters DG, Hoover RR, Gerlach MJ, et al. Activity of thefarnesyl protein transferase inhibitor SCH66336 against BCR/ABL-induced murine leukemia and primary cells from patientswith chronic myeloid leukemia. Blood. 2001;97:1404-1412.

21. Reichert A, Heisterkamp N, Daley GQ, Groffen J. Treatment ofBcr/Abl-positive acute lymphoblastic leukemia in P190transgenic mice with the farnesyl transferase inhibitorSCH66336. Blood. 2001;97:1399-1403.

22. Hoover RR, Mahon F-X, Melo JV, Daley GQ. OvercomingSTI571 resistance with the farnesyltransferase inhibitorSCH66336 [abstract]. Blood. 2001;98:617a.

23. Thomas D, Cortes J, O’Brien S, et al. R115777, a FarnesylTransferase Inhibitor (FTI), has significant anti-leukemiaactivity in patients with chronic myeloid leukemia [abstract].Blood. 2001;98:727a.

24. Rousselot P, Labaume S, Marolleau JP, et al. Arsenic trioxideand melarsoprol induce apoptosis in plasma cell lines and inplasma cells from myeloma patients. Cancer Res.1999;59:1041-1048

25. Wang ZG, Rivi R, Delva L, et al. Arsenic trioxide andmelarsoprol induce programmed cell death in myeloid leukemiacell lines and function in a PML and PML-RARalpha indepen-dent manner. Blood. 1998;92:1497-1504.

26. Perkins C, Kim CN, Fang G, Bhalla KN. Arsenic inducesapoptosis of multidrug-resistant human myeloid leukemia cellsthat express Bcr-Abl or overexpress MDR, MRP, Bcl-2, or Bcl-x(L). Blood. 2000;95:1014-1022.

27. Puccetti E, Guller S, Orleth A, et al. BCR-ABL mediates arsenic

trioxide-induced apoptosis independently of its aberrant kinaseactivity. Cancer Res. 2000;60:3409-3413.

28. La Rosee P, Johnson K, Moseson EM, O’Dwyer M, Druker BJ.Preclinical evaluation of the efficacy of STI571 in combinationwith a variety of novel anticancer agents [abstract]. Blood.2001;98:839a.

29. Porosnicu M, Nimmanapalli R, Nguyen D, Worthington E,Perkins C, Bhalla KN. Co-treatment with As2O3 enhancesselective cytotoxic effects of STI-571 against Brc-Abl-positiveacute leukemia cells. Leukemia. 2001;15:772-778.

30. Adams J, Palombella VJ, Sausville EA, et al. Proteasomeinhibitors: a novel class of potent and effective antitumoragents. Cancer Res. 1999;59:2615-2622.

31. Rothwarf DM, Karin M. The NF-kappa B activation pathway: aparadigm in information transfer from membrane to nucleus.Science’s STKE [Electronic Resource]: Signal TransductionKnowledge Environment. 1999;1999:RE1.

32. Hamdane M, David-Cordonnier MH, D’Halluin JC. Activationof p65 NF-kappaB protein by p210BCR-ABL in a myeloid cellline (P210BCR-ABL activates p65 NF-kappaB). Oncogene.1997;15:2267-2275.

33. Reuther JY, Reuther GW, Cortez D, Pendergast AM, BaldwinAS, Jr. A requirement for NF-kappaB activation in Bcr-Abl-mediated transformation. Genes Devel. 1998;12:968-981.

34. Dou QP, McGuire TF, Peng Y, An B. Proteasome inhibitionleads to significant reduction of Bcr-Abl expression andsubsequent induction of apoptosis in K562 human chronicmyelogenous leukemia cells. J Pharmacol Exper Therapeut.1999;289:781-790.

35. Gatto SR, Scappini B, Verstovsek S, et al. In vitro effects of PS-341 alone and in combination with STI571 in BCR-ABLpositive cell lines both sensitive and resistant to STI571[abstract]. Blood. 2001;98:101a.

36. Aguayo A, Kantarjian H, Manshouri T, et al. Angiogenesis inacute and chronic leukemias and myelodysplastic syndromes.Blood. 2000;96:2240-2245.

