Sequential treatment including high-dose chemotherapy with peripheral blood stem cell support in patients with high-risk stage II-III breast cancer: outpatient administration in community cancer centers.

The authors determined outcomes for patients with localized high-risk breast cancer undergoing sequential outpatient treatment with conventional-dose adjuvant therapy, chemotherapy, and growth factor mobilization of peripheral blood stem cells (PBSC) and high-dose chemotherapy (HDC) with PBSC support in community cancer centers. Ninety-six patients with stage II-IIIB noninflammatory breast cancer with 10 or more positive lymph nodes and a median age of 46 years (range, 22-60 years) were treated with: 1) doxorubicin, 5-fluorouracil, and methotrexate (AFM), four courses at 2-week intervals; 2) cyclophosphamide (4 g/m2) and etoposide (600 mg/m2) (CE), followed by filgrastim (6 microg/kg per day) and PBSC harvest; and 3) cyclophosphamide (6 g/m2), thiotepa (500 mg/m2), and carboplatin (800 mg/m2) (CTCb), followed by PBSC infusion. All 96 patients received AFM, 95 (99%) received CE, and 95 (99%) received CTCb with a median hospital stay of 12 days (5-34 days) for all phases of treatment. Sixty-nine patients (72%) are alive, 55 (57%) without relapse at a median follow-up of 53 months (range, 37-77 months). One patient (1%) died of acute myeloid leukemia and all other deaths were associated with recurrent breast cancer. The probabilities of event-free survival (EFS) at 4 years for patients with or without locally advanced disease were 0.37 and 0.69, respectively (p = 0.004), and 0.71 and 0.48 for patients who were estrogen/progesterone receptor (ER/PR) positive or ER/PR negative, respectively (p = 0.016). In multivariate analyses, locally advanced disease (relative risk, 2.3; p = 0.021) and ER/PR-negative hormone receptor status (relative risk, 2.2; p = 0.014) were the only adverse risk factors for EFS identified. Patients with zero, one, or two of these adverse risk factors had 4-year EFS of 0.80, 0.56, and 0.33, respectively. The sequential administration of AFM, CE, and CTCb followed by PBSC in an outpatient community setting was well tolerated in patients with high-risk stage II-III breast cancer. More intensive or more novel treatment strategies will be required to decrease relapses in patients who have ER/PR-negative tumors and/or have locally advanced disease.

High-dose chemotherapy with BUCY or BEAC and unpurged peripheral blood stem cell infusion in patients with low-grade non-Hodgkin's lymphoma.

Forty-nine patients with low-grade non-Hodgkin's lymphoma (NHL) received high-dose chemotherapy (HDC) with busulfan and cyclophosphamide (BUCY) or carmustine, etoposide, cytarabine and CY (BEAC) followed by unpurged autologous peripheral blood stem (PBSC) infusion. All patients had failed initial chemotherapy or progressed after an initial remission. Peripheral blood stem cells were mobilized with CY alone (n = 1), CY, etoposide (n = 19), or CY, etoposide and cisplatin (n = 29) followed by granulocyte colony-stimulating factor. Twenty-two patients received BU, 16 mg/kg, and CY, 120 mg/kg. Twenty-seven patients received carmustine 300 mg/m2, etoposide 600 mg/m2, cytarabine 600 mg/m2, and CY 140 mg/kg. Four patients (8%) died of non-relapse causes, two (9%) in the BUCY group and two (7%) in the BEAC group. Twenty-seven patients (55%) relapsed or progressed at a median of 9.4 months (2-38) from PBSC infusion. Ten patients who relapsed are alive a median of 31 months (range, 6-47) after relapse. The probabilities of relapse at 3.6 years for patients receiving BUCY or BEAC were 0.57 and 0.70, respectively (P = 0.92). Twenty-seven patients (55%) are alive at a median of 3.6 years (range, 1-5). The probabilities of survival at 3.6 years for patients receiving BUCY or BEAC were 0.58 and 0.55, respectively (P = 0.72). The probabilities of EFS at 3.6 years for patients receiving BUCY or BEAC were 0.36 and 0.28, respectively (P = 0.82). It was concluded that BUCY is an active regimen for the treatment of patients with low-grade NHL.

