Establishment and Characterization of Human Pancreatic ......Tissue Culture Characteristics. Tumor cells of the line SW-1990 grown in tissue culture appeared as large epithelial-like

[CANCER RESEARCH 43, 4393-4401, September 1983]

Establishment and Characterization of Human Pancreatic AdenocarcinomaCell Line SW-1990 in Tissue Culture and the Nude Mouse1

Andreas P. Kyriazis,2 William B. McCombs III, Avery A. Sandberg, Aikaterini A. Kyriazis, Nathan H. Sloane, and

Ross LÃ©pera

Department of Pathology, University of Medicine and Dentistry of New Jersey, New Jersey Medical School, Newark, New Jersey 07103 [A. P. K., A. A. K.]; The Scott andWhite Clinic, Temple, Texas 76501 [W. B, M.]; Roswell Park Memorial Institute, Buffalo, New York 14263 [A. A. S.Â¡;and The Department of Biochemistry, University ofTennessee, Memphis, Tennessee 38163 [N. H. S.]

ABSTRACT

A new tumor line derived from a human pancreatic ductaladenocarcinoma of a 56-year-old Caucasian male was estab

lished in tissue culture and the nude mouse. In tissue culture,the neoplastic cells grew as large, epithelial-like, mucin-producingcells. Injection s.c. of 1 x 106 cultured neoplastic cells into nude

mice resulted in tumor formation histologically closely resemblingthe original neoplasm. Ultrastructurally, the neoplastic cellsshowed characteristics of ductal epithelium. The allozyme phe-notypic profile of the line was established in 14 geneticallydetermined loci, and chromosome studies showed a near-tetra-

ploid pattern. Production of macromolecules such as lactatedehydrogenase and carcinoembryonic antigen were present inmeasurable amounts in culture media, tumor cell extracts, nudemouse-grown tumors, and the serum of tumor-bearing mice in

amounts relative to tumor size. Pancreatic enzymes were notdetected. These characteristics make tumor line SW-1990 a

valuable tool in studying various aspects of the biology of humanpancreatic cancer.

INTRODUCTION

Human pancreatic cancer, a malignant neoplasia primarily ofductal origin (7), has demonstrated an increased incidence in thelast 2 decades (4,5). It ranks fourth as a cause of cancer deaths,exceeded only by cancer of the lung, large bowel, and breast(28). More than 21,000 cases are diagnosed each year, andinvariably all patients will die as a result of their disease (29).This poor prognosis may be explained by the late detection ofthe neoplastic process, lack of effective treatment, and limitedknowledge of its biological characteristics.

In an effort to cope with some of these problems, attemptshave been made to develop suitable animal models for pancreaticcancer. For this purpose, pancreatic tumors have been inducedwith chemical carcinogens in guinea pigs (35), hamsters (32,33),and rats (10, 25). However, answers to certain questions pertaining to human pancreatic cancer, i.e., factors influencinggrowth patterns and metastatic behavior, production of variousbiological markers, and response to treatment, may be facilitatedby using suitable in vivo and in vitro experimental models thatpermit direct testing of human pancreatic cancer. Thus, established and well-characterized cell lines of tumors of the human

exocrine pancreas may become valuable tools in such studies.As of today, attempts to establish such lines have been limited

' Supported in part by National Cancer Institute Grant CA-26693.2To whom requests for reprints should be addressed, at Department of Pathol

ogy, University of Medicine and Dentistry of New Jersey, New Jersey MedicalSchool, 100 Bergen Street, Newark, N. J. 07103.

Received October 20,1982; accepted June 1,1983.

with only few successful cases reported (1, 6, 8, 14, 20, 24, 31,38, 40) and with fewer cases having been adequately characterized.

We are reporting the establishment and characterization of anew cell line that was derived from a pancreatic adenocarcinomaof ductal origin. The line has been studied in both tissue cultureand the nude mouse with regard to growth characteristics,histopathological appearance, ultrastructural morphology, andproduction of certain biological markers such as LDH3 and CEA.

The karyotype of the line and the allozyme phenotypic profilealso have been studied.

MATERIALS AND METHODS

Cell line SW-1990 was established at Scott and White Clinic, Temple,Texas, in November 1978 from a pancreatic adenocarcinoma of a 56-year-old Caucasian male. The neoplastic tissue used for the initiation of

tissue culture was removed at the time of an exploratory laparotomy,during which a pancreatic tumor located in the tail of the pancreas andextending into the spleen was identified as the only intraabdominalneoplasm. Part of the pancreas with the tumor, in addition to the spleen,and the surrounding fibroadipose tissue were removed at that time.Histopathologically, the tumor was characterized as adenocarcinomaGrade II deriving from the exocrine pancreas and extending into thespleen and the adjacent fatty tissue.

