Cancer Research...Cancer Research VOLUMK32 DECEMBER 1972 NUMHKR 12 [CANCER RESEARCH 32, 2593-2607,...

Cancer ResearchVOLUMK32 DECEMBER 1972 NUMHKR 12

[CANCER RESEARCH 32, 2593-2607, December 1972]

Combination Cancer Therapy: Presidential Address

Emil Frei, III1

The University of Texas, M. D. Anderson Hospital and Tumor institute, Texas Medical Center, Houston, Texas 77025

First I want to thank you for the privilege and honor ofserving as your President during this past year. It has been aturbulent and an important year for cancer research andaccelerated support is in process. I would like to take thisopportunity to present some of the conceptual and practicaladvances that have been made in cancer chemotherapy in thepast 15 to 20 years and to outline future projections.

In 1955 I arrived at the National Cancer Institute. The needfor the development of systemic treatment for cancer, theencouragement from my superiors, and the opportunity toobserve the very dramatic effect that could be achievedoccasionally in the leukemias through treatment developed byDr. S. Farber and Dr. J. H. Burchenal confirmed my interest inmedical oncology and chemotherapy.

I have been extraordinarily fortunate in the superiors,associates, and trainees with whom I have been associated. Iwould like to mention three specifically. When I arrived at theNational Cancer Institute, it was Dr. Gordon Zubrod whoprovided the friendliness, stimulation, guidance, andopportunity to work, and he has continued to do so. Fivemonths after my arrival, a gentleman appeared in my officeand announced that his name was Emil Freireich. I allowedthat, aside from being somewhat long, this was indeed a veryexcellent name and that anyone with a name like that had to

1Present address: Children's Cancer Research Foundation, Inc., 35

Binney Street, Boston, Mass. 02115.Received August 23, 1972;accepted August 23, 1972.

be good. I offered him a job on the spot, which was easily oneof the smartest things I ever did; we have been working,playing, and fighting together ever since. Last but not least Iwanted to introduce my wife Elizabeth. Unfortunately, shecould not attend because of the urgency of her work on behalfof attaining peace in Indo-China.

Seventeen years ago the chemotherapist was isolated fromhis fellow clinicians in other disciplines relating to cancermainly because he had relatively little to offer. He was alsoisolated from the bridging and from the basic scientists,partially because the field was in its very early stages ofdevelopment but largely because most scientists, clinical andotherwise, were sceptical concerning the future of cancerchemotherapy.

A schematic diagram of drug treatment categories ofdisseminated cancer is presented in Chart 1. Our goal is to ridthe patient of his neoplastic cells. Using rather crudetechniques, such as Chalkly counts, to determine the numberof neoplastic cells at autopsy, we found that patients withdisseminated neoplastic disease have approximately onetrillion or IO12 cells (8). Similar values have been obtained via

more sophisticated techniques in myeloma by dividing thetotal paraprotein production by the amount of paraproteinproduced by individual cells (23). The initial category is calledremission induction wherein the goal is to produce completeremission, that is, to eliminate all clinical evidence of thedisease. This necessitates at least a 99% reduction in neoplasticcells or, to use a colloquialism, a 2-log or greater reduction inthe number of neoplastic cells. Once this is achieved,treatment during remission is initiated. For relativelyhomogeneous populations of tumor cells, a Ist-order kineticeffect of treatment obtains. This means that it takes just asmuch treatment to reduce the number of neoplastic cells froma million to a thousand (less than 1 mg of tissue) as it does toreduce the number of neoplastic cells from IO12 to IO9 (1 kg

of tumor). Thus, the production of complete remissionrepresents only the destruction of the top of the "exponentialiceberg," and continued treatment in remission is necessary.

As the number of cells are reduced there is cytokinetic andmore direct experimental evidence that the slope of the curvewith treatment may in fact become more steep, mainlybecause a larger proportion of the cells enter cycle. There isalso experimental evidence that negotiating the final hurdlefrom almost complete eradication of neoplastic cells tocomplete eradication may necessitate an altered approach to

DECEMBER 1972 2593

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Emil Frei, III

12-

10-

o

o

o

(C

4-

2-

Complete Remission

Remission Erodicotion ofN Treotment

Induction Treotment \ During Remission Neoplostic Cells

Immune Response

TIME (from start of treatment)

Chart 1. Temporal categories of treatment of cancer.

treatment because of selected G0 cells, drug-resistant cells,pharmacological barriers, and other factors.

We get a variable amount of help from the host immuneresponse which sets the threshold at higher than zero cells. Aswill be discussed later, it is important to interfere with thatimmune response as little as possible and indeed to improve itwherever possible.

