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CISPLATIN BASED CHEMOTHERAPY WITH ACCELERATED, B.I.D. RADIATION THERAPY FOR ADVANCED NON-SMALL CELL LUNG CANCER (NSCLC) .J. PISCH, S. MALAMDD, E. BEATTIE, B. VIKRAM. BETH ISRAEL MEDICAL CENTER, N.Y., N.Y. 10003 The purpose of this study was to evaluate the efficacy and toxicity of simultaneous chemo-radiation using accelerated interrupted b.i.d. irradiation. Methods and Materials: Between December 1988 and June 1990 20 patients with locally advanced NSCLC (4 St. IIIA + 116 St. IIIB) were entered into this study. Patients received Cisplatin (30 mg/m2 Dl-3), VP16 (80 mglm2 Dl-3), 5FU(VOO mg/m2 Dl-4). Two-hundred cGy was delivered twice a day with 6 HR time gap between the two treatments. Cycle was repeated twice for resected or for borderline resect- able patient. Unresectable patients received 3 cycles and were followed by chest x-ray and CT scans. Results: -- Treatment was well tolerated in this patient population (30-72 yrs. old, Median 60. M:F=13:71)Dysphagia and leukopenia were the most frequent side effects requir- ing 3 instead of 2 weeks rest between the 2nd and 3rd cyc.te in 8 patients. Side-effects never were serious enough to command discontinuation of treatment. Twelve of the 16 Stage IIIB patients are evaluable for response. Five of the 12 had CR (41%). 2 had PR (16%). 4 “roaressed artside the treatment ;ield-(33%). Oni pa&nt’sh&d progression in and outside the treatment field. Three of the IIIB patients underwent surgery - 1 had pathologic CR, 1 microscopic disease and 1 patient had unresectable tumor. Median follow up for the 12 IIIB patients is 8 months. Fiw patients are clinically NED at 18-13-10-8-l months post treatment. Our experience suggests that the program is sell tolerated with moderate toxicity. The 41% CR rate seen in patients with Stage IIIB disease is most encouraging.

HIGH LOCAL RECURRENCE AND BRAIN METASTASES IN ‘JON-SMALL CELL LUNG CANCER (NSCLC) TREATED WITH CHEMO-RADIOTHERAPY. Engracio P. Cortes and Daitatreyudu Nori, Booth MemoriarMedical Center, Flushmg, NY I 1355.

Thrty-seven NSCLC (IllA- 4, IIIB- 33) patlent were prospectively treated with Platmol 75mglm’ IV x I + 5pu lOOOmg/m’ 24.hr contmuous Infusion x 5 days + radiotherapy CRT) to the tumor bed and node drainine areas withIn I week after the start of chemotherapy to a total dose of 32-50 Gy. Patients who relapsed were treated wth Platlnol 75mgim’ IV xl + Velban 6mglm’ IVP x I T every other month Mltomycin-C lOmg/m’ IVP x I plus further RT If needed. Objertwe tumor responses were seen ,n 29 of 37 patients (78%): 13 complete, 16 partlal. Response by histology: a) squamous cell, 16 of 17; b) adenocarclnoma, 8 of 14; c) large ceil, 5 of 6. Overall median survival for 37 patrznts was 11.9 months: for non-responders, 4.2 months: partial responders, Il.2 months; complete responders, 16.3 months. Of the 29 responders, recurrences were: a) local, 15 (51%); b) bran, 9 (31%); c) bones, 3 (10%). Bran metastases were seen equally ,n all lustologles. Combmed chemo-radiotherapy m Stage IIIA, IIIB NSCLC was not enough to make a major impact on palient suw~val despite Its high response rate beaux of lngh local recurrence and bran metastases. Future design of combmed chemo-radiotherapy for NSCLC must mcorporate surgery and brachytherapy + bran RT, hopmg to dlrmmsh local recurrence and brain metastases.