37. Gabrilovich DI, Chen HL, Girgis KR, et al. Production ofvascular endothelial growth factor by human tumors inhibits thefunctional maturation of dendritic cells. Nature Med.1996;2:1096-1103.

38. Gabrilovich DI, Ishida T, Nadaf S, Ohm JE, Carbone DP.Antibodies to vascular endothelial growth factor enhance theefficacy of cancer immunotherapy by improving endogenousdendritic cell function. Clin Cancer Res. 1999;5:2963-2970.

39. Verstovsek S, Kantarjian H, Manshouri T, et al. Prognosticsignificance of cellular vascular endothelial growth factorexpression in chronic phase chronic myeloid leukemia. Blood.2002;99:2265-2267.

40. Chen W, Peace DJ, Rovira DK, You SG, Cheever MA. T-cellimmunity to the joining region of p210BCR-ABL protein. ProcNatl Acad Sci U S A. 1992;89:1468-1472.

41. Yotnda P, Firat H, Garcia-Pons F, et al. Cytotoxic T cellresponse against the chimeric p210 BCR-ABL protein inpatients with chronic myelogenous leukemia. J Clin Invest.1998;101:2290-2296.

42. Pinilla-Ibarz J, Cathcart K, Korontsvit T, et al. Vaccination ofpatients with chronic myelogenous leukemia with bcr-abloncogene breakpoint fusion peptides generates specific immuneresponses. Blood. 2000;95:1781-1787.

43. Cathcart K, Pinilla-Ibarz J, Korontsvit T, et al. All CML patientsvaccinated with a multivalent bcr-abl peptide vaccine showspecific immune responses in a phase II trial [abstract]. Blood.2001;98:728a.

44. Molldrem J, Dermime S, Parker K, et al. Targeted T-cell therapyfor human leukemia: cytotoxic T lymphocytes specific for a

134 American Society of Hematology

peptide derived from proteinase 3 preferentially lyse humanmyeloid leukemia cells. Blood. 1996;88:2450-2457.

45. Molldrem JJ, Clave E, Jiang YZ, et al. Cytotoxic T lymphocytesspecific for a nonpolymorphic proteinase 3 peptide preferen-tially inhibit chronic myeloid leukemia colony-forming units.Blood. 1997;90:2529-2534.

46. Molldrem JJ, Lee PP, Wang C, et al. Evidence that specific Tlymphocytes may participate in the elimination of chronicmyelogenous leukemia. Nature Med. 2000;6:1018-1023.

47. Wieder ED, Kant S, Lu S, et al. Effective specific immunityinduced after PR1 peptide vaccination correlates with cytoge-netic remission [abstract]. Blood. 2001;98:407a.

48. Svingen PA, Tefferi A, Kottke TJ, et al. Effects of the bcr/ablkinase inhibitors AG957 and NSC 680410 on chronic myelog-enous leukemia cells in vitro. Clin Cancer Res. 2000;6:237-249.

49. Sun X, Layton JE, Elefanty A, Lieschke GJ. Comparison ofeffects of the tyrosine kinase inhibitors AG957, AG490, andSTI571 on BCR-ABL—expressing cells, demonstrating synergybetween AG490 and STI571. Blood. 2001;97:2008-2015.

50. Mow BM, Chandra J, Svingen PA, et al. Effects of the Bcr/ablkinase inhibitors STI571 and adaphostin (NSC 680410) onchronic myelogenous leukemia cells in vitro. Blood.2002;99:664-671.

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marrow transplantation for CML: a report from the Interna-tional Bone Marrow Transplant Registry. Bone MarrowTransplant. 1996;17(suppl 3):S5-S6.

2. Clift RA, Storb R. Marrow transplantation for CML: the Seattleexperience. Bone Marrow Transplant. 1996;17(suppl 3):S1-S3.

3. Clift RA, Radich J, Appelbaum FR, et al. Long-term follow-upof a randomized study comparing cyclophosphamide and totalbody irradiation with busulfan and cyclophosphamide forpatients receiving allogeneic marrow transplants during chronicphase of chronic myeloid leukemia. Blood. 1999;94(11):3960-3962.

