Inhibition of cyclooxygenase-2 indirectly potentiates antitumor effects of photodynamic therapy in mice.

PURPOSE: The aim of the present study was to potentiate the antitumor effectiveness of photodynamic therapy (PDT). A cDNA microarray analysis was used to evaluate the gene expression pattern after Photofrin-mediated PDT to find more effective combination treatment with PDT and inhibitor(s) of the identified gene product(s) overexpressed in tumor cells. EXPERIMENTAL DESIGN: Atlas Mouse Stress Array was used to compare the expression profile of control and PDT-treated C-26 cells. The microarray results have been confirmed using Western blotting. Cytostatic/cytotoxic in vitro assay as well as in vivo tumor models were used to investigate the antitumor effectiveness of PDT in combination with cyclooxygenase (COX) 2 inhibitors. RESULTS: PDT induced the expression of 5 of 140 stress-related genes. One of these genes encodes for COX-2, an enzyme important in the tumor progression. Inhibition of COX-2 in vitro with NS-398, rofecoxib, or nimesulide, or before PDT with nimesulide did not influence the therapeutic efficacy of the treatment. Administration of a selective COX-2 inhibitor after PDT produced potentiated antitumor effects leading to complete responses in the majority of treated animals. CONCLUSIONS: COX-2 inhibitors do not sensitize tumor cells to PDT-mediated killing. However, these drugs can be used to potentiate the antitumor effectiveness of this treatment regimen when administered after tumor illumination.     

Erythropoietin treatment in chemotherapy-induced anemia in previously untreated advanced esophagogastric cancer patients

Background

The impact of erythropoiesis-stimulating agents in chemotherapy-induced anemia has been a constant topic of debate over recent years. We prospectively assessed the efficacy of epoetin beta (Epo-b) in improving hemoglobin (Hb) levels and outcome in patients within an open label, randomized clinical phase II trial with advanced or metastatic gastric/esophagogastric cancer.

Methods

Previously untreated patients were randomized to receive 3-weekly cycles of capecitabine (1000 mg/m² bid) for 14 days plus on day 1 either irinotecan 250 mg/m² or cisplatin 80 mg/m². Epo-b (30000 IU once weekly) was initiated in patients with Hb <11 g/dl and continued until Hb ≥12 g/dl was reached. If after 4 weeks the Hb increase was <0.5 g/dl, Epo-b was increased to 30000 IU, twice weekly.

Results

Of 118 patients enrolled, 32 received Epo-b treatment; of these, 65 % achieved an increase in Hb levels of at least 2 g/dl, with 74 % achieving the target Hb of ≥12 g/dl. Within the study population, patients receiving Epo-b showed better overall survival (median 14.5 vs. 8.0 months, P = 0.056) as well as a significantly improved disease control rate (78 vs. 55 %, P = 0.025). Patients in the irinotecan group profited significantly (P < 0.05) in terms of progression-free survival and overall survival under Epo-b treatment (median 6.5 vs 4.1 months and median 15.4 vs 8.4 months, respectively).

Conclusions

Epo-b was effective in raising Hb levels in patients with advanced esophagogastric cancer. Patients receiving Epo-b had a significantly increased response to chemotherapy and a clear trend to improved survival.     

Erythropoietin restores the anemia-induced reduction in radiosensitivity of experimental human tumors in nude mice

PURPOSE: The effect of recombinant human erythropoietin (rhEPO) on the radiosensitivity of human tumor xenografts growing in anemic and nonanemic nude mice was studied. METHODS AND MATERIALS: Anemia was induced by total body irradiation ([TBI], 2 x 4 Gy) of mice before tumor implantation into the subcutis of the hind leg. The development of anemia was prevented by rhEPO (750 U/kg s.c.) given 3 times weekly starting 2 weeks before TBI. Fourteen days after fractionated TBI (tumor volume of approx. 40 mm(3)), single-dose irradiation of the tumor with varying doses was performed so that in full dose-response relationship for the probability of tumor cure was obtained. RESULTS: Radiation-induced anemia (hemoglobin concentration [cHb] = 9.9 g/dl) led to a reduced radiosensitivity compared to controls [49.4 vs. 40.1 Gy radiation dose to control 50% of the tumors (TCD50)]. Upon rhEPO treatment for anemia prevention (cHb = 13.3 g/dl), the TCD50 was 39.8 Gy, illustrating restored radiosensitivity compared to anemic mice. CONCLUSION: These data provide further experimental evidence for restored radiosensitivity upon prevention of anemia with rhEPO.     

