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• ••:•'• W äf.:v [CANCER RESEARCH 45, 4447-4453, September 1985] Intraperitoneal Recombinant «-Interferon for "Salvage" Immunotherapy ¡n Stage III Epithelial Ovarian Cancer: A Gynecologic Oncology Group Study1 Jonathan S. Berek,2 Neville F. Hacker, Alan Lichtenstein, Tyler Jung, Gelsa Spina, Reba M. Knox, Jeanine Brady, Teri Greene, Lupe M. Ettinger, Leo D. Lagasse, Eric M. Bonnern, Robert J. Spiegel, and Jacob Zighelboim Division of Gynecologic Oncology, the Departments of Obstetrics and Gynecology [J. S. B., N. F. H., T. J., R. M. K., T. G., L. M. E., L D. L], Medicine, and Microbiology and Immunology [A. L., C. S., J. Z.], the UCLA School of Medicine, and Jonsson Comprehensive Cancer Center [J. S. B., N. F. H., L. D. L], Los Angeles, California 90024; and the Schering Corporation ¡J.B., £.M. B., R. J. S.], Kenilworth, New Jersey 07033; and the Gynecologic Oncology Group [J. S. B., N. F. H., L. M. £.,L. D. L.], Philadelphia, Pennsylvania 19107 ABSTRACT Fourteen patients with persistent epithelial ovarian cancer documented at second look laparotomy after combination chem otherapy were treated with 146 cycles of a-recombinant interferon (r!FN-a2) administered i.p. The initial dose was 5 x 106 units which was escalated weekly to 50 x 106 units over 4 weeks and then continued weekly for a total of 16 weeks. Eleven patients underwent surgical réévaluation after therapy which confirmed four pathological complete responses (36%), one par tial response (9%), and disease progression in six patients (55%). Five of seven patients (71%) with residual tumor <5 mm had a surgically documented response, whereas there was no re sponse in the four patients whose tumors were >5 mm. Three patients were évaluablefor clinical response only: one patient who refused surgery had a complete clinical response with total resolution of ascites; one had stable disease; and one had disease progression. Fever >38°C was seen in 58%, fever >39.0°C was seen in 18%, vomiting in 37%, abdominal pain was reported in 22%, and one patient had infectious peritonitis. Peripheral white blood cell counts and i.p. washings were ob tained pretreatment and on days 1, 3, and 7 after treatment. While there was no consistent alteration in peripheral white blood cell counts, the numbers of i.p. monocytes and lymphocytes showed a significant boost on day 1 after each dose of rlFN-a2. Natural killer lymphocyte cytotoxicity was elevated in the i.p. cavity fluid obtained from most patients on day 1 after treatment, while blood natural killer lymphocyte cytotoxicity values showed considerable variability. Pharmacokinetic studies show that i.p. levels of rIFN-az were 30-1000 times blood levels. rlFN-«2i.p. may act by increasing concentrations of drug and augmenting regional host cells in patients with minimal residual ovarian cancer. INTRODUCTION IFNs3 are a diverse group of locally active, naturally occurring, regulatory glycoproteins which are currently undergoing exten sive clinical testing as antineoplastic agents (1-9). Their biological 1Supported by grants from the Schering Corporation, the Concern Foundation, and The National Cancer Institute (Grant CA 12800, NIH); a USPHS CRC Grant RR865; and Gynecologic Oncology Group Grant CA 13630-13. 2 To whom requests for reprints should be addressed, at Division of Gynecologic Oncology, Department of Obstetrics and Gynecology, UCLA School of Medicine, Los Angeles, CA 90024. 'The abbreviations used are: IFNs, interferons; rlFN-a2, a-recombinant inter feroni NK, natural killer lymphocyte cytotoxicity; MRD, minimal residual disease; BRM, biological response modifier. Received 3/18/85; revised 6/5/85; accepted 6/7/85. CANCER RESEARCH activities are diverse, and in addition to their ability to modify immunoregulatory systems (1, 2) they possess direct antineo plastic activity. These agents alter tumor cell function and struc ture and induce cellular inhibition of replication of many viruses (1)Therapeutic responses have been observed in a variety of tumor systems (1-5, 7, 8). Human a-interferon, a specific antigenie type produced mainly by leukocytes (2), is now produced in highly purified, large quantities because genetic recombinant DNA technology has permitted the production of specifically cloned products (8). In our experience patients treated with combination chemo therapy for epithelial ovarian cancer and in clinical remission have a 60-70% rate of persistent disease at second look laparotomy (10). Therefore there is a need for improved primary or