There is only one human disease wherein the subclinicalnumber of neoplastic cells can be measured with precision; thisis choriocarcinoma (17, 22) (Chart 2). This is asemilogarithmic plot of chorionic gonadotropin titers asaffected by treatment with methotrexate. The curves aregenerally consistent with those in Chart 1. During the initialphases of treatment there is, in general, a delay in the rate ofdecrease, followed by a more rapid reduction, a reverseGompertzian curve, if you will, and finally a decreasedeffectiveness of treatment as the patient's titer approaches and

falls within the normal range.Table 1 presents data for childhood acute lymphocytic

leukemia as of 1961 and is mainly of historical interest. Theseobservations had major influence on the direction ofchemotherapeutic research. With individual or single agents,complete remissions ranging from 21 to 57% were achieved.Combinations of 2 agents produced an additive or synergisticeffect and indeed, with today's use of prednisone and

vincristine, fully 90% of children with acute lymphocyticleukemia will enter complete remission (8). Contemplation ofthese data led to 3 major conceptual and practical advances.

The first was that agents with qualitatively differenttoxicity and different mechanisms of action could usually becombined at full doses with resultant additive or synergisticantitumor effect. This obtains for prednisone and vincristineand for prednisone and 6-mercaptopurine. In contrast, agentswith similar dose-limiting toxicity, such as the myelosuppres-sion for methotrexate and 6-mercaptopurine, can be combinedsafely only by reducing the doses. Because of this a lessertherapeutic effect occurs. In fact, the clinical basis for much ifnot most of the progress in combination chemotherapy hasresulted from this rationale.

The 2nd observation was that appropriate combinations ofagents, for this disease at least, might be capable of destroying

Chart 2. Hormone titers in 5 women with metastatic choriocarcinoma treated with methotrexate. Data are from the papers ofHertz et al. (17) and Ross et al. (22). M. U., mouse units.

Table 1Combination chemotherapy in acute lymphatic leukemia of childhood

Data are from Leukemia Chemotherapy Group B.

Agent(s)MtxÂ°6-MPVCRPred.Mtx

+6-MPPred.+6-MPPred.+ VCRpatients484381723915463Complete

remissionNo.

%10

2112273847415717

45(42)b127

82(69)5384 (77)Reference1111271112628

Â°Mtx, methotrexate; 6-MP, 6-mercaptopurine; VCR, vincristine;

Pred., prednisone.b Numbers in parentheses, complete remission rate (CRR) calculated

assuming independent drug action:CRR, B = CRRA CRRB 1(100 - CRRA)/100]

IO12 leukemia cells and effecting cellular cure. Thus, if eachagent producing complete remission effected a 3-log reduction,that is a 10"3 effect, and 4 such agents could be combined, a10~12 effect was theoretically achievable. With these data and

concepts in mind, the 1st treatment program, which had as itsprimary goal the cellular cure of childhood acute lymphocytic

2594 CANCER RESEARCH VOL. 32



Combination Cancer Therapy

leukemia, was initiated by Freireich et al. in 1963 (l 1) (Table2). This program involved five 10-day courses of the 4-drugcombination referred to as VAMP. This included vincristine,methotrexate, 6-mercaptopurine, and prednisone. Theduration of remission following the end of treatment, that is,the duration of unmaintained remission, was used as theparameter of response. It was clear that such treatmentprovided significantly longer durations of unmaintainedremission than single agents. If one assumed a 4-day generationtime for the leukemic cells and cellular homogeneity, a verysubstantial reduction in neoplastic cells was achieved (8).However, these same 4 drugs, with a somewhat modified doseand schedule, the POMP program, given for a total of 14months as compared to 6 months, resulted in a much longerduration of unmaintained remission (15). This could not beexplained on the basis of simple cytokinetic models andfocused attention on the need for studies of the cytokinetic,cellular, pharmacological, and other leukemic cell heterogeneity. It also indicated that the longer the duration oftreatment, at least up to 14 months, the longer is the durationof unmaintained remission This has resulted in moreprolonged treatment during remission, and the problem of theoptimal duration of treatment for patients in completeremission for most neoplastic diseases remains a difficult one.Finally, it has focused attention on the need for alteredapproaches to chemotherapy and perhaps immunotherapy forthe eradication of minimal residual disease.

A 3rd derivative of the fact that 90% of children with acutelymphocytic leukemia could be placed in complete remissionwas that one could now approach quantitatively the problemof treatment during remission (Chart 3). In a series of studiesby Holland, Pinkel, and others, extrapolated from Phase Iclinical investigations and experimental therapeutic studiesconducted by Skipper, Goldin, Schabel, Bruce, and others,different dose schedules, combinations of agents, the cyclicuse of agents, reinduction therapy, and other techniques haveprogressively prolonged the duration of remission and survival.Each of the curves in Chart 3 represents a study by the AcuteLeukemia Group B or St. Jude Hospital and the year the studywas started. The number of patients in the study is given inparentheses. Actual 5-year survivals of 20 to 30% haveoccurred in studies started in 1966 and a realistic 5-yearsurvival estimate for some of the more recent studies is 50%

(1, 18). Burchenal, in a retrospective study of 5-year survivors,has found that the risk of recurrent disease and deathfollowing the 5th year decreases with time and that 50% of5-year survivors are alive and in remission at the llth year.These patients have a minimal risk of subsequent relapse.Making relatively risk-free assumptions, one can multiply the50% by 50% and find that 25% of patients whose treatment isinitiated today will have long-term tumor-free survival.Moreover, there is every reason to believe that the rapidprogress that has been made will continue.

