Download Critical Evaluation of Lymphocyte Functions in

(CANCER
RESEARCH
36, 132-137,January1976)
Critical Evaluation of Lymphocyte Functions in Urological
Cancer Patients1
Mostafa M. Elhllali,2 Sven BillIon, Stanley Brosman, and John L. Fahey
Departments of Surgery/Urology, Microbiology, and Immunology, University of California, Los Angeles 90024 (M.M.E., Sv.B., St.B., J.L.F.J and Department
of Surgery/Urology,HarborGeneralHospital, Torrance,California90609[St.B.J
SUMMARY
One hundred three patients with varying stages of
urobogical cancer (bladder, prostate, kidney) were
investigated with regard to the following lymphocyte
functions. T-cells were assessed numerically (E rosettes),
their blastogenic response to phytohemagglutinin (PHA)
was determined, and their cytotoxic potential against
heterobogous target cells in short-term presence of PHA
(i.e., PHA-dependent cellular cytotoxicity) was evaluated.
Similarly, B cells were numerically assessed (EA rosettes),
and their function was evaluated by antibody-dependent
cellular cytotoxicity against antibody-coated heterobogous
target cells. The data on cancer patients, divided on the
basis of extent of disease and prior radiation therapy, were
compared to those of normal young and age-matched
controls.
Our investigations emphasize the importance of the fol
lowing factors: (a) comparison of data with age-matched
controls, since several lymphocyte functions appear to
change with age; (b) use of multiple controls to compensate
for the inherent variability found in certain tests; (c) mini
mized contamination by nonlymphoid cells in the purified
cell preparation; and (d) the influence of certain treatment
regimens (radiation, chemotherapy, etc.) on the results.
Radiotherapy significantly depressed T-cell number with
a depression of PHA blastogenic responses as well as PHA
dependent cellular cytotoxicity.
When all of these conditions were taken into account, the
urobogical cancer patients as a group were found to have a
lower proportional value of E rosettes (T-cells) and a re
duced PHA blastogenic responsiveness. Certain cancer pa
tients displayed an elevated PHA-dependent cellular cyto
toxicity
as compared
to age-matched
controls,
which may
indicate the presence of activated cells in the presence of
tumors. With this identification of a group of cancer pa
tients with markedly depressed E rosette values and PHA
responsiveness, it will now be possible to follow them clini
cally in comparison with a group of cancer patients with
normal T-ceII functions.
INTRODUCTION
Immunocompetence
persensitivity
I These
as reflected by normal delayed hy
skin test reactions
investigations
were
supported
tends to correlate
by
National
Bladder
Cancer
with the
Project
Grant CA 16880.
a To whom
requests
for
reprints
should
be addressed,
at Department
Urology, Sherbrooke University, Sherbrooke, Quebec, Canada.
Received June 18, 1975; accepted
132
October
2, 1975.
of
stage, clinical course, and prognosis of cancer (4, 6). Skin
tests of this type, however, involve immunological and in
flammatory
components.
Assessment
of individual
compo
nents of the immune response in relation to tumor burden
and modes of therapy is needed. Also, such information is
needed to assess changes in the immune system of bladder
cancer patients receiving nonspecific adjuvant immunother
apy. No comparable study using the battery of investiga
tions reported in this work in patients with cancer of the
genitourinary tract has been reported.
A broad assessment of lymphocyte functions, measured
by in vitro assays, was undertaken as a part of immunologi
cal and immunotherapy investigations in bladder cancer.
The influence of tumor burden and radiation therapy was
compared by dividing the patients into appropriate groups.
Also, salient pitfalls encountered in this type of investiga
tion are noted.
MATERIAL AND METHODS
Peripheral venous blood samples (50 ml heparinized
blood) were obtained from 3 different populations. These
included 41 young controls (113 specimens), 47 age
matched controls (52 specimens), and 103 patients with
documented cancer of the bladder, kidney, or prostate (139
specimens) (Table 1). The mean age of the young controls
was 26.7 years, that of the age matched controls was 58.7,
and that of the cancer patients was 65.3 years. The distribu
tion of the specimens obtained from the 3 cancer groups
included 77 specimens obtained from 56 bladder cancer
patients, 45 specimens from 32 prostatic cancer patients,
and 17 specimens from 15 renal carcinoma patients.
