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Evaluation of Biological Markers in Bladder Cancer
Vol. 114, December Printed in U.S.A.
THE JOURNAL OF UROLOGY
Copyright© 1975 by The Williams & Wilkins Co.
EVALUATION OF BIOLOGICAL MARKERS IN BLADDER CANCER ZEW WAJSMAN, CLAUDE E. MERRIN, T. MING CHU, ROBERT H. MOORE AND GERALD P. MURPHY*
From the Roswell Park Memorial Institute, New York State Department of Health, State University of New York at Buffalo, Buffalo, New York
ABSTRACT
The value of biological markers of bladder cancer was studied in 66 patients. The markers included serum and urine carcinoembryonic antigens, serum and urine fibrinogen degradation products, total lymphocyte counts, urine lymphocytes and urine cytology. A high degree of accuracy (90 per cent) was found in correlating cytology and urinary fibrinogen degradation products with the activity of the disease. Serum and urine carcinoembryonic antigens, serum fibrinogen degradation products, total lymphocyte counts and urine lymphocytes were found to have no value in screening bladder cancer patients. Urinary fibrinogen degradation products and cytology in combination are recommended for screening and followup of patients at high risk.
·
;! • L
)I
Current survival indicates that two-thirds of the people with cancer may have metastases when the primary tumor is diagnosed. 1 The goal of the National Cancer Program is to develop research that would enable the diagnosis to be made early enough so that at least 90 per cent of the patients would be free of metastasis at the time of diagnosis. 1 The second but equally important consideration is the early discovery of recurrent metastatic disease. We herein analyze the value of different tests used in bladder cancer patients and define which of several diagnostic tests may be of benefit in screening and followup evaluation of patients with bladder cancer. METHODS
'I•i lj
i( ~
ti !
Sixty-six consecutive patients (55 male and 11 female) with past or present diagnosis of bladder tumors underwent routine cystoscopic evaluation between 1973 and 1974. At the time of cystoscopy the urine and serum were collected for testing of carcinoembryonic antigen (CEA) and fibrinogen degradation products (FDP), and blood count was obtained. Fresh urine specimens were sent for cytological examination. We obtained 1 to 2 specimens of urine as well as blood from each patient. All results were analyzed, compared to the clinical findings and related to the presence of active disease as was determined by cystoscopic findings, tissue diagnosis and metastatic diagnostic study, which included chest x-rays, bone surveys and liver Accepted for publication May 16, 1975. Read at annual meeting of American Urological Association, Miami Beach, Florida, May 11-15, 1975. Supported in part by United States Public Health Service Grant RR-05648-08 of the National Institutes of Health. * Requests for reprints: Roswell Park Memorial Institute, 666 Elm St., Buffalo, New York 14263. 879
scans. All results were evaluated and corrected statistically by the chi-square method. 2 Total peripheral blood lymphocyte count was calculated from the percentage of lymphocytes in the total leukocyte count. The normal absolute lymphocyte count ranges between 1,490 and 3,930 per cu. mm. 3 The cytological examination of urine was done essentially by the Papanicolaou method as introduced by Harrison and associates.• CEA in plasma and urine was determined with. Hansen Z-gel radioimmunoassay method• with modification. 6 The urine was dialyzed first according to Hall and associates. 7 Normal individuals have plasma CEA levels of less than 2.5 ng. per ml.; the normal urinary CEA range is up to 25 ng. per ml. Urinary lymphocyte assays were determined by the method described previously, 8 which is based on the ability of the immunocompetent lymphocytes to be stained by pyronine. 9 • 10 FDP in urine and in the serum were determined by immunoassay described by Merskey and associates. 11 In the urine of normal individuals no FDP are found and in the serum the FDP levels are less than 5.0 µg. per ml., according to estimations made in 20 normal controls. These values also were normal in our laboratory. The disease status was described as active if a bladder tumor was seen and confirmed by pathology examination. No present evidence of bladder cancer was determined on the basis of negative cystoscopic examination and metastatic diagnostic study. RESULTS
