Adjuvant chemo-immunotherapy after curative resection of Stage II and IIIA primary lung cancer

LUNGI CANCER Lung Cancer 14 (1996) 301-314

Adjuvant chemo-immunotherapy after curative resection of Stage II and IIIA primary lung cancer Hideki Kimura”**, Yutaka Yamaguchib “Division

of Thoracic Diseases, bDepartment OS Surgery,

Chiba Cancer Center, Nitona-cho 666-2, institute of Pulmonary Cancer Research, Chiba University, Chiba. Japan

Chuo-ku, Chiba 260, Japan School of Medicine.

Abstract Out of 642 primary lung cancer patients who underwent surgery between 1986 and 1992, 82 cases who underwent curative resection were enrolled for a randomized prospective controlled study of postsurgical adjuvant immunotherapy using Interleukin 2 (IL-2) and lymphokine activated killer (LAK) cells. From 1986 to 1989 (the initial period), Stage IIIA patients were divided into three groups: group A (chemo-immunotherapy) received IL-2, LAK cell adoptive immunotherapy after two courses of anticancer (CDDP, VDS, MMC) chemotherapy, group B (control) received no adjuvant therapy, and group C (chemotherapy) received the same anticancer chemotherapy as group A. In the latter (1990-1992) period, group C was discontinued because of poor results and Stage II and IIIA cases were randomly assigned to group A or B. The 5- and ‘I-year survival rates of group A (33 cases) and B (36 cases) were 58.2% and 31.5%, respectively in Stage II and IIIA cases. The prognosis of group A was significantly better than that of group B (P = 0.0038 by the Cox-Mantel (C-M) test and 0.0033 by the generalized Wilcoxon (G-W) test). The 5- and S-year survival rates of each group for Stage IIIA cases were 53.4% (group A, 25 cases), 33.4% (group B, 26 cases), and 30.8%, 15.3% (group C, 13 cases). The prognosis of group A was significantly better than that of group B (P = 0.045 by the C-M and 0.036 by the G-W test). The difference between group A and B was also significant in NO, Nl (P < O.Ol), in Tl, T2 (P < 0.01) and T3 (P < 0.05) cases. These results indicate that adjuvant immunotherapy using IL-2 and LAK cells in combination with chemotherapy is significantly effective in improving the postsurgical prognosis of lung cancer patients. Keywords:

Lung cancer; Adjuvant

* Corresponding

therapy;

LAK

cells;

Immunotherapy;

author. Tel.: + 81 43 2645431; fax: + 81 43 2628680.

0169-5002/96/$15.00 0 1996 Elsevier Science Ireland Ltd. All rights reserved PII SO169-5002(96)00555-7

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1. Introduction The prognosis of lung cancer patients has improved slowly during the past decade; however, the results in advanced cases are discouraging. The 5-year survival rate of Stage IIIA curatively (or completely) resected cases is around 20-30X Numerous attempts to improve the prognosis of advanced cancer patients have been reported recently. Postsurgical adjuvant therapy using chemotherapy and/or radiotherapy has been undertaken, but no conclusive data that the trials were effective have been reported [1,2]. Here, we report a randomized controlled study of post-surgical adjuvant therapy using Interleukin 2 (IL-2) and lymphokine activated killer (LAK) cells in Stage II and IIIA lung cancer patients in combination with chemotherapy.