In March 1979, the patient was found to have disease metastatic tothe liver, and chemotherapy was initiated. There was no improvementand the patient died on June 6, 1979. Postoperatively, and during thefollowing months until shortly before death, repeated determinations ofCEA levels were as follows: December 21,1978, 6.8 ng/ml; February 6,1979, 8.7 ng/ml; April 27, 1979, 21.3 ng/ml; May 27, 1979, 91.1 ng/ml.The patient's ABO blood group was A+.

For the initiation of tissue culture, parts of surgically removed tumorwere freed of necrotic tissues and blood clots and were processed asdescribed previously (23). Originally, Tissue Culture Medium L-15-CI wasused (22). Later, as the cultured cells adapted to in vitro growth, L-15

medium was used (21). Cells were routinely passed by removing themedium and overlaying the monolayer with 0.5% trypsin:0.2% EDTAsolution in 0.1 M phosphate-buffered saline, pH 7.8. Initially, 1:2 splits

were made, but by the sixth passage cultures were split to 1:100. At thepresent time, cell line SW-1990 is being maintained in L-15 medium

(GIBCO, Grand Island, N.Y.), containing 15% fetal bovine serum (GIBCO),and supplemented with penicillin (100 units/ml) and streptomycin (100Mg/ml). At the time of the initiation of these experiments, the cell culturewas free of Mycoplasma contamination when tested in Mycoplasmamedium (GIBCO).

Plating Efficiency. Plating efficiency was determined by seeding 1 x105 viable cells in 25-cu cm tissue culture flasks using complete L-15

medium. Fifteen hr later, the medium was discarded, and the attachedcells were collected following trypsinization and counted.

3The abbreviations used are: GIBCO, Grand Island Biological Co.; CEA, carcinoembryonic antigen; LDH, lactate dehydrogenase; LDH-1, LDH-2, LDH-3, LDH-4, LDH-5, lactate dehydrogenase isoenzymes 1 to 5.

SEPTEMBER 1983 4393

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A. P. Kyriazis et al.

Cell-doubling Time. Cell-doubling time was determined by counting

the number of viable cells from freshly trypsinized monolayers. Twentyflasks each receiving 1 x 106 cells were used, and countings were

performed at 12- to 24-hr intervals for 10 days. Throughout the entire

procedure, cell viability was determined by means of the trypan blueexclusion method.

Determination of CEA, Amylase, Trypsin, and Chymotrypsin. Plastic25-cu cm tissue culture flasks were inoculated with 1 x 106 viable tumorcells in 5 ml complete L-15 medium and incubated at 37Â° without

refeeding until termination of the experiment. Supernatants and cellsfrom 3 flasks at a time were collected on Days 7, 14, 21, and 35. Thesupernatant from each flask was transferred to a 15-ml centrifuge tube

and spun for 10 min at 500 x g to remove floating cells. The supematantswere transferred to sterile, screw-cap test tubes for storage at -80Â°

until assayed; CEA assays were performed on the supernatant fluid foreach time period, by the Abbott CEA-EIA diagnostic kit (Abbott Labora

tories, North Chicago, III.). Amylase assays were performed at each timeperiod by the amylase azure procedure as described by Hall ef al. (16).Titrimetric determinations of the pancreatic enzymes Chymotrypsin andtrypsin were determined for each time period as described by Dyck (11).The remaining cells in each flask were harvested with trypsin:EDTA, andthe final viable cell population was determined. CEA production wascalculated for 106 cells as has been described previously (26).

Human LDH QuantitÃ¤ten. The Corning Agarose Universal Electro-

phoresis System (Corning, Palo Alto, Calif.) was used to differentiate andquantitate human LDH isoenzymes in tissue culture media, cell extracts,and tumor-bearing nude mouse plasma. Human and mouse LDH isoen-

zymes were separated on the basis of different electrophoretic mobilities(20), and quantitation was done on a colorimetrie densitometer (HelenaQuick Scan, Jr., Helena Laboratories, Beaumont, Texas).

Allozyme Genetic Signature. The allozyme phenotypic profile ofcultured cells was determined using the method described by O'Brien ef

al. (30). Briefly, harvested cells were suspended in hypotonie 0.01 M Tris-HCI.-0.001 M disodium EDTA, pH 7.1, at a concentration of approximately5 x 107 cells and disrupted by freeze-thawing. The suspension was

centrifuged at 15,000 rpm and the supernatant used to resolve theisoenzymes. For this, the vertical starch gel electrophoresis apparatus(Buchler Instruments, Inc., Fort Lee, N. J.) was used. Gel preparation,gel electrophoresis, buffers, and composition of enzyme stains were thesame as described previously (30). HeLa and mouse cells were runsimultaneously as controls.

