Biomedical subjects

R S Freedman

Publications and source records attributed to R S Freedman.

At least 55 records · Page 3Linked to original sources

PubMed1996

Immunotherapy for peritoneal ovarian carcinoma metastasis using ex vivo expanded tumor infiltrating lymphocytes.

Surgery and chemotherapy have contributed to a modest overall survival in patients with epithelial ovarian carcinoma. It is therefore important to pursue novel therapy strategies for this disease that are different from conventional chemotherapy. Tumor infiltrating lymphocytes (TILs) from patients with ovarian carcinoma may represent an active immune response of the host directed against the tumor cells. These TILs can be expanded in vitro in low concentrations of recombinant interleukin-2 (rIL-2) by a few thousandfold. The resulting T-cell lines comprise CD3+CD4+TCR alpha beta + or CD3+CD8+TCR alpha beta + cells, or mixtures of both. These T-cell lines may exhibit either tumor-specific cytotoxicity against autologous tumor cells, or produce cytokines (interferon-gamma, tumor necrosis factor, and granulocyte stimulating factor) either in antigen-dependent (tumor-specific) or an antigen-independent manner. T-cell lines exhibiting primarily autologous tumor-specific cytotoxicity were developed from approximately 50% of the patients. Blocking experiments using appropriate monoclonal antibodies revealed that the CD3/TCR complex on the effector cells and the MHC class I antigens on the tumor cells were involved in the cytolytic process. We have developed a four-step method for the expansion of TILs to large numbers (1 x 10(10) to 1 x 10(11)) sufficient for clinical trials in patients with ovarian cancer. We have conducted a pilot clinical trial to examine the feasibility and clinical effects of intraperitoneal TILs and low-dose rIL-2 in patients with advanced ovarian carcinoma who were refractory to platinum-based chemotherapy. More recently, procedures have been developed for obtaining large numbers of purified CD8+ rIL-2-expanded TILs for the treatment of patients with ovarian carcinoma. The evolution of clinical trials and correlative studies necessary to develop an effective adoptive immunotherapy approach were discussed.

Adoptive Transfer↗

PubMed1996

Effects of interleukin-1 alpha on ovarian carcinoma in patients with recurrent disease.

The aim of this study was to evaluate the safety and the biological effects of interleukin (IL-1 alpha) in patients' with recurrent ovarian carcinoma treated with carboplatin. In this phase I study, IL-1 alpha was administered by a continuous intravenous infusion at doses ranging 0.1-10 micrograms/m2 every 24 h for 4 days (96 h) 3 weeks before the first dose of carboplatin (400 mg/m2) in patients with potentially platinum-sensitive ovarian cancer. The maximum tolerated dose was 3 microgram/m2/day. Dose-limiting effects at 10 micrograms/m2/day were fever, chills, hypotension and fluid retention. Minor but objective antitumour effects were observed in 2 of 18 patients. 4 patients (including 1 with a minor response) had a decrease of the CA-125 serum level ranging from 33 to 39%. The trial design precluded evaluation of the duration of response to single-agent IL-1 alpha. Based on this trial design, there is evidence of minor antitumour effect to a single course of IL-1 alpha dose given prior to chemotherapy.

Adult↗

PubMed1996

Proposal for translational analysis and development of clinical radiolabeled immunoglobulin therapy.

