`
`
`
`G. Lümmen
`H. Sperling
`H. Luboldt
`T. Otto
`H. Rübben
`
`Department of Urology, University of Essen
`Medical School, Essen, Germany
`
`Original Paper
`
`Urol Int 1998;61:215–219
`
`Received: August 28, 1998
`Accepted: November 4, 1998
`
`Granulocyte-Macrophage
`Colony-Stimulating Factor and
`Interferon-Alpha 2B in Patients with
`Advanced Renal Cell Carcinoma
`
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`
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`
`Key Words
`Granulocyte-macrophage
`colony-stimulating factor
`Interferon-·2B
`Renal cell carcinoma
`
`Abstract
`Objective: Biological response modifiers such as interferon-·2B (IFN-·2B)
`have well-known clinical activities against renal cell carcinoma (RCC).
`Recombinant human granulocyte-macrophage colony-stimulating factor
`(GM-CSF) has antitumorigenic effects both in vitro and in vivo. Therefore, a
`phase-I/II trial of IFN-·2B and GM-CSF was performed in patients with met-
`astatic RCC. Methods: 21 patients in groups of 3 patients received GM-CSF
`at 7 different dose levels (15–300 Ìg) subcutaneously in combination with
`IFN-·2B at a fixed dose of 10 ! 106 IU s.c. three times weekly for 12 weeks.
`Results: Two complete remissions have been observed, both with lung metas-
`tases only. With increasing dose levels of GM-CSF a slight tendency to more
`toxicity was detectable. Due to grade-3 toxicities 5 patients (24%) dropped out
`of the treatment schedule. Increases in WBC, neutrophils, lymphocytes, and
`monocytes were noted but were not related to the dose levels of GM-CSF.
`Conclusions: Results demonstrate that simultaneous administration of GM-
`CSF and IFN-·2B is tolerated up to doses of 120–150 Ìg GM-CSF three times
`weekly. But there is no additional antitumorigenic effect of GM-CSF because
`the overall response rate of the combined administration of GM-CSF/IFN-
`·2B is similar to IFN-·2B alone and there is no obvious dose relationship
`between increasing doses of GM-CSF and the responses.
`
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`Introduction
`
`There is no satisfying therapy for patients with meta-
`static renal cell carcinoma (RCC). Since deKernion et al.
`[1] in 1983 reported on objective responses of 16% using a
`partially purified interferon-· (IFN-·) preparation, nu-
`merous subsequent phase-II trials with recombinant IFN-
`· have yielded reproducible response rates varying be-
`tween 14 and 26% [2, 3].
`
`The granulocyte-macrophage colony-stimulating fac-
`tor (GM-CSF) is generally indicated for bone marrow fail-
`ure secondary to administration of chemotherapeutic
`drugs or radiationtherapy, bone marrow transplantation,
`and a variety of congenital or iatrogenic neutropenias [4].
`Furthermore, there are preclinical data on the tumoricidal
`effects of GM-CSF. GM-CSF is a potent inducer of tumor
`necrosis factor-·, interleukin-6 (IL-6), and other cyto-
`kines, as well as an activator of antitumor macrophages
`
`ABC
`Fax + 41 61 306 12 34
`E-Mail karger@karger.ch
`www.karger.com
`
`© 1999 S. Karger AG, Basel
`0042–1138/98/0614–0215$17.50/0
`
`Accessible online at:
`http://BioMedNet.com/karger
`
`Gerd Lümmen
`Department of Urology, University Essen Medical School
`Hufelandstrasse 55
`D–45122 Essen (Germany)
`Tel. +49 201 723 3211, Fax +49 201 723 5902
`
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`Table 1. Patient characteristics
`
`Eligible patients
`Males
`Females
`Age, years
`Median (range)
`Body weight, kg
`Median (range)
`Karnofsky index, %
`70
`80
`90
`100
`Nephrectomy
`Sites of metastatic disease
`Lung only
`Lung and other
`Only other
`
`Number
`of patients
`
`21
`14 (67%)
`7 (33%)
`
`65 (39–75)
`
`80.5 (54–97)
`
`4 (19%)
`6 (29%)
`9 (43%)
`2 (9%)
`16 (76%)
`
`10 (48%)
`5 (23%)
`6 (28%)
`
`[5]. GM-CSF showed tumoricidal activity by stimulating
`antibody-dependent cytolysis of tumor cells by mature
`human neutrophils and eosinophils [6].
`The aim of this phase-I/II trial was to ascertain the effi-
`cacy and toxicity of different doses of GM-CSF in combi-
`nation with a unique dose of IFN-·2B in patients with
`metastatic RCC.