COMBINATION IMMUNO(‘HEMOTHERAP1- IN PATIENTS WITH IXNG CANCER. Ariel Hollinshead. Geo.Wsshington Mzd.(‘tr..D.C.. L’S.4. Rr:search of the immune machanisms involved in solid tumor progression included B comparison of patients on chemotherspy(CTx~. immunotherapy t ITxl. immunochemotherapy(ICTx~ and controls. Me.lsurements of the humorocellulAr immune ch.mges

before. during and after therap>- indlcatc an integration between induction of long-lasting cell-mediated immunit! ((~‘MI: up to IO-14 yearsi Jnd the induction of car11 1st year rises in serum antibody humoral immunity c HI I to selected lung tumor-associated .mtigen (TAAI epitope D36h6. separated b>, a\ffinit>- shromstograph! using TA4 monoclonsl sntlhodies. HI test was predlctivt: of response to therdpyisurvival at S-6 mos post-therAp)-.and test indicated that immuno- chemotherapy may he superior for pts with Jdeno- carcinoma. In responders there was a post-therap? dhsence of immune complexes !c’Ic’i. We h:lve examined failed pts for number and ah class of hands. ag type & biochemistry. type of complement binding and size in C’IC and data suggest a specific role C unclear! in . blocking the killing of cancer cells. The incidsnce of late non-regional metastvses in patients out past 10 yrs is consistent with the induction of quiescence by IC’Tx. and indicates a mechanism of induced dormancy-. thus justifying the usage of booster immunizations perhaps at about five years Jfter initial therspg. With further knowledge. we may improve upon the Phase 3 tri;lls results in 234 stage 1 and 2 pts with five year sut_Lival of 75% for pts on lung TAA Immunotherapy YS 49% for control pts.

A Novel Membrane Protein Induced During Commit- ment Stage of Squamous Differentiation in Normal Human Bronchial Epitheliai Cells

Mika Kakefuda, Yuk-Chor Wang, Godofredo Urbana, and Samuel D. Bernal. Section of Medical Oncology, University Hospital, Boston MA

Squamous differentiation in bronchial epithelium cells is a metaplastic change that occurs primarily after chronic injury and often precedes neoplasia. Commitment is a critical stage in squamous differentiation, but there is little information on the biochemical than chial epithelium. !v

es associated with this stage in normal bron- e now describe a 48 kD membrane protein

that is induced during the commitment stage of squamous differentiation of normal human bronchial epithelial (NBE) cells. After 24 hours of exposure to 5% fetal calf serum and 600uM calcium, approximately 40 % of NBE cells were com- mitted to terminal differentiation. By 36 hours, more than 95 % of the cells were committed. Between 24-36 hours, expression of SQMl (a 48kD protein) was increased on the surface membrane of NBE cells. During this time, there were no changes in mor- phology, keratins, mvolucrin, or thymidine incorporation, although by day 8, these committed cells showed changes in these various markers consistent with terminal differentiation. The commitment stage was associated with increased adhesion of NBE cells to extracellular matrix which was inhibited by anti- SQMl antibody. The cDNA sequence of the SQMl gene shows a domain homologous with several adhesion proteins of the integrin family including the human leukocyte adhesion protein, endothelial end platelet glycoprotein IIIa, and van Willebrand’s factor. In addition, SQMl expression was found to be directly correlated with the level of folate transport inlungepithelial cells. The expression of SQMl at the stage of commitment and its function related to cell adhesion and folate transport su est that SQMl may play a role in the regulation of squamous g# Ifferen- tiation in lung epithelial cells.

PHASE 11 TRIAL OF VERY HIGH DOSE CISPLATIN @DP) AND ETOPOSIDE (VP-16) AND MITOMYCIN-C (MITO-C) IN ADVANCED NON-SMALL CELL LUNG CANCER (NSCLC). P. Caguioa, P. Hesketh, S. Tansan, W. BadueI, A. Hesketh, D. Karp, N. DiMartino, R. Elanchard, K. Zaner and D. Camey. The Univ Hosp. Boston VA Med Ctr. Boston