BACKGROUND AND PURPOSE: Radiolabeled immunoglobulin therapy (RIT) can be a selective, effective, low-toxicity outpatient cancer therapy. A consensus on the best approach for the preclinical and clinical development of RIT reagents needs to be developed. We report the M.D. Anderson Cancer Center prior experience in translating RIT from laboratory to clinic for the treatment of Hodgkin's disease and propose a flow diagram for the development of RIT for other malignancies. MATERIAL AND METHODS: Three different animal models are described: nude mice bearing human tumor xenografts, normal beagle dogs, and normal rhesus monkeys. We produced and purified antibodies and prepared chelate-immunoconjugates reactive with six different human tumor-associated antigens. The Igs used were derived from rabbits, mice, and humans (human-derived RIT reagents being less immunogenic in human patients). Eighty patients with refractory Hodgkin's disease were treated with radiolabeled antiferritin. RESULTS: We recommend a two-injection scheme using, (1) an indium-111-labeled radioimmunoconjugate for diagnosis, pharmacokinetic studies, and dosimetry, and (2) a yttrium-90-labeled radioimmunoconjugate for therapy. The animal models provide useful data on tumor targeting, radiotoxicology, and undesirable biodistributions. A 70% response rate is obtained in patients with advanced recurrent Hodgkin's disease. More extensive preclinical testing allows for safer and more effective clinical RIT studies. CONCLUSIONS: We recommend, (1) preclinical optimization of chelation chemistry, Ig size, Ig origin, route of administration, and fractionation, (2) new clinical Phase I-III studies more appropriate for RIT development than the classical Phase I-III studies used for the development of chemotherapeutic agents, and (3) more extensive preclinical testing of RIT reagents.

Animals↗

PubMed1996

Preliminary report of the activity of docetaxel in advanced or recurrent squamous cell cancer of the cervix.

Eighteen patients with squamous cell cancer of the cervix were treated with i.v. docetaxel 100 mg/m2 over 1 h every 21 days. No patient received prior chemotherapy, except as a radiation sensitizer. Median age was 42 years (range 30-58) and Zubrod performance status was 1 (0-2). Ten (59%) patients had prior surgery and 11 (65%) had prior radiation therapy. Sixteen patients were evaluable for response. Two patients had a partial response (13%; 95% CI 0-32%) and eight (50%; 95% CI 23-77%) had stable disease. Dose reduction to 75 mg/m2 was required in 10 patients and to 55 mg/m2 in one patient. Granulocytopenia was the major hematopoietic toxicity (31% grade 3 and 44% grade 4). Docetaxel is active in patients with squamous cell cancer of the cervix and may be tolerable at this dose schedule.

Adult↗

PubMed1996

Prolonged stabilization of progressive squamous cell cancer of the cervix with interferon-alpha and 13-cis-retinoic acid: report of two cases and review of the literature.

Recurrent and metastatic cervical carcinoma has very poor prognosis, mainly because there is no effective systemic therapy which would increase the duration of survival. Biologic agents have recently been found to have activity in cervical carcinoma. The combination of interferon (IFN)-alpha and 13-cis-retinoic acid had additive and synergistic antitumor activity. Both have antiviral, immunoregulatory and antiangiogenic properties, and are known to modulate malignant cell differentiation and proliferation. We report two patients with recurrent squamous cell carcinoma (SCC) of the cervix who had small-volume progressive metastatic disease, and were treated with a combination of IFN-alpha and 13-cis-retinoic acid. The first patient had pelvic lymph node metastases and the other had lung metastases. The previously progressive diseases remained stable for a prolonged period of time, 3 and 4 years, with a good quality of life. These cases suggest the possibility of using IFN-alpha and 13-cis-retinoic acid as a treatment for small-volume residual disease or as postinduction therapy in patients at high risk for disease recurrence.

Adult↗

PubMed1996

Immunopharmacology and cytokine production of a low-dose schedule of intraperitoneally administered human recombinant interleukin-2 in patients with advanced epithelial ovarian carcinoma.