2. Materials

and methods

2.1. Patients Out of 642 primary lung cancer patients who underwent surgery at the Institute of Pulmonary Cancer Research, School of Medicine, Chiba University, between January 1986 and December 1992, 82 cases who underwent curative or resection of lung cancer and met the eligibility criteria (Table l), were enrolled for the study of postsurgical adjuvant therapy. Patients were stratified according to stage and histological type, and were randomized by block randomization using envelope methods. From 1986 to 1989 (the initial period), Stage IIIA curative cases were registered in the study, stratified according to histologic type, and thereafter Stage II and IIIA curative cases (the latter period; Fig. 1). In the initial period, patients were divided into three groups: Group A (chemo-immunotherapy) received IL-2, LAK cell adoptive immunotherapy after two courses of anticancer (CDDP 80 mg/m2 on day 1, VDS 3 mg/m2 on day 1 and 8, MMC 8 mg/m’ on day I) chemotherapy. Group B (control) received no adjuvant therapy except for the regular follow up identical to other groups. Group C (chemotherapy) received the same anticancer chemotherapy as group A. In the latter period, group C was discontinued because of the poor results (Fig. 2). 2.2. IL-2 and LAK therapy This therapy has been described in detail [3,4]. We used two kinds of LAK cells for adoptive immunotherapy. Lymphocytes obtained at surgery from regional lymph nodes of the patients were cultured in the presence of IL-2 (Shionogi S6820, Japan, 150 U/ml) for 2-3 weeks (Rn-LAK cells) and those obtained from peripheral blood with a blood processor (Haemonetics V50, Haemonetics Co, MA) were

H. Kimura. Table 1 Eligibility 1. 2. 3. 4. 5. 6. I.

8. 9.

criteria

for postoperative

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therapy

Non-small cell carcinoma Stage IIIA curative resection (1986- 19891 Stage II and IIIA curative resection (1990-1992) Age at the time of surgery: < 76 years Performance status: O-2 No simultaneous multiple cancers Anti-cancer therapy has to be started within 1 month after surgery Indices of major organ function fall within the following limits: WBC z 4000/mms PLT > 100 000/mm3 Hb > 10 g/d1 GOT < 100 U GPT < 100 U Cr < 1.5 mg/dl Ccr > 50 ml/min BUN < 25 mg/dl No cardiac or pulmonary insufficiency Informed consent to participate in this study

cultured in IL-2 for 1 week (Pb-LAK cells) [5,6]. These LAK cells were stored in liquid nitrogen until use. After completion of chemotherapy and when the WBC count exceeded 3000/mm3, 7 x lo5 units/day of IL-2 dissolved in 1000 ml of physiological saline was given intravenously for three consecutive days. On the second day, LAK cells (l-5 x lo9 cells/one shot) were suspended in 50 ml physiological saline and transfused intravenously. Rn-LAK cells were used during the first and second courses of the therapy whenever possible and Pb-LAK cells were used thereafter. When we could not obtain enough lymphocytes from regional lymph nodes, we used Pb-LAK cells from the first course. For the first and the second courses of treatment, the patients received continuous infusion of IL-2 intravenously for three consecutive days in the hospital. For the latter courses of

Experimental

Design

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1. Experimental

design

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Time course

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schedule Chemotherapy

Chemotherapy

Zcourses

ZCOIXS~S

CDDP,VDS,MMC

Fig. 2. Time

course

schedule.

the treatment, they first received LAK cells on day 1 and then received 3.5 x lo5 U/day of IL-2 subcutaneously for 3 days at the outpatient department. The patients received adoptive immunotherapy every 2-3 months after surgery for 2 years. When recurrence was found during the course of treatment, chemotherapy using other anticancer drugs selected on the basis of sensitivity tests [7] or radiotherapy was added to the immunotherapy. When recurrence was found after the end of the therapy, chemotherapy or radiotherapy and immunotherapy was started again.

2.3. Treatment

of other

groups

Treatment of other groups was almost the same as that of group A except they did not receive immunotherapy. Group B received no postsurgical adjuvant therapy, but otherwise they received the same care as group A, i.e. same follow-up interval, regular check-up consisting of a chest X-ray, blood cell count and blood chemistry, examination of tumor markers every 1 or 2 months, and a CT scan at least once a year. When recurrence was found, anticancer chemotherapy using CDDP, VDS, and MMC or radiotherapy was started immediately. When recurrence was found in group C, anticancer chemotherapy using other drugs selectedon the basis of sensitivity tests or radiotherapy was started.