Blood Separation. The blood samples were kept at room
temperature and were separated as soon as possible by the
Ficoll-Hypaque technique described by Boyum (2).
Analysis of Surface Markers. Separated lymphocytes
were analyzed for T- and B-cell markers by 2 rosetting
assays. The ability of peripheral resting T-cells to produce
rosettes (E rosettes) with sheep ABC was used as a marker
for T-cells. The non-T-celbs were assessed by their ability to
rosette with sheep ABC coated with a 1/200 dilution of
rabbit anti-sheep immunoglobubin (EA rosettes). The tech
nique described by Jondab et al. (11) was followed for both
the E and EA rosette assays. The lymphocytes and sheep
ABC suspension were incubated as a pellet overnight at 4°.
The E rosettes were then suspended very gently, whereas
the EA suspension was Vigorously agitated in a Van-whirl
mixer before being counted.
CANCERRESEARCHVOL. 36
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Lymphocyte Functions in Uro!ogica! Cancer Patients
Table 1
% released by IRS (on PHA)
Number and age distribution of urological cancer patients
Age (yr)Na.
mensYoung
of
subjectsNo.
% specific lysis =
of
speci
—
%
released
in
NRS
(or
MEM)
NRS
(or
MEM)
% released by Triton
—
%
released
in
The percentage released by Triton represents the total
lysable counts (80% average). The IRS was obtained by
giving a rabbit a weekly i.v. injection of 10@D6 mouse tumor
cells for 3 weeks. Three days after the last injection the
animal was bled, and this serum was used at a 1/5@
dilution
which had maximal cytotoxic activity with normal lympho
cancers65.3
±11.0103139a
cytes in the ADCC assay. Both the IRS and NRS were heat
inactivated at 56°
for 60 mm. In the absence of effector cells,
Mean ±S.D..
neither of the sera caused any increased lysis of the target
PHA3-proliferative Assay. Purified lymphocyteswere cub cells over that obtained with cells cultivated in MEM alone
tured in microplates (microtest-2 Falcon) (Van Water and (<10%). PHA was diluted in MEM and added at a 1/@@
Rogers, Los Angeles, Calif.) at a concentration of 10@ dilution (10 /Lg) in 0.1 ml.
cells/mb in MEM plus 10% AB serum. Three concentrations
Statistical Analysis. Student's t test was used in the
of PHA (Burroughs Wellcome and Co. , Research Triangle
analysis of all test results. One-sided x2analysis was used to
Park, N. C.) were added at the start of the culture (0.5, 1, evaluate the frequency of certain observations. The results
and 5 pg). Two days later 2 @Ci
[3H]thymidine (New England
were considered significant when p < 0.01.
Nuclear, Boston, Mass.) in 30 pi were added, and the cells
were incubated for a further 18 hr. The cells were harvested
RESULTS
on Whatman Grade 934 AH glass filters (Reeve Angel, Clif
ton, N. J.) in a manifold collector (MASH), and uptake of the
The results obtained in the cancer patient population
[3H]thymidine was determined in a f3 counter after the filters
were evaluated and will be presented as a function of the
were washed 5 times in distilled water. PHA proliferation
tumor load present at the time of testing. The patients were
index was calculated according to the formula:
controls26.7
±4.1@41113Age-matched
contrals58.7
±10.14752Localizedwithcancer62.9
±12.92732Localized
without evi
±10.94153dent
67.9
cancerMetastatic
±9.53554Allcancer64.4
separated into2 groups relatedto the stage of the disease,
i.e., localized to the organ of origin, or metastatic. The
PHA index = cpm in presence of PHA
cpm in absence of PHA
Cytotoxic Assays. A gross virus-induced mouse lym
phoma (D6) of thymic origin from C3H mice (courtesy of Dr.
