Influence of time interval between surgery and radiotherapy on tumor regrowth.

UNLABELLED: In order to evaluate the influence of time intervals between tumor cell injection and radiotherapy on tumor control and regrowth after surgery, we performed two kinds of experiments on C3D2F1 mice bearing a mammary carcinoma inoculated in the foot or leg. 1st experiment: tumor in foot. END POINT: Tumor Control Probability (TCP). Single dose radiation treatments (RT) were administered at different period times from injection time of tumor cells (day 1). 1st group: unirradiated control, 2nd group: RT on day 2 (TCP50 29 +/- 2.1 Gy), 3rd group: RT on day 7 (TCP 52.5 +/- 2.9 Gy), 4th group: RT on day 12 (TCP50 61.9 + 2.4 Gy). 2nd experiment: tumor in leg. END POINT: percentage of tumor regrowth. Mice were randomly assigned to three groups: 1st control group (tumor growth in all mice), 2nd surgical excision of macroscopically evident tumor on day 7-9 from injection (tumor regrowth in 85% of mice), 3rd as the previous group plus 30 Gy radiation treatment within 24 hours from excision (tumor regrowth in 33% of cases). The radiation dose was selected on the basis of TCP50 observed in the 1st experiment for mice with sub-clinical disease. These data indicate that the radiation dose able to control 50% of tumors increases with the time interval between tumor cells injection and RT. A short time interval between surgery and RT should increase the probability of local control, supporting the rationale of intraoperative radiation therapy (IORT) as adjuvant therapy after surgical resection, when subclinical residual cells are suspected.

Schedule dependent toxicity and efficacy of combined gemcitabine/paclitaxel treatment in mouse adenocarcinoma.

Increased interest in combining drugs with different targets has emerged over recent years. Our study aims at evaluating the effectiveness of combined gemcitabine/paclitaxel treatment taking into consideration doses, schedules, and toxicity. A spontaneous mammary carcinoma was transplanted into the right-hind foot of C3D2F1 mice. Paclitaxel (in doses from 20 to 80 mg/kg b.w.) and gemcitabine (in doses from 30 to 480 mg/kg b.w.) were administered i.p. in single or fractionated treatments. Toxicity and tumor growth delay (TGD) were the endpoints. TGDs for different gemcitabine doses in single administration (120, 240, and 360 mg/kg) overlapped (TGD approximately = 2.5 days). Toxicity was very high in daily administration. Results with gemcitabine alone showed the efficacy of treatments every 3 days. TGDs in fractionated treatments of 60 and 120 mg/kg x 4 were of approximately equals 16 days. Also in this case, tumor growth curves overlapped pointing out the uselessness of the high drug doses. For combined treatments, we used only fractionated protocols, administering gemcitabine every 3 days. Paclitaxel was administered alone in one or two fractions and with different sequences in respect to gemcitabine administration. With 120 mg/kg of gemcitabine all the protocols showed an increased unacceptable toxicity. The best result was obtained administering paclitaxel 40 mg/kg on days 1 and 15 and gemcitabine 60 mg/kg on days 3, 6, 9, and 12 (TGD = 38.2 days). The light toxicity and the high efficacy obtained with this protocol indicate the possible use of gemcitabine/paclitaxel treatment in clinical practice.

Hyperthermia enhances the response of paclitaxel and radiation in a mouse adenocarcinoma.

PURPOSE: The aim of our study was to investigate if the efficacy of paclitaxel and paclitaxel-radiation treatments in vivo could be enhanced by hyperthermia. MATERIALS AND METHODS: Paclitaxel was administered i.p. in doses from 30 to 60 mg/kg b.w. to (C3D2F1) mice bearing spontaneous mammary carcinoma. Local hyperthermia (41 degrees, 42 degrees, 43 degrees C) was carried out by immersing tumor-bearing legs in a water bath for 1 h. Single X-ray treatments from 10 to 90 Gy were performed. Tumor growth delay (TGD) or tumor control dose (TCD(50), radiation dose needed to induce local tumor control in 50% of irradiated animals) were the endpoints. RESULTS: A significant increase of dose-dependent growth delay was observed in paclitaxel and 43 degrees C hyperthermia combined treatments, and a superadditive effect was seen with paclitaxel 45 mg/kg. Combined treatments with hyperthermia at 41 degrees and 42 degrees C were less effective. Administration of paclitaxel 24 h, 4 h, and 15 min before or 15 min and 4 h after hyperthermic treatments produced similar results (TGDs varying from 22.1 to 17 days), and administering paclitaxel 48 h before or 24 h after hyperthermic treatments decreased TGDs (about 10 days). Trimodality treatment (paclitaxel 45 mg/kg, hyperthermia, and X-ray), with a TCD(50) of 14. 1 Gy, in respect to the TCD(50) of 53.1 obtained with X-ray alone, was the most effective. CONCLUSIONS: Hyperthermia enhanced the effectiveness of paclitaxel in all the tested protocols. Our results show a superadditive effect of paclitaxel 45 mg/kg combined with a hyperthermic treatment of 1 h at 43 degrees C. Trimodality treatment, evaluated in terms of percentage of cures, shows a very high enhancement ratio.