Karyotyping. Chromosome preparations for the cell line and tumorgrown in nude mice were made according to methods described in detailpreviously (37, 39). G-banding was performed on the specimens, and

the chromosomes were identified according to the Paris nomenclature.Marker chromosomes were those for which the origin could not beidentified with certainty, and they have been marked as such in thekaryotype.

Nude Mice. Female BALB/c athymic mice (Charles River BreedingLaboratories, Inc., Wilmington, Mass.) were maintained in a pathogen-

free environment. Before use, animals were tested for presence of LDHvirus, which affects the host's ability to clear LDH (36), according to the

method described previously (9, 20). Animals used in the present studywere found to be free of LDH virus infection.

Collection of Mouse Plasma Samples. For the determination of theamount of human LDH present in the plasma of tumor-bearing mice,

blood was drawn by orbital venipuncture with heparinized microcapillarytubes. Plasma was separated by centrifugation and stored at -80Â° until

assayed.Solid Tumors in Nude Mice. For injection into nude mice, cultured

tumor cells were dispersed with 0.5 % trypsin:0.2 % EDTA per liter inHanks' balanced salt solution (GIBCO) and adjusted to 1 x 106 viable

cells/0.2 ml. Cell viability was determined by the trypan blue dye exclusiontest. Initially, s.c. tumors were established by giving the animals injectionsof 1 x 106 viable tumor cells. In subsequent experiments, tumor propa

gation was accomplished by transplanting s.c. through a skin incision in

the anterior lateral thoracic region a small piece of tumor measuringapproximately 0.3 x 0.3 cm (18). Tumor size was measured in 2 dimensions with calipers, and tumor volume was calculated weekly by usingthe formula (2)

Length x (width)2 x 0.4

Nude mouse-grown tumors were removed, fixed in buffered formalin,

embedded in paraffin, and stained with hematoxylin and eosin, periodicacid-Schiff, and Mayer's mucicarmine.

For transmission electron microscopy, tumor tissue was cut intofragments 0.5 to 1 mm thick and fixed in cold 2.5% glutaraldehyde in0.1 M cacodylate buffer, pH 7.4, immediately after removal. They wereembedded in Spurr, and thin sections were prepared with a Reichertultramicrotome, stained with uranyl acetate and lead citrate, and examined in a Philips EM-300 electron microscope.

RESULTS

Tissue Culture Characteristics. Tumor cells of the line SW-1990 grown in tissue culture appeared as large epithelial-likemucin-producing cells. They were characterized by large, relatively dark-stained nuclei, and slightly basophilic, reticulated, or

vacuolated cytoplasm. Vacuoles were single or multiple, occasionally occupying the entire cytoplasm, resulting in the appearance of signet-ring cell forms (Figs. 1 to 3). Mucin was demon

strated histochemically within the cytoplasmic vacuoles as largedroplets or had a more diffuse distribution appearing in the formof fine cytoplasmic granules. The plating efficiency of the culturewas 28.6%, and the doubling time was calculated at 64 hr.

The allozyme phenotypic profile was studied with regard to 14genetically determined loci. The results of the study are presented in Table 1, and a comparison is made with the HeLa cellline. These findings confirmed the human characteristics of theline and made it distinct from other cell lines, thus providing aninvaluable internal control in affirming its purity.

Tumor Growth in Nude Mice. Transplantation of 1 x 106

viable tumor cells s.c. in nude mice resulted in tumor growthhaving a latency period between transplantation and first positiveevidence of tumor growth of 3 to 4 weeks. To establish thegrowth characteristics of nude mouse grown tumors, solid piecesof tumor measuring approximately 0.3 x 0.3 cm were transplanted s.c. as described in "Materials and Methods." The follow-

Table 1Allozyme genetic signature of cell line SW-1990

SODCell

lineSW-1990

HeLaPGM1"1-21PGDA APGM31-21GL011-2 2BSD1 1ADA1 1G6PDBAPEPAHuman1PEPCHumanHumanAK-111NP-LDHHumanHumanMP1HumanHuman1HumanHuman20 HumanPGM, phosphoglucomutase, EC 2.7.5.1 ; PGD, 6-phosphogluconate dehydrogenase, EC 1.1.1.44; GL01, glyoxalase I, EC 4.4.1.5; ESD, esterase D, EC 3.1.1.1 ; ADA,

adenosine deaminase, EC 3.5.4.4; G6PD, glucose-6-phosphate dehydrogenase, EC 1.1.1.49; PEPA, PEPC, peptidases A and C, EC 3.4.1.1; AK-1, adenylate kinase]EC 2.7.4.3; NP, purine-nucleoside phosphorylase, EC 2.4.2.1; LDH, EC 1.1.1.27; MP1, mannosephosphate Â¡somerase,EC 5.3.1.8; SOD Superoxide dismutase EC1.15.1.1.