H. Weben, Department of Microbiology and Immunology,
UCLA) was used as a target
serum. Immediately before testing,the D6 cellswere Ia
beled with 51Crfollowing the technique of Wigzell (17). We
used 0.4 p.Ci sodium chnomate (l.C.N. Corporation, Irvine,
Calif.), which was added to 10@tumor cells in 1 ml MEM
supplemented with 10% fetal calf serum. After incubation at
37°for 90 mm the labeled cells were washed 4 times and
were added to the effecton lymphocytes at the ratio of 1:20
(target:effector). The tumor cell number was always kept
constant at 10@ cells/tube
yielding3000 to 5000 cpm. PHA,
IRS prepared against the target cells, NAS was then added
at a volume of 0.1 ml. The mixture was centrifuged at 185 x
g for 5 mm at room temperature and the pellet was incu
bated for 2 to 3 hr at 37°.The reaction was stopped by the
addition of 1 ml cold MEM, and the mixture was centrifuged
in the cold for 8 mm at 185 x g. The supemnatant was
decanted for determination of radioactivity in a y counter
(Nuclear Chicago, Chicago, Ill.). Percentage of specific lysis
was calculated according to the formula:
abbreviations
used
are:
PHA,
phytohemagglutinin;
MEM,
minimal
essential medium; IRS, immune rabbit serum; NRS, normal rabbit serum;
ADCC, antibody-dependent cellular cytotaxicity; PDCC, PHA-dependent cal
lular cytotoxicity; PMN, polymarphonuclear leukocyte; DNCB, 2,4-dinitro
chlorobenzene.
JANUARY
1976
Lymphocyte Subpopulation
cell in these assays. It was
grown in suspension in Tissue Culture Medium MJLA (Re
heis, Los Angeles, Calif.) supplemented with 10% fetal calf
3 The
group with localized cancer was divided into 2 categories,
namely, with or without clinically evident cancer at the time
of testing, i.e., pre- and postoperatively.
T-dependent Lymphocytes (E Rosettes). The sponta
neous sheep RBC-rosetting test was used to obtain the
percentage of T-dependent lymphocytes in separated blood
(Table 2). There was no significant difference between the
percentage of E rosettes in the young controls (68.39) and
that in the age-matched controls (67.20). On the other hand,
the difference
between
the age-matched
controls
and the
mean score of patients with localized cancer (62.98), meta
static cancer (58.24), or all combined (61.37) was statisti
cabbysignificant.
Non-T-dependent Lymphocytes (Fc Rosettes). Using an
tibody-coated sheep ABC in the rosette assays, the percent
age of non-T-dependent lymphocytes in separated blood
was obtained (Table 2). There was no significant difference
between the percentage of EA rosettes in the young con
trols (19.45) and that in age-matched controls (19.34).
However, the difference between the age-matched controls
and the mean score of patients with localized cancer
(22.92), metastatic cancer (25.01), or both combined (23.36)
was significant.
PHA Proliferative Assay
The blastogenic response to PHA in the 3 populations is
presented
in Table 2 as PHA index. The PHA index of cancer
133
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M. M. E!hi!ali et a!.
Table2
Comparisonof in vitro lymphocytestudies between controls and cancer patients
lysis)Young
E rosettes (%)Fc
controls68.39
(32)Age-matched
controls67.20
(38)Localized
with cancer64.20
(9)Localized
withoutevident cancer62.98
±0.92a
(100)C19.45
±1.40
(49)19.34
±2.02
(31)21.12
±1•77b
±0.64
(100)275.83
±0.74
(49)151
±1.58
(31)159.68
±1 00b
(53)22.92
(53)67.65
(19)Metastatic
cancer58.24
±1,83b
(54)25.01
(22)All
cancers61.37
±1,1ob
(138)23.36
a Mean
b p
<
C Numbers
(%
(% lysis)PDCC
±1.14
±2,79
(32)6.39
±2.49
(38)7.00
±2.94
(9)11.46
±42.82
(50)24.91
.82 ±27.59
(41)32.88
±57.01
(15)27.83
±1612b
±1.01
±2.71
±1.91
±3.43
(19)7.97
(19)34.13
±1 29D
(54)86.30
±0,74b
(138)99.63
±1.03
.89 ±2.42
±22.95
(22)31
(22)3.78
±18@83b
±1.76
(56)31.94
(50)6.75
±1.04
(50)
± S.E.