City Hosp, Boston, MA 02118; Mater Misericordiae Hosp, Dublin, Ireland. Very high dose DDP alone has demonstrated a 36% response rate in advanced NSCLC (Gandara DR et al: JNCI 81:790-794, 1989). This regimen was relatively well tolerated with the dose limiting toxicity being peripheral neuropathy. As myelosuppression with very high dose DDP was minimal we attempted to determine whether the addition of VP-16 and Mito-C can augment very high dose DDP’s activity. We devised a 4 cycle regimen as follows: DDP lOOmglm2 (in 3% saline iv over 3 hours) on d 1 and 8 of a 28 d cycle; VP-16 6Omglm2 iv on d 1, 2, 8&9 of cycles 1 and 3; & MITO-C 8mg/m2 iv on d 1 of cycles 2 and 4. To date, pts entered: 37; M/F: 27/10; Median age: 56 years; AJCC Stage IIIB: 18 pts; Stage IV: 19pts; ECOG PS O-l: 23pts: PS 2: 14 pts. No pt had received prior chemotherapy. Presently, 34 pts are evaluable for response (3 pts < 1 cycle). Overall response rate is 47%: I complete (CR) and 1.5 partial (PR) responses. 13 pts had stable disease (SD) and 5 pts progressive disease (PD). Response rate was very significantly associated with performance status: PS O-1: 64% response rate (14/22 pts); PS 2: 17% response rate (202 pts). Median response duration: 20.5 wks. Median survival: CR+PR: 38 wks; SD:42wks; and PD:20 wks. Overall median survival: 35 wks. Renal toxicity was modest with the median highest serum creatinine 1.6mg/dl. Two pts had a creatinine > 3mg/dl. Neurologic toxicity was dose limiting with 22% of pts (8/37) developing a grade 2 peripheral neuropathy, an incidence comparable to that seen with very high dose DDP alone. The frequency of peripheral neuropathy and ototoxicity were directly related to cummulative DDP dose. Of pts receiving 2 600mg/m2 of DDP. 41% (7117) and 29.4% (507) developed grade 2 peripheral neuropathy and grade 2-3 ototoxicity respectively. Hematologic toxicity was significant with 14 pts (38%) having grade 3 or 4 leukopenia and/or thrombocytopenia. We conclude that this regimen has significant activity in NSCLC but is associated with considerably more myelosuppression than high dose DDP alone.

MECHANISMS OF DECREASED DRUG TRANSPORT IN CELLS RESISTANT TO METHOTREXATE AND CIS- PLATINUM

A. Godofredo Urbano, Yuk Chor Wong, Mika Kakefuda and Samuel D. Bernal

Section of Medical Oncology, University Hospital, Boston, MA 02118.

Resistance of carcinoma cells to chemotherapeutic drugs is multifactorial but altered membrane drug transport is an important mechanism that appears early during development of drug resistance in vitro. Decreased methotrexate (MTX) trans- portwas observed in manycell lines prior to development ofother mechanisms of resistance, including increased dihydrofolate reductase and decreased polyglutamylation. Marked differences in MTXuptakewereobserved incell lines derived from histologic types of lung cancer, prior to treatment. Cis-platinum (CDDP) resistance was also associated with decreased CDDP uptake. MTX resistance otten appeared along with CDDP resistance although the cells were not previously exposed to MTX. These drug resistant cells showed no changes in GP170 but had reduced 46 kD membrane protein (SQMl). Incubation of cells with anti-SQMl antibody interfered with the early kinetics of MTX transport. The MTX transport-deficient cell lines showed no changes in amounts, substrate affinity or molecular weight of folate-binding protein (FBP) and no changes in amounts or folate binding capacity of dihydrofolate reductase (DHFR). However SQMl forms a complex with FBP in solution. FBP and DHFR are also detected in association with SQMl in reconsti- tuted membrane vesicles. The cDNA structure of SQMl shows a domain with a heptadic repeat of positively charged amino acids,similarto thefolate-bindingregionofDHFRThesere.sults suggest that the interaction of SQMl with FBP may be important in modulating MTX transport and that DHFR may play a role in MTX transfer from the membrane to the cytoplasm.