4394 CANCER RESEARCH VOL. 43



Pancreatic Adenocarcinoma Line SW-1990

up of many consecutive transplant generations resulted in aconsistently reproducible growth pattern, shown in Chart 1.

Microscopically, s.c. growing tumors exhibited characteristicsof a Grade II adenocarcinoma similar to that of the originalneoplasm. They were made up of atypical and irregular glandularstructures and solid tumor masses supported by bands of fibro-

vascular stoma. The neoplastic cells were either cuboidal orcolumnar, showing nuclear polymorphism, hyperchromasia,moderate mitotic activity, and evidence of mucin production.Stratification of neoplastic cells with evidence of papillary formations were also seen (Fig. 5). The tumor invaded its bed andsurrounding tissues with evidence of micro- and macroinvasion.

Criteria for invasion were the same as those described previously(19).

Ultrastructurally, the tumor cell characteristics were those ofductal epithelium (12) and showed close similarity to thosereported for the Capan-1 tumor cell line (20). The apical plasmamembrane consisted of well-developed microvilli, and the adja

cent cytoplasm contained many microvesicles, some of themmarkedly dilated. The remainder of the cytoplasm containedshort profiles of rough-surfaced endoplasmic reticulum, few mi

tochondria, and vesicles containing flocculent material corresponding to mucin droplets. Adjacent cells showed extensiveinfoldings of the basal cytoplasm with the presence of well-

developed desmosomes (Figs. 6 and 7).Production of LDH, CEA, and Pancreatic Enzymes. We

further investigated the ability of cell line SW-1990 to produce

LDH and CEA under different environmental conditions. LDHwas present in measurable amounts in tissue culture media andcell extracts. The ability of neoplastic cells to synthesize LDHcontinued in the nude mouse where it was detected at a very

700

600-

IBOO-C

Ã• 400-

O

3OO-

cI

Â¡200-

100-

o \ i r234

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early stage following transplantation. Table 2 shows the LDHisoenzyme profile under varying conditions. The relationshipbetween tumor size in the nude mouse and circulating LDH isshown in Chart 2.

Under the experimental conditions described previously, CEAwas detected in measurable amounts in both tissue culture mediaand serum of tumor-bearing mice. The release of CEA in culture

media was more pronounced during the lag and death phasesof the cell cycle (Chart 3), an observation which was in accordwith previous finding in studies involving colon carcinoma celllines (26). The presence of CEA in the serum of tumor-bearing

animals was detected when tumors had reached an average sizeof approximately 500 cu mm. Thereafter, CEA increased proportionally to the tumor size. When comparing tumor size withthe corresponding levels of circulating human CEA, a highlystatistically significant correlation was observed (Chart 4).

Tests for amylase, trypsin, and chymotrypsin all were negative.Chromosome Studies. Examination of the cell line SW-1990

obtained in tissue culture revealed the chromosome counts torange from 40 to 97, with most of the numbers being in thetetraploid range (67 to 75 chromosomes). In addition, a population of cells with 2 or 3 times the number of chromosomes in thetetraploid range was present, but their number was rather small.A total of 17 (31%) of the 54 metaphases examined had dicentricchromosomes, their number ranging from 1 to 8 per metaphase.Of more interest was the fact that 33 (61%) of the 54 metaphaseshad double minute chromosomes. The number of double minutechromosomes was 1 or 2 per cell. The metaphases showedmany morphological abnormalities ranging from deleted chromosomes to extra material on chromosomes (Fig. 8).

550"

LUm

490-

LU 430-OOff

> 370HzLUO

LU 310-i-<

< 250-

r-0.60

u^

Chart 1. Growth curve of SW-1990 tumor grown in the nude mouse. Each pointrepresents the mean of 6 animals. Bars, S.D.

200 400 600 800 1,000 1,200 1.400 1,600

TUMOR VOLUME in cu mm

Chart 2. Relationship between tumor volume and circulating LDH in the nudemouse. Each point represents a single animal. U/L, units/liter.

Table 2LDH isoenzyme profile of cell line SW-1990

Relative percentagevaluesSourceCulture

mediumCell extractsSerum of tumor-bearing

miceControlLDH-55.45

Â±2.47'

6.9 Â±0.1411. 25Â±2.0511.