0.01
when
in
compared
parentheses,
to
results
obtained
in
age-matched
controls
by
Student's
t test.
n.
patients as a group (99.63) was significantly depressed corn
pared to that of young controls (275.83) and age-matched
controls (151.82). The difference between the group of local
ized cancer patients without evident disease and the age
matched controls is significant.
Because the PHA index does not take into consideration
the daily experimental Variation, the PHA index of the test
@
indexADCC
rosettes (%)PHA
60
50
x
40
(I)
z
w
@1\@
x@x
Cancer
Pct,ents(
@——-—.
Controls(n@83)
subject was divided by the average of the PHA index of the
(.)
controls studied the same day. We calculated the percent
age of specimens that gave a PHA index less than 50% of the
control value for that day for each group. In the localized
cancer group with evident disease, the percentage of pa
tientS having less than 50% PHA index was 60%; whereas in
the group without evident cancer, this was 63%. In meta
static cancer this was 72% and in the overall cancer group it
was 63%.
The PHA index was obtained on 3 or more occasions in 8
young controls. In 3 subjects the multiple determinations
were quite similar. In 2 subjects the difference between any
2 determinations was less than 1 S.D. , while in the other 3
‘I,
(youngandage-matched).
Thefrequency(%)in eachpopulationis plotted
againstthePHAindex.A biphasiccurveis notedforthecontrolpopulation.
subjects the difference
The incidence of high response was greatly reduced in cancer patients.
was more pronounced
.. These varia
tions, we believe, are inherent in the test itself. The changes
in results obtained on different days can be minimized by
comparing the result of the test subject to 2 or more con
trols tested under the same conditions on the same day.
When the results of the PHA index were plotted in cancer
patients and controls, the values for young controls were
noted to cluster into 2 distinct groups, one being less than
150 and the other group being above 350. The young con
trols tended to have a greater distribution in the high group,
whereas the age-matched controls had more values on the
bower side with few high values. In cancer patients this
distribution was even more pronounced, with most of the
values occurring on the lower side and only an occasional
high responder. When the number of specimens in cancer
patients and controls (percentage of the total) was plotted
against the PHA index in increments of 50 (Chart 1), the
difference in distribution was readily apparent.
The PHA index results in all groups were arbitrarily di
vided into 2 subgroups
with a PHA index value more or less
than 50. The 1st group (>50) was considered to be the high
responders, and the 2nd group (<50) were the low respond
134
30
U.
0
‘p
20
I0
x____.___@@,__.
0L50
!
_S,. @,
00-ISO ‘ 200-250
PHA
Chart 1. PHA proliferative
300-350 >350
INDEX
response in cancer patients and controls
ers (Table 3). The differences between the number of sub
jects in the low and high responder groups of the cancer,
age-matched, and young control populations were highly
significant (x2@
P < 0.005). The incidence of low response
was higher in the cancer patients as compared to the con
trois. When the PHA index/control data were analyzed, the
index was less than 0.5 in 35 cancer patients (mean 0.14 ±
0.01), whereas this value was higher than 0.5 in 21 cancer
patients (mean 1.79 ±0.31).
Cytotoxiclty Results
ADCC.
As shown
in Table
2, the
difference
between
me
suIts obtained with young controls (24.91) and those for
age-matched controls (32.88) is significant. The differences
between age-matched controls and cancer patients are not
statistically significant.
PDCC.
No significant
differences
were
found
between
the
different cancer groups and the control groups (Table 2).
CANCERRESEARCHVOL. 36
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Lymphocyte Functions in Urological Cancer Patients
Influence of Blood Separation Purity on Results
The incidence of PMN contamination of more than 15% in
separated blood was compared in young controls, age
matched controls, and cancer patients. In young controls
the incidence of contamination was 1% as compared to 43%
in age-matched controls and 40% in cancer patients. These
differences are highly significant. The influence of this con
tamination on the assay results in age-matched controls
and cancer patients was studied.