We studied 65 patients with transitional cell carcinoma of the bladder and 1 patient with
880
W AJSMAN AND ASSOCIATES
squamous cell carcinoma. At the time of study 46 patients had no evidence of disease but they did have a positive tissue diagnosis of bladder cancer in the 2 years preceding the study. There was evidence of active disease in 20 patients (table 1). Lymphocyturia and bladder cancer. No pyronine-positive lymphocytes were found in any of the 66 cases. Lymphocyte count as related to disease status demonstrated no statistical significance of lymphopenia in active or non-active disease states (table 2). Urinary CEA levels were found to be normal in 12 of 17 patients with active disease, a high falsely negative result (table 3). Serum CEA determinations related to the activity of disease showed insignificant correlations as well as falsely negative and falsely positive values (table 4). Urinary cytology showed an accuracy of diagnosis around 70 per cent with 20 per cent falsely negative results (table 5). No correlation was found between the levels of serum FDP and the activity of disease (table 6). Urine FDP showed highly significant values and indicated a tendency to be elevated in the presence of active disease (table 6). The correlation with activity of disease was 72 per cent. Falsely negative results were in the range of 26 per cent. Correlation of cytology results with the FDP values in urine is shown in table 7. When the test status was completely negative 28 of 30 (93.3 per cent) showed no evidence of disease. When the test status was completely positive 12 of 14 (85. 7 per cent) exhibited active disease. Further statistical evaluation suggested that the positive result of urine cytology and FDP were even more accurate with higher grades and stages of tumors but the number of patients in the different TABLE
1. Disease status in bladder cancer patients Cases No. (%) 46 (69.7) 20 (:30.3) 66 (100.0)
No clinical evidence of disease Active disease Total
groups was limited, preventing a statistical conclusion. DISCUSSION
Early diagnosis in any type of cancer is crucial for the final outcome. This is especially true in bladder cancer, which has the tendency to recur in early stages. 12 In the progressive stages the results of treatment, whether surgical, radiotherapeutic or chemotherapeutic, are somewhat limited. 13 Therefore it is important to find a test which may help in obtaining an early diagnosis in patients under risk, such as aniline workers, patients with previous history of bladder tumor and patients with a family history of cancer. Amin and Lich analyzed the value of peripheral lymphocyte counts as a test for a host defense mechanism, which may influence the final outcome in bladder cancer patients. 14 Papatestas and 3. Urine CEA relationship to disease status in
TABLE
bladder cancer Urine CEA
No Evidence of Disease
Active Disease
Totals
<25 (normal) 2:25 Totals
31 13 44
12 5 17
43 18 61 *
Chi-square (X')-0.00, not significant. * In 5 patients no urinary CEA determinations were performed.
4. Serum CEA relationship to disease status in
TABLE
bladder cancer Serum CEA
No Evidence of Disease
Active Disease
Totals
34
1:3 4 17
47 12 59*
0-2.5 (normal) >2.5 (abnormal) Totals
8
42
Chi-square (X')-0.15, not significant. * In 7 patients no serum CEA determinations were done. TABLE
5. Relationship between urinary cytology and
disease status in bladder cancer TABLE
2. Lymphocyte count relationship to disease status
in bladder cancer No Evidence of Disease
Active Disease
0-500 501-i,OOO 1,001-1,500 1,501-2,000 2,001-2,500 2,501-3,000 :l,001-4,000 >4,000 Totals
0 9 10 13 7 3
1 5 4
<1,500 2:1,500 Totals
19 27 46
Lymphocyte Count
8
:1
1 1 0
1 46
20
0 10 10
20
Chi-square (X')-0.43, not significant.
Totals No.(%) 1 (1.5) 14 (21.2) 14 (21.2) 21 (:ll.8) 8 (12.2) 4 (6.1) 3 (4.5) 1 ( 1.5) 66 (100.0) 29 37 66
Urine Cytology
.'.\lo Evidence of Disease
Active Disease
Totals
Negative All others (positive) Totals
36 10
5 15
41 25
46
20
66
p less than 0.00005, highly sig-nificant. TABLE
6. Urinary FDP relationship to disease status in
bladder cancer Urinary FOP
No Evidence of Disease
Active Disease
Totals
0 (normal) > 0 (abnormal) Totals
:36
6 14 20
42 24 66
10
46
p less than 0.001, highly significant.