Combined hyperfractionated radiotherapy and protracted infusion chemotherapy in bladder cancer for organ preservation.

PURPOSE: To test an innovative schedule of concurrent protracted intravenous infusion (PVI) of cisplatin (CDDP) and 5-fluorouracil (5-FU) and hyperfractionated radiotherapy (HFRT) with organ-sparing intent in bladder cancer. PATIENTS AND METHODS: Fifty-two patients (pts) were selected to receive an aggressive TURB followed by 2 MCV cycles, and HFRT with concomitant CDDP and 5-FU PVI (33 pts) or HFRT and concomitant CDDP and 5-FU PVI (20 pts). The 5-FU and CDDP doses ranged from 180 to 220 mg/sm/day and from 4 to 6 mg/sm/day, respectively. Radiotherapy was delivered as three 100 cGy fraction per day or two 150 cGy fraction per day to a total dose of 50 Gy to the pelvis and a 20 Gy boost to the bladder. RESULTS: Grade III toxicity in pts who received or not MCV was: rectal tenesmus 12/33 and 0/20, dysuria 6/33 and 4/20, leukopenia 3/33 and 0/20, thrombocytopenia 7/33 and 1/20 pts, respectively. A Grade IV toxicity was observed in 2 pts. Of the 28 evaluable patients treated with MCV, CR were observed in 23 (82%) and PR in 5 cases. Of the 18 evaluable patients treated without MCV, CR were observed in 18 cases (100%). Actually, 65% and 14% of the CR pts treated with or without HCV are alive and free of tumor. CONCLUSIONS: This bladder-sparing treatment shows an acceptable acute and late toxicity, similar to that observed with radiotherapy alone. The high CRs and bladder preservation rates observed deserve further clinical evaluation.

Greater antitumor efficacy of paclitaxel administered before epirubicin in a mouse mammary carcinoma.

Combined treatment with paclitaxel and anthracyclines is increasingly being tested in clinical practice. Epirubicin is in general administered before paclitaxel. We have investigated, using a murine mammary adenocarcinoma, whether the efficacy and toxicity of this combination is influenced by treatment sequence, different time intervals and dose intensity. The tumor was transplanted into the right hind foot of C3D2F1 female mice. Paclitaxel was administered i.p. in doses ranging from 15 mg/kg to 75 mg/kg and epirubicin (i.p. or i.v.) in doses from 9 mg/kg to 30 mg/kg. The hepatic and peritoneal toxicity observed with epirubicin administration increased in combined treatments (stronger with i.p. than i.v. epirubicin administrations) and was dose-dependent. When paclitaxel and epirubicin were administered simultaneously or paclitaxel was given 24 h before epirubicin, the same tumor growth delays were obtained in all groups. A smaller effect was observed when paclitaxel was administered 24 h after epirubicin. Increasing the epirubicin or paclitaxel dose led to higher tumor growth delays but also an increased toxicity. In conclusion, in this experimental model, the administration of 45 mg/kg paclitaxel before 15 mg/kg epirubicin was very effective and the increased toxicity can be limited by introducing an interval of 24 h between drug administrations. These results should be considered when designing clinical trials.

DNA/protein flow cytometry as a predictive marker of malignancy in dysplasia-free Barrett's esophagus: thirteen-year follow-up study on a cohort of patients.

Intestinal metaplasia identifies Barrett's esophagus (BE) and is associated with an increased risk for esophageal adenocarcinoma. Dysplasia occurs as an intermediate step. However, progression from metaplasia to neoplasia without the demonstration of dysplasia has been described. The role of dual-parameter flow cytometry (FC) as a predictor of neoplastic risk in dysplasia-free cases was evaluated. DNA/protein FC and histology were performed on 362 samples from 30 dysplasia-free BE patients, followed up since 1985 once every 1-2 years. Nine cases were aneuploid, five of which (group IV) were frankly aneuploid; in the other four cases (group III), aneuploidy was detectable by dual-parameter analysis only. Twenty-one patients were diploid. Twelve (group II) also had an abnormally high G1-phase protein content compared to group I (nine patients), which were diploid with a low-moderate protein content. In three patients of group IV an adenocarcinoma in situ was diagnosed, after 5, 6, and 10 years, respectively. In two patients of group III, a low- and a high-grade dysplasia were observed at 3 and 6 years follow-up, respectively. One patient of group I first acquired a high protein content, then an aneuploid DNA content, and then progressed to adenocarcinoma (12 years). None of the still diploid patients (17 cases) have progressed to dysplasia or cancer compared with 6 of 13 presently aneuploid patients (P < 0.01). In conclusion, DNA/protein FC is a marker of increased malignant potential and thus may be used to detect patients at higher risk in dysplasia-free BE and assist in understanding the various stages of malignant transformation in long-term follow-up studies.