We determined in the peritoneal cavity (p.c.) of epithelial ovarian carcinoma patients during a 4-day treatment cycle of low-dose recombinant human interleukin-2 (rIL-2): (a) pharmacokinetics of IL-2, (b) endogenous cytokine production, and (c) numbers and percentages of peritoneal exudate lymphocytes. We administered 6 x 10(5) IU/m2 of rIL-2 (0.03 mg/m2 Proleukin rIL-2) intraperitoneally (i.p.) over 30 min on each of 4 days. One and one-half liters of D5 0.25 NS was injected i.p. before each rIL-2 infusion. Multiple peritoneal fluid samples were obtained from each of four patients on day 1 and day 4 for detection of IL-2, endogenous cytokines, and soluble IL-2 receptor (IL-2R-alpha). IL-2 concentrations in the peritoneal fluid were determined by bioassay and interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, IL-10, transforming growth factor (TGF)-beta 2, and sIL-2R-alpha by enzyme-linked immunosorbent assay. Numbers of cells per microliter and lymphocyte subpopulation percentages after staining with a panel of monoclonal antibodies were determined on day 1, day 4, and subsequent off-treatment days. IL-2 disappearance in the p.c. was well described by a pharmacokinetic model having constant-rate infusion and biexponential disposition. About 90% of the IL-2 disappearance occurred during the beta-phase, during which IL-2 concentrations were sustained at approximately 10-30 ng/ml (day 1 and day 4) and the median t1/2 beta was 21.5 and 9.2 h on days 1 and 4, respectively. In four of four patients, p.c. production of IL-10 was observed on day 1 and day 4 (maximum 387 pg/ml). Maximum levels of IFN-gamma and sIL-2R-alpha were observed on day 4. (IFN-gamma 217 pg/ml; sIL2-R-alpha: 3486 U/ml). No increases in TNF-alpha or TGF-beta 2 were observed. Large increases in p.c. CD3+, CD4+, CD8+, CD16+, and CD56+ cells were observed. We conclude that biologically active levels of IL-2 are generated in p.c. fluids after i.p. administration of rIL-2 at 0.03 mg/m2.

Adult↗

PubMed1996

Phase II study of intravenous topotecan as a 5-day infusion for refractory epithelial ovarian carcinoma.

PURPOSE: To determine the efficacy and toxicity of topotecan administered as a 5-day intravenous infusion in patients with advanced ovarian cancer refractory to cisplatin-based chemotherapy. PATIENTS AND METHODS: Thirty patients with advanced epithelial ovarian cancer refractory to cisplatin-based chemotherapy received intravenous infusions of topotecan 1.5 mg/m2 delivered over 30 minutes each day for 5 days. A course was repeated every 21 days. The patient eligibility requirements included age > or = 18 years, Zubrod score < or = 2, measurable disease, adequate hepatic and renal function, neutrophil count > or = 1,500/microL, platelet count > or = 100,000/microL, and anticipated survival > or = 3 months. RESULTS: Twenty eight patients were assessable for response and toxicity. All patients were assessable for survival. The major toxicity from administration of topotecan at this dose schedule was myelosuppression; 21 patients required dose reductions. Four patients had neutropenic fever that required hospitalization, and seven patients required platelet transfusions. Maculopapular pruritic exanthema occurred in 20% of patients; gastrointestinal side effects were mild. No deaths were reported on the study. At dose levels of 1.5, 1.25, and 1.0 mg/m2, 61%, 31%, and 25% of patients, respectively, required dose reductions. Of 28 assessable patients, four (14%; 95% confidence interval [CI], 4% to 34%) achieved a partial response (PR) at a median of 1.4 months and lasting 8.9 months, and 17 had stable disease (SD). The overall median survival time was 10.0 months (95% CI, 8.1 to 13.5). CONCLUSION: Topotecan shows modest clinical activity against cisplatin-refractory ovarian cancer, although the dose-intensity is compromised by the depth of the granulocyte nadir and the duration of granulocytopenia. Further studies of topotecan may necessitate a reevaluation of optimal dose schedule, with the possible incorporation of multilineage cytokines, and its activity in taxane-resistant tumors.

Adult↗

PubMed1996

Paclitaxel in patients with platinum-resistant ovarian cancer: a selected review of literature and clinical experience.