881
BIOLOGICAL MARKERS IN BLADDER CANCER TABLE
7. Relationship among urinary FDP, urinary cytology and disease status in bladder cancer Disease Status
Urinary cytology neg., Urinary cytology neg., Urinary cytology pas., Urinary cytology pas., Totals
urinary FDP normal urinary FDP abnormal urinary FDP normal urinary FDP abnormal
No Evidence of Disease
Active Disease
28 8 8 2 46
2 3 3 12 20
Totals No. (%) 30 (45.4) 11 (16.7) 11 (16.7) 14 (21.2) 66 (100.0)
p less than 0.00001, highly significant.
Kark showed that the difference in lymphocyte counts is completely independent of modalities of treatment and reflects the host's responses to tumor growth. 15 In our study we have found no correlation between the lymphocyte counts and activity of disease. Our feeling is that this test has no value in screening patients at risk and no value in followup or prognosis. Lymphocytes are the important elements in cellular immunity as demonstrated by Porter• and play crucial roles in acute rejection of renal allografts. These lymphocytes in the rejection phenomenon may escape into the urine, 10 and by findings of pyronine-positive lymphocytes in urine one may predict and diagnose acute rejection. 8 In bladder cancer, which is as other tumors associated with tumor specific antigens, 16 local immunity might have caused lymphocyte reaction. We failed to demonstrate pyronine-positiveimmune competent cells in the urine of patients with bladder cancer in different stages of the disease and in those who showed no evidence of disease, which may possibly be owing to: 1) a failure of the immunological surveillance to control the developing cancer or 2) the tumor antigens being weak antigens and unable to elicit a strong immunologic response, such as seen in the rejection phenomenon. The value of CEA, the tumor associated antigen described by Gold and Freedman, 17 was analyzed in urogenital cancer by Reynoso and associates. 18 They observed that 30 per cent of 116 patients with urogenital cancer had elevated plasma CEA. No positive results were found in any of 17 patients with well controlled prostatic carcinoma, whereas 7 of 11 patients with progressive prostatic disease had positive findings. In carcinoma of the bladder 10 of 20 patients with active disease and 2 of 24 with inactive disease had positive findings. 18 Chu and associates reported the value of plasma CEA in renal cell carcinoma, indicating good correlation with activity of the disease and with response to surgery and chemotherapy. 19 Hall and associates have reported that CEA was found in significant amounts in urine of patients with bladder carcinoma. 7 They suggested that urinary CEA determinations may be of clinical diagnostic value in the early detection of bladder cancer. However, in the previous study from our institution in which no correlation between plasma and urinary CEA can be made in patients with gastrointestinal
malignancies 20 and in the present study we were unable to confirm the significance of CEA in plasma and urine of bladder cancer patients. There was no correlation with activity of disease. The falsely positive and negative results in our patients may be owing to the fact that the result represented total urinary substances which crossreacted with anti-CEA in radioimmunoassay. It has been reported that urinary immunoglobulin or degradation products from immunoglobulin also cross-reacted with anti-CEA antiserum m radioimmunoassay. 21 O'Meara had shown extravascular deposition of fibrin in and around human malignant neoplasms. 22 According to his theory this network of fibrin leads to vascularization and growth of the tumors. Secondary fibrinolysis of such deposits and microthrombi might give rise to FDP (see figure). 23 Ossowski and associates have shown that avian and mammalian fibroblasts, transformed to malignancy by oncogenic viruses or chemical carcinogens, produce a fibrinolytic activity. 24 Pathological modifications of clotting and fibrinolysis are commonly associated with malignant disease in man, and the findings of this group from Rockefeller University highly merit further investigation. As we have shown in our study there was no correlation between the plasma FDP and urine FDP, and there was no correlation between the plasma FDP and the activity of the disease. In contrast a high correlation between urinary FDP and the presence of active bladder cancer was established. The urinary FDP in bladder cancer patients with active disease might have appeared in the urine because of leakage through the tumor cell membranes, which have increased permeability. 25 Exfoliative cytology in bladder cancer is commonly used for early detection and in the followup of patients, and in our study the results of cytology were comparable to the results of FDP in the urine. The use of both tests raised the positive results to around 90 per cent. CONCLUSION
Our study has shown no significant value for total lymphocyte counts or for plasma and urinary levels of carcinoembryonic antigen in the detection of bladder cancer. No correlation between FDP in serum and activity of disease was found. However, urinary FDP estimations in combination with
882
WAJSMAN AND ASSOCIATES
FIBRINOGEN DEGRADATION PRODUCTS FROM TUMOR SITES
TISSUE THROMBOPLASTINS AND PLASMINOGEN ACTIVATORS
A-V ANASTOMOSIS
FIBRIN--~
TORTUOUS DILATED VESSELS ......,.,,.,,,)l!D''<,
SLUGGISH
FLOW
LOCAL DEFIBRINATION, EXUDATION AND DEPOSITION OF FIBRIN AROUND TUMOR CELLS AND AROUND TUMOR IN SURROUNDING NORMAL TISSUE. COMPENSATORY - 1 D P FIBRINOLYSIS ( . L )
REACTIVE HYPERFIBRINOGENEMIA
REACTIVE ANTIPLASMINEMIA
I I L - - - -TUMOR CELL NUTRITION?