20 Â±0.7LDH-412.6

+ 0.2826.65 Â±0.968.55 Â±8.415.35

Â±0.2LDH-351

.2 Â±0.2837.7 Â±2.1211.2Â±2.8215.4

Â±0.98LDH-228.55

Â±2.4725.85 Â±0.77

6.1 Â±2.435.05

Â±1.0LDH-12.1

5 Â±0.632.95 Â±0.492.85 Â±1.233.0

Â±0.42a Mean Â±S.D. All determinations were run in triplicate.

SEPTEMBER 1983 4395




uiu

2.62.5-

2.2.3-

2,2-

2.1-

2.0-

1,9-

1.8-

1.7-

1,6-

1.5-

1.4-

1.3-

1.2-

1,1-

1.0-

90

80

70

60 <luO

50E

40 I

30

20

10

47 11 14 18 21 25 28 32

DAYS

Charts. Release of CEA in tissue culture medium from tumor line SW-1990.

, number of viable tumor cells at any given time; , amount of CEApresent in the culture medium at the same time.

80~

70-

60-

50-

40-

30-

20-I r.0.97

10-1 - /â€¢â€¢ p<0.001

01c

LUO

i r500 1,000 1,500

TUMOR VOLUME Â¡neu mm

Chart 4. Relationship between circulating CEA and tumor volume in the nudemouse.

The material obtained from the nude mouse was not of thesame quality as that seen in the cell line in culture; almost all thecells had high chromosome numbers ranging from twice that ofthe modal number in the cell line to 3 times the number. Detailedanalysis could not be performed on these cells, although theyappear to have karyotypic changes similar to those observed inthe tissue culture material.

DISCUSSION

The origin of the line from a Caucasian patient was confirmedthrough studies of glucose-6-phosphate dehydrogenase identi

fying it as type B, and studies of the allozyme phenotypic profilemade it distinct from any other cell line, thus providing valuable

information against any accidental inter- or intraspecies contam

ination. In addition, its epithelial character was evident in bothtissue culture and in the nude mouse, the production of mucinin both systems identified it with a secretory epithelium. Furthermore, morphological studies showed that SW-1990 tumors

grown in nude mice exhibited characteristics identical with thoseof the tumor of origin, an observation in agreement with reportspublished previously on this subject (13, 20).

Of importance with regard to the cytogenetic examination isthe fact that the chromosomes observed, particularly followinggrowth in nude mice, were all of human origin. The basic kary-otype appeared to have chromosomes in the near-triploid range

with most of the cells containing between 67 and 75 chromosomes. Every group in the karyotype except chromosomes 5and 18 was affected by either morphological or numericalchanges. Since the number of adenocarcinomas of the pancreasin which detailed chromosome analysis has been performed isvery small (37), it is difficult to say how the findings obtainedrelate to those of other pancreatic cancers. Undoubtedly, non-

random, if not specific, changes may be shown to exist in thiscancer, as have been described in ovarian cancer, meningiomas,and small-cell carcinomas of the lung (37); but on the basis of

our findings, in which the cells contain a large number of karyotypic abnormalities, it may be difficult to pinpoint at the presenttime the one change which may characterize pancreatic cancers.It is hoped that as more data are accumulated in this field it willbecome more clear as to which changes are primary and whichare secondary. An additional finding in the karyotype of the lineis the absence of the Y-chromosome (Fig. 8), an observationwhich is not uncommon in abnormal and normal cells in long-

term tissue culture and occasionally in tumors in vivo.LDH was selected as a tumor marker because it is present in

all human cells, and it is easy to demonstrate and quantitate.Furthermore, since human and mouse LDH have different elec-trophoretic mobilities, they can be easily separated by electro-phoresis (20). This property makes LDH a useful marker inidentifying the human nature of any tumor grown in nude mice,and to a certain extent it may be used as an indicator of theamount of the viable tumor mass (9). The latter test, however,should be used with caution, since tumor necrosis, cystic degeneration, animal handing, and bleeding practices may adverselyinfluence the amount of the circulating LDH. Furthermore, thelow correlation coefficient between tumor volume and circulatingLDH (Chart 2) argues against meaningful direct correlation between tumor size and LDH. From Table 2, it becomes apparentthat line SW-1990 is characterized by a very low concentrationof LDH-1, a constant finding in all 3 media examined. The lowconcentration of LDH-5 in culture media and cell extracts asopposed to the values obtained from serum of tumor-bearing

animals and the control, a finding observed by us in a number ofhuman cell lines of different histogenetic background, most probably reflects a cumulative effect in the latter media. It should benoted that mouse LDH-5 moves electrophoretically betweenhuman LDH-4 and LDH-3 (20) and, quite frequently, in a mixture

of mouse and human LDHs overlapping of isozymes of differentorigin may be observed in that region. This is most probably thereason for the high value of LDH-4 in the serum of tumor-bearing