`
`Material and Methods
`
`From November 1995 to August 1996, 21 patients with advanced
`RCC were recruited into this phase-I/II trial. The eligibility require-
`ments included evidence of progressive disease with measurable
`metastases, life expectancy of at least 3 months with a performance
`status above 70% on the Karnofsky index, and adequate renal, hepat-
`ic and bone marrow function. The ineligibility criteria were patients
`with serious active infection, hepatic and brain metastases, and
`patients who had previous treatment with IFN, GM-CSF or other
`lymphokines as well as previous radiation therapy, chemotherapy or
`immunotherapy within the last 5 years. The patients characteristics
`are outlined in table 1.
`Patients received a unique dose level of IFN-·2B of 10 ! 106 IU
`and 7 different dose levels of GM-CSF (15, 30, 60, 90, 120, 150 and
`300 Ìg), three times weekly for 12 weeks. The first group of 3 patients
`began with the lowest dose level of GM-CSF, escalation to the next
`dose level occurred only if, after 3 weeks of therapy, a minimum of
`2 patients had leukocytes lower than 20,000/Ìl and no serious side
`effects. All patients self-administered the medication subcutaneously
`in the evening, on an outpatient basis. IFN-·2B (IntronA®) and GM-
`CSF (Leucomax®) was supplied by Essex Pharma GmbH/Schering
`Plough (Munich, Germany). The dose of IFN-·2B had to be reduced
`
`to 6 ! 106 IU and the dose of GM-CSF to 50% in any patient who
`experienced grade-3 toxicity (WHO) attributable to IFN-·2B or GM-
`CSF, respectively. If toxicity persisted after changing the dosage, the
`patient was withdrawn from the study.
`Patients were monitored three times in the first week of therapy,
`weekly for the next 3 weeks and every 4 weeks thereafter with physi-
`cal examination, complete blood cell count, and serum chemistry
`determination. Response evaluation (UICC criteria) was performed
`after every 3 months.
`Survival was calculated using the Kaplan-Meier estimate [7].
`
`Results
`
`21 consecutive patients with histologically confirmed
`progressive metastatic RCC entered this phase I/II trial.
`All patients had not received prior therapy.
`
`Tumor Response
`We were unable to evaluate the response in 6 of the 21
`patients, as their treatment was discontinued due to
`grade-3 side effects (WHO), and 1 patient died of tumor
`progress within 8 weeks. The remaining 15 patients
`received treatment over 12 weeks as a complete treatment
`cycle. Two of them had a complete remission (CR) (last-
`ing for 52+ and 66+ weeks), but there were no partial
`remissions (PRs). 9 patients (60%) had stable disease (SD;
`median 43, range 16–66 weeks), and in 4 (27%) patients
`the tumor showed initial progression. The overall objec-
`tive response rate was 13% (90% confidence interval 2–
`36%). The two objective responses occurred in patients
`with lung metastases only. Concerning the dose levels of
`GM-CSF the 2 CRs were seen with 90 and 150 Ìg GM-
`CSF. Patients with SD were evaluated at each dose level
`besides 300 Ìg. After a median follow-up of 38 (8–68)
`weeks the median survival time from the start of therapy
`amounted to 61 weeks. Death occurred in 11 patients
`after 8–61 (median 32) weeks. The median time to pro-
`gression was determined to 19 weeks.
`
`Toxicity
`Toxicity associated with the combined administration
`of IFN-·2B and GM-CSF is shown in table 2. 18 of 21
`patients (86%) experienced a flu-like syndrome with fe-
`ver, chills, and fatigue. In 7 patients (34%) the symptoms
`nausea, vomiting, and weight loss occurred. Neurotoxici-
`ty characterized by somnolence and disorientation was
`experienced by 1 patient (5%), but 8 patients (38%) devel-
`oped paresthesias of the hands and feet. Two patients
`(10%) suffered from dyspnea, in 1 case as grade-3 toxicity.