Diagrammatic explanation of different coagulation processes as they occur around tumor cells
cytology led to greater accuracy in screening and followup of patients with bladder cancer. We suggest that further research of FDP in the urine should be developed as a test for patients under high risk and as a marker for response to surgery, radiotherapy and chemotherapy. FDP in urine may be of value when used in patients with ileal loops in whom cytology examination is difficult and testing of CEA is of no value at all owing to the presence of intestinal mucosa of the loop. It is suggested that use of more specific methods may discover minimal amounts of FDP in the urine and, therefore, increase the accuracy of discovering clinically occult disease states, including carcinoma in situ. REFERENCES
1. Berlin, N. I.: Research str.ategy in cancer: screening, diagnosis, prognosis. Hosp. Prac., 10: 83, 1975. 2. Dixon, W. J. and Massey, F. J., Jr.: Introduction to Statistical Analysis, 2nd ed. New York: McGraw: Hill Book Co., 1957. 3. Wintrobe, M. M.: Clinical Hematology, 6th ed. Philadelphia: Lea & Febiger, 1967. 4. Harrison, J. H., Botsford, T. W. and Tucker, M. R.: The use of the smear of the urinary sediment in the diagnosis and management of neoplasm of the kidney and bladder. Surg., Gynec. & Obst., 92: 129, 1951. 5. Hansen, H. J., Lance, K. P. and Krupey, J. P.: Demonstration of an ion sensitive antigenic site on carcinoembryonic antigen using zirconyl phosphate gel. Clin. Res., 19: 143, 1971. 6. Chu, T. M. and Reynoso, G.: Evaluation of a new radioimmunoassay method for carcinoembryonic antigen in plasma, with use of zirconyl phosphate gel. Clin. Chem., 18: 918, 1972.
7. Hall, R.R., Laurence, D. J. R., Darcy, D., Stevens, U., James, R., Roberts, S. and Neville, A. M.: Carcinoembryonic antigen in the urine of patients with urothelial carcinoma. Brit. Med. J., 3: 609, 1972. 8. Hrushesky, W., Sampson, D. and Murphy, G. P.: Lymphocyturia in human renal allograft rejection. Arch. Surg., 105: 424, 1972. 9. Porter, K. A.: Rejection in treated renal allografts. J. Clin. Path., suppl. vol. 20, p. 518, 1967. 10. Caine, R. Y.: Renal transplantation in man: a review. Brit. J. Surg., 51: 282, 1964. 11. Merskey, C., Kleiner, G. J. and Johnson, A. J.: Quantitative estimation of split products of fibrinogen in human serum, relation to diagnosis and treatment. Blood, 28: 1, 1966. 12. Sarma, K. P.: Tumours of the Urinary Bladder. New York: Appleton-Century-Crofts, p. 273, 1969. 13. Harve_Y,_ R., Mirand, ~- !",· and M1:1rph,'.~G. P.: Inh1b1t10n of erythrop01etm product10n by· cyproterone acetate. Urology, 1: 121, 1973. 14. Amin, M. and Lich, R., Jr.: Lymphocytes and bladder cancer. J. Urol., 111: 165, 1974. 15. Papatestas, A. E. and Kark, A. E.: Peripheral lymphocyte counts in breast carcinoma. An index of immune competence. Cancer, 34: 2014, 1974. 16. Bubenik, J., Perlmann, P., Helmstein, K. and Moberger, G.: Cellular and humoral immune responses to human urinary bladder carcinomas. Int. J. Cancer, 5: 310, 1970. 17. Gold, P. and Freedman, S. 0.: Demonstration of tumor-specific antigens in human colonic carcinomata by immunological tolerance and absorption techniques. J. Exp. Med., 121: 439, 1965. 18. Reynoso, G., Chu, T. M., Holyoke, D., Cohen, E., Nemoto, T., Wang, J.-J., Chuang, J., Guinan, P. and Murphy, G. P.: Carcinoembryonic antigen in patients with different cancers. J.A.M.A., 220: 361, 1972. 19. Chu, T. M., Shukla, S. K., Mittelman, A. 0. and
?·
BIOLOGICAL MARKERS IN BLADDER CANCER
20.