Single-agent paclitaxel infused over 24 hours produces response rates of 10 per cent to 20 per cent and 48 per cent respectively, for doses of 135 mg/m2 and 250 mg/m2. This suggests a dose response relationship. However, only a randomized trial comparing the 135 mg/m2 and 250 mg/m2 doses can confirm this result. Unfortunately, the median survival is comparable despite the difference in response rates. This may be secondary to a low CR noted at all doses. The apparent lack of benefit in terms of increased survival for the high dose group, with its attendant increase in incidence of toxic effects and cost (owing to both the paclitaxel and the G-CSF), suggest that the role of higher doses remains to be proven. It may be most useful in alleviating the severe cancer induced symptoms of some patients with advanced platinum resistant ovarian cancer(19). Despite the extensive international experience with paclitaxel in the ovarian cancer clinic its role still needs to be better defined both for salvage therapy and in combination with platinum as a front-line treatment.

Adult↗

PubMed1996

Paclitaxel retreatment in patients with platinum and paclitaxel resistant ovarian cancer.

The optimal role of paclitaxel in the treatment of epithelial ovarian cancer has not been determined. Dose intense paclitaxel therapy is safe and often recommended as second-line treatment for platinum resistant ovarian cancer. Retreatment with a higher dose-intense schedule of paclitaxel is feasible and patients who had a prior dose of 135 mg/m2 or paclitaxel free interval of at least six months may respond. Sensitivity to initial doses and a prolonged paclitaxel-free interval are predictors of a successful reinduction. We report 10 patients who had paclitaxel as second-line treatment and were later retreated with paclitaxel. Three of these patients had a partial response (30%; 95% CI 6-66%). All but one patient had platinum resistant disease. Initial dose and the best response to initial paclitaxel were assessed in relation to reinduction response. Patients with at least a six-month paclitaxel-free interval and previously low dose paclitaxel (135 mg/m2), or a complete response with high dose paclitaxel (250 mg/m2) may respond to high dose paclitaxel retreatment. Prospective trials of paclitaxel retreatment are warranted.

Adult↗

PubMed1996

Gestational trophoblastic disease.

GTD occurs in fewer than 1 in 1200 pregnancies in the United States, but it is much more common in Asia and Latin America, where its incidence may be as high as 1 in 200 pregnancies. Risk factors for GTD include advanced or young maternal age, low socioeconomic status, and prior hydatidiform mole. Early diagnosis and prompt treatment are key to a favorable outcome, and thus recognition of the signs and symptoms of the disease is important for all physicians. Because these diseases have low incidences and occur after reproductive events, screening for them in the general population is not worthwhile. No chemopreventive agents have yet been studied in women at risk for GTD, but the oral contraceptive is a good candidate.

Biomarkers, Tumor↗

PubMed1996

Intraperitoneal indium-111- and yttrium-90-labeled human IgM (AC6C3-2B12) in nude mice bearing peritoneal carcinomatosis.

UNLABELLED: Radiolabeled monoclonal antibodies are utilized increasingly for the diagnosis and treatment of human cancer. Tumor targeting of radiolabeled human monoclonal IgM improves with compartmental administration and might be useful for the diagnosis or treatment of peritoneal carcinomatosis. METHODS: A human monoclonal antibody IgM lambda (AC6C3-B12) reactive with human adenocarcinomas was conjugated to isothiocyanato-2-benzyl-3-methyl-diethylenetriamine-penta-a cetic acid and labeled with either 111In or 90Y. Nude mice bearing intra-abdominal lumps of a human colorectal carcinoma cell line (SW620) were used as a model for peritoneal carcinomatosis. A human monoclonal antibody IgM lambda (CR4E8) reactive with human squamous-cell carcinoma was used as a control. RESULTS: Indium-111-IgM and 90Y-IgM immunoconjugates were compared in nude mice at 2, 24, 72, 120 and 144 hr after intraperitoneal administration. Both showed high specific tumor uptake. The tumor-effective half-lives of the immunoconjugates were 39 hr for indium and 46 hr for yttrium. Tumor-to-normal organ ratios were high and similar for both reagents. Only the femur uptake at later time points was relatively higher for the 90Y-IgM than for 111In-IgM. The tumor uptake of specific AC6C3-2B12 was about fourfold higher than the uptake of aspecific CR4E8 at 24 and 120 hr. CONCLUSION: The combination of 111In- and 90Y-labeled AC6C3-2B12 offers a new opportunity to develop safer and more effective methods for diagnosing and treating human patients with peritoneal carcinomatosis.