Quality control study of the Italian Group of Cytometry on flow cytometry DNA content measurements: II. Factors affecting inter- and intralaboratory variability.

A multicentric national quality control study has been organized under the auspices of the Italian Group of Cytometry to find a possible influence of some procedural steps in DNA flow cytometry measurements on DNA index (DI) values and to identify the main parameters affecting the interlaboratory variability. To 40 participating laboratories we provided suspensions containing unknown mixture of different cell types: an homogeneous thymocyte population used to check instrument linearity; one mixture composed of two cell types characterized by DI = 1.00 and 1.10; and another composed of three different cell types with relative DIs of 1.00, 1.26, and 1.62, respectively. Possible effects due to staining protocols were studied, allowing the participants to stain cellular DNA according to the procedure routinely adopted in each laboratory, in addition to a standardized procedure with a fixed PI solution. As far as the influence of instrument linearity on DI values is concerned, we did not find any correlation with the DI variability observed, even if the use of a standardized staining protocol could lead to a sensible gain in interlaboratory DI reproducibility. Twenty-five of 40 (65%) laboratories were able to discriminate the near-diploid subpopulation, and a coefficient of variation of less than 4% was the minimum condition necessary to recognize the DI = 1.1 population. In samples containing two aneuploid subpopulations, 25 of 35 (71.4%) laboratories showed a high reproducibility with the standard staining protocol and 22 of 38 (57.9%) with the free staining protocol. However, a sensible improvement in interlaboratory reproducibility emerged with respect to the previous trial.

[Role of computerized tomography in bladder carcinoma treated with simultaneous combined radiotherapy and chemotherapy]. / Ruolo della Tomografia Computerizzata nel carcinoma vescicale sottoposto a radio e chemioterapia contemporanee associate.

We report on the pneumocystography-CT follow-up of locally aggressive bladder carcinoma patients treated with combined and simultaneous irradiation and chemotherapy. Particular attention was paid to assess residual thickening of the wall involved by the tumor, which is the most critical parameter to discriminate a persistent from a resolved condition. We examined 16 patients (mean age: 66.5 years) with histologically locally advanced urinary bladder cancer staged T2-T3. All patients underwent the first CT exam of the bladder with the pneumocystography technique before therapy, for disease staging, another exam after chemotherapy and a third one after simultaneous irradiation and chemotherapy. The endoluminal mass completely regressed in all cases, with normal endoscopic findings. In 9 of 16 cases, the bladder wall was thickened in the tumor site, with normal mucosal surface. The histology of bioptic specimens obtained during cystoscopy showed a high frequency of phlogistic reactions from foreign bodies, corresponding to the bladder wall thickening seen on CT images. We conclude that, in extensive neoplastic bladder wall involvement, therapy-induced histologic regressive phenomena can cause a granulomatous reaction in the bladder wall, thus thickening it, which is the most frequent sign in these cases but that, in our experience, was never associated with tumor invasion. This should be always considered in the CT studies performed after this kind of combined treatment.

[Continuous sinusoid systemic Fudr infusion in advanced colorectal carcinoma. Preliminary experience]. / Infusione sistemica continua sinusoidale di Fudr nel carcinoma colorettale avanzato. Esperienza preliminare.

Sinusoidal circadian continuous infusion with a maximal flow rate in the afternoon (3-9 pm) reduces Fudr toxicity. In order to estimate if the reported lower toxicity is merely due to the quasi-intermittence of the daily dose or to the circadian rhythm of infusion. Ten patients with widespread cancer (9 colorectal and 1 renal) underwent sinusoidal continuous iv Fudr infusion with the peak level in antiphase (ie 68% of the dose from 3 to 9 am) as compared with the Römeling shape. An initial dose of 0.15 mg/kg/d for 14 days monthly has been given, escalating it every cycle by 0.025 mg/kg/d increments until toxicity. Mean (+/- SD) number of cycles has been 4.1 +/- 2.1 (range 2-8), maximal dose given has been 0.2 mg/kg/die in 5 patient and mean dose intensity of 0.570 +/- 0.04. Gastrointestinal toxicity consisted of nausea/vomiting WHO grade 1 in one patient and diarrhoea grade 1 in two, grade 2 and 3 in one and one case. Toxicity and dose intensity of both sinusoidal infusion seem to be similar and allow higher dose of Fudr than continuous constant infusion. Some other studies have to be done to include pharmacokinetics evaluation in order to estimate chronobiologic implication in continuous Fudr infusion.