4. Gratwohl A, Hermans J, Goldman JM, et al. Risk assessmentfor patients with chronic myeloid leukaemia before allogeneicblood or marrow transplantation. Chronic Leukemia WorkingParty of the European Group for Blood and Marrow Transplan-tation. Lancet. 1998;352(9134):1087-1092.

5. Gale RP, Hehlmann R, Zhang MJ, et al. Survival with bonemarrow transplantation versus hydroxyurea or interferon forchronic myelogenous leukemia. The German CML StudyGroup. Blood. 1998;91(5):1810-1819.

6. van Rhee F, Szydlo RM, Hermans J, et al. Long-term resultsafter allogeneic bone marrow transplantation for chronicmyelogenous leukemia in chronic phase: a report from theChronic Leukemia Working Party of the European Group forBlood and Marrow Transplantation. Bone Marrow Transplant.1997;20(7):553-560.

7. Craddock C, Szydlo RM, Klein JP, et al. Estimating leukemia-free survival after allografting for chronic myeloid leukemia: anew method that takes into account patients who relapse andare restored to complete remission. Blood. 2000;96(1):86-90.

8. Clift RA, Buckner CD, Thomas ED, et al. Marrow transplanta-tion for chronic myeloid leukemia: a randomized studycomparing cyclophosphamide and total body irradiation withbusulfan and cyclophosphamide. Blood. 1994;84(6):2036-2043.

9. Slattery JT, Clift RA, Buckner CD, et al. Marrow transplanta-tion for chronic myeloid leukemia: the influence of plasmabusulfan levels on the outcome of transplantation. Blood.1997;89(8):3055-3060.

10. Goldman JM, Szydlo R, Horowitz MM, et al. Choice of

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11. Lee SJ, Klein JP, Anasetti C, et al. The effect of pretransplantinterferon therapy on the outcome of unrelated donor hemato-poietic stem cell transplantation for patients with chronicmyelogenous leukemia in first chronic phase. Blood.2001;98(12):3205-3211.

12. Giralt S, Szydlo R, Goldman JM, et al. Effect of short-terminterferon therapy on the outcome of subsequent HLA-identicalsibling bone marrow transplantation for chronic myelogenousleukemia: an analysis from the international bone marrowtransplant registry. Blood. 2000;95(2):410-415.

13. Morton AJ, Gooley T, Hansen JA, et al. Association betweenpretransplant interferon-alpha and outcome after unrelateddonor marrow transplantation for chronic myelogenousleukemia in chronic phase. Blood. 1998;92(2):394-401.

14. Hehlmann R, Hochhaus A, Kolb HJ, et al. Interferon-alphabefore allogeneic bone marrow transplantation in chronicmyelogenous leukemia does not affect outcome adversely,provided it is discontinued at least 90 days before the proce-dure. Blood. 1999;94(11):3668-3677.

15. Beelen DW, Graeven U, Elmaagacli AH, et al. Prolongedadministration of interferon-alpha in patients with chronic-phase Philadelphia chromosome-positive chronic myelogenousleukemia before allogeneic bone marrow transplantation mayadversely affect transplant outcome. Blood. 1995;85(10):2981-2990.

16. Davies SM, DeFor TE, McGlave PB, et al. Equivalent outcomesin patients with chronic myelogenous leukemia after earlytransplantation of phenotypically matched bone marrow fromrelated or unrelated donors. Am J Med. 2001;110(5):339-346.

17. Hansen JA, Gooley TA, Martin PJ, et al. Bone marrowtransplants from unrelated donors for patients with chronicmyeloid leukemia. N Engl J Med. 1998;338(14):962-968.

18. Weisdorf DJ, Anasetti C, Antin JH, et al. Allogeneic bonemarrow transplantation for chronic myelogenous leukemia:comparative analysis of unrelated versus matched sibling donortransplantation. Blood. 2002;99(6):1971-1977.

19. McGlave PB, Shu XO, Wen W, et al. Unrelated donor marrowtransplantation for chronic myelogenous leukemia: 9 years’experience of the national marrow donor program. Blood.2000;95(7):2219-2225.