Generation of effective antitumor vaccines using photodynamic therapy

Preclinical studies have shown that photodynamic therapy (PDT) of tumors augments the host antitumor immune response. However, the role of the PDT effect on tumor cells as opposed to the host tissues has not been determined. To test the contribution of the direct effects of PDT on tumor cells to the enhanced antitumor response by the host, we examined the immunogenicity of PDT-generated murine tumor cell lysates in a preclinical vaccine model. We found that the PDT-generated tumor cell lysates were potent vaccines and that PDT-generated vaccines are more effective than other modes of creating whole tumor vaccines, i.e., UV or ionizing irradiation, and unlike other traditional vaccines, PDT vaccines do not require coadministration of an adjuvant to be effective. PDT vaccines are tumor specific and appear to induce a cytotoxic T-cell response. We have demonstrated that although both UV and PDT-generated tumor cell lysates are able to induce phenotypic DC maturation, only PDT-generated lysates are able to activate DCs to express IL-12, which is critical to the development of a cellular immune response. Our results show that PDT effects on tumor cells alone are sufficient to generate an antitumor immune response, indicating that the direct tumor effects of PDT play an important role in enhancing that host antitumor immune response. These studies also suggest that in addition to the role of PDT as a therapeutic modality, PDT-generated vaccines may have clinical potential as an adjuvant therapy.     

Systemic antitumor effect of intratumoral injection of dendritic cells in combination with local photodynamic therapy.

PURPOSE: Photodynamic therapy (PDT), which is used clinically for the palliative treatment of cancer, induces local tumor cell death but has no effect on tumors in untreated sites. The purpose of this study was to determine if local PDT followed by intratumoral injection of na?ve dendritic cells (IT-DC) induces systemic antitumor immunity that can inhibit the growth of untreated as well as PDT + IT-DC-treated tumors. EXPERIMENTAL DESIGN: BALB/c or C57Bl/6 mice were injected s.c. with CT26 colorectal carcinoma cells and B16 melanoma cells, respectively, and following 10 to 12 days of tumor growth, the tumors were treated with PDT alone or PDT followed by IT-DC or IT-PBS. In other studies, tumors were established simultaneously in both lower flanks or in one flank and in the lungs, but only one flank was treated. RESULTS: Whereas neither PDT nor IT-DC alone was effective, PDT + IT-DC eradicated both CT26 and B16 tumors in a significant proportion of animals and prolonged the survival of mice of which the tumors were not cured. The spleens of mice treated with PDT + IT-DC contained tumor-specific cytotoxic and IFN-gamma-secreting T cells whereas the spleens of control groups did not. Moreover, adoptive transfer of splenocytes from successfully treated CT26 tumor-free mice protected na?ve animals from a subsequent challenge with CT26, and this was mediated mainly by CD8 T cells. Most importantly, PDT plus IT-DC administered to one tumor site led to tumor regression at distant sites, including multiple lung metastases. CONCLUSIONS: PDT + IT-DC induces potent systemic antitumor immunity in mice and should be evaluated in the treatment of human cancer.     

Light delivery over extended time periods enhances the effectiveness of photodynamic therapy.

PURPOSE: The rate of energy delivery is a principal factor determining the biological consequences of photodynamic therapy (PDT). In contrast to conventional high-irradiance treatments, recent preclinical and clinical studies have focused on low-irradiance schemes. The objective of this study was to investigate the relationship between irradiance, photosensitizer dose, and PDT dose with regard to treatment outcome and tumor oxygenation in a rat tumor model. EXPERIMENTAL DESIGN: Using the photosensitizer HPPH (2-[1-hexyloxyethyl]-2-devinyl pyropheophorbide), a wide range of PDT doses that included clinically relevant photosensitizer concentrations was evaluated. Magnetic resonance imaging and oxygen tension measurements were done along with the Evans blue exclusion assay to assess vascular response, oxygenation status, and tumor necrosis. RESULTS: In contrast to high-incident laser power (150 mW), low-power regimens (7 mW) yielded effective tumor destruction. This was largely independent of PDT dose (drug-light product), with up to 30-fold differences in photosensitizer dose and 15-fold differences in drug-light product. For all drug-light products, the duration of light treatment positively influenced tumor response. Regimens using treatment times of 120 to 240 min showed marked reduction in signal intensity in T2-weighted magnetic resonance images at both low (0.1 mg/kg) and high (3 mg/kg) drug doses compared with short-duration (6-11 min) regimens. Significantly greater reductions in pO(2) were observed with extended exposures, which persisted after completion of treatment. CONCLUSIONS: These results confirm the benefit of prolonged light exposure, identify vascular response as a major contributor, and suggest that duration of light treatment (time) may be an important new treatment variable.     