Prognostic significance of flow cytometry in lung cancer. A 5-year study.

Flow cytometrically determined cellular DNA content has been measured on specimens from 101 patients affected by lung cancer (40 epidermoid cell carcinoma, 22 adenocarcinoma, 21 large cell carcinoma, 11 small cell carcinoma, and seven undifferentiated carcinoma), and one by mesothelioma. Ninety-eight of 102 (96%) patients with neoplastic disease evidenced the occurrence of at least one cytometrically aneuploid cell subpopulation. Fifty-five of 102 (54%) cases evidenced the occurrence of multiclonality, that is, the presence of more than one aneuploid stem cell line. However, the incidence of multiclonality in lung carcinoma was statistically different in surgical cases (where multiple site sampling from the primary and lymph nodes was possible) in comparison to the nonsurgical ones (e.g., bronchial washing): 48/77 (62%) and six of 24 (25%), respectively. Therefore, only the 77 surgical patients were used for further analysis. The cases were classified according to the DNA index (DI) in the following way: Group A (tumors with one or more stem lines with DI ranging from 1 to 2) and Group B (tumors with at least one stem line with DI less than 1 or greater than 2). A significant correlation has been found between the cytometric ploidy condition so defined (Groups A and B) and the tumor mass doubling time (DT), Group B being associated with fast growing tumors (DT lower than 90 days). A statistically better 12-month survival rate (5-year maximum follow-up) was observed in Group A (88%) in respect to Group B (47%) and is evident in the patient survival time course. A better prognostic indication can be achieved by stratifying the patients according to both the cytometric ploidy condition and the tumor DT. Flow cytometric data can usefully contribute to the prognostic assessment of lung carcinoma provided that representative cellular material is collected by multiple site sampling.

Role of delayed cutaneous hypersensitivity reaction in classifying patients with bronchial carcinoma.

Cell-mediated immune response was evaluated in 150 patients with histologically confirmed bronchopulmonary carcinoma using bacterial and fungal recall antigens injected intradermally (PPD, candida, trichophyton). In the study group negative skin test reaction was found in 51 of 150 patients (34.0%), whereas in the control population it was found in 5 of 33 cases (15.1%) (p less than 0.05). Histologic cell type and stage of disease were defined for each patient. It was possible to calculate the growth rate of the primary tumor only in 68 of 150 patients, and it was recorded as doubling time. Evaluation of the skin test reaction in each prognostic subgroup showed no statistically significant differences. The only statistically significant differences were found when each prognostic subgroup was compared with the control population according to the frequency of a negative response to the skin test, particularly in stage III M1 (p less than 0.05) and stage III M0 (p less than 0.02). The delayed cutaneous hypersensitivity studied with recall antigen stimulation was mainly correlated with the stage of disease, and it should not be considered as an independent prognostic factor.

A new multiparametric classification in lung cancer patients - S.M.I.G.

The classification of bronchogenic carcinoma as a function of the prognosis is still an open field. The evaluation of stage, by use of the TNM system, and histologic cell type is not sufficient to guarantee a correct prognosis. The growth rate of the neoplasm is another important parameter. We propose a classification that takes into account the stage (S), histologic cell type (M), immune status (I) and the growth rate of the primary tumor (G): S.M.I.G. We studied 90 lung cancer patients according to the S.M.I.G. classification and we observed that their prognoses were directly correlated with their S.M.I.G. scores (the higher the score, the higher the 10-month mortality rate). The mortality rates within the first 10 months of follow-up were respectively 0%, 0%, 36.36%, 68%, 90.9% for the 5 groups obtained by S.M.I.G. The difference is statistically significant (P less than 0.0075) and there is a linear correlation between the mortality rate and the score assigned to each group (R = 0.943; P less than 0.05). The S.M.I.G. classification can predict the prognosis more efficiently than the usual classification (TNM) and histological cell type.

Non-small-cell lung carcinoma: tumor characterization on the basis of flow cytometrically determined cellular heterogeneity.