A phase I-II study of high-dose melphalan, mitoxantrone and carboplatin with peripheral blood stem cell support in patients with advanced ovarian or breast carcinoma.

The purpose of this study was to develop a high-dose chemotherapy (HDC) and peripheral blood stem cell (PBSC) regimen for treatment of patients with ovarian carcinoma that could be administered in an outpatient setting. Fourteen patients with advanced ovarian (n = 9) or breast (n = 5) carcinoma, who had failed conventional chemotherapy, were entered into a dose-escalation trial to determine the maximum tolerated dose (MTD) of carboplatin that could be administered with fixed doses of melphalan (160 mg/m2) and mitoxantrone (50 mg/m2). Twenty-five additional patients were included in a phase II trial at the MTD. Two of two patients had grade 4 severe regimen-related toxicities (RRT), one fatal, at a dose level of 1600 mg/m2. Two of 29 patients (6.9%) treated at the MTD (carboplatin, 1400 mg/m2) died of RRT. All three patients who died of toxicity had a calculated AUC for carboplatin >30 mg/ml/min. Thirty-one patients with ovarian cancer who had failed chemotherapy were treated, 24 at the MTD. Fourteen of 20 patients (70%) with ovarian carcinoma with evaluable disease achieved a CR and seven (35%) are alive disease-free a median of 20 months (range, 7-26). Five of seven patients with ovarian cancer who had failed chemotherapy but were rendered clinically disease-free following surgery survive without progression a median of 13 months (range, 9-19). Eight of 16 (50%) platinum-resistant and 4/12 (33%) platinum-sensitive patients with ovarian cancer survive disease-free.

Treatment-related mortality in 1000 consecutive patients receiving high-dose chemotherapy and peripheral blood progenitor cell transplantation in community cancer centers.

High-dose chemotherapy (HDC) with autologous peripheral blood progenitor cell (PBPC) is being increasingly utilized as a therapeutic modality for patients with chemotherapy-sensitive disease. Several published HDC regimens have become relatively widely used. The purpose of this analysis was to determine treatment-related mortality (TRM) following administration of five different HDC regimens in community cancer centers. A retrospective evaluation of 1000 consecutive patients with leukemia, non-Hodgkin's lymphoma, Hodgkin's disease, multiple myeloma, sarcoma, ovarian cancer, or breast cancer who received one of five published HDC regimens followed by PBPC infusion over a 5-year period in community cancer centers was performed to determine TRM. Fifty-nine patients (5.9%) died within 100 days of PBPC infusion. Twenty-five patients (2.5%) died predominantly of causes related to disease progression. Thirty-four patients (3.4%) died of TRM, 15 patients (1.5%) died from infection and 19 (1.9%) died from regimen-related toxicities (RRT). In a logistic model, increasing age (P = 0.001) and lower numbers of CD34+ cells/kg (P = 0.003) were associated with an increased risk of 100-day TRM. High-dose cyclophosphamide, thiotepa, and carboplatin was associated with a lower risk of mortality than other regimens (P = 0.0001). High-dose chemotherapy and autologous PBPC support can be performed in community cancer centers with relative safety. Patient age, the type of preparative regimen and the number of CD34+ cells infused were important determinates of mortality.

Induction, mobilization of peripheral blood stem cells (PBSC), high-dose chemotherapy and PBSC infusion in patients with untreated stage IV breast cancer: outcomes by intent to treat analyses.