secondary therapies in order to improve survival and cure rates. One approach which has been suggested is to treat the residual disease with the i.p. administration of antitumor agents (11-13). Complete and partial responses have been surgically documented in response to the i.p. administration of Corynebacterium parvum as second line therapy in epithelial ovarian cancer patients (11,12). These responses have been documented only in patients with MRD (;.e., <5 mm maximum diameter of the residual tumor) at second look operations. However, since this BRM is relatively toxic when administered i.p., BRMs which induce a less intense peritonitis might be more practical and effective (13,14). The i.p. instillation of BRMs for ovarian cancer has clinical appeal because of the potential for increasing the drug concentration at the tumor site and decreasing systemic toxicity(11-15). This report is a trial of the i.p. administration of recombinant (rlFN-«2)in patients with persistent epithelial ovarian cancer documented at second look laparotomy. Evaluation of efficacy, toxicity, pharmacokinetics, and host cell parameters are pre sented. MATERIALS AND METHODS Patients. A total of 14 patients with persistent epithelial ovarÃ-ancancer were treated with 146 cycles of ¡.p.r!FN-a2. All patients were évaluable for clinical response and toxicity. Eleven patients were surgically reevaluated after therapy. Serial r!FN-a2 levels, studies of host cell populations, and lymphocyte cytotoxicity assays were performed on peritoneal wash ings and peripheral blood. Patients ranged in age from 42 to 66 years with a mean age of 53.4 years. All patients were classified as International Federation of Gyne cology and Obstetrics stage III. No patient had documented extraperitoneal disease at the initiation of immunotherapy. VOL. 45 SEPTEMBER 1985 4447 Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1985 American Association for Cancer Research. INTERFERON ¡.p.IN OVARIAN Residual tumor prior to rlFN-a2 treatments ranged from microscopic disease (positive cytology only) to a 4- x 4-cm palpable pelvic mass. Six patients (43%) had only positive peritoneal cytology (classes IV and V), two patients (14%) had measurable disease <5 mm in diameter, one patient (7%) had disease 5 to 15 mm in diameter, and five patients (36%) had disease >15 mm. Papillary serous cystadenocarcinorna was the histological type in 11 of the 14 patients. Two patients had grade I tumors, six had grade II lesions, and six had grade III lesions. Cytology of peritoneal fluids were graded as follows: class I or II, negative; class III, equivocal or suspicious; and class IV or V, positive for frankly malignant cells. Prior to i.p. rlFN-«2therapy each patient had undergone one to three exploratory laparotomies to resect tumor, including a total abdominal hysterectomy, bilateral salpingooophorectomy, as well as 5 to 13 (me dian, 9) cycles of combination chemotherapy with c/s-platinum, doxorubicin, and cyclophosphamide. Two patients previously had received ex ternal pelvic radiation therapy. Immediately prior to entry onto this study, a second look minilaparotomy was performed at which time two Tenckhoff catheters were implanted using standard techniques (11): the tip of one catheter was directed toward the pelvis; and the tip of the second catheter was directed cephalad up the right paracolic gutter. An operation to evaluate response to rlFN-«2were performed 1 to 6 weeks following the final course of r!FN-a2 as described previously (10,16). The protocol was approved by the Human Subject Protection Committee of the University of California, UCLA. Dialysis fluid was instilled into the peritoneal cavity (Dyalasate with 1.5% dextrose) after being warmed to 37°Cand was supplemented with sodium heparin (500 units/liter; Panheparin; Abbott Laboratories). i.p. Therapy. After catheter placement several peritoneal lavages were performed using 1 to 2 liters of dialysis fluid until peritoneal rinses were free of blood. Three consecutive blood free peritoneal lavages were required before the first rlFN-«2treatment. Catheters were flushed every other day with 10 ml of heparinized saline except for treatment days in which only saline solution was used. Once rlFN-«2treatment began peritoneal lavages were performed on days 2 and 7. Aliquots obtained prior to each lavage were examined cytologically for malignant cells, and total WBC differential counts and immunological assays of peritoneal host cells were performed. Interferon Administration. Human rlFN-«2(SCh 30500, Schering