These principles derived from studies of acute leukemiahave been applied to a number of other neoplastic diseaseswith notable success in some, such as Hodgkin's disease (Table

3). As in acute leukemia there are a number of agents thatindividually are effective, but the complete remission rate isonly 10 to 20% (20, 21). This was improved with thecombination of 3 agents (21) and particularly with the 4-drugcombination developed by DeVita et al. (5) known as theMOPP program. This combination involves mustargen,vincristine, procarbazine, and prednisone, all of which areactive in Hodgkin's disease and only 2 of which have

100

Percent

Surviving

75

50

25

1968 (27) ALG "best arm"

12345

Years From Onset Of Therapy

Chart 3. Acute lymphocytic leukemia in children improving survival.The data are from the Acute Leukemia Group B (ALG) and St. JudeHospital (SJ).

Table 2Duration of unmaintained remission in childhood acute lymphocytic leukemia

RemissioninductiontreatmentVaPVAMP(VP+Mtx+MP)POMP(VP+Mtx+MP)VPVPVPEarlyremissiontreatmentNoneNoneMtxMtx

- MP -> Cyt --BCNUMtx^ VPTotalduration

oftreatment(mo.)1.51.56142.53.58.0Durationofunmaintainedremission(days)4060140240110160315

0 V, vincristine; P, prednisone; Mtx, methotrexate; MP, 6-mercaptopurine; Cyt, Cytoxan;BCNU, 1,2-bis(chloroethyl)nitrosourea.

DECEMBER 1972 2595



Emil Frei, III

Table 3Remission induction in Hodgkin 's disease

All patients had Stage III or IV disease and 76% had Stage IVBdisease.

SingleagentsPred.0Nitrogen

mustardCycloVCRVLBProcarb.No.

ofpatients389238848Completeremission(%)


No.Deaths

ce=1co

oetLuCL

D Unmatntained

CR. not randomized

A Partial responders

O Non responders

0 25 50 75 100 125 150 175

SURVIVAL (Wks.) FROM ONSET OF STUDY

Chart 4. Effect of remission and maintenance treatment on survival. CR, complete remission; Pts., patients.

End of TreotmentNCI MOPPx6â€” Unmointomed

SWCCSG

Nopis35 o33 Ã»

MOPPq2mox9 35 o

I 2 3 4 56

Years from End of Treotment or Randomization

Chart 5. Effect of the duration of MOPP therapy on the remission ofHodgkin's disease (all Stage III and IV disease; 76% Stage IVB). NCI,

National Cancer Institute;S!VCCSG, Southwest Group; q 2 mo, every 2months.

where relapses occur in untreated sites, the relapses followingcombination chemotherapy occur with a high degree ofpredictability at sites of major pretreatment involvement.Considering the Ist-order kinetic effects of chemotherapy andthe vascular, cytokinetic, pharmacological, and other adversefeatures of large tumors with respect to response tochemotherapy, this correlation is not surprising. It has a majortherapeutic implication which is currently under study. Sinceradiotherapy is capable of sterilizing local Hodgkin's lesions, a

major quantitative study has been undertaken by Coltman and

the Southwest Group, wherein radiotherapy is delivered tosites of major pretreatment involvement in those patients whohave entered complete remission on MOPP therapy.

The marked effectiveness for radiotherapy in patients withlocalized Hodgkin's disease, and the increasing effectiveness of

chemotherapy for patients at the other end of the spectrum,that is, patients with disseminated symptomatic disease, hasbrought the disciplines of radiotherapy and chemotherapytogether. The results of this collaboration are indicated inChart 7.

In this randomized study Kaplan and Rosenberg comparedtotal nodal radiotherapy to total nodal radiotherapy followedby 6 courses of MOPP chemotherapy in patients withintermediate stages of Hodgkin's disease. This included

patients with Stages I1B, IIIA, and IIIB disease. The percentageof relapse-free patients for the 2 treatment programs areplotted from the end of treatment. Even with limitedfollow-up, the combination, that is the addition of MOPPchemotherapy to total nodal radiotherapy, is significantlysuperior to radiotherapy alone (4). Thus, it is probable that a2nd disease has been identified (the 1st one was Wilm's tumor)

wherein chemotherapy added to radiotherapy and/or surgeryimproves the cure rate.

The scientific basis for the construction of therapeutic trialsis becoming increasingly broad and relevant. I would like toillustrate a few areas of investigation in this area relating tocytokinetics.

Vincristine arrests cells during metaphase. Thus, following asingle dose of vincristine the mitotic index of human tumorsincreases sharply with the peak at 6 to 12 hr (Chart 8) (10).