Effects of photodynamic therapy with hypericin in mice bearing highly invasive solid tumors.

The tumoricidal properties of photodynamic therapy (PDT) with hypericin (HY) were evaluated in a highly metastatic adenocarcinoma (DA3Hi) and anaplastic squamous cell carcinoma (SQ2) tumors in vivo. Photosensitization of the tumor site with hypericin (HY-PDT) reduced primary tumor development and significantly prolonged the survival of tumor-bearing (TB) mice. Of these two tumors the squamous cell carcinoma emerged as more sensitive to HY-PDT compared with DA3Hi adenocarcinoma both in vitro and in vivo. HY-PDT caused extensive tumor necrosis that was followed by local, intratumoral, and systemic inflammatory reactions. Analyses of cytokine mRNA profiles reveal increases in mRNA levels of expression confined to inflammation-related cytokines both within the tumor and also systemically (measured in spleens). However, there was no evidence for any HY-PDT-induced antitumoral immune reactions. Our results suggest that PDT with hypericin can be considered as a supplementary treatment in the management of some invasive and metastatic cancers such as squamous carcinoma and similar tumors.     

Erythropoietin for the treatment of anemia associated with hematological malignancy

Anemia is common in patients with hematological malignancy. Most patients will have their anemia attributed to the anemia of chronic disease. The anemia of chronic disease is caused by cytokine mediated suppression of erythropoiesis and low serum erythropoietin levels are found in the majority of patients with cancer. Many of these anemic patients will be symptomatic with fatigue. Data from many studies indicates that treatment of anemic patients with erythropoietin will increase their hemoglobin concentration, decrease transfusion need and also improve their quality of life. A recent study also suggests that improving the hemoglobin level may improve the patients' prognosis but this finding needs to be confirmed.     

Erythropoietin restores the anemia-induced reduction in cyclophosphamide cytotoxicity in rat tumors

The aim of this study was to examine the impact of anemia prevention by recombinant human erythropoietin (rHuEPO) treatment on the cytotoxicity of cyclophosphamide in solid experimental tumors. Anemia was induced using a single dose of carboplatin (50 mg/kg i.v.) resulting in a long-lasting reduction (30%) of the hemoglobin concentration. In a second group, the development of anemia was prevented by rHuEPO (1000 IU/kg) administered s.c. three times/week starting 7 days before carboplatin application. Four days after carboplatin treatment, tumors (DS-sarcoma of the rat) were implanted s.c. onto the hind food dorsum. Neither carboplatin nor rHuEPO treatment influenced tumor growth rate per se. When tumors were treated with a single dose of cyclophosphamide (60 mg/kg i.p.) 5 days after implantation, a growth delay with a subsequent regrowth of the tumors was observed. In the anemia group, the growth delay was significantly shorter compared with nonanemic controls (13.3 days versus 8.6 days). In the group where anemia was prevented by rHuEPO treatment, growth delay was comparable with that of nonanemic controls (13.3 days). These results suggest that chemotherapy-induced anemia reduces cytotoxicity of cyclophosphamide in tumors, whereas correction of anemia by rHuEPO treatment (epoetin alpha) increases the sensitivity, probably as a result of an improved oxygen supply to tumor tissue.     