Corporation, Kenilworth, NJ) was administered i.p. in 250 ml normal saline after 1750 ml of dialysis solution were instilled i.p. via Tenckhoff catheters. Ten patients in the first dose schedule of the study received an escalating weekly dosage of r!FN-a2. The dosage of rlFN-a2 was initially 5 x 10" units, which was escalated to 10 x 10" and then 20 x 10" units and then followed by five consecutive courses at 50 x 106 units. The second dose schedule initially called for patients to be infused with rlFN«2three times a week (every other day) at 50 x 106 units. This regimen was to be followed every other week for 16 weeks. However, because of toxicity the second dosage schedule was modified to 50 x 10" units a week for 8 more weeks. Patients were premedicated with 25 mg diphenhydramine hydrochlo- ride and 650 mg acetaminophen p.o. The former was discontinued during the second dose schedule because no major allergic reactions from rlFN«2were observed and the patients complained of somnolence. Acetami nophen was given every 4 h for the duration of the hospital visit (usually 24 h) and was then taken by patients upon discharge as needed. When 24 h had elapsed after rlFN-«2treatment, 2 liters of warmed dialysis fluid were instilled into the peritoneal cavity and recovered for cytological and immunological assays. The exposed ends of the Tenck hoff catheters were resealed aseptically. Response Criteria. Response was defined as follows: complete re sponse, no evidence of disease at the completion of therapy, including the absence of malignant cells in the peritoneal fluid cytological evaluation in those patients whose cytologies had been positive; partial response, the presence of a positive cytology only at the completion of therapy in CANCER RESEARCH CANCER those patients who had had macroscopic disease, or the regression of macroscopic disease by at least 50% in tumor dimension; stable disease, no change in the positive cytology and/or the tumor dimension; progres sive disease, evidence of tumor growth during or at the completion of treatment. Toxicity. Toxicity was monitored and graded according to the criteria outlined by the Gynecologic Oncology Group (15) and modified to adapt to the expected effects of rlFN-a2. rlFN-«2routinely causes fevers which at times are above 39°Cand this was not considered a toxicity requiring dose reduction. For rlFN-«2dose-limiting toxicity is usually a cumulative one based on the patients' general condition and symptoms, e.g., fatigue and malaise, rather than objective laboratory parameters (1, 2). Toxicities which may be encountered by the patients receiving rlFN<*2include: (a) fatigue and malaise. If a subject experienced fatigue to the extent that she was unable to ambulate, the rlFN-«2was held. One to 2 weeks after the patient was again ambulatory, the rlFN-a2 was resumed on the regular schedule at the last maximum tolerated dosage, or reduced by 25%, whichever dose was higher; (ù) weight loss. The guidelines listed below were followed for weight loss believed to be solely induced by the rlFN-«2(for >15 pounds reduce dosage by 50%; for >20 pounds, hold treatment); (c) abdominal pain if the pain was intermittent over a time period of <24 h and required no analgesics, it was considered mild (grade I), if it lasted >24 h and necessitated p.o. analgesics which relieved the pain it was grade II, and severe (grade III) pain included that which was continuous weekly and not responsive to p.o. analgesics; (d) chills: grade I, those that lasted <10 min at a time and did not restrict patient activity in any way; grade II, those that lasted >10 but <30 min at a time and did not confine the patient to bed; and grade III, rigorous shivering that required the patient to stay in bed and that lasted continuously for a period of >30 min; (e) headache: grade I, those lasting <24 h relieved by p.o. analgesics; grade II, those that lasted >24 h and <48 h and are temporarily relieved by p.o. analgesics; and grade III, those lasting >48 h and temporarily relieved by p.o. analgesics; (f) temporary anorexia: grade I, <24 h; grade II, >24 h and <72 h; and grade III, >72 h; (g) pruritis: grade I, <24 h; grade II, >24 h and <48 h; and grade III, >48 h. Blood Lymphocyte Analysis. Ten ml of blood were drawn before treatment for base-line analysis and one to three times/week during treatment. Samples were assayed for differential WBC, lymphocyte populations, and cytotoxic activity. Peritoneal Cell Analysis. Peritoneal lavage fluid specimens were obtained two to three times/week during rlFN-«2treatment. An