animals.Increased levels of CEA have been reported in approximately

57% of patients with pancreatic cancer (3). Information, however,




Pancreatic Adone-carcinoma Line SW-1990

on the production of CEA by established pancreatic cancer linesis scanty, with only few lines having been reported as producingCEA in vitro (17, 27). There is no information as to the behaviorof all these lines, however, when transplanted into nude mice,and no attempt has been made to investigate the correlationbetween tumor size and circulating levels of CEA. The data thatwe have presented in this report indicate that line SW-1990

preserves its ability to elaborate CEA in the nude mouse. It isworth emphasizing that the levels of circulating CEA reflectedthe tumor size as indicated by the highly statistically significantcorrelation coefficient between levels of CEA and the corresponding tumor volume (Chart 4). This observation and a comparison between the levels of CEA in the patient from whom thetumor originated and those in the tumor-bearing mouse indicatethat nude mouse-xenotransplanted tumors continue to elaborate

biological products at an amount and rate comparable to thatobserved in the patient.

Pancreatic enzymes, amylase, trypsin, and chymotrypsin werenot produced by SW-1990 cells. This finding is in agreement

with previous reports indicating that pancreatic carcinomas ofductal origin do not elaborate pancreas-related enzymes (15)

and further corroborates the origin of the present line from thepancreatic ductal system.

In conclusion, line SW-1990 derived from a human pancreatic

carcinoma of ductal origin shows morpological, biological, andbiochemical characteristics similar to that of the original tumor.It grows both in tissue culture and the nude mouse and expresses a distinct enzyme phenotypic profile. Furthermore, theexpression of some differentiated characters (Figs. 4 and 5) maybe of value to those investigators who may undertake studies ofthe differentiation markers of cancers of the digestive system.Because of these characteristics, tumor line SW-1990 may con

tribute positively to improving our understanding of the variousaspects of human pancreatic cancer.

ACKNOWLEDGMENTS

We are grateful to Dr. Walter Dyck and John Lacester for their assistance inperforming the amylase, trypsin, and chymotrypsin assays.

REFERENCES

1. Akaki, T., and Kimoto, T. Establishment and characteristics of a humanpancreatic cancer cell line (HGC-25). Acta Pathol. Jpn., 27: 51-58, 1977.

2. Attia, M. A., and Weiss, D. W. Immunology of spontaneous mammary carcinomas in mice. V. Acquired tumor resistance and enhancement in strain Amice infected with mammary tumor virus. Cancer Res., 26: 1787-1800,1966.

3. Bender, R. A., Weintraub, B. D., and Rosen, S. W. Prospective evaluation oftwo tumor-associated proteins in pancreatic adenocarcinoma. Cancer (Phila.),43:591-595, 1979.

4. Carter, S. K., and Comis, R. L. The integration of chemotherapy into acombined modality approach for cancer treatment. VI. Pancreatic adenocarcinoma. Cancer Treat. Rev., 2:193-214,1975.

5. Carter, S. K., and Comis, R. L. Adenocarcinoma of the pancreas: currenttherapeutic approaches, prognostic variables and criteria of response. In: M.J. Staquet (ed.), Cancer Therapy, pp. 237-253. New York: Raven Press, 1975.

6. Courtenay, V. O., and Mills, J. An in vitro colony assay for human tumorsgrown in immune-suppressed mice and treated in vivo with cytotoxic agents.Br. J. Cancer, 37: 261-269, 1978.

7. Cubilla, A. L., and Fitzgerald P. J. Morphological patterns of primary nonen-docrine human pancreas carcinoma. Cancer Res., 35:2234-2247,1975.

8. Dexter, D. L., Matook, G. M., Meitner, P. A., Bogaars, H. A., Jolly, G. A.,Turner, M. D., and Calabresi, P. Establishment and characterization of twohuman pancreatic cancer cell lines tumorigenic in athymic mice. Cancer Res.,42:2705-2714,1982.

9. DiPersio, L., Kyriazis, A. P., Michael, J. G., and Pesce, A. J. Monitoring thetherapy of human tumor xenografts in nude mice by the use of lÃ¡clatedehydrogenase. J. Nati. Cancer Inst., 62. 375-379,1979.

10. Dissin, J., Mills, L. R., Main, D. L., Black, 0., and Webster, P. D. Experimentalinduction of pancreatic adenocarcinoma in rats. J. Nati. Cancer Inst., 55: 857-864, 1975.

11. Dyck. W. P. Titrimetric measurements of fecal trypsin and chymotrypsin incystic fibrosis with pancreatic exocrine insufficiency. Am. J. Dig. Dis., 72:310-317,1967.

12. Ekholm, R., Zelander, T., and Edlund, Y. The ultrastructural organization ofthe rat exocrine pancreas: II. Centroacinar cells, intercalary and intralobularducts. J. Utrastruct. Res., 7: 78-83, 1962.