20. Bensinger WI, Martin PJ, Storer B, et al. Transplantation ofbone marrow as compared with peripheral-blood cells fromHLA-identical relatives in patients with hematologic cancers. NEngl J Med. 2001;344(3):175-181.

21. Elmaagacli AH, Beelen DW, Opalka B, Seeber S, Schaefer UW.The risk of residual molecular and cytogenetic disease inpatients with Philadelphia-chromosome positive first chronicphase chronic myelogenous leukemia is reduced aftertransplantation of allogeneic peripheral blood stem cellscompared with bone marrow. Blood. 1999;94(2):384-389.

22. Elmaagacli AH, Basoglu S, Peceny R, et al. Improved disease-free-survival after transplantation of peripheral blood stem cellsas compared with bone marrow from HLA-identical unrelateddonors in patients with first chronic phase chronic myeloidleukemia. Blood. 2002;99(4):1130-1135.

23. Remberger M, Ringden O, Blau IW, et al. No difference ingraft-versus-host disease, relapse, and survival comparingperipheral stem cells to bone marrow using unrelated donors.Blood. 2001;98(6):1739-1745.

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27. McGlave PB, De Fabritiis P, Deisseroth A, et al. Autologoustransplants for chronic myelogenous leukaemia: results fromeight transplant groups. Lancet. 1994;343(8911):1486-1488.

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34. Lion T, Henn T, Gaiger A, Kalhs P, Gadner H. Early detectionof relapse after bone marrow transplantation in patients withchronic myelogenous leukaemia. Lancet. 1993;341(8840):275-276.

35. Mensink E, van de Locht A, Schattenberg A, et al. Quantitationof minimal residual disease in Philadelphia chromosomepositive chronic myeloid leukaemia patients using real-timequantitative RT-PCR. Br J Haematol. 1998;102(3):768-774.

36. Miyamura K, Tahara T, Tanimoto M, et al. Long persistent bcr-abl positive transcript detected by polymerase chain reactionafter marrow transplant for chronic myelogenous leukemiawithout clinical relapse: a study of 64 patients. Blood.1993;81(4):1089-1093.

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38. Pichert G, Roy DC, Gonin R, et al. Distinct patterns of minimalresidual disease associated with graft-versus-host disease afterallogeneic bone marrow transplantation for chronic myelog-enous leukemia. J Clin Oncol. 1995;13(7):1704-1713.

39. Radich JP, Gehly G, Gooley T, et al. Polymerase chain reactiondetection of the BCR-ABL fusion transcript after allogeneicmarrow transplantation for chronic myeloid leukemia: resultsand implications in 346 patients. Blood. 1995;85(9):2632-2638.

40. Radich JP, Gooley T, Bryant E, et al. The significance of bcr-ablmolecular detection in chronic myeloid leukemia patients“late,” 18 months or more after transplantation. Blood.2001;98(6):1701-1707.

41. Roth MS, Antin JH, Ash R, et al. Prognostic significance ofPhiladelphia chromosome-positive cells detected by thepolymerase chain reaction after allogeneic bone marrowtransplant for chronic myelogenous leukemia. Blood.1992;79(1):276-282.

42. Sawyers CL, Timson L, Kawasaki ES, Clark SS, Witte ON,Champlin R. Molecular relapse in chronic myelogenousleukemia patients after bone marrow transplantation detectedby polymerase chain reaction. Proc Natl Acad Sci U S A.1990;87(2):563-567.

43. Branford S, Hughes TP, Rudzki Z. Monitoring chronic myeloidleukaemia therapy by real-time quantitative PCR in blood is areliable alternative to bone marrow cytogenetics. Br JHaematol. 1999;107(3):587-599.

44. Mughal TI, Yong A, Szydlo RM, et al. Molecular studies inpatients with chronic myeloid leukaemia in remission 5 yearsafter allogeneic stem cell transplant define the risk of subse-quent relapse. Br J Haematol. 2001;115(3):569-574.

45. Higano CS, Chielens D, Raskind W, et al. Use of alpha-2a-interferon to treat cytogenetic relapse of chronic myeloidleukemia after marrow transplantation. Blood.1997;90(7):2549-2554.

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