`This patient chose to withdraw from further participation
`
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`Urol Int 1998;61:215–219
`
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`in the study and died due to tumor progress within
`8 weeks. One patient (5%) developed cardiovascular com-
`plications resulting in discontinuation of treatment. The
`maximum severity of adverse events reached grade-3 tox-
`icity (WHO), and was reported for 7 patients (33%). As to
`be expected in treatment with IFN-·2B, 6 of 9 (66%)
`grade-3 toxicities (WHO) consisted of nausea/vomiting,
`neurologic and flu-like symptoms. 13 patients or 62% suf-
`fered from grade-2 toxicity (WHO) and 1 patient (5%)
`from grade-1 toxicity. Clinical side effects were consis-
`tently controlled with the use of antipyretics, antiemetics,
`and administration of the trial drugs in the evening when
`possible. However, due to grade-3 side effects (WHO),
`5 patients dropped out of the treatment schedule (1
`patient with 30 Ìg, 1 with 120 Ìg, 1 with 150 Ìg, and 2
`with 300 Ìg GM-CSF, respectively). With increasing
`dose-levels of GM-CSF a slight tendency to more toxicity
`was found. At dose levels of 120 Ìg GM-CSF and higher,
`5 of 9 patients suffered grade-3 toxicity compared to 2 of
`12 patients in the first 4 groups. At the final dose level of
`300 Ìg GM-CSF, grade-3 toxicity occurred in 2 patients
`after a short time of treatment. One patient suffered an
`angina attack immediately after the first drug application
`and the second patient dropped out on day 15 of treat-
`ment. Therefore, the maximum tolerable dose of GM-
`CSF is estimated to be below 300 Ìg three times weekly.
`Leukocyte elevation above 15 ! 109/l, attributable to
`increasing GM-CSF doses, was reported in 4 patients
`(19%) only: in 2 patients at a dose level of 150 Ìg leuko-
`cytes elevated up to 25 ! 109/l, and in 2 patients at doses
`of 60 and 90 Ìg GM-CSF. There was no obvious dose
`relationship between increasing doses of GM-CSF and the
`leukocyte counts. Low hemoglobin values due to WHO
`toxicity grade 2 requiring transfusions occurred in only
`3 patients (14%). No platelet values below 100,000/Ìl
`were reported. Concerning chemistry values of renal and
`liver function there were no significant differences detect-
`able between baseline and controls during treatment.
`
`Discussion
`
`GM-CSF and other cytokines are widely used to re-
`duce the period of neutropenia following chemotherapy
`to reduce infectious complications and allow the treat-
`ment to be given at full dose [8]. Concerning side effects of
`GM-CSF, Steward et al. [9] reported on pyrexia at doses
`above 1 Ìg kg–1 in all patients. These were clinically insig-
`nificant and resolved within 1–2 h. At doses of 10 Ìg kg–1
`day–1 the total leukocyte count elevated to more than
`
`Table 2. WHO toxicity of treatment with IFN-·2B and GM-CSF
`(n = 21)
`
`Side effect
`
`WHO grade
`
`Total
`
`Fever, chills, fatigue
`Nausea/vomiting
`Weight loss
`Dyspnea
`Cardiac ischemia
`Neurologic
`Somnolence
`Parethesias
`Local
`
`I
`
`4
`2
`2
`1
`0
`
`0
`5
`2
`
`II
`
`10
`4
`4
`0
`0
`
`0
`3
`1
`
`III
`
`4
`1
`1
`1
`1
`
`1
`0
`0
`
`18 (86%)
`7 (34%)
`7 (34%)
`2 (10%)
`1 (5%)
`
`1 (5%)
`8 (38%)
`3 (14%)
`
`250–400% of the starting values. The count fell to pre-
`treatment levels within 72 h of discontinuing therapy.
`Serious toxicity occurred at a dose level of 60 Ìg kg–1 with
`an acute onset of left-sided chest pain in 3 patients due to
`pericarditis [9]. Compared to the present study the maxi-
`mal dose of 300 Ìg GM-CSF three times weekly is nearly
`half the dose of 10 Ìg kg–1 day–1 and the therapy discon-
`tinued for 48 h. At these low dose levels, a leukocyte ele-
`vation up to 25 ! 10–9/l was detected in only 4 patients
`and the leukocyte counts increased maximally up to 100%
`above the starting values. The present data showed no
`obvious dose relationship between increasing doses of
`GM-CSF and the leukocyte counts. One patient devel-
`oped an acute anginal attack at the first GM-CSF admin-
`istration (300 Ìg). The symptoms resolved after discon-
`tinuation of GM-CSF. At dose levels above 90 Ìg GM-
`CSF, a modest increase in side effects was apparent. 5 of
`9 patients suffered grade-3 toxicity compared to 2 of 12
`patients in the first 4 groups. At the final dose level 2 of
`3 patients experienced unacceptable toxicity after only a
`short time of treatment, thus clearly indicating that the
`maximum tolerable dose had been surpassed when ap-
`plying 300 Ìg of GM-CSF.