Erythropoietin treatment in patients with myelodysplastic syndrome and anemia

Hematological disorders are commonly complicated by anemia, and the symptoms of red cell deficiency adversely affect the quality of life. Erythropoietin is a glycoprotein which controls red blood cell production. Recombinant human erythropoietin, 50 U/kg/day, was given subcutaneously to 16 patients with myelodysplastic syndrome and anemia. All but one patient was transfusion dependent. Diverse pretreatment endogenous serum erythropoietin levels were noted and ranged from 17 to 3616 IU/l. Two patients (12.5%) demonstrated an improvement in hemoglobin levels obviating the need for transfusions. Their responses lasted 5+ and 7 months with maintenance erythropoietin treatment. The responders had endogenous serum erythropoietin levels of 44 and 170, respectively. Treatment was generally tolerated without constitutional side-effects. However, three patients developed thrombocytopenia and one developed joint pain and leukocytosis on treatment. Overall, six patients showed changes in non-erythroid cells: two patients had an increase in platelet counts; three patients, a decrease in platelet counts; and one patient, an increase in white blood cell counts. Most of these changes reversed rapidly once erythropoietin was stopped. It is concluded that (a) serum erythropoietin levels are extremely variable in anemia patients with myelodysplastic syndrome, (b) only a minority of patients benefit from treatment with recombinant human erythropoietin, and (c) erythropoietin can affect cells of the myeloid and megakaryocytic lineage in a small proportion of patients.     

Erythropoietin treatment of tumor-associated anemia in patients with multiple myeloma

Anemia of malignancy is a complication of neoplastic disease which causes impairing symptoms and often requires blood transfusions. In this clinical trial, we have treated 13 patients suffering from chronic anemia of malignancy and multiple myeloma with recombinant human erythropoietin (rHuEPO) three times a week. Eleven patients responded to the treatment by appropriate increases of their hemoglobin levels and the eventual correction of the anemic state, one non-responding patient had to terminate the treatment early because of transfusion requirements. Under rHuEPO therapy, the evaluated parameters of iron metabolism indicated the enhanced synthesis of hemoglobin. The symptoms of anemia subsided in the responding patients and most of them reported a hightened subjective sense of well-being. No adverse side effects, particularly no episodes of hypertension, were observed in any patient.     

Anti-tumour activity of photodynamic therapy in combination with mitomycin C in nude mice with human colon adenocarcinoma.

The interaction of photodynamic therapy (PDT) and a chemotherapeutic drug, mitomycin C (MMC), was investigated using WiDr human colon adenocarcinoma tumours implanted on Balb/c athymic nude mice. The WiDr tumours were treated with PDT alone, MMC alone or with both. It was found that the combined treatment produced a greater retardation in the growth of the WiDr tumour than monotherapy with MMC or PDT. The synergistic effect was especially prominent when PDT was used in combination with a low dose of MMC (1 mg kg-1), since treatment of 1 mg kg-1 MMC alone had no effect on the tumour. The anti-tumour activity of PDT was found to be increased with MMC of 5 mg kg-1. The response of normal skin on mice feet to PDT slightly greater when PDT was combined with 5 mg kg-1 MMC than when PDT was applied alone, while no detectable additional effect on skin photosensitivity was observed when PDT was combined with 1 mg kg-1 MMC. An enhanced uptake of Photofrin in tumours was found 12 h and 24 h after administration of MMC. The effect of MMC on the cell cycle distribution of cell dissociated directly from the tumours was studied. The results suggest that the increased susceptibility to photoinactivation of Photofrin-sensitised tumours may be due to MMC-induced accumulation of the tumour cells in S-phase.     

Erythropoietin ameliorates chemotherapy-induced fibrosis of the lungs in a preclinical murine model

Organ toxicity induced by chemotherapeutic drugs is a serious obstacle in the effective treatment of patients suffering from cancer and autoimmune disease. A strong association exists between pulmonary toxicity, particularly fibrosis, and chemotherapeutic drugs. Attempts have been made to identify compounds capable of suppressing fibrosis. In addition to its erythropoietic activity, erythropoietin (EPO) has been shown to have effects on nonhemopoietic cells. Therefore, we postulated that EPO may exert beneficial effects on lung tissue during chemotherapy. To test our hypothesis, we investigated pulmonary changes caused by bleomycin, a fibrosis-inducing agent, in animals treated with the drug alone and in combination with EPO. Fibrosis, cellular alterations and structural changes were assayed by blind analysis of the lung sections. A 6-fold decrease in the number of prominent endothelial cells--suspected to be indicative of cellular activation and inflammatory response--was observed in lung sections derived from mice treated with bleomycin and EPO compared to animals injected with bleomycin alone (p < 0.008). Additionally, there was twice the number of ICAM1-positive endothelial cells in animals treated with bleomycin alone compared with the number in the bleomycin and EPO-treated group (p < 0.05). Alveolar mononuclear phagocytic hyperplasia was reduced by as much as 100% in animals treated with bleomycin and EPO compared to animals treated with bleomycin alone (p < 0.03). Finally, a 5-fold decrease in interstitial fibrosis was observed in lung sections obtained from animals treated with bleomycin and EPO (p < 0.02). We conclude that EPO can ameliorate drug-induced fibrosis and endothelial damage caused by chemotherapeutic agents.     