In vitro cytokinetic studies of the antitumor antibioticbleomycin are presented in Chart 9. In Chinese hamster cellssynchronized in various stages of the mitotic cycle, it was

DECEMBER 1972 2597



Emil Frei, III

Table 4Risk of relapse after radiotherapy in Hodgkin's disease

AuthorPetersMusshoffKaplanKligermanStagesI

andIIAllI

andIIIand IIAccrual

time1932-19601953-1956

19681957-No.

of "

patients36533214578Proportion

ofsubsequent00.670.510.500.4210.540.250.360.3420.420.190.220.21relapsesat following time interval from treatment(yr)30.36'0.120.080.1240.320.080.050.08(*)"(164)(200)(25)(23)50.260.0400.0660.200.0400.0670.140.0300.06

1(*), no. of patients at risk at the 4th year.

feiMHh

ÃŒÃŒÃŒ1ÃŒÃŒÃŒ(J h M MPrÂ«

CORRELATION

Exact 9

Good 6

Poor 0

Chart 6. MOPP in Hodgkin's disease. Correlation between treatment

and relapse sites of involvement.

found that bleomycin compromised the viability of the cellsmuch more markedly when it was present during the processof mitosis (2). This suggested that, if one produced mitoticarrest with vincristine and if bleomycin were delivered at theappropriate interval following vincristine, a much greaterdestruction of cells would result. This was demonstrated invivo in the Lewis lung system where vincristine followed bybleomycin was found to be synergistic (F. Schabel, Jr., and H.E. Skipper, unpublished observations). In a clinical study inpatients with metastatic lung cancer, a program involvingvincristine followed in 6 hr by bleomycin twice weekly hasbeen developed (Table 5). These preliminary results are beingextended.

Similarly, 2 agents that are commonly used in combinationin man are vincristine and ara-C.3 In general, little or no

attention is paid to the intimate time relationship of the doseschedule. That such is important is indicated in Chart 10

3 The abbreviation used is: aia-C, 1-0-D-arabinofuranosylcytosine.

wherein vinblastine, which is a metaphase arrester similar tovincristine, and ara-C, a DNA synthesis inhibitor, are examinedin the L1210 mouse leukemia system (27). The antileukemiceffect is presented in the ordinate. Vinblastine and ara-C weregiven at fixed doses every other day alone or in close timesequence. The effect of the agents used alone is indicated.When the 2 drugs are given simultaneously there is an actualreduction in the antileukemic effect. When ara-C is given 16 hrafter vinblastine, the maximum effect is achieved. In cyto-kinetic studies it was demonstrated that vinblastine results inmaximum synchrony in the S period by 16 hr. This is theperiod most sensitive to ara-C. Clearly, such information isimportant in the development of dose schedules forcombination chemotherapy in man.

One of the most important problems in chemotherapy hasbeen the selection of new agents and treatment programs forclinical trial. While the transplanted tumor systems,particularly the LI 210 mouse leukemia, are effective anduseful, they are dissimilar to their clinical counterpart in avariety of ways. This may compromise the correlation betweenthe experimental and clinical systems and thus reduce thepredictability value. While agrÃ©Ã¢tdeal of work has occurred inthis area, I would like to emphasize particularly thespontaneous AKR mouse leukemia system shown in Table 6(24, 25). Like the human disease, acute myelogenousleukemia, it is spontaneous in contrast to the transplantedsystems. It is diagnosed by palpation of the spleen and likehuman leukemia is widely disseminated at the time ofdiagnosis. Cytokinetically, it has a long leukemia cell-doublingtime which corresponds to the clinical disease. This is related,in considerable part, to the fact that the growth fraction inboth systems is relatively low in contrast to the LI210. Therecovery time for the normal bone marrow in the mouse is 3to 4 days following cell cycle-specific agent administration incontrast to man in whom it is 9 to 15 days. After a series ofstudies, Schabel and Skipper have observed that the besttreatment programs with cell cycle-specific agents such asara-C for both the LI210 and the AKR consists of 24- to 48-hrcourses of continuous treatment followed by interruptionbetween courses sufficient to allow for complete bone marrowrecovery (25). Translated cytokinetically to man, this wouldrepresent roughly 5-day courses of continuous ara-C every 14days. In noncomparative studies such interrupted courses oftreatment are superior to various types of daily treatment, forboth complete remission induction and maintenance and forsurvival (Table 7) (Ref. 7; J. Bickers and E. J Freireich,manuscript in preparation). The spontaneous AKR system hasalso been used to evaluate combination chemotherapy (Chart





IÂ§

80

60

0'S 40

20

J_

p = 0.04â€¢¿�XRT '-MOPP

o XRTA individual patients at risk.

l i l i l l l J I I I I I L4 8 12 16 20 24 28 32

Time in Months After End of Treatment

Chart 7. Effect of combined radiotherapy and chemotherapy onStages HB, IIIA, and IHB of Hodgkin's disease. XRT, total nodal

radiotherapy.

I40 -T

1 Lymphosarcomo

2 Melanoma

3 Cancer of the Breast

4 Myeloma5 Acute Lymphatic

Leukemia

26 8Hours

12 468Days

24 48

TIME AFTERVINCRISTINE

Chart 8. The effect of vincristine on the mitotic index in the bonemarrow. Mitotic index, no. of mitoses/1000 cells.