EPITHELIAL ANTIGEN EXPRESSION AND DRUG- SENSITMTY IN LUNG CANCER. Ma. Teresa Alhambra-Banaga, Teresita Tuazon, Diosespaz

Soriano, k Godofredo Urbano, Mika Kakefuda, Yuk Chor Wong and Samuel Bernal. Lung Center of the Philippines and Section of Medical Oncology, University Hospital, Boston, MA 02118.

The histologic types of lung cancer differ in drug respon- siveness in viva. Fresh cells and cell lines from lung cancer types also show different drug responses. To assess drug sensitivity in vitro, we developed a dye assay, Rhodamine- @h-123), used in conjunction with fluorescent anti-tumor antibodies, which allowsselective assays of tumor cells in mixed cell populations. We confirmed that fresh cells of small cell carcinoma of the lune (SCCL) are highly drug-responsive, followed by well differen- tiated squamous carcinoma (WD-SqCL). Poorly differentiated squamous carcinoma (PD-SqCL) and adenocarcinomas (AdCL) showedintermediatesensitivity. Largecellcarcinomaofthelung (LCCL) was least responsive. Cell lines also followed approxi- mately the same order of drug sensitivity. We then determined whether the expression of epithelial differentiation antigens correlated with drug sensitivity. No correlation was found between drug sensitijity and keratin or involucrin expression. However. a new euithelial differentiation marker. SOMl. was associated with sdnsitivlty to cis-platinum (CDDPj a3 m&ho- trexate (MTX). Isolates with highest numbers of SQMlereactive cells were most sensitive to MTX and CDDP and showed highest levels of drug transport. SCCL cell lines resistant to MTX or CDDP showed reduced SQMl. There was no significant corre- lation between SQMl and resistance to anthracyclines or aIka- loids. Other membrane markers, including N-CAM, sialylated glycoproteins and fucosylated glycolipids, were expressed more in SCCL but did not change with development of CDDP or MTX resistance in SCCL cell lines. The relationship between lung cancer differentiation and drug resistance is complex and may involve multiple mechanisms. -However, decreased drug tra&- oort is an imnortant mechanism of resistance to MTX and %DP. Decre&d drug transport was associated with reduced SQMl, but was not correlated with other known epithelial markers, indicating that drug resistance in lungcancercells was not related to differentiation per se but to specific membrane alterations that may play a role in drug transport.

CHARACTERIZATION OP A GENE ASSOCIATED WITH PROLIFERATION-ARRBSTBD RAT ALVEOLAR TYPE 2 CELLS.

Kevin Mihal. Martin Joyce-Brady, and Norbert Riedel. moon-enter. Department of Medicine, and Department of Biochemistry, Boston University School of Medicine, 80 E. Concord Street, Boston, MA 02118. USA.

The alveolar epithelial type 2 (T2) cell plays an essential proliferative role during fetal and neonatal lung development. Although T2 cell turnover is minimal in the adult lung. it can be induced in response to lung injury. The mechanisms regulating the T2 cell’s proliferative capacity are unclear. To investigate whether adult T2 cells express genes specifically associated with their growth-arrested state, a lambda gtl I cDNA library was constructed from primary cultures of nonproliferating adult T2 cells and was differentially screened with cDNA probes from proliferating neonatal and nonproliferating adult T2 cells. Four genes were isolated, one of which, designated 8S, was found to be identical to rat Clara cell secretory IO kDa protein

Clone 8S is expressed in adult lun a

and T2 cells, but i? not detectable in alveolar macrop ages. an enriched population of adult Tl cells, primary kidney and .ntestinal epithelial cells, and a variety of other rat organs. Consistem with its potential role in T2 cell proliferation-arrest. 8S is highly expressed in nonproliferating adult T2 cells and serum-deprived, nonproliferating neonatal T2 cells, weakly in proliferating neonatal T2 cells. and not at all in SV40T- and El A-immortalized T2 cells. Furthermore, 8S is developmentally regulated. By Northern Ihlot analysis of whole fetal lung,. we detect 8S from IX dav gestation throueh birth. The gerie is then down-regulatei &ring neonaLl lung develooment and re-exoressed m the adult lung. The ahilitv of 8S to ‘induce growth-&rest is currently being-studied b&t& using recombinant protein and transfection assays.