2.4. Statistics The primary end point was survival. For the study of survival, death due to causes other than lung cancer was included in the calculation if the patients had tumor recurrence. Patients who died without tumor recurrences were censored. The survival rate was calculated by the actuarial methods of Culter and Ederer [8,9] and statistical significance was evaluated by the Cox-Mantel test and generalized Wilcoxon test using a statistical computer program ‘Statistica’ (Stat Soft Co., Inc., OK). For the multivariate statistical analysis of prognostic factors, we also employed this program. Survival time was used as a dependent variable with T, N factor, stage, histological type as well as adjuvant LAK therapy as independent variables.

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design

This study was designed to have 40 Stage IIIA curatively resected patients in each group at the beginning of the study because we estimated a 15-200/o difference in 5-year survival rate. However, in the course of the study, we realized that the result of group A (chemo-immunotherapy) was much better than that of group B (control) although there was no statistically significant difference and that the results of group C (chemotherapy alone) were either not better or were worse than those of group B (control) at the end of 1989. Therefore we eliminated group C and enrolled Stage II curative cases in the randomized study from the beginning of 1990.

3. Results 3.1. Follow-up

of the patients

and treatment

compliance

A total of 82 eligible patients were enrolled in the study, 33 cases in group A, 36 cases in group B and 13 cases in group C. There were no cases lost to follow-up and the study was completed 3 years ago. All the patients in groups A and C received at least two courses of anticancer chemotherapy and patients in group A received at least three courses of IL-2, LAK immunotherapy. The background of the patients in groups A and B was similar (Table 2). Group C consisted only of Stage IIIA cases (Table 2). All group A patients received at least three courses (mean 6.09 k 2.4) of IL-2 LAK therapy and LAK (mean, 95 + 53 x 10’: min, 20 x 10’; max, 256 x 10’) cells. Six patients (18.2%) received three courses, four patients (12.1%) four courses, eight patients (24.2%) five courses, and the rest of the patients (45.5%) received more than five courses of LAK therapy. Out of a total of 195 courses of LAK therapy, there were no severe side,effects. Fever, chills and tremor were observed in most of the patients, but they recovered to a normal condition within a few days. 3.2. Postoperative

survival

of curatively

resected Stage II and IIIA

cases

Of the total of 69 cases enrolled in the study, 33 cases were in group A and 36 cases were in group B. The 5- and 7-year survival rates of group A and B were 58.2 f 14.5% ( + standard error: 95% confidence interval C.I.: 85.7-30.6%) and 31.5 + 12.0% (C.1: 54.3-8.7%) respectively (Fig. 3). The outcome of group A was significantly better than that of group B (P = 0.0038 by the Cox-Mantel (C-M) test, and 0.0033 by the generalized Wilcoxon (G-W) test). 3.3. Postsurgical

survival of Stage IIIA

curative cases

A total of 64 Stage IIIA cases were enrolled in the study, 25 cases in group A, 26 cases in group B and 13 cases in group C (Fig. 4). The outcome of group A was

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significantly better than that of group B (P = 0.036 by the G-W test, P = 0.044 by the C-M test). The 5- and 7-year survival rates were 53.4 & 14.6% (C.I.: 80.0-25.6%) in group A, 33.4 f 14.3% (C.I.: 60.0-6.8%) in group B and 30.8%, 15.3 f 12.6% in group C. The median survivals of group B and C were 25.2 and 25.8 months, respectively while that of group A has not yet been reached. 3.4. Postsurgical

survival of Stage II and IIIA

cases in adenocarcinoma

There were 23 group A and 20 group B adenocarcinoma cases (Fig. 5). In group A there were five Stage II and 18 Stage IIIA cases, compared to four Stage II and 16 Stage IIIA cases in group B. A trend in favor of group A (P = 0.044 by the C-M test, 0.056 by the G-W test) in terms of survival was observed. The 5- and 8-year survival rates were 69.4% and 46.2 i- 19.1% (C.I.: 82.3-10.1%) in group A and 37.8% and 18.9 f 14.5% (C.I.: 45.5-O%) in group B, respectively. In other histological groups, the numbers of cases were insufficient to evaluate the efficacy of treatment.