aliquot of 50 ml of fluid was examined cytologically for malignant cells. The remaining dialysis fluid was centrifugea at approximately 700 rpm for 5 min and resuspended with minimal essential media. Total WBC/ml were counted, Cytoprep slides were made, and differential cell counts/ml were performed. Separation of peritoneal cells was performed with a FicolhHypaque density gradient as described previously (18). Specimens which had high concentrations of macrophages were treated with GAF iron fillings and removed with a magnet (19). Differential WBC were performed. Lympho cyte populations were isolated and a specimen was assayed by the UCLA Clinical Immunological Research Laboratory for lymphocyte cytctoxicity. Natural Killer Cell Activity. A constant number of "Cr-labeled target cells (1 x 104) was mixed with varying numbers of lymphocyte effector cells at an effectortarget ratio of 25:1. Cultures were incubated in V-well mterotiter plates for 3 h at 27°C. Supernatant was collected and 51Cr release was assayed by -y-émissionaccording to the method of Jondal ef al. (20). The percentage of lysis was calculated as [cpm of target + effector] - [cpm of target alone] % of lysis Addition [cpm of target + detergent (Triton X)] - [cpm of target alone] of detergent VOL. 45 SEPTEMBER to targets evaluated x 100 maximal release (always 1985 4448 Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1985 American Association for Cancer Research. INTERFERON Lp. IN OVARIAN greater than 85% of incorporated cpm) while culture of targets alone served as background release (always less than 20% of incorporated cpm). The myeloid line K562 served as the target cell of the NK assay. Neutralizing Antibodies. Frozen serum samples from each patient were obtained before and after treatment and were sent to the Schering Corporation Clinical Laboratories to assay for generation of antibodies specific for the rIFN-az. The procedure for detection of anti-IFN-aj antibody in human sera is a radioimmunological technique to screen for antibodies as described previously (21). Pharmacokinetics. From each patient 10 ml of blood were drawn prior to rIFN-az (50 x 106) and 24 h afterward. Additionally five patients were studied after the administration of a rlFN-«2treatment at time 0 (pretreatment = base line), 4, 8, 12, 20, and 24 h from initiation of the i.p. infusion. Blood samples were centrifugea and subsequent frozen serum specimens were sent to the Schering Corporation Clinical Labo ratories to determine rIFN-az levels. The method for measuring rIFN-az levels utilized a human foreskin diploid cell line, the murine EMC virus, and a crystal violet stain to visually detect cytopathic effects as described (22). Approximately 50 ml of dialysis fluid were obtained from peritoneal lavages at pretreatment (time = 0) and at 4 and 24 h following 50 million units of rIFN-az infused at time 0. Additional peritoneal fluid samples (48 and 72 h) were obtained by peritoneal lavages consisting of approxi mately 1 liter of dialysis fluid. Specimens were frozen immediately and sent to the Schering Corporation Clinical Laboratories to determine rlFN<i2levels. RESULTS CANCER and progressed, one developed a 2- x 2-cm retroperitoneal tumor nodule which was removed. After whole abdominal radiation treatment she remained clinically free of disease for 12 months before abdominal recurrence. The other patient with microscopic disease was found to be free of disease by laparoscopy following her first eight cycles of treatment, but progressive abdominal disease was documented at the completion of the protocol by laparotomy where 1-2 mm studding on the small bowel mesen tery was documented. Aliquots of peritoneal fluid were obtained weekly during the course of therapy and examined cytologically for malignant cells (Chart 1). The six patients who were clinically free of disease and evaluated surgically at the completion of therapy retained evidence of malignant cells throughout treatment (class IV or V cytology), whereas those four patients who had surgically doc umented complete responses developed negative cytology by week 8 of treatment. This corresponds to the week after the patients had received their fourth treatment cycle at the maxi mum dose of 50 x 106 units/week. Of the three patients who were only clinically évaluable,one had a complete resolution of ascites during treatment and re mains clinically free of disease 16 months posttreatment. One patient had stable disease during treatment and remains stable after 4 months with 2 cm disease palpable in