21.
22. 23.
Murphy, G. P.: Plasma carcinoembryonic antigen in renal cell carcinoma patients. J. Urol.. 111: 742. 1974. Karakousis, C. P., Holyoke, E. D. and Chu, T. M.: Carcinoembryonic antigens in the urine. Clin. Res., 21: 972, 1973. Guinan, A., Ablin, R. J., Bruns, G. R., Guinan, P. and Bush, I. M.: Urinary excretion of immunoglobulins in prostatic cancer. Clin. Res., 22: 490, 1974. O'!Vleara, R. A.: Coagulative properties of cancers. Irish J. Med. Sci., 394: 474, 1958. Mink, I. R and Ambrus, J.: A diagrammatic expla-
883
nation of the different coagulation processes as they occur around the tumor cells. Personal communication, 1975. 24. Ossowski, L., Unkeless, J.C., Tobia, A., Quigley. J. P., Rifkin, D. B. and Reich. E.: An enzymatic function associated with transformation of fibroblasts by oncogenic viruses. II. Mammalian fibroblast cultures transformed by DNA and RNA tumor viruses. J. Exp. Med., 137: 112, 197'.3. 25. Worowski, K. and Farbiszewski, R.: Studies on the proteolytic enzyme inhibitors in malignant neoplastic diseases. Pol. Med. J ., l 502, 1972.
THE JOURNAL OF UROLOGY
Copyright© 1975 by The Williams & Wilkins Co.
EVALUATION OF BIOLOGICAL MARKERS IN BLADDER CANCER ZEW WAJSMAN, CLAUDE E. MERRIN, T. MING CHU, ROBERT H. MOORE AND GERALD P. MURPHY*
From the Roswell Park Memorial Institute, New York State Department of Health, State University of New York at Buffalo, Buffalo, New York
ABSTRACT
The value of biological markers of bladder cancer was studied in 66 patients. The markers included serum and urine carcinoembryonic antigens, serum and urine fibrinogen degradation products, total lymphocyte counts, urine lymphocytes and urine cytology. A high degree of accuracy (90 per cent) was found in correlating cytology and urinary fibrinogen degradation products with the activity of the disease. Serum and urine carcinoembryonic antigens, serum fibrinogen degradation products, total lymphocyte counts and urine lymphocytes were found to have no value in screening bladder cancer patients. Urinary fibrinogen degradation products and cytology in combination are recommended for screening and followup of patients at high risk.
·
;! • L
)I
Current survival indicates that two-thirds of the people with cancer may have metastases when the primary tumor is diagnosed. 1 The goal of the National Cancer Program is to develop research that would enable the diagnosis to be made early enough so that at least 90 per cent of the patients would be free of metastasis at the time of diagnosis. 1 The second but equally important consideration is the early discovery of recurrent metastatic disease. We herein analyze the value of different tests used in bladder cancer patients and define which of several diagnostic tests may be of benefit in screening and followup evaluation of patients with bladder cancer. METHODS
'I•i lj
i( ~
ti !