Animals↗

PubMed1996

Leuprolide acetate for treating refractory or persistent ovarian granulosa cell tumor.

OBJECTIVE: To evaluate leuprolide acetate for treating refractory or persistent ovarian granulosa cell tumor (GCT). STUDY DESIGN: We treated six patients who had recurrent or persistent ovarian GCT with monthly intramuscular injections of a depot formulation of leuprolide acetate, 7.5 mg. Four patients had received prior cisplatin-based chemotherapy. Initially these patients' tumors had responded to the cisplatin, but then the disease had progressed in all cases. One of these four patients also had received prior therapy with tamoxifen that had resulted in three months of stable disease. Two patients had received no prior chemotherapy. RESULTS: Responses could be evaluated in five of the six patients; in the sixth patient, only survival was evaluable. Among the five evaluable patients, two achieved partial responses lasting 3 and 11 months, with disease progression-free intervals of 4 and 12 months. The other three patients (60%; 95% confidence interval [CI], 20-95%), including two whose disease had progressed on chemotherapy, had stable disease with disease progression-free intervals of 3+, 3+ and 13+ months. The objective response rate was 40% (two of five patients; 95% CI, 10-90%). Cessation of disease progression was noted in all five of the evaluable patients (100%; 95% CI, 40-100%). The sixth patient, whose disease could not be evaluated, was treated with leuprolide acetate for 24 months and at this writing had no clinical evidence of disease. The therapy was convenient to administer, and no major side effects were noted. Leuprolide acetate thus appears to have activity in patients with refractory GCT, and it may prolong the disease progression-free interval. Further trials of this relatively nontoxic and convenient therapy are warranted.

Adult↗

PubMed1996

Transcatheter arterial embolization for the control of life-threatening pelvic hemorrhage in a patient with locally advanced cervix carcinoma.

A woman with a refractory locally advanced recurrent cervical cancer was brought to the emergency room because of a life-threatening vaginal hemorrhage from the tumor. She presented with a compromised hemodynamic status that necessitated multiple blood transfusions and large volume of fluid resuscitation. An arteriogram revealed the site of bleeding from the uterine branch of the right internal iliac artery. The branches of anterior and posterior internal iliac artery were embolized with Gelfoam, Ivalon and coils. This emobolization successfully controlled the bleeding and stabilized the hemodynamic status of the patient. The treatment options in patients with massive pelvic hemorrhage are reviewed and discussed.

Angiography↗

PubMed1996

Phase II study of docetaxel in patients with epithelial ovarian carcinoma refractory to platinum.

We analyzed the efficacy and toxicity of docetaxel in patients with ovarian cancer who failed previous chemotherapy with platinum. Fifty-five patients with measurable ovarian cancer were entered in this Phase II study at The University of Texas M. D. Anderson Cancer Center. Treatment consisted of 100 mg/m2 docetaxel given i.v. every 3 weeks. Because of hypersensitivity reactions, premedication with steroids and antihistamine was initiated during the study. Twenty-two (40%) patients responded (there were 3 complete responders and 19 partial responders). Twenty-one (38%) patients had stable disease. The median survival was 10 months. The main toxicity was neutropenia (98% of patients), with 13 episodes of neutropenic fever. Cumulative fluid retention was the main reason for dose modification and required a combination of diuretics and steroids for palliation. Other side effects were alopecia (100%); anemia (87%); dermatitis (67%); gastrointestinal disorders (53%); stomatitis (49%); neurotoxicity (45%); excessive lacrimation (33%); and hypersensitivity reactions (11%), which in one case were life threatening (loss of consciousness, fluid resuscitation). Docetaxel as a single agent proved to be active in heavily pretreated ovarian cancer patients but is associated with significant side effects. Objective toxicity consisted mainly of neutropenia and fluid retention. Neutropenia was dose limiting and required therapy with granulocyte colony-stimulating factor. Fluid retention was improved but not eliminated by diuretics and corticosteroids. Additional studies of docetaxel in ovarian carcinoma are indicated to define the activity in relation to paclitaxel and in platinum combination therapy.