Table 2 Characteristics

of eligible

patients Group

Age Sex Male Female Histology Adeno Ca sq Ca Large Ca Others Stage II HIA T factor Tl T2 T3 N factor NO Nl N2 Total “No

significant

difference

A

Group

B

Group

62.3 (K-12)

60 (43-73)

23 10

26” IO”

23 9 1 0

20" 12” 3” 1”

8 25

10” 26”

10 12 I1

8” 20” 8”

5 IO 18

6” 12” 18”

4 0 9

33

36

13

between

groups

A and B.

C

60 (43-73) 8 5

0 13

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307

58.2%(n.33)

7 p
“.

0

Fig. 3. Postsurgical group B.

3.5. Postsurgical

i

survival

of Stage

6

II. and Stage

IIIA,

curative

8 Years cases;

m---m

group

A, 0-O

survival and T factor

There were 22 cases in group A and 28 cases in group B with a T factor of 1 or 2 (Fig. 6). Group A consisted of one NO, seven Nl, and 14 N2 cases and group B consisted of 10 N 1 and 18 N2 cases. The 4- and 6-year survival rates were 81.4% and 40.7 f 28.3% (C.I.: 93.2-O”/o) in group A and 42.6% and 21.3 + 15.0% (C.I.: 48.0-O%) in group B. The prognosis of group A was significantly better than that of group B (P = 0.013 by the C-M test, P = 0.0068 by the G-W test). When the prognosis in T3 cases was compared, group A consisted of four NO, three Nl and four N2 cases and group B consisted of six NO and two Nl cases. The 5- and 7-year survival rates of group A (11 cases) were 58.3 f 15.4% (C.I.: 86.5-28.7%) and of group B (eight cases) 25.7 _+ 17.0% (C.I.: 57.330%) (Fig. 7). The difference was statistically significant (P = 0.026 by the G-W test, P = 0.038 by the C-M test).

3.6. Postsurgical

survival and N factor

There were 15 cases in group A and 18 cases in group B in which the N factor was 0 or 1 (Fig. 8). Group A consisted of eight Tl or T2 cases and seven T3 cases and group B consisted of 10 Tl or T2 cases and eight T3 cases. The 5- and 7-year survival rates were 64.2 & 14.5% (C.I.: 91.7-36.6%) in group A and 32.1% and 0 + 16.4% (C.I.: 31.1-O%) in group B. The median survival time of group B was 19.5 months but that of group A has not yet been reached. The difference between groups A and B in terms of survival was statistically significant (P = 0.0082 by the C-M test and 0.0094 by the G-W test). In N2 cases, a trend in favor of group A was

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observed but it was not statistically significant (P = 0.077 by the C-M and 0.0884 by the G-W test, Fig. 9). There were 18 N2 cases in each group. Group A consisted of 14 Tl or T2 and four T3 cases and group B 18 Tl or T2 cases. The 5- and 6-year survival rates were 78.4% and 58.7 + 18.2% (CL: 92.9-24.1%) in group A and 49.8% and 24.9 f 19.1% (C.I.: 61.0-O%) in group B.

3.7. Multivariate

statistical analysis

To analyze the effect of adjuvant therapy, we employed multivariate statistical analysis by Cox’s proportional hazard model (Table 3). When we used T, N factor, stage, histology and adjuvant therapy (LAK therapy) as independent variables, adjuvant LAK therapy was the only significant independent variable for survival (Table 3; t-value 2.93 and P < 0.05). In other combinations of prognostic factors, when LAK therapy was included in the analysis of independent variables, it was the only significant independent variable for prognosis (data not shown).

4. Discussion The results presented in this paper clearly demonstrate that postsurgical adjuvant IL-2 LAK chemo-immunotherapy is effective for the improvement of postsurgical survival of lung cancer patients. In a previous paper, we demonstrated the efficacy of adjuvant IL-2 LAK therapy in noncuratively resected lung cancer patients [4]. In the present study, we demonstrated its efficacy in cases treated by curative resection.

20-

1

153%(n=13)

v-

0

2

4

8

s

Years Fig. 4. Postsurgical survival A-A group C.

of Stage IIIA

relatively

curative

cases; n -m

group

A; LO

group

B;

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2018.9%(n=20) 04

Fig. 5. Postsurgical group B.