the rectovaginal septum. The third had a 3-4-cm mass palpable in the rectovaginal septum progress to 5-6-cm disease after eight treat ments. Response Toxicity Four of 11 surgically evaluated patients demonstrated com plete responses to rlFN-<v2treatment. Tumor disappearance was documented by a negative reassessment laparotomy and at least six consecutive negative (class I or II) peritoneal cytological washings at the completion of rlFN-«2therapy. Three of the complete responders entered the program with microscopic disease as documented by positive cytology only, and the other had 5-10 residual tumor nodules, approximately 1-2 mm in diameter in the pelvic peritoneum. All four patients remain alive without recurrence from 5 to 14 months posttreatment. An additional patient had a partial response. This patient began the study with 6-10 peritoneal tumor nodules of 2-3-mm tumors in the pelvic cul-de-sac. Upon reassessment laparotomy the patient had no macroscopic evidence of disease, but a specimen for cytological evaluation obtained from the upper abdomen was positive for persistent malignant cells. Surgically documented responses by residual disease are listed in Table 1. Six surgically evaluated patients had disease progression dur ing treatment. Four of these patients entered treatment with intraabdominal tumor nodules >1.5 cm in diameter, and two patients had only microscopic disease following optimal debulking. Of the two patients who entered with microscopic disease Objective and subjective systemic reactions were monitored throughout the study in all patients (Table 2). Of the 146 courses monitored for subjective side effects, fatigue was noted in 65%, chills in 53%, temporary anorexia in 39%, headaches in 38%, nausea and/or vomiting in 37%, diarrhea in 27%, abdominal pain in 22%, and total body pruritis in 12%. Other common complaints included "abnormal" taste, "dry" mouth, back pain, and muscle aches, all of which were mild to moderate. Vital signs were monitored during the first 24 h after treatment in 132 courses. Fever (>38°C) was documented in 77 courses Non-responders I ffiII . n- Responders Itti i- IFN Dose*5 t LAP IO 20 SOxIcf uiits • Table 1 Surgically documented response by residual disease Maximum diameterCytology positive 00 of patients5a 2»4 1 04Partial0 11ResponseComplete3 CANCER RESEARCH 2 3 4 5 6 7 Time («neks) 8 9 IO II I2 I3 I4 I5 I6 Chart 1. Plot of mean of the cytologicalclasses of peritonealwashings by grade for responders and nonresponders in a group of 11 surgically évaluable patients treated with i.p. rlFN-.»2. Cytological grades I and II are negative, grade III is equivocal, and grades IV and V are frankly positive for malignant cells. Following two baseline(time = 0) cytological assays, specimens were obtained weekly while on treatment. The maximum dose of 50 x 10* units r!FN-a2was reached by week 4 and repeated weekly thereafter. LAP, laparotomy. 1 100 01%60 0 45 * Five of seven (71%) of patients with <5 mm disease responded to therapy. <5 mm >5mm TotalNo. I VOL. 45 SEPTEMBER 1985 4449 Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1985 American Association for Cancer Research. INTERFERON i.p. IN OVARIAN CANCER Table 2 Toxicity of patients receiving i.p. % of patients with side effects Leukopenia0Day Grade of Toxicity1. Paina38°C: Fever8 40 Mild >39°C:17 II. Moderate >40°C: 111 III. SevereAbdominal Total 58 and vomiting21 Chills19 anorexia31 131 1347 Day 794 16 10 33 1Malaise30 2Headache3 36 16 16Diarrhea19 8Pruritis6 6Temporary7 22 24 1Nausea 1Anemia0 5Day 4 26 9 1Day 22 53 36 65 38 37 12 27 39 58 62 13 * Fever assessed in 132 of 146 cycles. " Anemia and leukopeniaassessed in 140 of 146 cycles. (58%). The degree of fever is listed in Table 2. Response could not be correlated with the fever because both nonresponding and responding patients had systemic temperature evaluations. A slight increase in pulse without tachycardia was measured in some patients during this first 24-h period; however, in all cases pulse rates returned to normal values within 24 h. Respiration and blood pressure remained normal during treatment. Hematological analysis in 140 courses was performed pre treatment and 24 h and 7 days after treatment. Base-line (pre treatment = time 0) blood hemoglobin levels showed only 5% anemia, but by day 1, 31 % had grade I anemia, 22% had grade II anemia, and 5% had grade III anemia. By day 7, 34% had grade I toxicity, 24% had grade II toxicity, and 4% had grade III toxicity. With base-line (pretreatment = time 0) values of leuko penia in only 1% of