During recent years, we have cultured SCLC from lymph nodes, pleural effusion, peritoneal 
effusion, and bone marrow from 36 patients at the Miami V.A. Medical Center. From six patients we 
obtained samples twice (at the time of diagnosis and relapse), from one patient we obtained two 
samples, from bone barrow and from pleural fluid, and from one patient we obtained three samples, 
two from bone marrow at diagnosis and at relapse, and one from peritoneal fluid. We were able to 
establish 6 permanent cell lines and 2 short term cultures (6 months) from 1 1 samples with histologically 
or cytologically confirmed diagnosis of SCLC in the bone marrow or malignant effusion. Three bone 
marrow samples with histological confirmed diagnosis of SCLC did not grow; one of these samples 
from a previously treated patient. On the other hand, we were able to establish cultures from three 
patients with extensive SCLC whose bone marrow biopsies appeared normal. Similar results also have 
been published by Hunter et al. (72). He reported 22% positive tumor colony growth in histologically 
negative marrows. Furthermore, studies using one to three different monoclonal antibodies have 
documented a 28%-43% increased sensitivity in detecting bone marrow metastasis over conventional 
light microscopy technique. I 
! 
A summary of our culture results (73-75) is shown in Table 6 (see Appendix C). The characteristics 
of our SCLC lines are summarized in Table 7 (see Appendix C). 
All cell lines show abnormalities in 3p chromosome and homozygosity loss at the DNF15S1 (3p21) 
loci by Southern analysis, in association with other chromosomal structural abnormalities. The 
chemosensitivity of these cell lines has been described (76,77). 
1.6 M1HC expression of and lymphokine production by transfected SCLC 
To date we have analyzed four recently established SCLC lines, one of which grows as adherent 
cell type and the other three as loose aggregates. Fig. 3, 4 (see Appendix C) shows the FACS analysis 
for MHC class I and class II expression in comparison to nonspecific IgG binding. Constant region 
recognizing MHC antibodies were used. All four SCLC lines express class I. Class II MHC is also 
present except in line one, where it is barely detectable above background. It is of interest to note the i 
resemblance to the murine LLC also with regard to the apparent membrane shedding and the ability to 
grow semi adherent. 
Transfection and selection in G418 does not significantly effect the cell surface expression of MHC 
molecules Fig. 4 (Appendix C). IL2 production in SCLC#1 was 5720U/ml 5xl0 6 cells x 24hr. 
SCLC-hI12#l was irradiated at 0, 1500, 3000, 6000, and 12000 rad and plated in microtiter plates j 
at 5X10 4 cells/well. For the next ten days parallel samples were counted tested for 112 (Table 9a, b; see 
Appendix C). We found that by day 10 112 production had dropped to zero in parallel with loss of 
viability in cells irradiated with 6000 and 12000rad (Table 9b, Appendix C). For the first four days little 
decline in 112 production was observed (350U/well). By day 8 the production was still 90U/well. These 
studies indicate that proliferation is not necessary for 112 production and hence that lethally irradiated, 
transfected tumor cells ought to function well in the induction of an immune response. 
These data indicate that SCLC a. is transfectable similar to LLC; b. that transfection and selection 
does not change the surface expression of MHC; c. that SCLC produces high levels of lymphokines; 
and d. that 12000rad irradiation results in slow cell death, allowing the production of lymphokine for 
at least eight days. SCLC cell lines thus fulfill the necessary criteria for gene therapy. 
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