13. Fogh, J., Bean, M. A., Bruggen, J., Fogh, H., Fogh, J. M., Hammar, S. P.,Kodera, Y., Loveless, J. D., Sorg, C., and Wright, W. C. Comparison of ahuman tumor cell line before and after growth in the nude mouse. In: J. Foghand B. C. Giovanella (eds.), The Nude Mouse in Experimental and ClinicalResearch, pp. 215-234. New York: Academic Press, Inc., 1978.

14. Grant, A. G., Duke, D., and Hermon-Taylor, J. Establishment and characterization of primary human pancreatic carcinoma in continuous cell culture andin nude mice. Br. J. Cancer, 39: 143-151, 1979.

15. Grant, A. G., McGlashan, D., and Hermon-Taylor, J. A study of pancreaticsecretory and intracellular enzymes in pancreatic cancer tissue, other gastrointestinal cancers, normal pancreas and serum. Clin. Chim. Acta, 90: 75-82,1978.

16. Hall, F. F., Gulp, T. W., Hayakawa, T.. Ratliff, C. R, and Hightower, N. D. Animproved amylase assay using a new starch derivative. Am. J. Clin. Pathol.,53: 627-634,1970.

17. Kaku, M., Nishiyama, T., Yagawa, K., and Abe, M. Establishment of a carci-noembryonic antigen-producing cell line from human pancreatic carcinoma.Gann, 77:596-601,1980.

18. Kyriazis, A. A., and Kyriazis, A. P. Preferential sites of growth of human tumorsin nude mice following subcutaneous transplantation. Cancer Res., 40: 4509-4511. 1980.

19. Kyriazis, A. P., Kyriazis, A. A., McCombs, W. B., Ill, and Kereiakes, J. G.Biological behavior of human malignant tumors grown in the nude mouse.Cancer Res., 47: 3995-4000, 1981.

20. Kyriazis, A. P., Kyriazis, A. A., Scarpelli, D. G., Fogh, J., Rao, S., and LÃ©pera,R. Human pancreatic adenocarcinoma line Capan-1 in tissue culture and thenude mouse. Morphologic, biologic and biochemical characteristics. Am. J.Pathol., 706:250-260,1982.

21. Leibovitz, A. The growth and maintenance of tissue-cell cultures in free gasexchange with the atmosphere. Am. J. Hyg., 78: 173-180,1963.

22. Leibovitz, A., McCombs, W. B., Ill, Johnston, D., McCoy, C. E., and Stinson,J. C. New human cancer cell culture lines. I. SW-13, small cell carcinoma ofthe adrenal cortex. J. Nati. Cancer Inst., 57: 691-697,1973.

23. Leibovitz, A., Stinson, J. C., McCombs, W. B., Ill, McCoy, C. E., Mazur, K. C.,and Mabry, N. D. Classification of human colorectal adenocarcinoma cell lines.Cancer Res., 36: 4562-4569,1976.

24. Lieber, M., Massella, J., Nelson-Rees, W. A., Kaplan, M., and Todaro, G.Establishment of a continuous tumor-cell line (PANC-1) from a human carcinoma of the exocrine pancreas. Int. J. Cancer, 75: 741-747, 1975.

25. Longnecker, D. S., and Curphey, T. J. Adenocarcinoma of the pancreas inazaserine-treated rats. Cancer Res., 35: 2249-2258, 1975.

26. McCombs, W. B., Ill, Leibovitz, A., McCoy, C. E., Stinson, J. C., and Berlin, J.D. Morphologic and immunologie studies of a human colon tumor cell line (SW-48). Cancer (Phila.), 38: 2316-2327,1976.

27. Morgan, R. T., Woods, L. K., Moore, G. E., Quinn, L. A., McGavran, L., andGordon, S. G. Human cell line (COLO 357) of metastatic pancreatic adenocarcinoma. Int. J. Cancer, 25: 591-598,1980.

28. National Cancer Institute, Biometry Branch. National cancer survey adv. three-year report (excluding carcinoma in situ), Bethesda, Maryland, 1969-1971.Department of Health, Education, and Welfare Publication No. (NIH) 74-637.Washington, D. C.: Government Printing Office, 1974.

29. National Cancer Institute. Third National Cancer Survey: Cancer Statistics.CAâ€”Bull. Cancer Prog., 26: 14-29, 1976.

30. O'Brien, S. J., Shannon, J. E., and Gail, M. H. A molecular approach to the

identification and individualization of human and animal cells in culture: isozymeand allozyme genetic signatures. In Vitro (Rockville), 76:119-135,1980.