patients, leukopenia was noted in 36% of cases on day 1 and 13% of cases on day 7. In 90% of the courses the total WBC on day 1 was less than pretreatment value, but only 36% were <3000/ml (10% grade III), and those values returned to normal by day 3 in most cases. Additionally in 88% of these courses the WBC on day 7 was greater than the WBC on day 1, with only 13% having WBC <3000/ml on day 7. The mean base-line WBC for the 14 treated patients was 5350/ml. This level decreased slightly during treatment with mean values of 4400 on day 1 and 4900/ml on day 7. WBC for responders and nonresponders were not significantly different. Blood chemistry tests performed every week of treatment revealed no elevation of liver enzymes, renal function tests, or other abnormalities. The dose limiting toxicity occurred in the second dose sched ule phase of the study during which patients received 50 x 106 units rlFN-a2 three times every other week, but after two cycles persistent moderate toxicity (fatigue, abdominal pain, and head aches) required that the treatment schedule be modified so that the rlFN-a2 was given weekly thereafter. In an additonal two patients Tenckhoff catheters were inserted with the intention of treating the patients with i.p. r!FN-o2. One patient developed a clotted catheter and decided not to continue the study, and the catheter was removed prior to any r!FN-a2 treatment. Another patient had a catheter placed at the time of laparotomy but developed a postoperation thrombophlebitis and was thus excluded from the protocol. Immunological Results Peripheral Blood. Differential counts revealed that lympho cytes decreased each cycle for a mean base-line value of 1530/ ml to 550/ml on day 1 and then rapidly returned to near baseCANCER 100r •Total A Mono o Lymph •Poly IE 50 r 0 1 """ ;> "3 4 5 6 7 Time (days) Chart 2. Fluid WBC of patients treated with i.p. r!FN-a2 are presented. After three baseline values (pretreatment = time 0), specimens were obtained on days 1, 3, and 7 after most weekly treatments. Day 7 also represents the time immedi ately prior to the subsequenttreatment. Valuesrepresent the mean integrated sum of all treatment cycles (oars, SD) showing a rapid rise in total WBC, lymphocytes, and mononuclear cells by day 1 after treatment, and a gradual return to below baselineby day 7. Total WBC are plotted, as well as polymorphonuclearleukocytes (Poly),lymphocytes, and monocytes. line levels of 1300/ml by day 7. Blood monocytes and polymor phonuclear leukocytes remained relatively constant throughout treatment. Peritoneal Fluid. The mean base-line peritoneal WBC for treated patients was 48,100/ml. A consistent increase was seen in the WBC by day 1 in all patients (83,100/ml), a mean value which returned to levels slightly below base line by day 7, i.e., 22,000/ml (Chart 2). The absolute numbers of peritoneal mono cytes and lymphocytes increased on day 1 of each cycle after treatment and returned to base-line values by day 7. Monocyte numbers increased from a mean base-line value of 16,300/ml to 52,600/ml on day 1, while lymphocytes increased from a mean base-line value of 12,300/ml to 26,500/ml on day 1 and returned to 6,600/ml by day 7. By contrast polymorphonuclear leukocyte levels dropped from a base line of 19,500/ml to 4,000/ml on day 1 and rose to 7,700/ml on day 7. There were no significant differences in WBC counts and differential populations between responders and nonresponders. Some increase in total cell counts were observed by the seventh and eighth weeks of therapy. NK Evaluation of NK (effectortarget ratio, 25:1) in blood and peritoneal fluid is presented (mean ±SE) in Chart 3. The NK RESEARCH VOL. 45 SEPTEMBER 1985 4450 Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1985 American Association for Cancer Research. INTERFERON i.p. IN OVARIAN CANCER io" i 40 id1 • 30 k •H20 io1 • 10 iplFN io'O 2 3 4 Time (days) Chart 3. NK. Points, the mean percentage of specific lysis of each patient; bars, SE. The peripheral blood lymphocytes ( ) and the peritoneal cavity lymphocytes ( ) are presented. Time zero = baseline levels prior to initial treatment, and day 1 is the mean of NK 24 h after i.p. administration of rlFN-a2- Specimens taken 7 days after treatment are also those taken immediately prior to subsequent treat ment. base line in the blood was 27 ±3, which was 24 ±3 by day 1, and rose slightly to 32 ±2 by day 7. In contrast the i.p. NK values were markedly boosted from a base line of 6 ±2 to 31 ±4 on day 1 and fell to 10 ±2 by day 7. Over the successive treatment weeks alterations in NK were relatively homogeneous with no major variation over time; I.e., there was no cumulative boost or depression in the NK over 8 to 16 weeks. Boosting of NK did not uniformly relate to response; i.e., while all patients who had a documented response had significant elevation of i.p. NK on day 1, five of the eight patients who did not have a response also demonstrated some NK elevation. 