Some 150 tumor specimens from 49 patients with non-small-cell carcinoma of the lung (23 epidermoid, 14 adenocarcinoma, 12 large-cell carcinoma) and three with nonneoplastic lung disease were analysed for cellular DNA content by flow cytometry. Monodispersed cells were stained with ethidium bromide and mithramycin. Normal specimens and samples from patients with nonneoplastic disease constantly yielded a single cell population with diploid DNA content. Twenty of 23 epidermoid carcinomas exhibited one or more than one aneuploid subpopulation. Ten of 12 large-cell carcinomas were characterized by one aneuploid clone and 2/12 by two aneuploid clones. Adenocarcinoma exhibited multiclonal cell subpopulations (one to five aneuploid clones). Further information has been obtained on the differential presence of clones in various tumor areas and in infiltrated lymph nodes. These tumors appear characterized by a remarkable degree of cellular heterogeneity. The cytometric ploidy level(s) and the cell population multiclonal structure yield, in comparison with, and in addition to, pathology, indications of possible clinical interest. A correlation between the clonal DNA content and a prognostic parameter such as the tumor mass doubling time has been demonstrated.

TNM staging in lung cancer: role of computed tomography.

Current procedures to determine the clinical staging of disease in patients with lung cancer are lacking in accuracy, particularly regarding the presence of metastatic disease. We have evaluated the use of computed tomography (CT) of the chest, brain, and upper abdomen for clinical staging of the extent of disease in 113 consecutive patients with histologically confirmed carcinoma of the lung. Comparisons with mediastinoscopy and surgical findings were made regarding the extent of primary tumor in 47 patients and nodal involvement in 41 patients. The CT scan showed a sensitivity of 86.9%, a specificity of 91.6%, and an accuracy of 89.3% for extrapulmonary extension of the primary tumor and a sensitivity of 50%, a specificity of 96.5% and an accuracy of 82.9% for mediastinal node involvement. Thirty-two of the 85 patients studied by total body CT scan had distant metastasis, of which 24 (75%) were clinically silent. Thus 28.2% of the 85 patients studied had asymptomatic metastatic disease. We conclude that CT of the chest, brain, and upper abdomen is a reliable procedure for staging lung cancer.

Cytometric evidence of cytogenetic and proliferative heterogeneity of human solid tumors.

Flow cytometry is a widely recognized method of rapidly assessing the ploidy and proliferation status of experimental and solid tumors. In the present work, a variety of human cancers from various sites (lung, head and neck, etc.), of traditional interest in our laboratory, have been analyzed. In agreement with a number of recent reports, a general heterogeneity of human solid tumors can be evidenced. In particular: (a) solid tumors are characterized by a variable degree of aneuploidy; (b) the internal structure of solid tumors is highly heterogeneous especially with respect to the fraction of aneuploid malignant cells and their distribution through the cycle phases; and (c) some solid tumors are also characterized by the presence, to a variable extent, in the tumor of mass of multiple cell clones. Static fluorimetry of Feulgen-stained (mitotic) single cells offers a way to confirm this kind of observations.

[Elective radiotherapy in breast carcinoma]. / Trattamento radiante elettivo del carcinoma mammario.

From March 1972 to November 1979 a total of 157 patients with stage I to III primary breast cancers have been irradiated after segmental resection (12 cases), tilectomy (89 cases) or biopsy (56 cases). Complete local control was achieved in all Stage I lesions, in 97% of Stage II lesions and in 68% of Stage III lesions. Non recurrence has been observed in patients previously operated by segmental resections, while local failures occurred in 6/89 and in 16/56 patients operated by tilectomy or biopsy, respectively. Of the 28 Stage I patients, 24 (86%) are alive, one with distant metastases. Four patients of this group are dead, 2 of intercurrent disease and 2 of breast cancer. Of the 61 (Stage II) patients, 38 (62%) are alive, 5 of these with distant metastases. Twenty-three patients are dead, 15 with active disease, and 8 suffered intercurrent death. Of the 68 Stage III patients, 21 (31%) are alive, 6 of these with distant metastases. Fourty-seven patients are dead, 43 of breast cancer and 4 of intercurrent disease. The high probability of initial subclinical deposits is evidenced by the fact that 49 of the 68 patients in this group developed distant metastases. The patients with T1 lesions appear to comprise the most favourable group with a relapse free survival at five years of 76%. The 5 years relapse free survival was 62% for T2 lesions and 25% for T3 and T4 lesions. N0 status does not confer the same favourable prognosis as T1 status. N+ status, however, resulted in a definitely negative prognostic factor. Cosmetic results after our treatment approach appear to be extremely good. A marked difference between the irradiated and controlateral breast occurred only in 10 of the 125 patients after a minimum of 2 years observation.