Adult↗

PubMed1996

Activity of paclitaxel in advanced or recurrent squamous cell cancer of the cervix.

Twenty-six patients with squamous cell cancer of the cervix were treated with i.v. paclitaxel, 250 mg/m2 over 3 h every 21 days. They received steroid, H1 and H2 blocker premedications, and granulocyte-colony-stimulating factor (G-CSF) support (5 microgram/kg/day). No prior chemotherapy, except as a radiation sensitizer, was allowed. The median age was 50 (range, 36-81) years, and performance status Zubrod was 1 (range, 0-2). Eight (33%) patients had prior surgery, and 22 (92%) had prior radiation therapy. Twenty-four patients were evaluable for response; 2 were later found to be ineligible. Five patients had partial responses (21%; 95% confidence interval, 6-40%), and 14 (58%; 95% confidence interval, 35-78%) had stable disease. The median duration of response was 10 (range, 3-27+) weeks. The responses were within the radiation port (four responses) and outside of it (one response). The median interval from the start of irradiation to the start of paclitaxel in responding patients was 94 weeks, whereas in patients with stable disease it was 68 weeks, and in patients whose disease progressed it was 46 weeks. Eighty-eight percent of the 105 cycles of paclitaxel were administered at a dose of 250 mg/m2 or higher. Granulocytopenia was brief and noncumulative, with grades 3 and 4 experienced by 5 and 3 patients, respectively. G-CSF was used for a median of 7 (range, 2-14) days/cycle. Anemia was mild, with G3 noted in 3 patients, and thrombocytopenia was not significant. Infections and musculoskeletal pain were mild and infrequent. Sensory (14 patients G1 or G2 and 2 patients G3) and motor (4 patients G1 or G2 and 1 patient G3) neurotoxicity was noted. There was no significant cardiovascular toxicity. Paclitaxel is active in patients with squamous cell cancer of the cervix and is well tolerated at this dose schedule with G-CSF support.

Adult↗

PubMed1995

Preclinical analysis of intraperitoneal administration of 111In-labeled human tumor reactive monoclonal IgM AC6C3-2B12.

An IgM lambda human tumor cell-reactive monoclonal antibody was developed that reacts with cells of ovarian cancer, colorectal cancer, breast cancer, and certain other malignancies. The monoclonal antibody AC6C3-2B12, which was obtained from a recent recloning, was purified from tissue culture supernatants and analyzed by high-performance liquid chromatography and sodium dodecyl sulfate-PAGE. An animal model was developed in which human tumors grew either as solid peritoneal metastases or as s.c. nodules utilizing the human colorectal carcinoma cell line SW620. The biodistribution of 111In-labeled IgM conjugate was studied after i.v. or i.p. administration in nude mice bearing an s.c. xenograft or peritoneal tumor lumps of a human colorectal carcinoma (SW620). IgM administered i.v. cleared rapidly from blood and was deposited mainly in the liver [50% injected dose/g (ID)/g)], pancreas (20% ID/g), and kidney (10% ID/g) at 24 h. Tumor deposition was low (< or = 1.0% ID/g) in the s.c. tumor xenograft. In contrast, high tumor targeting (29% ID/g) was found in peritoneal tumor lumps after i.p. administration of 111In-labeled IgM. The biological half-life of IgM in the tumor was 100 h. Long peritoneal residence time (t 1/2 = 67 h) and low liver uptake (7% ID/g) were observed after i.p. administration. Blood activity was < 1% of the injected activity. Tumor:normal organ ratios were high (range, 2-290) from 2 to 144 h after i.p. administration. Whole body autoradiograms at 24 h after i.p. 111In-labeled IgM administration confirmed the biodistribution results. In normal beagle dogs, 75% of the i.p.-administered 111In-IgM decayed in the peritoneal cavity. The majority of the remaining radioactivity was taken up by mediastinal lymph nodes. Biological half-life in both locations was approximately 137 h. The i.p. administration of intact, specific radiolabeled IgM provides prolonged retention of radioactivity in tumor, low normal tissue uptake, a long peritoneal residence time, and very limited spillover of IgM into the circulation. This approach offers a promising new method for the diagnosis and treatment of certain patients with peritoneal carcinomatosis.