We investigated the outcomes of patients with breast cancer undergoing induction chemotherapy, mobilization of peripheral blood stem cells (PBSC) and high-dose chemotherapy (HDC) with PBSC infusion. One hundred and fourteen patients with untreated stage IV breast cancer, with a median age of 46 years (range 24-62), were entered on a phase II trial consisting of; (1) doxorubicin, 5-flurouracil, methotrexate (AFM) x 4 courses at 2 week intervals; (2) cyclophosphamide (4 g/m2), etoposide (600 mg/m2), cisplatin (105 mg/m2) (CEP), filgrastim (6 micrograms/kg/day) and PBSC collection; (3) cyclophosphamide (6 g/m2), thiotepa (500 mg/m2), carboplatin (800 mg/m2), (CTCb) followed by PBSC infusion. All patients received AFM, 107 (94%) received CEP, 93 (82%) received CTCb and PBSC as per protocol and 99 (87%) ultimately received HDC and PBSC. There was one infectious death after AFM and all other deaths were associated with progressive disease. Fifty-two patients (46%) are alive, 21 (18%) without progression, at a median 31 months (range 22-47). The probabilities of survival and progression-free survival at 3.5 years were 0.40 and 0.17, respectively. All 62 patients with visceral disease and/or a prior history of doxorubicin adjuvant therapy have relapsed or progressed. We conclude that the sequential administration of AFM, CEP and CTCb followed by PBSC resulted in long-term PFS only in patients who were NED, had bone-only disease or had lymph node or soft tissue disease with or without bone disease. Other strategies, aimed at improving responses to initial therapy, improving HDC regimens and/or developing immunomodulatory therapies, will be necessary to improve PFS for patients who fail doxorubicin adjuvant or who have visceral disease.

High-dose chemotherapy and peripheral blood stem cell infusion in patients with non-Hodgkin's lymphoma: results of outpatient treatment in community cancer centers.

The outcomes for patients with non-Hodgkin's lymphoma (NHL) treated with high-dose chemotherapy (HDC) and peripheral blood stem cell (PBSC) infusion by practicing oncologists in community cancer centers in the United States were determined. Eighty-three patients with NHL, who had failed conventional chemotherapy, underwent mobilization of PBSC with chemotherapy and a recombinant growth factor in an outpatient facility. At a median of 40 days (range 26-119) after mobilization chemotherapy all received carmustine (300 mg/m2 x 1), etoposide (150 mg/m2 twice a day x 4 days), cytarabine (100 mg/m2 twice a day x 4 days) and cyclophosphamide (35 mg/kg x 4 days) (BEAC) followed by infusion of unmanipulated PBSC in an outpatient facility. The probabilities of treatment-related mortality, relapse/progression, overall survival (OS) and event-free survival (EFS) at 3 years for all 83 patients were 0.07, 0.57, 0.49 and 0.38, respectively. The probabilities of relapse/progression, OS and EFS at 3 years for 28 patients who had failed primary induction chemotherapy were 0.55, 0.42 and 0.38, respectively. The probabilities of OS and EFS for 27 patients in untreated first relapse were 0.52 and 0.44, respectively, as compared to 0.56 and 0.32, respectively, for 18 patients who had reinduction attempts prior to receiving mobilization chemotherapy (P = 0.81 for OS and 0.99 for EFS). No significant risk factors for the outcomes of TRM, relapse/progression, OS or EFS could be identified. These data demonstrate that approximately 40% of patients with NHL who have failed conventional chemotherapy become long-term disease-free survivors after mobilization chemotherapy, high-dose BEAC and PBSC infusion administered in an outpatient setting in community cancer centers, with the major cause of failure being relapse. Results obtained in this study are comparable to published data in similar patient populations receiving therapy as inpatients, suggesting that clinical trials involving well-tested HDC regimens can be carried out safely in this setting.

Biological activity, binding site and affinity of monoclonal antibodies to the fusion protein of respiratory syncytial virus.

The neutralizing activity and fusion-inhibition activity per unit weight of immunoglobulin were determined for each of a panel of 20 monoclonal antibodies (MAbs) to the fusion (F) protein of respiratory syncytial (RS) virus. Neutralization did not correlate with fusion-inhibiting activity, suggesting that the F protein plays at least two independent, antibody-sensitive roles in viral infection. Antibodies with the highest biological activity against A2, a subgroup A strain of RS virus, neutralized a subgroup B strain (8/60) poorly, suggesting a degree of antigenic variation that may be important in human infection. All but one fusion-inhibiting MAb bound to protein blots and binding was mapped to two areas on overlapping F protein fragments. One MAb with relatively poor fusion-inhibiting activity bound only to fragments C-terminal of amino acid 384, the remainder bound only to fragments containing residues 253 to 289. MAbs directed to the latter site were heterogeneous in neutralizing activity, subgroup specificity and fusion-inhibiting activity. These variations between MAbs could not be accounted for by differences in their binding avidities. We suggest that this binding site is not the complete antibody epitope which probably includes conformation-dependent elements.