31. Owens, R. B., Smith, H. S., Nelson-Rees, W. A., and Springer, E. L. Epithelialcell cultures from normal and cancerous human tissues. J. Nati. Cancer Inst.,56: 843-849, 1976.

32. Pour, P., Althoff, J., Kruger, F. W., and Mohr, U. A potent pancreatic carcinogenin Syrian hamsters: A/-nitrosobis(2-oxopropyl)amine. J. Nati. Cancer Inst., 58:1449-1453,1977.

33. Pour, P., Kruger, F. W., Althoff, J., Cardesa, A., and Mohr, U. Cancer of thepancreas induced in the Syrian golden hamster. Am. J. Pathol., 76: 349-358,

1974.34. Povlsen, C. 0., Fialkow, P. J., Klein, E., Klein, G., Rygaard, J.. and Wiener, F.

Growth and antigenic properties of a biopsy-derived Burkitt's lymphoma in

thymusless (nude) mice. Int. J. Cancer, 77: 30-39,1973.

35. Reddy, J. K., and Rao, M. S. Pancreatic adenocarcinoma in inbred guinea pigsinduced by N-methyl-N-nitrosourea. Cancer Res., 35: 2269-2276,1975.

36. Riley, V., Loveless, J. D., Fitzmaurice, M. A., and Siler, W. M. Mechanism oflactate dehydrogenase (LDH) evaluation in virus infected hosts. Life Sci., 4:487-507,1965.

SEPTEMBER 1983 4397




37. Sandberg, A. A. The Chromosomes in Human Cancer and Leukemia. Amster- mosomes and causation of human cancer and leukemia. XLIV A method fordam: Elsevier/North-Holland Biomedicai Press, 1980. chromosome analysis of solid tumors. Cancer Genet. Cytogenet., 3: 1-10,

38. Schmidt. M., Deschner, E. E., Taler, H. T., Clements, L, and Good, R. A. 1981.Gastrointestinal cancer studies in the human to nude mouse heterotransplant 40. Yunis, A. A., Arimura, G. K., and Russin, D. J. Human pancreatic carcinomasystem. Gastroenterology, 7: 829-840, 1977. (MIA PaCa-2) in continuous culture: sensitivity to asparaginase. Int. J. Cancer,

39. Wake, N., Slochum, H., Rustum, Y., Matsui, S., and Sandberg, A. A. Chro- ?9.' 128-135, 1977.

Fig. 1. Cell line SW-1990 grown in tissue culture is characterized by large epithelial-like cells with large rounded or ovoid nuclei, chromatin clumping, and abundantreticulated or vacuolated cytoplasm containing mucin. H & E, x 250.

Fig. 2. Same as in Fig. 1, showing characteristic signet-ring formation. H & E, x 400.

Fig. 3. Same as in Fig. 1, showing a syncytial-looking cell aggregate with multivacuolated, mucin-containing cytoplasm. H & E, x 250.

Fig. 4. Original tumor. Nuclear polymorphism, mitotic activity, and a tendency to form broad-based papillary structures are evident. Cytoplasmic mucin in the form ofsmall or larger vacuoles is seen in many cells. H & E, x 250.

Fig. 5. Tumor cell line SW-1990 grown in the nude mouse showing morphological characteristics similar to those of the original tumor. Compare with Fig. 4. H & E,x250.

Fig. 6. Electron micrograph showing a portion of SW-1990 tumor growing in the nude mouse. The neoplastic cells are characterized by well-developed microvilli,numerous mucin-filled microvesicles, and poorly developed rough endoplasmic reticulum. Desmosomes connecting adjacent cells are also seen. Inset, higher magnificationof a desmosome. Uranyl acetate-lead citrate, x 12,600; msef, x 157,500.

Fig. 7. Higher magnification of same tumor as in Fig. 8. Well-developed microvilli, numerous mucin-containing vesicles, and interdigitation of cell membranes ofadjacent cells are characteristics of this type of tumor. Inset, higher magnification of microvilli with distinct glycocalyx. Uranyl acetate-lead citrate, x 22,500; inset, x67,500.

Fig. 8. Karyotype of cell line SW-1940 in the near-triploid range. The Y-chromosome is missing; however, X-chromosomes, including a shortened X-chromosome anda double minute chromosome (DMS) are present, mar, marker.





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1983;43:4393-4401. Cancer Res Andreas P. Kyriazis, William B. McCombs III, Avery A. Sandberg, et al. Nude MouseAdenocarcinoma Cell Line SW-1990 in Tissue Culture and the Establishment and Characterization of Human Pancreatic

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Establishment and Characterization of Human Pancreatic ......Tissue Culture Characteristics. Tumor cells of the line SW-1990 grown in tissue culture appeared as large epithelial-like