In age-matched controls no significant effect was demon
strated on E or Fc rosettes or on PDCC. The PHA index with
PMN less than 15% was 236.82 ±57.43 (mean ±S.E.) as
compared to 137.01 ±26.91 when PMN was more than 15%.
This difference was not statistically significant. The ADCC
results were 29.12 ±3.41 with PMN less than 15% and 39.17
±4 .06 with
PMN
more
than
1 5% . This
difference
was
statisti
to nonradiotherapy group. There were no significant differ
ences in the ADCC results.
The PHA index results were analyzed in the nonradiated
group after elimination of data from patients with a higher
than 15% polymorphonuclear contamination. The group of
patients with localized but evident cancer (170.95 ±71.64)
and the group with metastatic cancer (144.71 ±60.00) were
not significantly lower than the age-matched controls
(236.82 ±57.43). The differences between the group of
patients with localized but no evident cancer (51.37 ±
14.29) and the age-matched controls were significant. Simi
larly, the PHA index results in the overall cancer group
(120.17 ±31.85) were significantly different from those
obtained in age-matched controls.
Correlation of Laboratory Results and Delayed Hypersen
sitivity
cabbysignificant.
In the group of cancer patients only the PDCC assay was
significantly elevated when the PMN contamination was
less than 15% (10.48 ±2.21) as compared to a higher than
15% contamination (4.25 ±0.95).
Reactions
The cancer patients studied were concomitantly skin
tested as part of the investigative protocol. The results of 97
patients sensitized to DNCB were divided into positive and
negative responders and were compared to the laboratory
data.
Radiotherapy Effect
A group of patients who had received radiotherapy (39
patients) was divided into 3 subgroups. The 1st group had
completed radiation within 3 months of testing, the 2nd
group within 6 months to 1 year, and the 3rd group more
than 1 year prior to testing. The radiation was confined to
the pelvis in all but 2 patients, who received therapy to the
spine or breasts. The dose of radiotherapy ranged between
1600 and 7000 rads.
Most patients
received
Levels of E or Fc rosettes, ADCC, or PDCC were not found
to correlate with the presence or absence of positive DNCB
response. There was a significant reduction in the PHA
index, however, in the DNCB nonresponders (34.70 ±9.28)
as compared to the DNCB-positive group of patients (169.76
± 43.86).
A more
complete
account
of
other
skin
testing
results and their correlation to laboratory tests and other
variables will be published separately.
1600 rads,
which was administered prior to cystectomy.
There was a significant reduction in E rosettes in the
group of patientswhose radiationwas completed more than
1 year prior to testing as compared to either the age
matched controls or the group with no radiation (Table 4).
In the same group, the Fc rosettes were significantly higher
than in age-matched controls on the group without radia
tion. The PHA index shows a tendency to decrease after the
onset of radiation which is even more depressed at the 6month to 1-year period but recovers slightly 1 year after
radiation. The results of PDCC showed a pattern similar to
that of the PHA index, with significant reduction within 3
months and 6 months to 1 year postradiation as compared
DISCUSSION
The studies undertaken included a broad assessment of
the lymphocyte functions in a population of patients with
different stages of unobogical cancer. The influence of tu
momload and the effect of radiation therapy were studied.
The choice of tests has been partly empirical and partly
influenced by our own experience with certain assays. The
T- and B-cell surface markers are accepted as a means of
numerical evaluation of peripheral T- and B-cells. The PHA
proliferation assay has also been widely used as a measure
of T-cell function. Although the cells were exposed to var
ious doses of PHA, the data reported were obtained with the
Table 3
Distribution of blastogenic response in high and low responders
(56)Young
(41)High
controls
responders>50<50>50<50>50<50PHA
respondersLow
index
315.79±47.06a
(43)b.
a Mean
b Numbers
@
r
<
r30.25
ContraIs (91)Cancer
patients
(50)Age-matchedcontrols
respondersHigh
±4.95
([email protected]
respondersLow
±33.59
(28)C18.76
respondersHigh
±3.17
(13)―179.30
respondersLow
±29.50
(29)C14.50
±2.34
(27)@
± SE.
in parentheses,
by 1-sided
n.
x2 analysis
for the frequency
of law and high response
in each
group.