5 IO 15 20 25 30 35 40 45 50 55 60 65 70 75 Tim« (hours) Chart 4. Mean values of blood ( ) and ¡.p.fluid ( ) r!FN-a2 levels following the i.p. administration of 50 x 10' r!FN-«2;oars, SE. DISCUSSION This is the first report demonstrating efficacy for the i.p. administration of rlFN-a2 in patients with persistent MRD epithe lial ovarian cancer detected at second look laparotomy. The documentation of complete responses in 4 of the 11 surgically evaluated patients suggests that this biological response modi fier may prove useful in the treatment of MRD ovarian cancer. Responses appeared to occur after four to six cycles of 50 x 106 units of rlFN-a2, as indicated by the results obtained with serial peritoneal fluid cytologies performed on these patients. The toxicity of the i.p. rlFN-a2 was moderate; the principal problem related to the use of the i.p. catheters. In this study the rlFN-a2 pharmacokinetic data suggest that very high i.p. concentrations of the agent are present for at least 24 h, at which time dialysis solution is withdrawn for study. Neutralizing Antibodies However, a low level of rlFN-a2 is detectable 48 and 72 h after treatment. Peripheral blood levels remained elevated for 5 to 6 days. These data might help to explain the biological efficacy of Multiple serum samples obtained prior to treatment and on rlFN-02. Also the serum levels of rlFN-«2are quite high in days 1 and 7 after treatment were tested for neutralizing anti bodies specific to the human rlFN-<*2. No antibodies were de comparison to the usual levels achieved with standard i.m. or s.c. administration, which usually ranges approximately 200 to tected at any time. 300 units/ml (1-8). Thus the peritoneal cavity may be acting as a reservoir, leading to both constant surface exposure and to a IFN Pharmacokinetics persistent serum level. This feature has implications for cell cycle studies as indicated by in vitro data which demonstrate that Serum. Serum r!FN-a2 levels rose from pretreatment base line constant exposure to r!FN-«2produces more marked cytotoxic of <38 units/ml to levels of 75-300 units/ml 8 h after treatment and cytostatic effect than does pulse exposure (3, 4). This (Chart 4). By 24 h serum rlFN-«2levels consistently ranged from antitumor activity might result from exposure of the ovarian cancer cells to high concentrations of rlFN-a2. 150 to 600 units/ml. Serum samples taken from day 2 through day 7 were scarce due to the logistics of patient travel to and It is of interest that only patients with MRD (<5 mm maximum from the hospital on "off" days; however, from the small number tumor diameter) achieved a surgically documented response and of samples that we were able to obtain, it appears that rlFN-a2 that patients with larger disease did not respond. Indeed five of levels remained high or above base-line values up to day 6. seven surgically evaluated patients (71%) with MRD had a Serum levels returned to base line by day 7. response. As such, future trials of this agent should probably be Peritoneal Fluid. Peritoneal fluid rlFN-o2 levels rose from a confined to patients with MRD. These data are similar to our pretreatment level of <38 units/ml up to approximately 38,400 previous experience using i.p. immunotherapy (11,12), chemo units/ml during the first 24 h of treatment at Interferon dosage therapy (13), and whole abdominal radiation therapy (23) and of 50 x 106 units of rlFN-a2 (Chart 4). Thereafter coinciding with with data obtained using a murine ovarian cancer model (14,24removal of dialysis fluid at the 24-h mark, levels of rlFN-a2 in the 26). Therefore it would appear that the tumor burden prior to the peritoneal cavity returned to base line. Measurements of rlFN-«2 initiation of therapy is critical in these patients; thus surgical levels in lavages at day 2 (48 h), day 3 (72 h), and day 7 (168 h) resection of tumor might be important if larger tumors are present were equal to base-line levels of <38 units/ml. at the time of exploratory surgery (27). CANCER RESEARCH VOL. 45 SEPTEMBER 1985 4451 Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1985 American Association for Cancer Research. INTERFERON