0

2

4

survival

of Stage II and IIIA

1 8 Years

6

adenocarcinoma

cases;

W--m

group

A, 0-O

In lung cancer, the results of curative resection in Stage IIIA cases are discouraging since the 5-year survival rate is approximately 20-30%. In the initial period, from 1986 to 1989, we aimed at evaluating the effect of postsurgical adjuvant

40.7%(n=22) -is .E 2 c3

40t

0

2

4

6

8 Years

Fig. 6. Postsurgical

survival

of Tl,

T2 cases;

U---H

group

A, LO

group

B.

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58.3%(n=ll)

pco.05 257%(n=8)

”

4

0

s

ii Years

Fig. 7. Postsurgical

survival

of T3 cases;

m---W

group

A, LO

group

B

chemotherapy and chemo-immunotherapy with control groups. At the end of 1989, however, we had to discontinue the chemotherapy group arm because of the poor outcome. The 3-year survival rate of group C (chemotherapy) was 35.0% and the median survival time (MST) was 30 months, while that of group A (chemo-im-

Years

Fig. 8. Postsurgical

survival

of NO, Nl

cases;

U--W

group

A, O-O

group

B.

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311

i40: -L 20

24.9%(n=lB)

I

0-l

2

0

Fig. 9. Postsurgical

survival

4

6

of N2 cases: n -m

group

a Years A, O--O

group

B.

munotherapy) was 71.5% with the MST not yet reached and group B (control) was 39.5%, MST 30 months. At the same time, a controlled randomized study by a Japanese lung cancer study group, using the same chemotherapy regimen also reported negative data concerning postsurgical adjuvant chemotherapy [lo]. From 1990, the latter period, we added Stage II curative patients to the randomized study, stratified them according to stage and histologic type and divided them into two groups; group A (chemo-immunotherapy) and group B (control). As stated elsewhere, there are four major differences between our regimen and other methods referred to as adoptive immunotherapy [I 1,121. We used immunotherapy as an adjuvant to other therapeutic approaches. In this series, surgery and chemotherapy were the main treatments, to which adjuvant immunotherapy was added. Although chemotherapy by itself did not improve the outcome in curatively resected cases, combination with immunotherapy significantly improved Table 3 Multivariate

statistical

Independent

variable

analysis

Regression

T factor N factor Stage Histology LAK therapy NS, not significant; 0.04950; calculated

of prognostic

0.417399 0.180122 -0.255290 -0.223486 1.156143 dependent by Cox’s

variable: proportional

coefficient

factors (B)

Standard

Error

0.3275309 0.2855326 0.5056054 0.2566913 0.3932996 survival time (days); hazard model.

x’

= 11.0979;