Binding of neutralizing monoclonal antibodies to regions of the fusion protein of respiratory syncytial virus expressed in Escherichia coli.

cDNA containing the entire coding sequence of the respiratory syncytial (RS) virus fusion (F) protein gene (574 amino acids) and two large PstI restriction fragments, encoding amino acids 18 to 212 and 214 to 574, were expressed in Escherichia coli as C-terminal chimeras with beta-galactosidase (beta-gal) in the pEX expression vector system. A further cDNA fragment, overlapping the PstI restriction site and encoding amino acids 190 to 289, was derived by PCR and expressed in a similar manner. Polyclonal rabbit serum raised against RS virus bound to all four chimeric proteins but most strongly to those containing C-terminal sequences. Two monoclonal antibodies (MAbs), 1E3 and RS348, capable of neutralizing the virus and inhibiting the viral fusion function, bound to all chimeras except that derived from the N-terminal PstI fragment, suggesting that their binding sites were located between amino acids 214 and 289. Further analysis of binding to expressed fragments from restriction enzyme digests and PCR amplification demonstrated that both antibodies bound to amino acids 253 to 289. MAb RS348 bound to 12-mer overlapping synthetic peptides containing the sequence 265 to 272 (PITNDQKK) but MAb 1E3 failed to bind to any 12-mer peptide derived from the F protein sequence. Immunization of mice with chimeric proteins containing the whole F protein coding sequence or amino acids 253 to 384, which includes the binding site of the two MAbs identified here, failed to induce antibodies that recognized the native RS virus F protein or could neutralize the virus. This suggests that either the beta-gal partner inhibits the immune response to the protein or that elements missing from the protein expressed in E. coli, perhaps conformational or added post-translation, contribute to the neutralizing antibody epitope.

Inpatient continuous-infusion interleukin-2 in 788 patients with cancer. The National Biotherapy Study Group experience.

BACKGROUND: Interleukin-2 (IL-2), used alone or in combination with adoptive cellular therapy, is one of the most promising biologic therapeutic agents for cancer treatment. METHODS: The National Biotherapy Study Group conducted 15 trials of continuous-infusion IL-2 involving 788 patients with cancer, 638 of whom were evaluable for tumor response. The protocols included administration of IL-2 and lymphokine-activated killer (LAK) cells, IL-2 and cyclophosphamide, IL-2 and tumor-infiltrating lymphocytes, IL-2 and alpha-interferon (IFN), IL-2 and tumor necrosis factor, and IL-2 and LAK alternating with combination chemotherapy. RESULTS: Responses were detected in 33 of 188 patients (18%) with melanomas, 13 of 167 (8%) with renal cell carcinomas, and 1 of 76 (1%) with colorectal cancers. The median survival times in patients with melanoma and renal cell cancer were 9.6 and 9.3 months, respectively. The proportion of patients surviving 1 year were 35% and 43%, respectively. There were responses in 8 of 51 patients (16%) with lung cancer, but many of these patients received IL-2 and LAK alternating with platinum-based chemotherapy. Four of 23 patients (17%) responded who had breast cancer and received IL-2 and IFN. The protocols involving IL-2 plus adoptive cellular therapy produced a higher response rate than those not involving activated cells (48 of 312 [15%] versus 24 of 326 [7%], P = 0.003); however, there was no difference in survival. There was a 1.8% mortality rate attributed to the complications of IL-2 itself. CONCLUSIONS: IL-2 produces durable tumor responses in some patients, especially in those with melanoma and renal cell carcinoma. Because the drug's toxicity is significant and the overall response rates are low, patient selection may be the most important factor in the clinical use of continuous-infusion IL-2 therapy.

Continuous interleukin-2 and lymphokine-activated killer cells for advanced cancer: a National Biotherapy Study Group trial.