11). The best combination with appropriate attention toschedule has been cyclophosphamide combined with ara-C andalso with vincristine and prednisone. The survival curve ofuntreated (control) mice with spontaneous AKR leukemia isdescribed. For combined treatment the survival curve levels offat about the 50% survival level and, by bioassay, no tumorcells could be demonstrated in the thymus in 50% of theseanimals following treatment (H. E. Skipper and F. J. Schabel,Jr., unpublished observations). After 4 weeks, mortality fromleukemia again appears, probably because of backgroundreinduction by Gross virus. That cellular cure in all probabilitycan be achieved by chemotherapy in this spontaneous tumorsystem is of signal importance. These animals have 100 millionleukemic cells at the time of treatment, and it is reliablyestimated that a greater than 1,000-fold destruction of normalstem cells would be fatal. Thus, this chemotherapy wascapable of delivering a 100,000-fold selectivity against theneoplastic cells. When this combination was subjected to arandomized study by Bodey and the Southwest Group andcompared directly to ara-C, it was highly effective and wassignificantly superior in terms of the complete remission rate,

the duration of complete remission, and survival (Table 8).Additional studies by Goldin and Schabel indicate that thespontaneous AKR leukemia generally responds to agents thatare effective in the leukemias and lymphomas in man. Thus,the development of this system represents major progress inthe critical area of selecting new agents and new treatmentprograms for clinical trial.

As already indicated, there is evidence, both in experimentaland in clinical tumor systems, that intensive short-term coursesof treatment are superior to continuous treatment. In man thiswould appear to be particularly true in Burkitt's lymphoma,choriocarcinoma, myeloma, Hodgkin's disease, and adult acute

leukemia. Perhaps one of the most compelling arguments infavor of intermittent treatment relates to chemicalimmunosuppression. As our treatment becomes more effectivein terms of remission induction and as we move to morelong-term treatment, the relative effects of various treatmentprograms on suppression of immune response will becomeincreasingly important.

G,Cells

I10 25 50

DOSE OF BLEOMYCIN (fÂ»g/mlfor 30 minutes)

Chart 9. Effect of bleomycin on survival of mammalian cellssynchronized in various stages of the cell cycle.

Table 5VCRa-Bleo sequence for lung cancer

PartialresponseBleoVCR

-> BleobMrÂ»

r*fcases7713No.24%331

" VCR, vincristine; Bleo, bleomycin.b VCR followed in 6 hr by Bleo twice a week.

DECEMBER 1972 2599



Emil Frei, III

Hersh has studied immunosuppression in patients receiving5-day courses of intensive chemotherapy (Chart 12). Theimmunological parameters evaluated included macrophageentry into experimental inflammatory sites (the skin window),response to primary antigenic stimulation, and lymphocyteblastogenesis to phytohemagglutinin (3). There is a markeddecrease in all of these parameters while the patient is ontreatment. However, within 2 to 3 days following suchtreatment there is complete or nearly complete recovery inimmunological response and indeed, in many patients,immunological "overshoot" occurs during the recovery period.

Thus, when such 5-day courses of treatment are given every 2to 4 weeks, the patient's immunological apparatus is normal

most of the time. This contrasts to what is seen in continuoustreatment wherein a lesser initial degree of immunologicaldepression occurs but is sustained and tends to be progressive.Indeed, there is evidence that intermittent chemotherapy has agreater effect on circulating antibody response as compared tocontinuous treatment, which has a relatively greater effect oncellular immunity. These data are presented in Charts 13 and14 wherein kidney transplant patients receiving continuousdaily chemotherapy are compared to patients with cancerreceiving intermittent chemotherapy. In Chart 13 the humoral

150 r

100

S -E

50

nVLB Aro-C

only0 246 8 16

VLB-Aro-C

Interval (Mrs) Between Injections

Chart 10. Schedule of vinblastine (VLB) and ara-C combination inL1210 cells. Vinblastine (1 mg/kg) and ara-C (20 mg/kg) given every 2

days.

antibody response to keyhole limpet hemocyanin issubstantially impaired as compared to controls in both groupsof patients, but more so in those receiving intermittentchemotherapy. In contrast (Chart 14) lymphocyteblastogenesic response to keyhole limpet hemocyanin, ameasure of cellular immunity, is markedly impaired in patientsreceiving continuous chemotherapy as compared to thosereceiving intermittent chemotherapy. While the manydifferences between the 2 groups of patients is recognized, thediscordance in the cellular and humoral immune response as afunction of the schedule of chemotherapy is of note. Sincecellular, i.e., T-cell, immunity is the major tumor-restrainingfacet of immune response and since humoral immunity may be"enhancing or blocking," intermittent chemotherapy not only

provides a lesser overall immunosuppression but tends topreserve cellular immunity.

There is ample evidence in experimental systems that thepreservation and indeed the augmentation of immunity andparticularly cellular immunity is important to the antitumoreffect of chemotherapy. There is also compelling evidence inman that response to chemotherapy is highly correlated with apatient's immune response prior to treatment. This is

illustrated in Table 9 in which the response of adults withacute leukemia to established skin test antigens and theblastogenic response to phytohemagglutinin prior to treatmentand at the time of recovery following the 1st course oftreatment was evaluated (16). For these patients who had aremission as a result of chemotherapy, there was a significantlysuperior cellular immunity as compared to those patients whodid not -respond; this difference was particularly true after

recovery from the 1st course of chemical immunosuppression.Thus, those patients destined to have a remission (major

Table 7Effect of dose schedule on ara-C in acute myelogenous leukemia

Separate studies.