t-value

P-value

1.274381 0.630827 -0.504919 - 0.870640 2.939598

NS NS NS NS to.05

degree

of freedom,

5; P =

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survival. Since it is questionable whether immunotherapy alone would be as effective as chemo-immunotherapy, a subsequent comparative study of IL-2, LAK immunotherapy and chemo-immunotherapy may be necessary for the evaluation of the efficacy of these therapeutic modalities. Secondly, we treated postsurgical patients with relatively good prognosis, the target of our therapy being residual micrometastasis. No study on immunotherapy has shown that presently available methods can save the lives of patients in the terminal stage [13,14]. In the terminal stage, host immune mechanisms have already been demolished beyond recovery or suppressor cells or suppressive factors [ 15,161 have overcome anticancer responses, and therefore, immunotherapy would not be effective. Thirdly, we used Rn-LAK cells as much as possible because Rn-LAK cells, produced from regional lymph node lymphocytes, consist mainly of activated T cells and have higher and more specific anticancer activity against autologous tumor cells than Pb-LAK cells [5,6,17,18]. Pb-LAK cells are produced from peripheral blood lymphocytes and consist mainly of activated NK cells. Finally, we designed our study to treat patients every 2-3 months for 2 years. It is well documented that an immunologically enhanced state lasts for at least 2 months following IL-2 LAK immunotherapy [11,12] and that the possibility of tumor recurrence is highest in the initial 2 years. There was a statistically significant difference in survival between group A and group B on the whole (P < 0.01 Stage IIIA and II curative resection), as well as in Stage IIIA curative resection (P < 0.05), in NO, Nl (P < O.Ol), in Tl, T2 (P < 0.01) and in T3 (P < 0.05) cases. On the other hand, there was a tendency in favor of the immunotherapy group but without a statistically significant difference in N2 (P = O.l), and in adenocarcinoma (P = 0.04-0.05) cases. It seems that the lower the stage and T, N, M factor, the greater the difference of prognosis. This is because IL-2 and LAK immunotherapy are more effective for patients in earlier stage than in advanced stages. In early stage patients. theoretically the numbers of residual tumor cells or micrometastatic lesion cells are small and the possibility of total elimination of tumor cells by chemotherapy and immunotherapy should be greater. There appears to be two reasons why adjuvant chemotherapy is not effective for the improvement of postsurgical prognosis. In curative resection cases, most of the recurrences are distant metastases which were already present at the time of surgery. Therefore the target of postsurgical therapy should be distant micrometastasis. Since these micrometastatic foci are biologically heterogeneous, the sensitivity to anticancer drugs of each focus is also heterogeneous. The elimination of some sensitive metastatic foci may be possible by postsurgical chemotherapy, but total elimination of all micrometastatic foci may be impossible by chemotherapy alone. Furthermore, anticancer drugs are toxic not only to tumor cells but also to host immune cells. Host immune cells, such as macrophages, neutrophils, and lymphocytes are especially important in eliminating micrometastatic or damaged tumor cells. Adoptive immunotherapy using IL-2 and LAK cells is useful to overcome the defects of as well as to enhance the effects of chemotherapy. LAK cells, especially

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Rn-LAK cells have specific cytotoxic activity against autologous tumor cells, and may well be toxic to chemo-resistant micrometastatic foci [13]. Adoptive transfer of killer cells can promote the recovery of the host immune responses suppressed by chemotherapy. Furthermore, there is also an advantage of chemotherapy. Immunosuppressive agents such as anticancer drugs inhibit the production of or eliminate suppressor cells or suppressive factors (antigen-antibody complex) which inhibit the anticancer activity of LAK cells [19,20]. IL-2 LAK immunotherapy administered after chemotherapy can effectively eliminate micrometastatic tumor cells. It has become increasingly clear that apoptosis plays an important role in tumor cell death induced by chemotherapy or radiotherapy [21]. Furthermore, apoptosis is employed as the destruction mechanisms of target cells in killer T cells or NK cells [22,23]. It is conceivable that killer cells and anticancer drugs cooperate synergistically to induce apoptosis of micrometastatic tumor cells leading to their elimination. For further improvement of this therapy, analysis of each case will be necessary. The total number of, the activity of, the route of, the kind of LAK cells transferred, and the recurrence pattern and stage and histological, biological analysis will help us to further understand precisely the reason for treatment success and failure. Progress in chemotherapy, of immunotherapy with more specific and effective activity, and improvement through combination with other cytokines will help us to save more advanced stage patients.

Acknowledgements This work was supported in part by smoking research foundation (NO 56) and a Grant-in-Aid Health Sciences Research Grants and Funds for Highly Advanced Medical Research from the ministry of Welfare. We are also grateful to Professor J. Patrick Barron of the International Medical Communications Center, Tokyo Medical College, for his help in the preparation of this manuscript.

References [I] [2] [3] [4] [5]

[6]