JANUARY1976
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135
M. M. Elhi!ali et a!.
Table4
Effect of radiation therapy on various lymphocytestudies
lysis)Age-matched
E rosettes (%)Fc
controls67.20
(38)No
radiation63.10
(26)Radiation
within 3 mo.63.66
(8)Radiation
6 mo.—1
yr
.82 ±27.59
(41)7.00
±29.63
±1.55
(32)9.41
(74)21.04
(74)134.40
±1.52
±16.92
±2.17
(11)2.00
(25)24.98
(25)60.47
±26.21
±2.55
±2.89
(7)
(14)
(14)
52.37 ±2@62@@,d
29.79 ±1,72@@d71.78±48.82
(6)4.69
(24)38.71
(24)21.92
Radiation >1 yr62.14
a Mean
<
0.01
d p < 0.01
±0.74
(49)151
±0.86
±1 @4Q
(49)b19.34
in
parentheses,
(% lysis)ADCC (%
.63 ±2.69
±1.01
(38)31
±1.65
±4.40
(26)30.45
±2.92
±2.38e
(8)31.34
±1.32'@
±5.64
(9)
(10)
5.04 ±0.73
32.88 ±2.49
(7)
(6)32.20
n.
when
compared
to
when
compared
to results
results
obtained
in
obtained
in age-matched
“No radiation―
lowest dose (1 i@g)found to give a consistent positive prolif
eration index in normal controls.
The 2 cytotoxic assays (ADCC and PDCC) were chosen
because of previous work in this laboratory (1, 18). The
ADCC measures a non-T-ceIb function (cytotoxicity) against
antibody-coated target cells (16). The PDCC measures cyto
toxic potential of lymphocytes in the presence of PHA. In
the short-term assay as we use it
, 2 hr), PHA is unlikely
to induce a cytotoxic potential in ‘
virgin―lymphocytes as
previously suggested (10). Instead, it is more likely to meas
ure a cytotoxic potential of in vivo activated effector cells,
probably of T-cell origin (B. Bonavida and A. Robins, per
sonal communication).
A significant reduction was found in the number of circu
bating T-cells (E rosettes) in cancer patients as compared to
the control groups. This was associated with a concomitant
reduction in PHA proliferative response as compared to the
control population. This depression of blastogenic re
sponse cannot be totally explained by the decrease in the
percentage of T-cebls. Such a conclusion is substantiated in
the age-matched controls where there was a discrepancy
between a normal T-celI number but a depressed PHA probif
erative response compared to the young controls. This
would lead us to assume that subclasses of E-rosetting T
cells exist and that those capable of proliferative responsive
ness to PHA are primarily
affected
in old age.
The variability in PHA response in the same subject tested
sequentially necessitates the use of multiple control speci
mens for these assays. To learn more about the reproducibil
ity of the PHA response, the data from the young control
population were plotted against variables such as PMN
contamination, percentage of lymphocytes, and age. The
responses spread across a wide range. This wide range
accounts for the large S.D. seen in this assay. The majority
of the young controls were high responders. The older
controls (58.7 ±10.1 years) had a greater percentage of low
responders than did the young controls. In the cancer popu
lation there was a wide distribution of responses, but the
increase in the number of bow responders was significantly
accentuated in the cancer group.
The various assays were evaluated to determine whether
they were affected by PMN contamination. The results were
136
indexPDCC
± S.E.
b Numbers
Cp
rosettes (%)PHA
group
controls
by
Student's
by Student's
t test.
t test.
arbitrarily divided into 2 groups with more or less than 15%
contamination. In the age-matched controls there is an
indication that the PHA index may be depressed when the
PMN count is high; however, too few patients were studied
to provide statistical significance. The major difference in
the control group was seen in the ADCC assay. The en
hanced ADCC response may be explained by the participa
tion of PMN in this reaction. In the cancer group the P0CC
assay was the only test significantly affected. The lowered
response seen with the PMN-contaminated specimens
(more than 15%) could be explained by the presence of fewer
available lymphocytes, which are required for this assay.