We conducted a multicenter, phase II trial of continuous-infusion recombinant interleukin-2 (rIL-2) and lymphokine-activated killer (LAK) cells. Patients had advanced cancer, measurable disease, and a good performance level. Treatment included a 5-day continuous infusion of 18 x 10(6) IU/m2/d of rIL-2 followed by 1 day of rest, 4 days of leukapheresis to collect cells for in vitro augmentation of cellular cytotoxicity, and 5 more days of rIL-2 infusion with reinfusion of LAK cells for 3 successive days. Therapy was repeated after 2 weeks. There were 117 patients enrolled: 63% were males, with a median age of 51 years. Eighty-two percent were managed in oncology units, and 18% were in intensive care units. Six patients died within 1 month of initiating therapy. In renal cell carcinoma, the response rate was one of 31 patients (3%), with a median survival of 10.7 months. In melanoma, the response rate was four of 33 patients (12%), with a median survival of 6.1 months. For all other histologies, response rate was three of 53 patients (5%), with a median survival of 7.4 months. All responders were asymptomatic when therapy was initiated. This trial confirms the feasibility of administering continuous rIL-2 and LAK cells outside the intensive care unit environment. Antitumor activity in melanoma was similar to that seen in multicenter trials of bolus rIL-2 and LAK cells. Activity in renal cell cancer was disappointing.

Recombinant interleukin-2 and adoptive immunotherapy alternated with dacarbazine therapy in melanoma: a National Biotherapy Study Group trial.

We evaluated adoptive cellular therapy with recombinant interleukin-2 (rIL-2) plus lymphokine-activated killer (LAK) cells alternating with sequential dacarbazine chemotherapy in 27 patients with metastatic melanoma. rIL-2 was given to the patients as a 5-day continuous-infusion priming cycle followed by 1 day of rest, 4 days of leukapheresis for in vitro LAK cell expansion, and then 4 1/2 days of continuous rIL-2 infusion in conjunction with reinfusion of LAK cells during the first 3 days of the continuous infusion. Two weeks later, patients received dacarbazine (1,200 mg/m2) chemotherapy. Two patients achieved complete remission, and five achieved a partial remission for a response rate of 26% (95% confidence interval = 12%-47%). Three patients had mixed responses. The partial and mixed responses were brief, ranging from 1 month to 6 months, whereas the two complete responses have been sustained for 13+ and 24+ months. There were no additive toxic effects except for thrombocytopenia, which delayed treatment in two patients. Alternating adoptive immunotherapy and dacarbazine chemotherapy appear to be reasonably tolerated by patients, but the response rate is not clearly better than that achieved with other rIL-2 regimens or with chemotherapy alone.

Biodistribution and plasma survival in mice of anti-melanoma monoclonal antibody cross-linked to OKT3.

Anti-melanoma monoclonal antibody XMMME-001 was cross-linked to anti-CD3 monoclonal antibody OKT3 with succinimidyl 3(2-pyridyldithio)propionate (SPDP), and 2-iminothiolane. The dimer heteroconjugate was purified by HPLC gel filtration, labeled with 131I, and 10 micrograms was injected into each of 24 BALB/c mice. The dimeric heteroconjugate's blood survival in sequentially bled mice (n = 3) and its biodistribution in organs of sacrificed mice (n = 21) were studied. In plasma, the heteroconjugate showed an alpha phase with a half-life of 4 h, and a beta phase with a half-life of about 18 h. Electrophoretic analysis of labeled heteroconjugate in plasma showed that the half-life of disulfide bonds linking the monoclonal antibodies was approximately 7-8 h. Radioactive heteroconjugate accumulated primarily in the liver; significant uptake was also seen in white blood cells and spleen. Very little radioactivity accumulated in kidney, lung, or colon. As a comparison, SPDP-derivatized anti-melanoma antibody was studied by the same methods. It showed an average alpha-phase half-life of 12.5 h; its maximum accumulation in liver or white blood cells was less than 30% of that of the heteroconjugate. Very low levels accumulated in other tissues. The results imply that the shorter half-life in plasma of the heteroconjugate is primarily determined by clearance due to its larger size and conformation, not the lability of cross-linking disulfide bonds.