Green MR. Dose adjuvant chemotherapy decreases distant metastasis formation in patients with non-small cell lung cancer? Lung Cancer 1994; 10 Suppl: S187-S194. Payne DC. Is preoperative or postoperative radiation therapy indicated in non-small cell cancer of the lung? Lung Cancer 1994; 10 Suppl: S205-S212. Kimura H, Yamaguchi Y. Controlled randomized study on adoptive immunotherapy for resected primary lung cancer patients (in Japanese). Jpn J Chest Dis 1995; 54: 451-457. Kimura H, Yamaguchi Y. Adjuvant immunotherapy with interleukin 2 and lymphokine-activated killer cells after noncurative resection of primary lung cancer. Lung Cancer 1995; 13: 3144. Kimura H, Yamaguchi Y, Fujisawa T. Cytotoxicity of autologous and allogenic lymphocytes against cultured human lung cancer cells: optimal conditions for the production of cytotoxic lymphocytes. Gann (Jpn J Cancer Res) 1984; 75: 1006-1016. Kimura H, Yamaguchi Y. Cytotoxic activity of various LAK cells against autologous lung cancer. K-562 and Daudi cells. Jpn J Clin Oncol 1989; 19: 222-228.

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[7] Sekiya S, Iijima N, Oosaki T et al. A newly developed in vitro chemosensitivity test (Nuclear Damage Assay): application to ovarian cancer. Gynecol Oncol 1991; 40: 138-143. [8] Cuber SJ, Ederer F. Maximum utilization of the life table methods in analyzing survival. J Chron Dis 1958; 8: 699. [9] Anderson RP, Bonchek LI, Grunkemeier SL, et al. The analysis and presentation of surgical results by actuarial methods. J Surg Res 1974; 16: 224-230. [lo] Ohta M. Tsuchiya R, Shimoyama M, et al. Adjuvant chemotherapy for completely resected stage III non-small cell lung cancer. J Thorac Cardiovasc Surg 1993; 106: 703-708. [ll] Rosenberg SA, Packard BS, Aebersold PM, et al. Use of tumor-infiltrating lymphocytes and interleukin-2 in the immunotherapy of patients with metastatic melanoma. A preliminary report. New Eng J Med 1988; 319: 1676-1680. [12] Rosenberg SA, Lotze MT, Muul LM, et al. A progress report on the treatment of 157 patients with advanced cancer using lymphokine activated killer cells and interleukin-2 or high dose interleukin-2 alone. New Engl J Med 1987; 316: 889-897. [13] Melief CJM. Tumor eradication by adoptive transfer of cytotoxic T lymphocytes. Adv Cancer Res 1992; 58: 1433175. [14] Kedar E, Klein E. Cancer immunotherapy; are the results discouraging? Can they be improved? Adv Cancer Res 1992; 59: 2455322. [15] Kadoyama C, Kimura H, Yamaguchi Y. Inhibition of cytotoxicity to autologous tumor cells by the regional lymph node cells of patients with primary lung cancer. Jpn J Clin Oncol 1987; 17: 29939. [I61 Naor D, Klein BY. Tarcic N, Duke-Cohen JS, editors. Immunosuppression and human malignancy. Clifton, NJ: Humana, 1989. [17] Watanabe Y. Hashizume Y, Shimizu J, et al. Functional character and augmentation of lymphocytes in regional lymph nodes of patients with lung cancer. Am Rev Respir Dis 1990; 142: 7699774. [ 181 Nakamura H. Ishiguro K. Mori T. Different immune functions of peripheral blood. regional lymph node, and tumor infiltrating lymphocytes in lung cancer patients. Cancer 1988; 62: 248992497. [19] Greenberg PD, Klarnet JP, Kern DE, et al. Therapy of disseminated tumors by adoptive transfer of specifically immune T cells. Prog Exp Tumor Res 1988; 32: 104-127. [20] Berd D, Mastrangelo MJ. Effect of low dose cyclophosphamide on the immune system of cancer patients: depletion of CD4 + , 2H4 + suppressor-inducer T-cells. Cancer Res 1988; 48: 1671-1675. [21] Eastman A, Grant S, Lock R, et al. Cell death in cancer and development - AACR special conference in cancer research. Cancer Res 1994; 54: 2812-2818. [22] Hanabuchi S, Koyanagi M, Kawasaki A, et al. Fas and its hgand in a general mechanism of T-cell-mediated cytotoxicity. Proc Nat1 Acad Sci 1994; 91: 4930-4934. [23] Rouvier E. Luciani MF, Golstein P. Fas involvement in Ca’+ independent T cell-mediated cytotoxicity. J Exp Med 1993; 177; 1955200.