In studies of this nature it is important to consider the
influence of various therapeutic modalities. In the group of
patients receiving radiotherapy, we detected a significant
reduction in the percentage of E rosettes 1 year after com
pletion of treatment. The PHA blastogenic response was
found to be reduced to a greater extent in patients studied
within 3 months of radiation therapy, although the percent
age of E rosettes was not significantly reduced.
Stratton et al. (15) studied the effects of radiation therapy
delivered to the thymus on the numbers and functional
capacities of certain subpopulations of circulating bympho
cytes. They compared these results with pelvic radiation.
Using rosette formation , membrane immunofluorescence,
and blastogenic response to mitogens, they found that both
T- and B-cells undergo rapid depletion during radiotherapy,
whether or not the thymus is included in the treatment field.
The T-cells disappeared more rapidly and their eventual
drop is somewhat greater than that of B-cells. On the other
hand, Stjernsward et a!. (14) felt that radiation of the thymus
per se is the primary cause of the alterations in circulating
lymphocyte populations. Stratton et a!. (15) also demon
strated that the decrease in the ability to synthesize DNA in
response to mitogen stimulation is much greater than the
decrease in number of lymphocytes during radiotherapy, a
finding in line with our observation. The recovery of this
depression was biphasic, showing an improvement in 1 to 3
months followed by a depression in 4 to 6 months and
partial recovery by 7 to 12 months. This correlates
well with
the trend we noticed in the progression of PHA inhibition in
our results in a different patient population.
CANCERRESEARCHVOL. 36
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1976 American Association for Cancer Research.
Lymphocyte
Chee et a!. (5) concluded that the reduction in PHA
response following radiation was primarily due to the de
creased numbers of a subpopulation of circulating lympho
cytes normally responsive to PHA. The rapidly dividing bym
phocytes are probably the cells most vulnerable to radia
tion.
When the results of the cytotoxic assays were analyzed,
we noticed a normal number of cytotoxic effector cells
against antibody-coated target cells in cancer patients. This
finding is in contrast to what has been reported in patients
with hematobogical tumors (9). We do not have an explana
tion for this difference but the normal values for ADCC in
cancer patients corresponded well with the higher than
normal numbers of cells with Fc receptors (EA rosettes)
which seem to represent the effector cells in this cytotoxic
reaction (12). When the samples with more than 15% contami
nation
of PMN were eliminated,
the cancer
patients
as a
group displayed an increased cytotoxicity against heterobo
gous target cells in the presence of PHA. This kind of
cytotoxicity
(PDCC) could be a function
of activated
cyto
toxic cells in vivo that will express the cytotoxicity if brought
in contact with target cells by means of PHA. The effector
cells appearto be an activated thymus-denived cell (8). Such
cells have been found in the spleen of sarcoma-beaning
mice (16). An attractive possibility is that these effector cells
have specificity for the tumor target cell in vivo, and we
intend to compare the P0CC data in these patients with the
results of tests for specific cytotoxicity against human blad
dem cancer cells in Vitro. The presence of cell-mediated
cytotoxicity
associated
with
bladder
and kidney
cancer
seems to correlate with the stage and course of the disease
(3, 7, 13).
The value of studying immunocompetency in cancer pa
tients has been shown by correlations between the immune
state and stage of cancer (4). Additional value may be
gained by monitoring therapeutic modalities known to de
press the immune system. The possible value of these tests
in objectively assessing immunotherapy and designing opti
mal treatment regimens is still to be determined.
ACKNOWLEDGMENTS
The authors wish to express their thanks to Laurie Buckley, Ulla Claeson,
Candace Vescera, Pam Cohn, Wanda Hale, Neal Groch, Pablo Villanueva,
Functions
in Uro!ogical
Cancer Patients
and Joe Rosenblatt for their expert technical assistance. We are grateful to
Dr. B. Bonavida for his assistance in the establishment of the PDCC assay.
Dr. Virginia Clark, professor of biostatistics (UCLA, School of Public Health),
assisted in the statistical analysis of the data.
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JANUARY1976
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137
Critical Evaluation of Lymphocyte Functions in Urological
Cancer Patients
Mostafa M. Elhilali, Sven Britton, Stanley Brosman, et al.
Cancer Res 1976;36:132-137.
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