Activation and expansion of tumor-derived activated cells for therapeutic use.

We obtained tumor specimens from patients with cancer in an effort to activate and expand the tumor-infiltrating lymphocytes for therapeutic use. With the use of finely minced tumor preparations from eight different tumor types, recombinant interleukin-2, and lymphokine-activated killer cell-conditioned medium, lymphocytes were expanded in vitro. After 4 weeks, the tumor cells were virtually absent from the cultures. At this point, the lymphocytes were termed "tumor-derived activated cells" (TDACs). Over 90% of the TDACs from each of the different tumor types were T lymphocytes, and the percentage of cells expressing either CD4 or CD8 varied considerably from population to population. The lymphocytes showed specific cytolytic activity in melanoma and colon and renal cell carcinomas. Continued expansion and long-term growth of the TDACs, as well as maintenance of the cytolytic activity, were achieved by periodic stimulation of the TDACs with irradiated autologous tumor cells. In a clinical study of 28 patients with cancer, we generated a mean number of 1.2 X 10(11) TDACs in an average time in culture of 69 days. These TDACs were subsequently infused into the patients with cancer. TDACs appear to represent an important resource for biotherapy of patients with cancer.

Tumor-derived activated cells: preliminary laboratory and clinical results.

It is well known that T lymphocytes can mediate significant anti-tumor responses. A limiting factor has always been the ability to expand T cells, whether from the peripheral blood, spleen, or tumor. The recent availability of recombinant interleukin-2 (r-IL2) has demonstrated the feasibility of expanding T cells and the clinical efficacy of these cells as anti-tumor effectors in murine models. Concomitantly, researchers discovered that lymphokine-activated killer cells--peripheral blood cells functionally distinct from T cells--could be cultured, expanded, and re-infused in patients, with significant clinical effects. For many years, the infiltrating lymphocytes have been recognized in tumor biopsies and known to be cytolytically active. Major limiting factors were the ability to culture large numbers of these infiltrating cells and the limited understanding of the tumor antigens involved for T-cell stimulation. Restimulation by antigen (tumor cells) appears to provide the ongoing antigen stimulation needed to maintain selective killing of tumor cells. By defining various factors in the medium that support and enhance T-cell growth and activation, the components are becoming available to develop a broad attack on advanced cancer by using this laboratory-based technology of stimulation and expansion of tumor-derived activated cells.

IL-2: a review of current knowledge.

A limiting factor in the use of T-cell therapy for cancer has always been the ability to expand T-cells, whether derived from the peripheral blood, spleen or tumor. The availability of r-IL-2 has confirmed the feasibility of expanding T-cells and LAK cells for clinical trials after confirmation of their anti-tumor activity in murine models. While the most promising results using IL-2/LAK cells have been in patients with melanomas and renal cancer, anti-tumor effects have been seen in patients with a wide variety of cancers, even those with bulky tumors. This form of adoptive cellular biotherapy has confirmed that an expanded and activated cell population using the cancer research laboratory can provide a method by which clinicians can effectively treat advanced cancer. In tumor biopsies, the infiltrating lymphocytes have been recognised and are known to be cytolytically active for many years. Using IL-2 stimulation of growth and an ongoing antigen stimulation (tumor cells), we have maintained selective killing of tumor cells. Accessing cells and various factors in the medium which support and enhance T-cell growth and activation are being defined. The role of antigen stimulation is also basic to further progress with this technology. Tumor cell chunks and cultures, nude mouse xenografts, or purified antigen all represent potential sources of repeated antigen stimulation. Thus, the components are now available to develop a broad attack on advanced cancer using this laboratory-based technology of tumor-derived activated cell (TDAC) stimulation, expansion and therapy. These approaches and preliminary results point to the dramatic change in technology which has allowed the cancer research laboratory to be a substantial component in new clinical approaches to cancer treatment. Laboratory scientists have become a major component in the design and conduct of clinical trials using adoptive biotherapy. These techniques are laboratory based and it is only with close and effective communication between the laboratory scientists and the clinician that rapid and effective translation of these technologies to the patient will occur (20).