Treatment of chemotherapy-induced anemia with recombinant human erythropoietin in cancer patients

Thirty patients with chemotherapy-induced anemia were treated with recombinant human erythropoietin for 4 weeks. In this dose-escalation study, cohorts of five to eight patients were treated per dose level. The doses of erythropoietin were 25, 50, 100, 200, or 300 IU/kg/d given intravenously for 5 days each week. Of 30 patients, 15 (50%) had a greater than 10% increase of their hemoglobin (Hb) values and were considered responders. At the two highest dose levels, 11 of 13 patients (85%) responded. In the 15 responding patients, the mean Hb level increased by 1.7 g/dL from baseline compared with an average decrease of 1.5 g/dL in the previous cycles of chemotherapy without erythropoietin administration. Recombinant human erythropoietin is effective in ameliorating chemotherapy-induced anemia when administered in adequate doses.     

Sequential therapy with lithium in chemotherapy-induced vomiting

Having noticed psychotic traits in some patients showing incoercible vomiting due to antineoplastic drugs we have thought of establishing a therapy with lithium in the days preceding the therapeutic cycle in order to reduce the emetic events. The effectiveness of lithium carbonate (600 mg/mq p.o./day for one week) in the prevention or reduction of vomiting induced by antiblastic therapy has been checked in comparison with metoclopramide and domperidone in 40 patients. In the group pretreated with lithium, 80% of the cases showed favorable results. In the control groups, on the contrary, the efficacy of the antiemetic therapy has shown to be of lesser importance (55%). The undesirable side-effects of lithium appear to be irrelevant. We therefore think that pretreatment with lithium may become, in selected cases not affected by traditional antiemetics, of great importance in the control of emetic symptomatology.     

Systemic toxicity in mice induced by localized porphyrin photodynamic therapy

An unexpected high level of acute lethality has been documented following Photofrin II-mediated photodynamic therapy (PDT) treatments which were localized to the hind leg of normal and tumor-bearing mice. Doses of PDT which induced lethality (10 mg/kg Photofrin II, 200-500 J/cm2) were in the range of doses required to obtain murine tumor cures. The percentage of lethality was proportional to the total light dose but was inversely proportional to the dose rate of delivered light. Comparable levels of acute toxicity were observed in four pigmented mouse strains (C57BL/6J, C3H/HeJ, DBA/1, and DBA/2) and in two albino mouse strains (BALB/c and Swiss Webster). Decreased sensitivity to PDT-induced lethality was observed in two pigmented mouse strains (B10D2/OSN and B10D2/NSN). The administration of warfarin, aspirin, indomethacin, or antihistamine had significant protective effects in terms of decreasing PDT-induced lethality. However, injection of cobra venom factor (to deplete C3 and C5 of the complement system) did not alter the lethality mediated by PDT. Histological profiles obtained 24 h following PDT demonstrated vascular congestion in the liver, kidney, lung, and spleen. Significant decreases in removable blood volume, core temperature, and spleen weight were also observed within 24 h of localized PDT treatment. These results indicate that PDT-induced lethality is consistent with a traumatic shock syndrome and suggest that endogenous vasoactive mediators of shock such as prostaglandins, thromboxanes, and histamine are associated with the lethality induced by localized PDT in mice.     