ScheduleDaily

5-day infusionsevery 2-3 wkComplete

No. of remissionpatients(%)98

8-2585 38Duration

Remissionmedian

mo.7

12Survival50%2.5225%8 14

Table 6Spontaneous AKR mouse leukemia model

TumorL1210Spontaneous

AKRAcute

myelogenousleukemiaOriginTransplanted"Spontaneous"Gross

virustolerant"Spontaneous"Status

attimeofinitialtreatmentVariableDisseminated10Â»

cellsDisseminated10"

cellsDoublingtime(days)0.5>5>5Tc(days)0.51.02-47"s(hr)0.91120Growthfraction10.50.2-0.4Normal

marrowrecovery(days)"33-49-15Bestschedule*1forcellcycle-specific

agent24+-hr

coursesevery4days24+-hr

coursesevery4days5-day

coursesevery14 days

a After suppression with cell cycle-specific agents.b Best schedule for cell cycle-specific agent consisted of courses of treatment for at least twice the Tc with treatment-free internal between

courses to allow for normal marrow recovery.





100

o

oV)

80-

60 -I

40-

Â°- 20-

O(E

0

Drug

VCR

Pred

Cyt

P-ara-C

mg/kg/dose Schedule

221% ILS

0.5 day 0 only

30.0 qd x5, day 03

150.0 day 4 only

25.0 qdx5, days 711

1970 Control (636 mice)

33 Mice Treated with:

Backgroundreinduction rate

I

0 20 40t

60 80 IOO I20

Bioassav revealed viablelymphoma cells in thethymus in only 50%

LIFESPAN, Days Postdiagnosis

Chart 11. Bifunction survival of AKR lymphoma after "curative" chemotherapy. ILS, increased life-span.

Table 8ara-C vs. COAP" in adult acute leukemia

Randomized comparative study.

Treatment"ara-C

COAPbNo.

ofpatients57

64Complete

remission23%

50%Duration

ofremissionmedian(mo.)9

14Survival50%2825%1325

p < 0.01 p = 0.05 p < 0.01

" COAP, Cytoxan, Oncovin (vincristine), ara-C, and Prednisone.b Both treatment programs given as 5-day courses every 2 to 3

weeks.

response) recovered completely whereas recovery was faulty orlacking in the patients destined not to respond tochemotherapy. In summary, proper attention to thescheduling of chemotherapeutic agents, with emphasis onintensive short-term courses of intermittent therapy, tends topreserve immune response and, particularly, cellular immuneresponse. As active immunotherapeutic programs arecombined with chemotherapy, the active immunization can beadministered at the time of immunological recovery or"overshoot." Finally, there is evidence that esoteric infections

such as fungus, viral, and Pneumocystis infections are lessfrequent in patients maintained on long-term intermittentchemotherapy than in those on continuous chemotherapy.

The discussion up to this point has dealt with hematologicalcancers. I would like to turn now to selected solid tumors.Some of the cytokinetic, pharmacological, vascular, and otherfeatures of solid tumors as compared to the leukemias thatmight adversely affect response have been emphasized. Iwould like to indicate another difference which I don't believe

has been sufficiently recognized (Table 10). In the acute

ANTIBODY TITER(ANTIGEN 2)

ANTIBODY TITER(ANTIGEN I )

21 28DAYS

ANTIGENI 2

Chart 12. Effect of chemotherapy on immune response.

leukemias, while cytogenetic abnormalities occur in theleukemic cells in about 50% of patients, these usuallyrepresent a single clone, are pseudodiploid or near diploid,and, most important, usually remain stable throughout thedisease (14). For these patients, responses are frequent and,with appropriate treatment, prolonged. Similarly, in the early

DECEMBER 1972 2601



Emu Frei, III

phase of chronic myelogenous leukemia where aneuploidy orpseudodiploidy is slight, response to treatment is excellent. Inthe blast phase, however, aneuploidy, which may be unstable,is common and response to treatment is poor (26). While

Table 9Effect of various types of chemotherapy on the delayedhypersensitivity response to keyhole limpet hemocyanin

Keyhole limpet hemocyanin skin test response

I

01234

WEEKS AFTER IMMUNIZATION

Chart 13. Total antibody response of various groups to keyholelimpet hemocyanin.

101

WEEKS AFTER IMMUNIZATION

Chart 14. Lymphocyte response of various groups to keyhole limpethemocyanin.

GroupContinuous

therapyIntermittent therapyControlPositive/

tested2/16

16/2013/14%

positive12.5

80.092.8Diameter

ofpositivereactions7.5

8.59.5

metastatic solid tumors in general have been studiedinsufficiently, the existing data suggest that they aresubstantially different from the leukemias. Thus for metastaticmelanoma, marked aneuploidy with hyperdiploid up throughhypertetraploid lines are common, stem cell lines are multiple,and clonal evolution (i.e., instability) is very common (28).This heterogeneity should be theoretically ideal for thedevelopment or selection of drug resistance.