Sixty-six consecutive patients (55 male and 11 female) with past or present diagnosis of bladder tumors underwent routine cystoscopic evaluation between 1973 and 1974. At the time of cystoscopy the urine and serum were collected for testing of carcinoembryonic antigen (CEA) and fibrinogen degradation products (FDP), and blood count was obtained. Fresh urine specimens were sent for cytological examination. We obtained 1 to 2 specimens of urine as well as blood from each patient. All results were analyzed, compared to the clinical findings and related to the presence of active disease as was determined by cystoscopic findings, tissue diagnosis and metastatic diagnostic study, which included chest x-rays, bone surveys and liver Accepted for publication May 16, 1975. Read at annual meeting of American Urological Association, Miami Beach, Florida, May 11-15, 1975. Supported in part by United States Public Health Service Grant RR-05648-08 of the National Institutes of Health. * Requests for reprints: Roswell Park Memorial Institute, 666 Elm St., Buffalo, New York 14263. 879
scans. All results were evaluated and corrected statistically by the chi-square method. 2 Total peripheral blood lymphocyte count was calculated from the percentage of lymphocytes in the total leukocyte count. The normal absolute lymphocyte count ranges between 1,490 and 3,930 per cu. mm. 3 The cytological examination of urine was done essentially by the Papanicolaou method as introduced by Harrison and associates.• CEA in plasma and urine was determined with. Hansen Z-gel radioimmunoassay method• with modification. 6 The urine was dialyzed first according to Hall and associates. 7 Normal individuals have plasma CEA levels of less than 2.5 ng. per ml.; the normal urinary CEA range is up to 25 ng. per ml. Urinary lymphocyte assays were determined by the method described previously, 8 which is based on the ability of the immunocompetent lymphocytes to be stained by pyronine. 9 • 10 FDP in urine and in the serum were determined by immunoassay described by Merskey and associates. 11 In the urine of normal individuals no FDP are found and in the serum the FDP levels are less than 5.0 µg. per ml., according to estimations made in 20 normal controls. These values also were normal in our laboratory. The disease status was described as active if a bladder tumor was seen and confirmed by pathology examination. No present evidence of bladder cancer was determined on the basis of negative cystoscopic examination and metastatic diagnostic study. RESULTS
We studied 65 patients with transitional cell carcinoma of the bladder and 1 patient with
880
W AJSMAN AND ASSOCIATES
squamous cell carcinoma. At the time of study 46 patients had no evidence of disease but they did have a positive tissue diagnosis of bladder cancer in the 2 years preceding the study. There was evidence of active disease in 20 patients (table 1). Lymphocyturia and bladder cancer. No pyronine-positive lymphocytes were found in any of the 66 cases. Lymphocyte count as related to disease status demonstrated no statistical significance of lymphopenia in active or non-active disease states (table 2). Urinary CEA levels were found to be normal in 12 of 17 patients with active disease, a high falsely negative result (table 3). Serum CEA determinations related to the activity of disease showed insignificant correlations as well as falsely negative and falsely positive values (table 4). Urinary cytology showed an accuracy of diagnosis around 70 per cent with 20 per cent falsely negative results (table 5). No correlation was found between the levels of serum FDP and the activity of disease (table 6). Urine FDP showed highly significant values and indicated a tendency to be elevated in the presence of active disease (table 6). The correlation with activity of disease was 72 per cent. Falsely negative results were in the range of 26 per cent. Correlation of cytology results with the FDP values in urine is shown in table 7. When the test status was completely negative 28 of 30 (93.3 per cent) showed no evidence of disease. When the test status was completely positive 12 of 14 (85. 7 per cent) exhibited active disease. Further statistical evaluation suggested that the positive result of urine cytology and FDP were even more accurate with higher grades and stages of tumors but the number of patients in the different TABLE
1. Disease status in bladder cancer patients Cases No. (%) 46 (69.7) 20 (:30.3) 66 (100.0)
No clinical evidence of disease Active disease Total
groups was limited, preventing a statistical conclusion. DISCUSSION
Early diagnosis in any type of cancer is crucial for the final outcome. This is especially true in bladder cancer, which has the tendency to recur in early stages. 12 In the progressive stages the results of treatment, whether surgical, radiotherapeutic or chemotherapeutic, are somewhat limited. 13 Therefore it is important to find a test which may help in obtaining an early diagnosis in patients under risk, such as aniline workers, patients with previous history of bladder tumor and patients with a family history of cancer. Amin and Lich analyzed the value of peripheral lymphocyte counts as a test for a host defense mechanism, which may influence the final outcome in bladder cancer patients. 14 Papatestas and 3. Urine CEA relationship to disease status in
TABLE
bladder cancer Urine CEA
No Evidence of Disease
Active Disease
Totals
<25 (normal) 2:25 Totals
31 13 44
12 5 17
43 18 61 *
Chi-square (X')-0.00, not significant. * In 5 patients no urinary CEA determinations were performed.