`Initial reserves about the possibility of growth-promot-
`ing activity of GM-CSF on renal tumor cells in vitro at
`physiological concentrations [10] were later dissipated
`[11]. Other studies have ruled out an antitumor effect of
`GM-CSF, because it induces macrophage tumoricidal
`activity in vitro [12] and is capable of stimulating anti-
`body-dependent cytolysis of tumor cells by mature human
`neutrophils and eosinophils [6]. Hill et al. [5] demon-
`strated in a murine tumor model that GM-CSF (120 Ìg/
`kg) administered subcutaneously for 7 days was associ-
`
`Biological Response Modifiers in Renal Cell
`Carcinoma
`
`Urol Int 1998;61:215–219
`
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`randomized trial performed by Muss et al. [15], 97
`patients with metastatic RCC received recombinant IFN-
`·2B by either the subcutaneous or intravenous route.
`Approximately 60% of patients receiving intravenous
`therapy had grade-3 toxicity, while only 30% of patients
`on subcutaneous therapy had such difficulties. The subcu-
`taneous dosage was 2 ! 106 IU/m2 three times weekly
`escalating to dosages of 10 ! 106 IU/m2. In all patients, a
`flu-like syndrome occurred and in 33% nausea and vomit-
`ing. Somnolence was noted in 5% of the patients, compa-
`rable to the present study, and cardiovascular toxicity
`experienced as symptomatic tachycardia was limited to
`1 patient on intravenous therapy [15]. Therefore, the toxic
`profile of the present study is predominantly formed by
`the IFN-·.
`In conclusion, our results demonstrate that simulta-
`neous administration of GM-CSF and IFN-·2B is well
`tolerated up to doses of 120–150 Ìg GM-CSF three times
`weekly. But there is no additional antitumorigenic effect
`of GM-CSF because the overall response rate of the com-
`bined administration of GM-CSF/IFN-·2B is similar to
`IFN-·2B alone.
`
`Acknowledgements
`
`We would like to thank Dr. Axel Hinke for his statistical analysis.
`Our thanks also go to Stephanie Hendann for her time-consuming
`work with the monitoring.
`
`ated with a significant inhibition of tumor growth. The
`cytotoxicity of macrophages was enhanced by GM-CSF
`treatment with a parallel increase in TNF-· and IL-6 pro-
`duction of macrophages [5].
`The use of biologic response modifiers such as IFN-·
`as anti-tumor treatment has been explored throughout the
`last decade [1, 2]. Horoszewicz and Murphy [3] in 1989
`reviewed 16 separate trials that examined IFN-· in meta-
`static RCC in a total of 573 patients and reported about a
`combined response rate of 14% (83 of 573 patients). The
`best response rates were gained with intermediate doses of
`IFN-· (3–10 ! 106 U/day) [3]. IFN’s mechanism of anti-
`tumor activity is not completely understood. However,
`Kosmidis et al. [13] have shown that the administration
`of IFN-· in RCC patients results in the augmentation of
`T-cell responses and cytokine production in vitro. Be-
`cause of the in vitro and in vivo evidence of synergism
`between different biologic response modifiers [2], we used
`GM-CSF as an inducer of biological response modifiers in
`combination with the well-known IFN-·2B in patients
`with advanced RCC. The overall objective response rate
`in the present study was 13% (2 CRs), which is compara-
`ble to the combined response rate of 14% found by Horo-
`szewicz and Murphy [3]. Kosmidis et al. [13] using an
`equivalent IFN-· dose schedule of 15 ! 106 IU/week
`reported about 15% objective responses (4 PR of 26
`patients). The median survival time of 61 weeks or 14
`months in our study supports information from a pre-
`vious study of cytokine treatment in metastatic RCC per-
`formed at our institute. In this randomized trial of IFN-Á
`versus IFN-· and IL-2, the median survival time of 60
`patients amounted to 13 months [14]. Comparable to the
`above-mentioned IFN-· studies both CRs in the present
`study occurred in patients with lung metastases only. In
`regard to the dose levels of GM-CSF the 2 CRs were seen
`with 90 and 150 Ìg GM-CSF. The 9 patients with SD
`were evaluated at each dose level beside the highest dose
`level suggesting no clear dose-response relationship to
`increasing GM-CSF doses. Because of the missing dose
`relationship between increasing doses of GM-CSF and the
`evaluated remissions, it is obvious that GM-CSF did not
`contribute to treatment outcome in this combination.
`The toxicity of IFN-· is dose-related and reversible.