The effects of this cytogenetic heterogeneity of humanmelanoma mÃ©tastaseson chemotherapy are indicated in Charts15 and 16. Four clones of melanoma cells were isolated fromthe same metastasis, and their responsiveness in vitro to ara-Cand to 1,2-bis(chloroethyl)nitrosourea was determined. Forara-C, under the conditions of the experiment, the clonesvaried in responsiveness from a 0 to 50% reduction in thesurvival fraction (Chart 15). A similar or even greater variationwas seen for 1Â¿-bis(chloroethyl)nitrosourea (Chart 16; S.Barranco and R. Humphrey, unpublished observations). Inview of this it is not surprising that ara-C is not particularlyeffective in the treatment of metastatic melanoma. Thirteenpatients with metastatic melanoma to the skin were treatedwith ara-C and are ranked in order of response (Table 11).The pretreatment tumor size is expressed as 100%. There wasonly 1 really good response (Patient V.M.) which lasted for 7months. Two marginal partial responses occurred which lastedonly 2 months. With the marked variability inchemotherapeutic response of clones within a given metastasis,one would expect the rapid selection of the resistant clonesupon treatment with ara-C. ara-C is activated by the enzymedeoxycytidine kinase (K) to the nucleotide, at which level itinhibits DNA polymerase and is substantially inactivated bydeoxycytidine (or ara-C) deaminase (D). It was postulatedthat, for a given tumor, the ratio of these 2 enzymes shoulddetermine the activation and the tissue susceptibility to ara-C.This proved to be the case. The K/D ratio was much higher inresponding patients (Table 11) (p = 0.04) (13). These resultshave 2 implications. The first is that, with modern techniques,it should become increasingly possible to determine in advancein vivo susceptibility to chemotherapeutic agents. The isolationof different clones from a given tumor may further assist inthis goal. Most particularly, this heterogeneity constitutes astrong argument for a multifaceted approach to metastaticdisease including combination chemotherapy andimmunotherapy.

In spite of the above limitations and problems, intenseinvestigation of the chemotherapy of various solid tumors isunderway and progress is being made. In the past, predictive orscreening systems have emphasized transplanted leukemias andlymphomas. This has, at least in part, contributed to the fact





Table 10Cytogenetic abnormalities and response to chemotherapy

Cytogenetic abnormalities

Aneuploidy

Acute leukemias

Chronic myelocyticleukemia

Frequency Stem Response to(%) Severity Stability lines chemotherapy

Frequent and50 Slight Stable 1 prolonged

Early phase

Late (blast)phaseMetastatic

solidtumors (melanoma)8080+90+pH'SlightMajor0Stable

UnstableUnstable1SeveralMultipleFrequent

andprolonged

PoorPoor

' Hyperdiploid, triploid, tetraploid, and hypertetraploid cell lines are common.

IOO-Â»

50-

o

5

LU

Â£Â£

10

?6-5 1

49-5

19-4

20 80 10040 60DOSE ARA-C (pq/ml for I hr.)

Chart 15. Effect of ara-C on 4 strains of human malignant melanomagrowing asynchronously in vitro.

that agents selected for clinical trial have tended to be mostactive in the hematological cancers. Increasing emphasis isbeing given to experimental solid tumor models such as thoseshown in Table 12. These data, largely from the SouthernResearch Institute, indicated the limited activity of agents thattend to be effective in the hematological cancers and that are,for the most part, cell cycle specific. However, some of the

19-4

30 50 70

DOSE BCNU (pg/ml for I hr.i

100

Chart 16. Effect of 1,2-bis(chloroethyl)nitrosourea (BCNU) on 4strains of human malignant melanoma growing asynchronously in vitro.

new agents such as bleomycin, the anthracyclines, thetriazenoimidazoles, and particularly the nitrosoureas alone orin combination with cyclophosphamide have markedantitumor activity. These studies are currently being translatedto man with positive results in some instances, as we shall seeshortly. I would predict that major progress will soon be seenfor breast cancer. There are at least 6 classes ofchemotherapeutic agents capable of producing objective

DECEMBER 1972 2603



Emil Frei, III

Table 11Correlation of cellular pharmacology and response for ara-C in me tasta tic melanoma

Enzyme assays(nmoles product/g tissue 1 hr)

ResponseResponseFailureadequatetrialPatientV.M.E.

D.D.H.G.

J.E.H.W.

M.J.T.J.L.G.J.D.C.L.

M.G.S.L.