4. Serum CEA relationship to disease status in
TABLE
bladder cancer Serum CEA
No Evidence of Disease
Active Disease
Totals
34
1:3 4 17
47 12 59*
0-2.5 (normal) >2.5 (abnormal) Totals
8
42
Chi-square (X')-0.15, not significant. * In 7 patients no serum CEA determinations were done. TABLE
5. Relationship between urinary cytology and
disease status in bladder cancer TABLE
2. Lymphocyte count relationship to disease status
in bladder cancer No Evidence of Disease
Active Disease
0-500 501-i,OOO 1,001-1,500 1,501-2,000 2,001-2,500 2,501-3,000 :l,001-4,000 >4,000 Totals
0 9 10 13 7 3
1 5 4
<1,500 2:1,500 Totals
19 27 46
Lymphocyte Count
8
:1
1 1 0
1 46
20
0 10 10
20
Chi-square (X')-0.43, not significant.
Totals No.(%) 1 (1.5) 14 (21.2) 14 (21.2) 21 (:ll.8) 8 (12.2) 4 (6.1) 3 (4.5) 1 ( 1.5) 66 (100.0) 29 37 66
Urine Cytology
.'.\lo Evidence of Disease
Active Disease
Totals
Negative All others (positive) Totals
36 10
5 15
41 25
46
20
66
p less than 0.00005, highly sig-nificant. TABLE
6. Urinary FDP relationship to disease status in
bladder cancer Urinary FOP
No Evidence of Disease
Active Disease
Totals
0 (normal) > 0 (abnormal) Totals
:36
6 14 20
42 24 66
10
46
p less than 0.001, highly significant.
881
BIOLOGICAL MARKERS IN BLADDER CANCER TABLE
7. Relationship among urinary FDP, urinary cytology and disease status in bladder cancer Disease Status
Urinary cytology neg., Urinary cytology neg., Urinary cytology pas., Urinary cytology pas., Totals
urinary FDP normal urinary FDP abnormal urinary FDP normal urinary FDP abnormal
No Evidence of Disease
Active Disease
28 8 8 2 46
2 3 3 12 20
Totals No. (%) 30 (45.4) 11 (16.7) 11 (16.7) 14 (21.2) 66 (100.0)
p less than 0.00001, highly significant.