Animals↗

PubMed1995

A trial of lobaplatin (D-19466) in platinum-resistant ovarian cancer.

Long-term survival in epithelial ovarian cancer remains problematic despite multimodality therapy. A fundamental difficulty is the development of tumor resistance to platinum compounds. Analogs have been developed that demonstrate activity in platinum-resistant cell lines both in vitro and in vivo. Lobaplatin (D-19466), a third-generation compound, demonstrates significant activity in carboplatin and cisplatin-resistant cell lines. Lobaplatin was given to 17 assessable patients with platinum-refractory ovarian cancer. The drug was initially administered at a dose of 50 mg/m2 but was later reduced to 40 mg/m2 because of excessive thrombocytopenia. Nine patients required red cell transfusions during therapy. Cycles were repeated every 21-35 days (median cycle length 28 days). No objective responses were observed. Lobaplatin has no activity in platinum-resistant epithelial ovarian cancer.

Adult↗

PubMed1995

Transduction of rIL-2 expanded CD4+ and CD8+ ovarian TIL-derived T cell lines with the G1Na (neor) replication-deficient retroviral vector.

We have expanded ovarian tumor-infiltrating lymphocytes (TIL) in low concentrations of recombinant interleukin-2 (rIL-2) to conduct intraperitoneal adoptive immunotherapy trials in patients with ovarian cancer. We have previously demonstrated that certain T cell lines and clones derived from ovarian TIL exhibit in vitro autologous tumor-specific cytotoxicity and/or cytokine production (interferon-gamma, tumor necrosis factor-alpha) preferentially in response to autologous tumor cells. Studies that utilize a marker gene introduced into the DNA of TIL can provide useful information on specific uptake or localization of TIL at tumor sites and on the survival of TIL in vivo. We have conducted a series of preclinical experiments in which we have successfully transfected TIL with G1Na, which encodes the gene for neomycin phosphotransferase (neoR). NeoR was detected in at least 10% of CD8+ cells (mean = 10.4%) and between 2.5 and 20% of CD4+ TIL (mean = 8.5%). Transduction of ovarian TIL with G1Na caused no substantial changes to the T cell phenotypes or in vitro cytotoxicities against ovarian and hematogenous tumor cell targets, or on the rIL-2 requirements of TIL for growth and proliferation. In addition, the intact G1Na provirus in transduced TIL cells was rescuable by replication-competent retrovirus and was transferred into the genome of NIH-3T3 fibroblasts, which were rendered resistant to G418. An enhanced polymerase chain reaction (PCR) procedure utilizing detection by ethidium bromide staining was developed. The enhanced PCR detected 1 in 100,000 neoR-labeled cells. Furthermore, detection of the G1Na genome in transduced TIL by in situ hybridization with an RNA probe provided evidence for expression of the neoR gene in transduced TIL. Results obtained from these studies suggest that ovarian TIL-derived T cell lines transduced with the neoR gene post infection with the G1Na retroviral vector can be utilized to examine the in vivo trafficking pattern of ovarian TIL-derived T cell lines expanded in low concentrations of rIL-2 and their survival.

3T3 Cells↗