F.No.

ofpatients12334223332113Reductionof

tumor size(%)"55050100100120150150150150150150200ara-Ckinase(K)50634329537611723350302320ara-C

deaminase(D)27874648450124012211292441871059402419296(K/D)X10018246511269614270.30550

0 Pretreatment size, 100%.

Table 12Activity of various agents against a spectrum of solid tumors

AgentMethotrexate6-Mercaptopurine5-Fluorouracilara-CHydroxyureaVincristinePrednisoneDaunomycinActinomycin

DBleomycinNitrogen

mustard(HN2)DL-SarcolyÃ¡nCytoxan1

,2-Bis(chloroethyl)nitrosoureaH2-Chloroethyl)-3-cyclohexyl-l-nitrosoureaMethylcyclohexylureaMethylcyclohexylurea

+ CytoxanSarcomaTumor

wt.T/C(%)"321436269461355701000180Cures0/60/63/60/80/100/100/71/100/618/200/618/2014/20Ca755Tumor

wt.T/C(%)2701386020401350Cures0/109/100/100/100/100/102/102/201/100/100/10C3HILS)009312701104710016155B16melanoma(%ILS)0230?1864225075-1004775-100>100Lewis

lungEarly

EstablishedT/C (%)rating6552518763565566532000â€”_â€”_+â€”â€”+-H-++++

' T/C, tumor size in treated animal/tumor size in control animal; ILS, increase in life-span of treated animals/control animals.

response. Several of these, such as the nitrosoureas andadriamycin, have been identified in the above screens onlywithin the past several years. This situation resembles that forHodgkin's disease 7 years ago and for childhood leukemia 11

years ago. The appropriate application of clinical investigationand basic science techniques and extrapolations, as well as thedevelopment of further new agents, will almost certainly resultin improved control of metastatic breast disease. While there isconclusive evidence that combinations will increase theremission rate, the overall effect of this on the disease remainsto be determined.

An example of recent progress in a tumor category thatwas almost totally unresponsive to chemotherapy just 3 yearsago is indicated in Table 13. Soft tissue and bone sarcomashave been highly resistant to chemotherapy. This table

represents data from the Southwest Group. Soft tissue andbone sarcomas have a 15% objective response rate to thetriazenoimidazole and a 31% response rate to the newantibiotic, adriamycin. In experimental tumors thiscombination was found to be synergistic and to be tolerated incombination at full doses without an increase in dose-limitingtoxicity. This was found also to be true for the combination inman with a resultant response rate of 41% and a medianduration of response of over 5 months (12).

It is apparent that a great deal of progress has been madeand is continuing in the chemotherapy of cancer. However,new and more effective drugs are needed. Perhaps the rate ofprogress in chemotherapy is best illustrated by the increasingrate with which clinically effective antitumor agents are beingdiscovered (Chart 17). The 23 clinically effective drugs are





Table 13Soft tissue and hone sarcomas

TreatmentNo. of

patientsResponse

No. ofcompleteremissions

Durationof response

median(mo.)

Diethyltriazenoimidazole carboximide0 55 15 1Adriamycin0 108 31 1Diethyltriazenoimidazole carboximide 100 41 5 >5

+ adriamycinb

0 Collected series.b Ref. 12.

AdriamycinTGAra-CBleomycinAsparaginaseDIGStreptozotozinBCNUDaunarubicinProcarbazineMithramycinHydroxyureaVCRMelphalanCytoxanVLBFluorouracilChlorambucilBusulfanAct DMPMTXHN,

Chart 17. Rate of discovery of clinically effectiveantitumor agents exclusive of hormones. TgG,thioguanine; DIC, dimethyltriazenoimidazole car-boxamide; BCNU, l,2-bis(chloroethyl)nitrosourea;VCR, vincristine; VLB, vinblastine;/4c/Â£), actinomy-cin D; MP, 6-mercaptopurine; MTX, methotrexate.

1943 1950 1955 1960 1965

YEAR EFFECTIVENESS ESTABLISHED

1969

OH

COCH3

H3CO

DaunorubicinNSC-82151

AdriamycinNSC-123127

Chart 18. The anthracycline antibiotics.

DECEMBER 1972 2605



Emil Frei, III

Table 14Comparative antitumor effects of anthracycline antibiotics

Daunorubicin Adriamycin

Experimental antitumoractivity

Ehrlich carcinoma 2+Spontaneous mammary 0-1+

carcinomaL1210 0Sarcoma 180 1+

4+3+

1-2+2-3+

Human antitumoractivityLymphomaAcute

myelogenousleukemiaAcute

lymphaticleukemiaBreast

carcinomaSarcomasBladder

carcinoma20

(87)Â°37(469)27

(388)12

(29)13(46)64

(44)Â°27(51)43(121)43

(23)31(108)37

(19)

Surgery -

Radiotherap

CytokineticsBiostatistics

ImmunologyScreening

Serendipity _..

PATIENT WITH CANCER*'


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P., and Ben, M. The Stathmokinetic Effect of Vincristine. CancerRes., 24: 1918-1925,1964.

11. Freireich, E. J., Karon, M., and Frei, E., III. Quadruple CombinedChemotherapy (VAMP) for Acute Lymphocytic Leukemia inChildren. Proc. Am. Assoc. Cancer Res., 5: 20, 1964.

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DECEMBER 1972 2607



1972;32:2593-2607. Cancer Res Emil Frei III Combination Cancer Therapy: Presidential Address

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