The effectiveness of darbepoetin alfa administered every 3 weeks on hematologic outcomes and quality of life in older patients with chemotherapy-induced anemia

Chemotherapy-induced anemia (CIA) may substantially impact the health-related quality of life (HRQoL) of older cancer patients. This exploratory analysis evaluated the effect of darbepoetin alfa administered as a fixed dose (300 microg) every 3 weeks (Q3W) on hematologic outcomes, HRQoL, and safety in older (> or =65 years old) versus younger (<65 years old) patients with CIA (hemoglobin <11 g/dl). Patients were categorized by age at screening: <65, > or =65 to <70, > or =70 to <75, > or =75 to <80, and > or =80 years old. Patients who received at least one dose of darbepoetin alfa were included in the analysis; of 1,493 patients, 724 were > or =65 years old. Age did not appear to influence hematologic outcomes after treatment with darbepoetin alfa; in all age categories, similar percentages of patients (78%-80%) achieved the target hemoglobin in approximately the same time (4-5 weeks). Also, the percentage of patients in each age category who received RBC transfusions was reduced from 10%-13% in month 1 to 2%-4% in month 4. Although younger patients reported the greatest improvement in HRQoL scores, approximately one half in each older age category reported clinically significant improvement in fatigue, and improvement in the Energy and Overall Health Assessment and Work Productivity and Activity Impairment scales. There were no treatment-related deaths. Treatment-related thromboembolic events were reported by <1% of patients <65 years old and <1% of patients > or =65 to <70 and > or =70 to <75 years old. Darbepoetin alfa Q3W appeared well tolerated and effective for treating older patients with CIA.     

Effects of photodynamic therapy using mono-L-aspartyl chlorin e6 on vessels and its contribution to the antitumor effect.

The effect of photodynamic therapy (PDT) on the vascular system has a significant role in tumor tissue destruction. We investigated the contribution of vascular damage to the antitumor effects of PDT and analyzed the quantitative vascular changes after PDT. Fibrosarcoma-bearing BALB / c male mice were injected with mono-L-aspartyl chlorin e6 (NPe6) at a dose of 0.25, 5 or 15 mg / kg, and photoradiation was performed with a diode laser 10 min, 2 h or 24 h after injection, respectively. Ten minutes after injection of 0. 25 mg / kg, NPe6 was found to be present only in plasma, while at 2 h after injection of 5 mg / kg it was present in both plasma and tumor, and 24 h after injection of 15 mg / kg it was present only in the tumor. The antitumor effects observed in the 5 mg / kg-2 h and 0. 25 mg / kg-10 min groups were virtually the same, whereas the effect in the 15 mg / kg-24 h group was weaker. The damage to the tumor vasculature and tumor cells in the 15 mg / kg-24 h group occurred later than under the other conditions, and vascular damage in the tumor-surrounding tissue was also less marked even 24 h after PDT. These results suggested that the plasma NPe6 concentration during laser irradiation contributed more than the tumor NPe6 concentration to the antitumor effect, and that the minimal damage to blood vessels around the tumor at the low plasma NPe6 concentration may be one reason for the failure to obtain a marked antitumor effect.     

Erythropoietin is effective in improving the anemia induced by imatinib mesylate therapy in patients with chronic myeloid leukemia in chronic phase

BACKGROUND: Myelosuppression occurs in up to 50% of patients with chronic myeloid leukemia (CML) who are treated with imatinib and > or = Grade 3 myelosuppression is reported in approximately 10% of patients. METHODS: The authors investigated the prognostic significance of anemia occurring during therapy with imatinib in patients with CML in chronic phase. RESULTS: Of 338 patients treated with imatinib (150 patients after interferon failure and 188 patients with newly diagnosed CML), 230 (68%) developed anemia. In a multivariate analysis, factors associated with an increased probability of developing anemia were a starting hemoglobin level < 12 g/dL, age > or = 60 years, female gender, higher imatinib dose, and intermediate or high Sokal risk group. Of these 230 patients, 102 patients received treatment with 40,000 U of recombinant human erythropoietin administered subcutaneously once weekly. An increase in the hemoglobin level of > or = 2 g/dL was achieved in 69 patients (68%) and 22 patients (22%) had an increase of 1-1.9 g/dL. Patients who developed anemia had a trend toward a lower probability of complete cytogenetic remission compared with patients without anemia (68% vs. 77%; P = 0.14), as well as a trend for inferior survival. Patients with anemia and other manifestations of myelosuppression were found to have a significantly worse outcome than those with isolated anemia. CONCLUSIONS: The authors concluded that erythropoietin is safe and effective in patients in chronic-phase CML who develop anemia with imatinib therapy.