`Daily doses of 1–9 ! 106 U/day are tolerated well by most
`patients [2, 15]. In the present trial, 86% of the patients
`experienced a flu-like syndrome with fever, chills, and
`fatigue and 34% of the patients developed nausea, vomit-
`ing, and weight loss. Three of 5 patients which were with-
`drawn from study due to grade-3 toxicity suffered from
`flu-like syndrome, nausea, vomiting, and weight loss. In a
`
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`218
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`Urol Int 1998;61:215–219
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`Lümmen/Sperling/Luboldt/Otto/Rübben
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`References
`
`1 deKernion JB, Sarna G, Figlin R, Lindner A,
`Smith RB: The treatment of renal cell carcino-
`ma with human leukocyte alpha-interferon. J
`Urol 1983;130:1063–1066.
`2 Choudhury M, Efros M, Mittelman A: Interfer-
`ons and interleukins in metastatic renal cell
`carcinoma. Urology 1993(suppl 41):67–72.
`3 Horoszewicz JS, Murphy GP: An assessment of
`the current use of human interferons in therapy
`of urological cancers. J Urol 1989;142:1173–
`1184.
`4 Hansen F: Hematopoietic growth and inhibito-
`ry factors in treatment of malignancies. Acta
`Oncol 1995;34:453–468.
`5 Hill ADK, Redmond HP, Naama HA, Bou-
`chier-Hayes D: Granulocyte-macrophage colo-
`ny-stimulating factor inhibits tumor growth
`during the postoperative period. Surgery 1996;
`119:178–185.
`6 Metcalf D, Begley CG, Johnson GR, Nicola
`NA, Vadas MA, Lopez AF, Williamson DJ,
`Wong GG, Clark SC, Wang EA: Biological
`properties in-vitro of a recombinant human
`granulocyte-macrophage
`colony-stimulating
`factor. Blood 1986;67:37–45.
`
`7 Kaplan EL, Meier P: Nonparamatic estimation
`from incomplete observations. J Am Stat Assoc
`1958;53:457–481.
`8 Ferguson JE, Dodwell DJ: High-dose dose-
`intensive chemotherapy with doxorubicin and
`cyclophosphamide for the treatment of ad-
`vanced breast cancer. Br J Cancer 1993;67:
`825–829.
`9 Steward WP, Scarffe JH, Austin R, Bonnem E,
`Thatcher N, Morgenstern G, Crowther D: Re-
`combinant human granulocyte macrophage
`colony-stimulating factor (rh GM-CSF) given
`as daily short infusions – A phase I dose-toxici-
`ty study. Br J Cancer 1989;59:142–145.
`10 Asano S, Sato N, Mori M, Ohsawa N, Kosaka
`K, Ueyama Y: Detection and assessment of
`human tumours producing granulocyte-macro-
`phage colony-stimulating factor (GM-CSF) by
`heterotransplantation into nude mice. Br J
`Cancer 1980;41:689–694.
`11 De Riese W, Allhoff EP, Werner M, Stief CG,
`Liedke S, Kirchner H, Atzpodien J: Effects of
`cytokines on growth in vitro of primary human
`renal cell carcinoma. Urol Res 1992;20:369–
`373.
`
`12 Grabstein KH, Urdal DL, Tushinski RJ, Mo-
`chizuki DY, Price VL, Cantrell MA, Gillis S,
`Conlon PJ: Induction of macrophage tumorici-
`dal activity by granulocyte macrophage colony-
`stimulating factor. Science 1986;232:506–508.
`13 Kosmidis PA, Baxevanis CN, Tsavaris N, Pa-
`panastasiou M, Anastasopoulos E, Bacoyiannis
`C, Mylonakis N, Karvounis N, Bafaloukos D,
`Karabelis A: The prognostic significance of im-
`mune changes in patients with renal cell carci-
`noma treated with interferon alpha-2b. J Clin
`Oncol 1992;10:1153–1157.
`14 Lümmen G, Goepel M, Möllhoff S, Hinke A,
`Otto T, Rübben H: Phase II study of interfer-
`on-Á versus interleukin-2 and interferon-·2b in
`metastatic renal cell carcinoma. J Urol 1996;
`155:55–58.
`15 Muss HB, Costanzi JJ, Leavitt R, Williams
`RD, Kempf RA, Pollard R, Ozer H, Zekan PJ,
`Grumberg SM, Mitchell MS, Caponera M,
`Gavigan M, Ernest ML, Venturi C, Greiner J,
`Spiegel RJ: Recombinant alpha interferon in
`renal cell carcinoma: A randomized trial of two
`routes of administration. J Clin Oncol 1987;5:
`286–291.
`
`216.185.156.28 - 11/14/2016 73333 PM
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`Carcinoma
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