Kark showed that the difference in lymphocyte counts is completely independent of modalities of treatment and reflects the host's responses to tumor growth. 15 In our study we have found no correlation between the lymphocyte counts and activity of disease. Our feeling is that this test has no value in screening patients at risk and no value in followup or prognosis. Lymphocytes are the important elements in cellular immunity as demonstrated by Porter• and play crucial roles in acute rejection of renal allografts. These lymphocytes in the rejection phenomenon may escape into the urine, 10 and by findings of pyronine-positive lymphocytes in urine one may predict and diagnose acute rejection. 8 In bladder cancer, which is as other tumors associated with tumor specific antigens, 16 local immunity might have caused lymphocyte reaction. We failed to demonstrate pyronine-positiveimmune competent cells in the urine of patients with bladder cancer in different stages of the disease and in those who showed no evidence of disease, which may possibly be owing to: 1) a failure of the immunological surveillance to control the developing cancer or 2) the tumor antigens being weak antigens and unable to elicit a strong immunologic response, such as seen in the rejection phenomenon. The value of CEA, the tumor associated antigen described by Gold and Freedman, 17 was analyzed in urogenital cancer by Reynoso and associates. 18 They observed that 30 per cent of 116 patients with urogenital cancer had elevated plasma CEA. No positive results were found in any of 17 patients with well controlled prostatic carcinoma, whereas 7 of 11 patients with progressive prostatic disease had positive findings. In carcinoma of the bladder 10 of 20 patients with active disease and 2 of 24 with inactive disease had positive findings. 18 Chu and associates reported the value of plasma CEA in renal cell carcinoma, indicating good correlation with activity of the disease and with response to surgery and chemotherapy. 19 Hall and associates have reported that CEA was found in significant amounts in urine of patients with bladder carcinoma. 7 They suggested that urinary CEA determinations may be of clinical diagnostic value in the early detection of bladder cancer. However, in the previous study from our institution in which no correlation between plasma and urinary CEA can be made in patients with gastrointestinal
malignancies 20 and in the present study we were unable to confirm the significance of CEA in plasma and urine of bladder cancer patients. There was no correlation with activity of disease. The falsely positive and negative results in our patients may be owing to the fact that the result represented total urinary substances which crossreacted with anti-CEA in radioimmunoassay. It has been reported that urinary immunoglobulin or degradation products from immunoglobulin also cross-reacted with anti-CEA antiserum m radioimmunoassay. 21 O'Meara had shown extravascular deposition of fibrin in and around human malignant neoplasms. 22 According to his theory this network of fibrin leads to vascularization and growth of the tumors. Secondary fibrinolysis of such deposits and microthrombi might give rise to FDP (see figure). 23 Ossowski and associates have shown that avian and mammalian fibroblasts, transformed to malignancy by oncogenic viruses or chemical carcinogens, produce a fibrinolytic activity. 24 Pathological modifications of clotting and fibrinolysis are commonly associated with malignant disease in man, and the findings of this group from Rockefeller University highly merit further investigation. As we have shown in our study there was no correlation between the plasma FDP and urine FDP, and there was no correlation between the plasma FDP and the activity of the disease. In contrast a high correlation between urinary FDP and the presence of active bladder cancer was established. The urinary FDP in bladder cancer patients with active disease might have appeared in the urine because of leakage through the tumor cell membranes, which have increased permeability. 25 Exfoliative cytology in bladder cancer is commonly used for early detection and in the followup of patients, and in our study the results of cytology were comparable to the results of FDP in the urine. The use of both tests raised the positive results to around 90 per cent. CONCLUSION
Our study has shown no significant value for total lymphocyte counts or for plasma and urinary levels of carcinoembryonic antigen in the detection of bladder cancer. No correlation between FDP in serum and activity of disease was found. However, urinary FDP estimations in combination with
882
WAJSMAN AND ASSOCIATES
FIBRINOGEN DEGRADATION PRODUCTS FROM TUMOR SITES
TISSUE THROMBOPLASTINS AND PLASMINOGEN ACTIVATORS
A-V ANASTOMOSIS
FIBRIN--~
TORTUOUS DILATED VESSELS ......,.,,.,,,)l!D''<,
SLUGGISH
FLOW
LOCAL DEFIBRINATION, EXUDATION AND DEPOSITION OF FIBRIN AROUND TUMOR CELLS AND AROUND TUMOR IN SURROUNDING NORMAL TISSUE. COMPENSATORY - 1 D P FIBRINOLYSIS ( . L )
REACTIVE HYPERFIBRINOGENEMIA
REACTIVE ANTIPLASMINEMIA
I I L - - - -TUMOR CELL NUTRITION?
Diagrammatic explanation of different coagulation processes as they occur around tumor cells
cytology led to greater accuracy in screening and followup of patients with bladder cancer. We suggest that further research of FDP in the urine should be developed as a test for patients under high risk and as a marker for response to surgery, radiotherapy and chemotherapy. FDP in urine may be of value when used in patients with ileal loops in whom cytology examination is difficult and testing of CEA is of no value at all owing to the presence of intestinal mucosa of the loop. It is suggested that use of more specific methods may discover minimal amounts of FDP in the urine and, therefore, increase the accuracy of discovering clinically occult disease states, including carcinoma in situ. REFERENCES
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BIOLOGICAL MARKERS IN BLADDER CANCER
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