Lp. IN OVARIAN The only previous studies of the use of systemically adminis tered IFNs for the treatment of ovarian cancer have been carried out in Scandinavia (2, 9) and the United States (28). The first group of authors reported partial clinical responses in 2 of 18 previously treated patients, 3 patients with "minimal" responses, and 3 patients who had stable disease for up to 1 year. The second group of authors (27) reported no responses, while 4 of their 13 patients had stable disease (median, 6 months). In our experience ovarian cancer often remains confined to the peritoneal cavity, even after clinical recurrence (10). Intracav- patients after primary cytoreductive surgery in combination with chemotherapy or in an adjuvant setting after a negative second look laparotomy. REFERENCES itary therapies are appealing because of their potential for in creasing the concentration of the agent and bringing the agent into direct contact with the residual tumor (6,11 -15). Historically BRMs have been most effective in patients in whom the agent has been administered intralesionally or intracavitarilly (6,11-15, 24, 25). Also the presence of tumor cytotoxic chemotherapy appears to be immunosuppressive in patients with ovarian can cer (19). BRMS have generally been unsuccessful in the treatment of ovarian cancer (13). However, some studies have suggested that combinations of systemically administered Bacillus Calmette-Guer/n (18) and C. parvum (30) administered concomitantly with cytotoxic chemotherapy induce a higher proportion of clini cally documented complete responses than the use of combi nation chemotherapy alone. The i.p. administration of C. parvum in patients with MRD documented at second look has produced surgically documented responses (11,12). NK and antibody dependent cell mediated cytotoxicity may be partially responsible for tumor control and/or proliferation in the peritoneal cavity (11,12,19,31). These effector cells have been shown to be deficient in the peritoneal cavities of women with ovarian cancer previously treated with combination chemother apy (18). The i.p. administration of C. parvum has been reported to augment peritoneal cavity cytotoxic effector lymphocytes, both NK (12,31) and antibody-dependent cell mediated cytotox icity (11, 12), in responding patients. Since rlFN-<*2 has been shown to be a potent stimulator of NK (6), its i.p. administration to patients with persistent epithelial disease thus seems rational. Our measurement of both blood and i.p. fluid NK in relationship to the rlFN-02 treatments shows that i.p. NK is significantly boosted after 1 day, while blood values showed considerable variability. No cumulative boosting or suppression is demon strated. Also the values do not correlate with the objective response. The temporal dynamics of NK effectors relative to the rlFN-(>2 pharmacokinetics are the subject of a separate report.4 Although mononuclear cells were not tested for cytotoxicity in these patients, future studies will focus on these potentially important cells. IFNaS have been shown to have an antiproliferative effect against ovarian cancer cells grown in human tumor stem cell assays (32). Also there is evidence that IFNs may act synergistically with cytotoxic chemotherapeutic agents in vitro (33-36). These data suggest that the exposure of cancer cells to rlFN-a2 prior to or concomitant with cytotoxic agent(s) can increase cell kill. Therefore i.p. rlFN-<*2may be potentially useful in addition to cytotoxic chemotherapy in patients with ovarian cancer. Indeed there is the potential for the use of rlFN-a2 in the treatment of 4 C. Spina, A. Uchtenstein, J. S. Berek. and J. Zkjhelboim. 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Intraperitoneal Recombinant α-Interferon for ''Salvage'' Immunotherapy in Stage III Epithelial Ovarian Cancer: A Gynecologic Oncology Group Study Jonathan S. Berek, Neville F. Hacker, Alan Lichtenstein, et al. Cancer Res 1985;45:4447-4453. Updated version E-mail alerts Reprints and Subscriptions Permissions Access the most recent version of this article at: http://cancerres.aacrjournals.org/content/45/9/4447 Sign up to receive free email-alerts related to this article or journal. To order reprints of this article or to subscribe to the journal, contact the AACR Publications Department at [email protected]. To request permission to re-use all or part of this article, contact the AACR Publications Department at [email protected]. Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1985 American Association for Cancer Research.